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Sample records for arg-gly-asp rgd tripeptide

  1. Arg-Gly-Asp (RGD) Modified Biomimetic Polymeric Materials

    Institute of Scientific and Technical Information of China (English)

    Xufeng NIU; Yuanliang WANG; Yanfeng LUO; Juan XIN; Yonggang LI

    2005-01-01

    The new generation of biomaterials focuses on the design of biomimetic polymeric materials that are capable of eliciting specific cellular responses and directing new tissue formation. Since Arg-Gly-Asp (RGD) sequences have been found to promote cell adhesion in 1984, numerous polymers have been functionalized with RGD peptides for tissue engineering applications. This review gave the advance in RGD modified biomimetic polymeric materials,focusing on the mechanism of RGD, the surface and bulk modification of polymer with RGD peptides and the evaluation in vitro and in vivo of the modified biomimetic materials.

  2. Molecular Interactions Between the Active Sites of RGD (Arg-Gly-Asp with its Receptor (Integrine

    Directory of Open Access Journals (Sweden)

    E. Jauregui

    2000-03-01

    Full Text Available A study of the molecular interactions between the active sites of RGD (Arg-Gly-Asp with it Receptor using simultaions is reported. Our calculations indicate that the guanidine-carboxylate complex is energetically favourd with respect to the guanidine-methyl tetrazole complex.

  3. The use of synthetic analogues of Arg-Gly-Asp (RGD) and soluble receptor of tumor necrosis factor to prevent acute and chronic experimental liver injury.

    Science.gov (United States)

    Bruck, R; Hershkoviz, R; Lider, O; Shirin, H; Aeed, H; Halpern, Z

    1997-01-01

    In chronic viral hepatitis, autoimmune hepatitis, and some chronic cholestatic liver diseases, T-lymphocytes serve as effector cells of the immunostimulatory processes. Cellular interactions of immune cells with extracellular matrix (ECM) components are regulated primarily via the beta 1 subfamily of integrin receptors. The target epitope of several such integrin receptors is the Arg-Gly-Asp (RGD) sequence, a cell adhesion motif shared by several matrix-associated adhesive glycoproteins. We review the use of synthetic nonpeptidic analogues of RGD and of soluble receptor of tumor necrosis factor (TNF)-alpha in the prevention of immune-mediated, concanavalin A-induced liver damage in mice and of RGD analogues in inhibiting the development of liver cirrhosis in rats. The concanavalin A-induced elevation of serum transaminases and TNF-alpha, and the infiltration of liver tissue by inflammatory cells, were inhibited by pretreatment of the mice with the synthetic RGD mimetics and soluble TNF receptor. In rats, the progression of thioacetamide-induced liver cirrhosis was markedly inhibited by the coadministration of the RGD mimetic SF-6,5. The compounds described here may be examined therapeutically for pathological conditions in the liver, manifested as necroinflammation, cholestasis and fibrosis. PMID:9626759

  4. Synthesis of Fluorine-18 Labeled Glucose-Lys-Arg-Gly-Asp-D-Phe as a Potential Tumor Imaging Agent

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Kyo Chul; Kim, Ji Sun; Sung, Hyun Ju; Jung, Jae Ho; An, Gwang Il; Chi, Dae Yoon [Korea Institute of Radiological and Medical Sciences, Seoul (Korea, Republic of); Lee, Byung Chul; Moon, Byung Seok; Choi, Tae Hyun; Chuna, Kwon Soo [Inha Univ., Inchon (Korea, Republic of)

    2005-07-01

    The {alpha}{sub v}{beta}{sub 3} integrin is an important receptor affecting tumor growth, metastatic potential on proliferating endothelial cells as well as on tumor cells of various origin, tumor-induced angiogenesis could be blocked by antagonizing the {alpha}{sub v}{beta}{sub 3} integrin with RGD. Therefore, {alpha}{sub v}{beta}{sub 3} integrin is a target for angiogenesis imaging that might be useful in assessing tumor-induced angiogenesis and identifying tumor metastasis. To design potent radiotracer for imaging angiogenesis containing a cRGD moiety should include low hepatic uptake in vivo. Tripeptide Arg-Gly-Asp (RGD), naturally existed in extracellular matrix proteins, is known to be the primary binding site of the {alpha}{sub v}{beta}{sub 3} integrin. The imaging of {alpha}{sub v}{beta}{sub 3} receptor expression will give the information of the metastatic ability of the tumor which is not available by [{sup 18}F]FDG. Our interest in developing new radiopharmaceuticals for in vivo visualization of angiogenesis has led us to synthesize derivatives of cRGD (cyclic arginineglycine-aspartic acid) that contains glucose moiety. Because sugar-protein interaction is a key step in metastasis and angiogenesis, it has also been proposed to play an intriguing role in imaging of tumor. We designed and synthesized two fluorine-18 labeled RGD glycopeptides . N-fluorobenzyl-diaminobutane-N'-glucose-Lys-Arg-Gly-Asp-D-Phe ([{sup 18}F]fluorobenzyl-glucose-KRGDf, and Nfluorobenzoyl- diaminobutane-N'-glucose-Lys-Arg-Gly-Asp-D-Phe ([{sup 18}F]fluorobenzoyl-glucose-KRGDf, from same precursor as a diagnostic tumor imaging agent for positron emission tomography (PET). Fluorine-18 labeled cRGD glycopeptides were prepared using two different simple labeling methods: one is reductive alkylation of an amine with [{sup 18}F]fluorobenzaldehyde and the other is amide condensation with [{sup 18}F]fluorobenzoic acid.

  5. Protease-catalyzed Synthesis of Bz-Arg-Gly-Asp-OMe in Full Aqueous Medium

    Institute of Scientific and Technical Information of China (English)

    HOU Rui-zhen; LI Hong-mei; LIU Yun-jia; ZHANG Long; XU Li; ZHANG Xue-zhong

    2007-01-01

    Synthesis of N-benzoyl-argininylglycylasparagine methyl ester(Bz-Arg-Gly-Asp-OMe), a precursor tripeptide of Arg-Gly-Asp) was catalyzed by papain under kinetic control, at alkaline pH, in a full aqueous medium. The substrates were N-benzoyl-argininylglycine ethyl ester and asparagine dimethyl ester. An aqueous solution of 0. 1 mol/L KCl/NaOH containing 8 mmol/L EDTA and 2 mmol/L DTT was selected as the reaction medium. The synthesized hydrophilic tripeptide was soluble in the reaction medium during the reaction process, however, the secondary hydrolysis of the tripeptide product was not considerable. The effects of different factors, including water content, temperature, reaction time, and molar ratio of the substrates, on the yield of Bz-Arg-Gly-Asp-OMe were examined. The optimal reaction conditions were 0.05 mol/L Bz-Arg-Gly-OEt and 0.15 mol/L Asp(-OMe)2 · HCl in 0.1 mol/L KCl/NaOH solution(pH 8.5), at 40 ℃, and a reaction time of 60 min, with a maximum conversion yield of 62.4%.

  6. Cellular Adhesion Tripeptide RGD Inhibits Growth of Human Ileocecal Adenocarcinoma Cells HCT-8 and Induces Apoptosis

    Institute of Scientific and Technical Information of China (English)

    WANG Hua; ZENG Hong-bin; YANG Shao-juan; GAO Shen; HUANG Yi-bing; HOU Rui-zhen; ZHAO Mi-feng; XU Li; ZHANG Xue-zhong

    2007-01-01

    The tripeptide, Arg-Gly-Asp(RGD) motif is an integrin-recognition site found in adhesive proteins present in extracellular matrices(ECM) and in the blood. HCT-8 cells were treated with cellular adhesion tripeptide RGD at various concentrations. MTT assay was performed to examine the growth and proliferation of HCT-8 cells after treatment with RGD for 48 h. Haematoxylin and Eosin(HE) staining and electromicroscope were used to observe the morphology of apoptotic cells. Survivin and flow cytometry were also used to analyze the HCT-8 apoptosis. Cellular adhesion tripeptide RGD significantly inhibits the growth and proliferation of HCT-8 cells in a dose-dependent manner and induces apoptosis of HCT-8. These results indicate that cellular adhesion tripeptide RGD inhibits the growth and proliferation of tumor HCT-8 cell, probably by the aid of inducing apoptosis of HCT-8 cell.

  7. A cell surface receptor complex for collagen type I recognizes the Arg- Gly-Asp sequence

    OpenAIRE

    1987-01-01

    To isolate collagen-binding cell surface proteins, detergent extracts of surface-iodinated MG-63 human osteosarcoma cells were chromatographed on affinity matrices of either type I collagen- Sepharose or Sepharose carrying a collagen-like triple-helical peptide. The peptide was designed to be triple helical and to contain the sequence Arg-Gly-Asp, which has been implicated as the cell attachment site of fibronectin, vitronectin, fibrinogen, and von Willebrand factor, and is also present in ty...

  8. Repetitive Gly-Leu-Lys-Gly-Glu-Asn-Arg-Gly-Asp peptide derived from collagen and fibronectin for improving cell-scaffold interaction.

    Science.gov (United States)

    Chaisri, Patcharaporn; Chingsungnoen, Artit; Siri, Sineenat

    2015-03-01

    Suitable scaffolds for tissue engineering should provide a microenvironment for cell dwelling and directing cell behavior that resemble the native environment. Three-dimensional geometry of electrospun scaffolds well supports cell deposition, but they often lack biomacromolecules to induce cell responses. In this work, the repetitive collagen and fibronectin motif (rCF) peptide containing multiple repeats of Gly-Leu-Lys-Gly-Glu-Asn-Arg-Gly-Asp sequence derived from the cell adhesion motifs of collagen and fibronectin was produced as the alternative agent to induce cell-scaffold interaction. The DNA fragment encoding rCF peptide was amplified by a polymerase chain reaction using overlap primers without a DNA template, cloned into a protein expression vector, and expressed as a His-tag fusion peptide in Escherichia coli. The purified rCF peptide possessed cell adhesion activity about 1.5-fold of the commercial RGD peptide. The rCF peptide was grafted onto the electrospun PCL scaffold via RF plasma of Ar/O2 discharge and acrylic acid treatment. The immobilized rCF peptide significantly increased surface hydrophilicity and enhanced cell proliferation of the electrospun PCL scaffold. These findings suggest the potential application of rCF peptide for improving the biomimetic functions of polymeric scaffolds for tissue engineering.

  9. Technetium-99m-labeled Arg-Gly-Asp-conjugated alpha-melanocyte stimulating hormone hybrid peptides for human melanoma imaging

    Energy Technology Data Exchange (ETDEWEB)

    Yang Jianquan; Guo Haixun [College of Pharmacy, University of New Mexico, Albuquerque, NM 87131 (United States); Miao Yubin, E-mail: ymiao@salud.unm.ed [College of Pharmacy, University of New Mexico, Albuquerque, NM 87131 (United States); Cancer Research and Treatment Center, University of New Mexico, Albuquerque, NM 87131 (United States); Department of Dermatology, University of New Mexico, Albuquerque, NM 87131 (United States)

    2010-11-15

    Introduction: The purpose of this study was to examine whether {sup 99m}Tc-labeled Arg-Gly-Asp (RGD)-conjugated alpha-melanocyte stimulating hormone ({alpha}-MSH) hybrid peptide targeting both melanocortin-1 (MC1) and {alpha}{sub v{beta}3} integrin receptors was superior in melanoma targeting to {sup 99m}Tc-labeled {alpha}-MSH or RGD peptide targeting only the MC1 or {alpha}{sub v{beta}3} integrin receptor. Methods: RGD-Lys-(Arg{sup 11})CCMSH, RAD-Lys-(Arg{sup 11})CCMSH and RGD-Lys-(Arg{sup 11})CCMSHscramble were designed to target both MC1 and {alpha}{sub v{beta}3} integrin receptors, MC1 receptor only and {alpha}{sub v{beta}3} integrin receptor only, respectively. The MC1 or {alpha}{sub v{beta}3} integrin receptor binding affinities of three peptides were determined in M21 human melanoma cells. The melanoma targeting properties of {sup 99m}Tc-labeled RGD-Lys-(Arg{sup 11})CCMSH, RAD-Lys-(Arg{sup 11})CCMSH and RGD-Lys-(Arg{sup 11})CCMSHscramble were determined in M21 human melanoma-xenografted nude mice. Meanwhile, the melanoma uptake of {sup 99m}Tc-RGD-Lys-(Arg{sup 11})CCMSH was blocked with various non-radiolabeled peptides in M21 melanoma xenografts. Results: RGD-Lys-(Arg{sup 11})CCMSH displayed 2.0 and 403 nM binding affinities to both MC1 and {alpha}{sub v{beta}3} integrin receptors, whereas RAD-Lys-(Arg{sup 11})CCMSH or RGD-Lys-(Arg{sup 11})CCMSHscramble lost their {alpha}{sub v{beta}3} integrin receptor binding affinity by greater than 248-fold or MC1 receptor binding affinity by more than 100-fold, respectively. The melanoma uptake of {sup 99m}Tc-RGD-Lys-(Arg{sup 11})CCMSH was 2.49 and 2.24 times (P < .05) the melanoma uptakes of {sup 99m}Tc-RAD-Lys-(Arg{sup 11})CCMSH and {sup 99m}Tc-RGD-Lys-(Arg{sup 11})CCMSHscramble at 2 h post-injection, respectively. Either RGD or (Arg{sup 11})CCMSH peptide co-injection could block 42% and 57% of the tumor uptake of {sup 99m}Tc-RGD-Lys-(Arg{sup 11})CCMSH, whereas the coinjection of RGD+(Arg{sup 11})CCMSH peptide mixture

  10. Technetium-99m-labeled Arg-Gly-Asp-conjugated alpha-melanocyte stimulating hormone hybrid peptides for human melanoma imaging

    International Nuclear Information System (INIS)

    Introduction: The purpose of this study was to examine whether 99mTc-labeled Arg-Gly-Asp (RGD)-conjugated alpha-melanocyte stimulating hormone (α-MSH) hybrid peptide targeting both melanocortin-1 (MC1) and αvβ3 integrin receptors was superior in melanoma targeting to 99mTc-labeled α-MSH or RGD peptide targeting only the MC1 or αvβ3 integrin receptor. Methods: RGD-Lys-(Arg11)CCMSH, RAD-Lys-(Arg11)CCMSH and RGD-Lys-(Arg11)CCMSHscramble were designed to target both MC1 and αvβ3 integrin receptors, MC1 receptor only and αvβ3 integrin receptor only, respectively. The MC1 or αvβ3 integrin receptor binding affinities of three peptides were determined in M21 human melanoma cells. The melanoma targeting properties of 99mTc-labeled RGD-Lys-(Arg11)CCMSH, RAD-Lys-(Arg11)CCMSH and RGD-Lys-(Arg11)CCMSHscramble were determined in M21 human melanoma-xenografted nude mice. Meanwhile, the melanoma uptake of 99mTc-RGD-Lys-(Arg11)CCMSH was blocked with various non-radiolabeled peptides in M21 melanoma xenografts. Results: RGD-Lys-(Arg11)CCMSH displayed 2.0 and 403 nM binding affinities to both MC1 and αvβ3 integrin receptors, whereas RAD-Lys-(Arg11)CCMSH or RGD-Lys-(Arg11)CCMSHscramble lost their αvβ3 integrin receptor binding affinity by greater than 248-fold or MC1 receptor binding affinity by more than 100-fold, respectively. The melanoma uptake of 99mTc-RGD-Lys-(Arg11)CCMSH was 2.49 and 2.24 times (P 99mTc-RAD-Lys-(Arg11)CCMSH and 99mTc-RGD-Lys-(Arg11)CCMSHscramble at 2 h post-injection, respectively. Either RGD or (Arg11)CCMSH peptide co-injection could block 42% and 57% of the tumor uptake of 99mTc-RGD-Lys-(Arg11)CCMSH, whereas the coinjection of RGD+(Arg11)CCMSH peptide mixture could block 66% of the tumor uptake of 99mTc-RGD-Lys-(Arg11)CCMSH. Conclusions: Targeting both MC1 and αvβ3 integrin receptors enhanced the melanoma uptake of 99mTc-RGD-Lys-(Arg11)CCMSH in M21 human melanoma xenografts. Flank M21 human melanoma tumors were clearly visualized by single

  11. 精氨酸-甘氨酸-天冬氨酸肽类似物的固相合成%Solid-phase synthesis of Arg-Gly-Asp related peptoids

    Institute of Scientific and Technical Information of China (English)

    邓昌辉; 邓桂茹; 郝鹤; 韩香

    2011-01-01

    Objective Peptides involving the Arg-Gly-Asp motive(arginine-glycine-aspartic acid,RGD) recognize members of the integrin receptor family,but those small RGD peptides are readily degraded in vivo.Since structural modifications including peptoids were recently found to improve the tolerance to peptidase significantly,five RGD related peptides were designed and synthesized with the purpose to improve the therapeutic index of this antitumoral drug family.[Methods]These compounds were structural featured by using Ida (imino-diacetic acid) residue and N-methyl(or ethyl) amide moiety as the surrogates for Asp and C-terminal carboxyl group respectively.All synthesis were performed by N-Boc protecting chemistry and stepwise coupling strategy on Merrifield resin support.[ Results]All N-substituted peptoids were obtained with total yields over 85%,purities over 95.1%,and characterized by LC-MS analysis.[Conclusion]It would be meaningful that an economical method for the synthesis of peptoids is possibly revealed by present synthesis.%[目的]含有Arg-Gly-Asp(精氨酰甘氨酰天冬氨酸,RGD)片段的多肽可以识别整合素受体家族,但是这些含RGD小肽在体内很容易被蛋白酶降解.本研究设计并合成RGD肽类似物,通过结构改造克服这一缺点,以期开发新的高效的抗肿瘤药物.[方法]分子设计采用N取代甘氨酸(NSG)残基替代法,即用NSG残基Ida代替Asp、用酰甲胺或酰乙胺代替C末端的羧基,生成N取代类肽型的产物;化学合成采用Boc保护策略,在Merrifield树脂上逐步固相合成N取代类肽型的产物.[结果]全部RGD肽类似物结构均经LC-MS分析证明,总收率在85%以上,纯度>95.1%.[结论]通过本合成工作,为建立经济、可行的类肽合成提供了一种有效途径.

  12. Chapter 20: In vitro and in vivo evaluation of ['99mTc(CO)3-cyclo[Arg-Gly-Asp-D-Tyr-Lys(PZ1)

    International Nuclear Information System (INIS)

    As part of a coordinated research project, Development of 99mTc Based Small Biomolecules Using Novel 99mTc Cores, with the International Atomic Energy Agency, we report the design and development of a new cyclic Rd analogue cyclo[Arg-Gly- Asp-D-Tyr-Lys(P Z1)] (P Z1 3,5Me2-Pz(CH2)2N((CH2)3COOH)(CH2)2NH2) that can be radiolabelled with the 99mTc(CO)3 metal fragment. Radiochemical evaluation, including synthesis, in vitro uptake analyses and in vivo pharmacokinetics in normal mice, is discussed. (author)

  13. Covalent attachment of cell-adhesive peptide Gly-Arg-Gly-Asp (GRGD) to poly(etheretherketone) surface by tailored silanization layers technique

    Energy Technology Data Exchange (ETDEWEB)

    Zheng, Yanyan [Chengdu Institute of Organic Chemistry, Chinese Academy of Sciences, Chengdu 610041 (China); University of Chinese Academy of Sciences, Beijing 100049 (China); Xiong, Chengdong [Chengdu Institute of Organic Chemistry, Chinese Academy of Sciences, Chengdu 610041 (China); Li, Xiaoyu [State Key Laboratory of Oral Diseases, West China Hospital of Stomatology, Sichuan University, Chengdu 610041 (China); Zhang, Lifang, E-mail: zhanglfcioc@163.com [Chengdu Institute of Organic Chemistry, Chinese Academy of Sciences, Chengdu 610041 (China)

    2014-11-30

    Highlights: • The carbonyl groups on PEEK surface were effectively reduced to hydroxyl groups using sodium borohydride. • Silanization layers technique was employed to immobilize the cell-adhesive peptide Gly-Arg-Gly-Asp (GRGD) on hydroxylation-pretreated PEEK sheet surface by covalent chemical attachment. • XPS, surface profiler and water contact angle measurements proved the presence of GRGD on PEEK surface. • Osteoblast-like cells (MC3T3-E1) attachment and proliferation were improved effectively on GRGD-modified PEEK surface. - Abstract: Poly(etheretherketone) (PEEK) is a rigid semicrystalline polymer that combines excellent mechanical properties, broad chemical resistance and bone-like stiffness and is widely used in biomedical fields. However, PEEK is naturally bioinert, leading to limited biomedical applications, especially when a direct bone-implant osteointegration is desired. In this study, a three-step reaction procedure was employed to immobilize the cell-adhesive peptide Gly-Arg-Gly-Asp (GRGD) on the surface of PEEK sheet by covalent chemical attachment to favor cell adhesion and proliferation. First, hydroxylation-pretreated PEEK surfaces were silanized with 7-Oct-1-enyltrichlorosilane (OETS) in dry cyclohexane, resulting in a silanization layer with terminal ethenyl. Second, the terminal ethylenic double bonds of the silanization layer on PEEK surface were converted to carboxyl groups through acidic potassium manganate oxidation. Finally, GRGD was covalently attached by carbodiimide mediated condensation between the carboxyl on PEEK surface and amine presents in GRGD. X-ray photoelectron spectroscopy (XPS), attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy, surface profiler and water contact angle measurements were applied to characterize the modified surfaces. The effect of cells attachment and proliferation on each specimen was investigated. Pre-osteoblast cells (MC3T3-E1) attachment, spreading and proliferation

  14. (99m)Tc-Galacto-RGD2: a novel 99mTc-labeled cyclic RGD peptide dimer useful for tumor imaging.

    Science.gov (United States)

    Ji, Shundong; Czerwinski, Andrzej; Zhou, Yang; Shao, Guoqiang; Valenzuela, Francisco; Sowiński, Paweł; Chauhan, Satendra; Pennington, Michael; Liu, Shuang

    2013-09-01

    This study sought to evaluate [(99m)Tc(HYNIC-Galacto-RGD2)(tricine)(TPPTS)] ((99m)Tc-Galacto-RGD2: HYNIC = 6-hydrazinonicotinyl; Galacto-RGD2 = Glu[cyclo[Arg-Gly-Asp-D-Phe-Lys(SAA-PEG2-(1,2,3-triazole)-1-yl-4-methylamide)

  15. Evaluation of Osteoblast-Like Cell Viability and Differentiation on the Gly-Arg-Gly-Asp-Ser Peptide Immobilized Titanium Dioxide Nanotube via Chemical Grafting.

    Science.gov (United States)

    Kim, Ga-Hyun; Kim, Il-Shin; Park, Sang-Won; Lee, Kwangmin; Yun, Kwi-Dug; Kim, Hyun-Seung; Oh, Gye-Jeong; Ji, Min-Kyung; Lim, Hyun-Pil

    2016-02-01

    This study examined the effect of the immobilization of the Gly-Arg-Gly-Asp-Ser (GRGDS) peptide on titanium dioxide (TiO2) nanotube via chemical grafting on osteoblast-like cell (MG-63) viability and differentiation. The specimens were divided into two groups; TiO2 nanotubes and GRGDS-immobilized TiO2 nanotubes. The surface characteristics of GRGDS-immobilized TiO2 nanotubes were observed by using X-ray photoelectron spectroscopy (XPS) and a field emission scanning electron microscope (FE-SEM). The morphology of cells on specimens was observed by FE-SEM after 2 hr and 24 hr. The level of cell viability was investigated via a tetrazolium (XTT) assay after 2 and 4 days. Alkaline phosphatase (ALP) activity was evaluated to measure the cell differentiation after 4 and 7 days. The presence of nitrogen up-regulation or C==O carbons con- firmed that TiO2 nanotubes were immobilized with GRGDS peptides. Cell adhesion was enhanced on the GRGDS-immobilized TiO2 nanotubes compared to TiO2 nanotubes. Furthermore, significantly increased cell spreading and proliferation were observed with the cells grown on GRGDS-immobilized TiO2 nanotubes (P nanotubes and TiO2 nanotubes. These results suggest that the GRGDS-immobilized TiO2 nanotubes might be effective in improving the osseointegration of dental implants.

  16. A fluorescent Arg-Gly-Asp (RGD) peptide-naphthalenediimide (NDI) conjugate for imaging integrin α(v)β(3) in vitro.

    Science.gov (United States)

    Hu, Zhiyuan; Arrowsmith, Rory L; Tyson, James A; Mirabello, Vincenzo; Ge, Haobo; Eggleston, Ian M; Botchway, Stanley W; Pantos, G Dan; Pascu, Sofia I

    2015-04-25

    We have developed a fluorescent peptide conjugate (TrpNDIRGDfK) based on the coupling of cyclo(RGDfK) to a new tryptophan-tagged amino acid naphthalenediimide (TrpNDI). Confocal fluorescence microscopy coupled with fluorescence lifetime imaging (FLIM) mapping, single and two-photon fluorescence excitation, lifetime components and corresponding decay profiles were used as parameters able to investigate qualitatively the cellular behavior regarding the molecular environment and biolocalisation of TrpNDI and TrpNDI-RGDfK in cancer cells. PMID:25647279

  17. Effects of Arg-Gly-Asp-modified elastin-like polypeptide on pseudoislet formation via up-regulation of cell adhesion molecules and extracellular matrix proteins.

    Science.gov (United States)

    Lee, Kyeong-Min; Jung, Gwon-Soo; Park, Jin-Kyu; Choi, Seong-Kyoon; Jeon, Won Bae

    2013-03-01

    Extracellular matrix (ECM) plays an important role in controlling the β-cell morphology, survival and insulin secretary functions. An RGD-modified elastin-like polypeptide (RGD-ELP), TGPG[VGRGD(VGVPG)(6)](20)WPC, has been reported previously as a bioactive matrix. In this study, to investigate whether RGD-ELP affects β-cell growth characteristics and insulin secretion, β-TC6 cells were cultured on the RGD-ELP coatings prepared via thermally induced phase transition. On RGD-ELP, β-TC6 cells clustered into an islet-like architecture with high cell viability. Throughout 7days' culture, the proliferation rate of the cells within a pseudoislet was similar to that of monolayer culture. Under high glucose (25mM), β-TC6 pseudoislets showed up-regulated insulin gene expression and exhibited glucose-stimulated insulin secretion. Importantly, the mRNA and protein abundances of cell adhesion molecules (CAM) E-cadherin and connexin-36 were much higher in pseudoislets than in monolayer cells. The siRNA-mediated inhibition of E-cadherin or connexin-36 expression severely limited pseudoislet formation. In addition, the mRNA levels of collagen types I and IV, fibronectin and laminin were significantly elevated in pseudoislets. The results suggest that RGD-ELP promotes pseudoislet formation via up-regulation of the CAM and ECM components. The functional roles of RGD-ELP are discussed in respect of its molecular composition.

  18. Improving the affinity of fibroblasts for bacterial cellulose using carbohydrate-binding modules fused to RGD

    OpenAIRE

    Andrade, Fábia K; Moreira, Susana Margarida Gomes; Domingues, Lucília; Gama, F. M.

    2010-01-01

    The attachment of cells to biomedical materials can be improved by using adhesion sequences, such as Arg-Gly-Asp (RGD), found in several extracellular matrix proteins. In this work, bifunctional recombinant proteins, with a Cellulose-Binding Module (CBM), from the cellulosome of Clostridium thermocellum and cell binding sequences - RGD, GRGDY - were cloned and expressed in E.coli. These RGD-containing cellulose binding proteins were purified and used to coat bacterial cellulose fibres. Its ef...

  19. Graphene oxide-stimulated myogenic differentiation of C2C12 cells on PLGA/RGD peptide nanofiber matrices

    Science.gov (United States)

    Shin, Y. C.; Lee, J. H.; Kim, M. J.; Hong, S. W.; Oh, J.-W.; Kim, C.-S.; Kim, B.; Hyun, J. K.; Kim, Y.-J.; Han, D.-W.

    2015-07-01

    During the last decade, much attention has been paid to graphene-based nanomaterials because they are considered as potential candidates for biomedical applications such as scaffolds for tissue engineering and substrates for the differentiation of stem cells. Until now, electrospun matrices composed of various biodegradable copolymers have been extensively developed for tissue engineering and regeneration; however, their use in combination with graphene oxide (GO) is novel and challenging. In this study, nanofiber matrices composed of poly(lactic-co-glycolic acid, PLGA) and M13 phage with RGD peptide displayed on its surface (RGD peptide-M13 phage) were prepared as extracellular matrix (ECM)-mimicking substrates. RGD peptide is a tripeptide (Arg-Gly-Asp) found on ECM proteins that promotes various cellular behaviors. The physicochemical properties of PLGA and RGD peptide-M13 phage (PLGA/RGD peptide) nanofiber matrices were characterized by atomic force microscopy, Fourier-transform infrared spectroscopy and thermogravimetric analysis. In addition, the growth of C2C12 mouse myoblasts on the PLGA/RGD peptide matrices was examined by measuring the metabolic activity. Moreover, the differentiation of C2C12 mouse myoblasts on the matrices when treated with GO was evaluated. The cellular behaviors, including growth and differentiation of C2C12 mouse myoblasts, were substantially enhanced on the PLGA/RGD peptide nanofiber matrices when treated with GO. Overall, these findings suggest that the PLGA/RGD peptide nanofiber matrices can be used in combination with GO as a novel strategy for skeletal tissue regeneration.

  20. Chemoenzymatic Synthesis of Cellular Adhesion Tripeptide RGD Precursor in Organic Media

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    Chemoenzymatic synthesis of tripeptide Bz-RGD-(OEt)2 was conducted in this study. First, the free dipeptide Gly-Asp was synthesized via a novel chemical method, wherein only L-aspartic acid was used and was followed by the esterification of Gly-Asp. The formation of the linkage between the third amino acid Bz-Arg-OEt and Gly-Asp-(OEt)2 was completed by using the enzymatic method in organic media. The effects of several factors such as pH, the water content, triethylamine(TEA), the molar ratio of the substrates, and the reaction time on the yield of Bz-RGD-(OEt)2 were examined. It was obtained that the optimum conditions for Bz-RGD-(OEt)2 synthesis in an ethanol/Tris-HCl buffer system(volume ratio 93:7) were as follows; pH=8.0; temperature, 30 ℃; reaction time, 7 h. The tripeptide yield was 75.2%.

  1. Dimer of the peptide cycle (Ar-Gly-Asp-D-Phe-Lys) radiolabeled with {sup 99m}Tc for the integrin s over-expression image: formulation, biokinetics and dosimetry; Dimero del peptido ciclo(Arg-Gly-Asp-D-Phe-Lys) radiomarcado con {sup 99m}Tc para la imagen de sobre-expresion de integrinas: formulacion, biocinetica y dosimetria

    Energy Technology Data Exchange (ETDEWEB)

    Ortiz A, Z.

    2013-07-01

    In breast cancer, α(v)β(3) and/or α(v)β(5) integrin s are over-expressed in both endothelial and tumour cells. Radiolabeled peptides based on the RGD (Arg-Gly-Asp) sequence are radiopharmaceuticals with high affinity and selectivity for those integrin s. The RGD-dimer peptide (E-[c(RGDfK)]{sub 2}) radiolabeled with {sup 99m}Tc has been reported as a radiopharmaceutical with 10-fold higher affinity for the α(v)β(3) integrin as compared to the RGD-monomer. EDDA (Ethylenediamine-N,N-diacetic acid) is a hydrophilic molecule that may favours renal excretion when used as coligand in the {sup 99m}Tc labelling of HYNIC-peptides and can easily be formulated in a lyophilized kit. Aim: Establish a biokinetic model for {sup 99m}Tc-EDDA/HYNIC-E-[c(RGDfK)]{sub 2} prepared from lyophilized kits and evaluate the dosimetry as breast cancer imaging agent. Methods: {sup 99m}Tc labelling was performed by addition of sodium pertechnetate solution and 0.2 M phosphate buffer ph 7.0 to a lyophilized formulation containing E-[c(RGDfK)]{sub 2}, EDDA, tricine, mannitol and stannous chloride. Radiochemical purity was evaluated by reversed phase HPLC and ITLC-SG analyses. Stability studies in human serum were carried out by size-exclusion HPLC. In-vitro cell uptake was tested using breast cancer cells (MCF7, T47D and MDA-MB-231) with blocked and non-blocked receptors. Biodistribution and tumour uptake were determined in MCF7 tumour-bearing nude mice with blocked and non-blocked receptors, and images were obtained using a micro-SPECT/CT. Whole-body images from seven healthy women were acquired at 1, 3, 6 and 24 h after {sup 99m}Tc-EDDA/HYNIC-E-[c(RGDfK)]{sub 2} administration obtained with radiochemical purities of >94 %. Regions of interest (ROIs) were drawn around source organs on each time frame. Each ROI was converted to activity using the conjugate view counting method. The image sequence was used to extrapolate {sup 99m}Tc-EDDA/HYNIC-E-[c(RGDfK)]{sub 2} time-activity curves in each

  2. Roughened titanium surfaces with silane and further RGD peptide modification in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Wen-Cheng, E-mail: wencchen@fcu.edu.tw [Advanced Medical Devices and Composites Laboratory, Department of Fiber and Composite Materials, College of Engineering, Feng Chia University, Taichung 40724, Taiwan, ROC (China); Ko, Chia-Ling [School of Dentistry, College of Dental Medicine, Kaohsiung Medical University, Kaohsiung 807, Taiwan, ROC (China)

    2013-07-01

    The strategy to achieve osteoregeneration of dental implants during early-stage regeneration is strongly related to surface conditions for achieving highly successful effects after implantation. Surface modifications, namely, mechanical ground, silanization, bonded and sandblasted with pentasequence Gly-Arg-Gly-Asp-Ser (GRGDS) peptide, and acid-etched with Arg-Gly-Asp (RGD) peptide, were compared for their ability to support cell attachment, proliferation, and differentiation on titanium surfaces. The characteristics and comparative in vitro bio-interactions toward osteoprogenitor cells were tested in the four groups with various surface modifications. Compared with the other groups, the sandblasted and acid-etched, and silane with subsequent RGD peptide modified surfaces had the smallest wetting angle, absence of a significant cell viability difference, and largest quantity of alkaline phosphatase production during the expressions of early-stage cell differentiation. The method of synthesizing GRGDS peptides on roughened titanium surfaces has the potential to provide a combination of early bone regeneration and implant of long-term anchored capabilities. Highlights: • The osteoregeneration during early-stage is strongly related to surface conditions. • The wettability with RGD peptide treated surfaces can be enhanced. • Rougher surface binding with RGD peptide can achieve better osseogeneration. • Surfaces with RGD peptide accelerate the progenitor bone cell mineralization.

  3. Interfacial adsorption of fibrinogen and its inhibition by RGD peptide: a combined physical study

    Energy Technology Data Exchange (ETDEWEB)

    Armstrong, Johanna [Biological Physics Group, Department of Physics, UMIST, PO Box 88, Manchester M60 1QD (United Kingdom); Salacinski, Henryk J [Biomaterial and Tissue Engineering Centre, Department of Surgery, Royal Free and University College Medical School, UCL, Pond Street, London, NW3 2QG (United Kingdom); Mu Qingshan [Biological Physics Group, Department of Physics, UMIST, PO Box 88, Manchester M60 1QD (United Kingdom); Seifalian, Alex M [Biomaterial and Tissue Engineering Centre, Department of Surgery, Royal Free and University College Medical School, UCL, Pond Street, London, NW3 2QG (United Kingdom); Peel, Louise [Farfield Sensors Ltd, Salford University Business Park, Salford, M6 6AJ (United Kingdom); Freeman, Neville [Farfield Sensors Ltd, Salford University Business Park, Salford, M6 6AJ (United Kingdom); Holt, Cathy M [Cardiac Physiology Unit, University of Manchester, Manchester Medical School, Stopford Building, Oxford Road, Manchester, M13 9PT (United Kingdom); Lu, Jian R [Biological Physics Group, Department of Physics, UMIST, PO Box 88, Manchester M60 1QD (United Kingdom)

    2004-07-07

    The Arg-Gly-Asp (RGD) peptide sequence is known as a cell recognition site for numerous adhesive proteins present in the extracellular matrix (ECM) and in blood. Whilst surface immobilized RGD groups enhance cell attachment, RGD components present in solution can effectively inhibit cell attachment by competing with endogenous ligands for the same recognition site. In contrast to the widely reported binding to cell integrin, this study demonstrates a new RGD feature: its inhibitive effect on fibrinogen adsorption. Through a combined analysis of spectroscopic ellipsometry, neutron reflection and dual polarization interferometry, we show that the kinetic process of fibrinogen adsorption as a model pro-coagulant at the silica/solution interface and in the absence of any cells can be substantially reduced by the addition of RGD in solution and that the extent of the reduction is dependent on the relative concentration of RGD.

  4. Improving Tumor Uptake and Pharmacokinetics of 64Cu-Labeled Cyclic RGD Peptide Dimers with Gly3 and PEG4 Linkers

    OpenAIRE

    Shi, Jiyun; Kim, Young-Seung; Zhai, Shizhen; Liu, Zhaofei; Chen, Xiaoyuan; Liu, Shuang

    2009-01-01

    Radiolabeled cyclic RGD (Arg-Gly-Asp) peptides represent a new class of radiotracers with potential for the early tumor detection and non-invasive monitoring of tumor metastasis and therapeutic response in cancer patients. This report describes the synthesis of two cyclic RGD peptide dimer conjugates, DOTA-PEG4-E[PEG4-c(RGDfK)]2 (DOTA-3PEG4-dimer: DOTA = 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid; PEG4 = 15-amino-4,7,10,13-tetraoxapentadecanoic acid) and DOTA-G3-E[G3-c(RGDfK)]2 ...

  5. Status and Advances of RGD Molecular Imaging in Lung Cancer

    Directory of Open Access Journals (Sweden)

    Ning YUE

    2014-12-01

    Full Text Available Lung cancer has been one of the most common and the highest mortality rates malignant tumors at home and abroad. Sustained angiogenesis was not only the characteristic of malignant tumors, but also the foundation of tumor proliferation, invasion, recurrence and metastasis, it was also one of the hot spots of treatments in lung cancer biology currently. Integrins played an important part in tumor angiogenesis. Arg-Gly-Asp (RGD peptides could combine with integrins specifically, and the application of radionuclide-labeled RGD molecular probes enabled imaging of tumor blood vessels to reflect its changes. The lung cancer imaging of RGD peptides at home and abroad in recent years was reviewed in this article.

  6. Interaction between integrin a Ⅱbβ 3 and synthesized cyclic hexapeptide containing RGD

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    The RGD sequence generally exists in the extracellular matrix proteins and can be recognized by many integrin proteins. The binding ability of immobilized biotinylated cyclic hexapeptide [cyclo(-Arg-Gly-Asp-D-Phe-Lys-Gly-)] containing RGD to integrin a ab β3 was tested by the methods of ELISA and SPR. Results showed that a spacer of 1.48-2.2 nm between the peptide and the biotin residue was long enough to send the RGD sequence into the binding center embedded within a Ⅱbβ 3, and the equilibrium dissociation constant was 1.1 μm. The work provides an ideal model system for the research of cell adhesion on solid surfaces.

  7. RGD修饰的聚乳酸-羟基乙酸骨组织工程材料的研究进展%RGD-modified polylactide-co-glycolic acid tissue engineering scaffolds for bone regeneration: an advance

    Institute of Scientific and Technical Information of China (English)

    陶春; 陈琰; 钟延强

    2012-01-01

    Polylactide-co-glycolic acid (PLGA) tissue engineering scaffolds play an important role in the regeneration of bone and cartilage. Due to the poor hydrophilicity of PLGA, it is difficult for cells to attach to the scaffolds. Modification by RGD (Arg-Gly-Asp) peptides can effectively improve the cellular affinity of PLGA and adhesion and proliferation of the seed cells. This review summarizes the recent progress in PLGA tissue engineering scaffolds modified by RGD peptides.%聚乳酸-羟基乙酸(PLGA)骨组织工程支架在骨损伤修复和再造方面有着重要的应用,但由于PLGA亲水性差,不利于种子细胞在支架上的黏附和增殖.RGD(精氨酸-甘氨酸-天冬氨酸,Arg-Gly-Asp)肽修饰PLGA支架后,材料的细胞亲和性得到了有效改善,促进了种子细胞黏附和增殖.本文就近年来RGD修饰的PLGA骨组织工程材料的相关研究作一综述.

  8. The RGD finger of Del-1 is a unique structural feature critical for integrin binding

    Energy Technology Data Exchange (ETDEWEB)

    Schürpf, Thomas; Chen, Qiang; Liu, Jin-huan; Wang, Rui; Springer, Timothy A.; Wang, Jia-huai (Harvard-Med)

    2012-11-13

    Developmental endothelial cell locus-1 (Del-1) glycoprotein is secreted by endothelial cells and a subset of macrophages. Del-1 plays a regulatory role in vascular remodeling and functions in innate immunity through interaction with integrin {alpha}{sub V}{beta}{sub 3}. Del-1 contains 3 epidermal growth factor (EGF)-like repeats and 2 discoidin-like domains. An Arg-Gly-Asp (RGD) motif in the second EGF domain (EGF2) mediates adhesion by endothelial cells and phagocytes. We report the crystal structure of its 3 EGF domains. The RGD motif of EGF2 forms a type II' {beta} turn at the tip of a long protruding loop, dubbed the RGD finger. Whereas EGF2 and EGF3 constitute a rigid rod via an interdomain calcium ion binding site, the long linker between EGF1 and EGF2 lends considerable flexibility to EGF1. Two unique O-linked glycans and 1 N-linked glycan locate to the opposite side of EGF2 from the RGD motif. These structural features favor integrin binding of the RGD finger. Mutagenesis data confirm the importance of having the RGD motif at the tip of the RGD finger. A database search for EGF domain sequences shows that this RGD finger is likely an evolutionary insertion and unique to the EGF domain of Del-1 and its homologue milk fat globule-EGF 8. The RGD finger of Del-1 is a unique structural feature critical for integrin binding.

  9. Angiogenesis Imaging Using (68)Ga-RGD PET/CT: Therapeutic Implications.

    Science.gov (United States)

    Eo, Jae Seon; Jeong, Jae Min

    2016-09-01

    Angiogenesis imaging is important for diagnostic and therapeutic treatment of various malignant and nonmalignant diseases. The Arg-Gly-Asp (RGD) sequence has been known to bind with the αvβ3 integrin that is expressed on the surface of angiogenic blood vessels or tumor cells. Thus, various radiolabeled derivatives of RGD peptides have been developed for angiogenesis imaging. Among the various radionuclides, (68)Ga was the most widely studied for RGD peptide imaging because of its excellent nuclear physical properties, easy-to-label chemical properties, and cost-effectiveness owing to the availability of a (68)Ge-(68)Ga generator. Thus, various (68)Ga-labeled RGD derivatives have been developed and applied for preclinical and clinical studies. Clinical trials were performed for both malignant and nonmalignant diseases. Breast cancer, glioma, and lung cancer were malignant, and myocardial infarction, atherosclerosis, and moyamoya disease were nonmalignant among the investigated diseases. Further, these (68)Ga-labeled RGD derivatives could be applied to assess the effects of antiangiogenic treatment or theragnosis or both, of cancers. In conclusion, the angiogenesis imaging technology using (68)Ga-labeled RGD derivatives might be useful for the development of new therapeutic assessments, and for diagnostic and theragnostic applications. PMID:27553467

  10. Status and Advances of RGD Molecular Imaging in Lung Cancer%RGD分子影像在肺癌的研究现状与进展

    Institute of Scientific and Technical Information of China (English)

    岳宁; 袁双虎; 杨国仁

    2014-01-01

    肺癌是国内外最常见、死亡率最高的恶性肿瘤之一。持续的新生血管生成是恶性肿瘤的特征,是肿瘤增殖、浸润、复发和转移的基础,也是目前肺癌生物学治疗热点之一。肿瘤血管生成过程中,整合素的作用至关重要。精氨酸-甘氨酸-天冬氨酸(Arg-Gly-Asp, RGD)肽能特异地与整合素结合,应用放射性核素标记的RGD分子探针,可使肿瘤血管显像,能反映肿瘤血管的变化。本文对近年来国内外RGD肽的肺癌显像研究进展进行综述。%Lung cancer has been one of the most common and the highest mortality rates malignant tumors at home and abroad. Sustained angiogenesis was not only the characteristic of malignant tumors, but also the foundation of tumor proliferation, invasion, recurrence and metastasis, it was also one of the hot spots of treatments in lung cancer biology currently. Integrins played an important part in tumor angiogenesis. Arg-Gly-Asp (RGD) peptides could combine with integrins speciif-cally, and the application of radionuclide-labeled RGD molecular probes enabled imaging of tumor blood vessels to relfect its changes. hTe lung cancer imaging of RGD peptides at home and abroad in recent years was reviewed in this article.

  11. Cyclic RGD peptide-modified liposomal drug delivery system: enhanced cellular uptake in vitro and improved pharmacokinetics in rats

    Directory of Open Access Journals (Sweden)

    Chen Z

    2012-07-01

    Full Text Available Zhongya Chen,1,2 Jiaxin Deng,1,2 Yan Zhao,1,2 Tao Tao1,21National Pharmaceutical Engineering Research Center, 2Shanghai Institute of Pharmaceutical Industry, China State Institute of Pharmaceutical Industry, Shanghai, People's Republic of ChinaBackground: Integrins αvβ3 and αvβ5, both of which specifically recognize the Arg-Gly-Asp (RGD motif, are overexpressed on many solid tumors and in tumor neovasculature. Thus, coupling the RGD motif to the liposomal surface for achieving active targeting can be a promising strategy for the treatment of tumors.Methods: Cyclo(Arg-Gly-Asp-D-Phe-Cys (cRGD was covalently coupled with the liposomal membrane surface, followed by coating with poly(ethylene glycol (PEG using the post-insertion technique. The coupling efficiency of cRGD was determined. Doxorubicin as a model anticancer drug was loaded into liposomes using an ammonium sulfate gradient method to investigate the encapsulation efficiency, cellular uptake by the integrin-overexpressing human glioma cell line U87MG in vitro, and pharmacokinetic properties in Sprague-Dawley rats.Results: cRGD was conjugated to the liposomal surface by a thiol-maleimide coupling reaction. The coupling efficiency reached 98%. The encapsulation efficiency of doxorubicin in liposomes was more than 98%. The flow cytometry test result showed that cRGD-modified liposomes (RGD-DXRL-PEG had higher cell uptake by U87MG cells, compared with nontargeted liposomes (DXRL-PEG. The cellular uptake was significantly inhibited in the presence of excess free cRGD. Both the targeted (t1/2 = 24.10 hours and non-targeted (t1/2 = 25.32 hours liposomes showed long circulating properties in rat plasma. The area under the curve of the targeted and nontargeted liposomes was 6.4-fold and 8.3-fold higher than that of doxorubicin solution, respectively.Conclusion: This study indicates preferential targeting and long circulating properties for cRGD-modified liposomes in vivo, which could be used as

  12. RGD peptide-displaying M13 bacteriophage/PLGA nanofibers as cell-adhesive matrices for smooth muscle cells

    Energy Technology Data Exchange (ETDEWEB)

    Shin, Yong Cheol; Lee, Jong Ho; Jin, Oh Seong; Lee, Eun Ji; Jin, Lin Hua; Kim, Chang Seok; Hong, Suck Won; Han, Dong Wook; Kim, Chun Tae; Oh, Jin Woo [Pusan National University, Busan (Korea, Republic of)

    2015-01-15

    Extracellular matrices (ECMs) are network structures that play an essential role in regulating cellular growth and differentiation. In this study, novel nanofibrous matrices were fabricated by electrospinning M13 bacteriophage and poly(lactic-co-glycolic acid) (PLGA) and were shown to be structurally and functionally similar to natural ECMs. A genetically-engineered M13 bacteriophage was constructed to display Arg-Gly-Asp (RGD) peptides on its surface. The physicochemical properties of RGD peptide-displaying M13 bacteriophage (RGD-M13 phage)/PLGA nanofibers were characterized by using scanning electron microscopy and Fourier-transform infrared spectroscopy. We used immunofluorescence staining to confirm that M13 bacteriophages were homogenously distributed in RGD-M13 phage/PLGA matrices. Furthermore, RGD-M13 phage/PLGA nanofibrous matrices, having excellent biocompatibility, can enhance the behaviors of vascular smooth muscle cells. This result suggests that RGD-M13 phage/PLGA nanofibrous matrices have potentials to serve as tissue engineering scaffolds.

  13. RGD-conjugated gold nanorods induce radiosensitization in melanoma cancer cells by downregulating αvβ3 expression

    Directory of Open Access Journals (Sweden)

    Pang B

    2012-02-01

    Full Text Available Wencai Xu1, Teng Luo2, Ping Li1, Chuanqing Zhou2, Daxiang Cui3, Bo Pang4, Qiushi Ren4, Shen Fu11Department of Radiation Oncology, Shanghai Sixth People's Hospital, 2School of Biomedical Engineering, and 3National Key Laboratory of Nano/Micro Fabrication Technology, Key Laboratory for Thin Film and Microfabrication of Ministry of Education, Institute of Micro-Nano Science and Technology, Shanghai Jiao Tong University, Shanghai, 4Department of Biomedical Engineering, College of Engineering, Peking University, Beijing, People's Republic of ChinaBackground: Melanoma is known to be radioresistant and traditional treatments have been intractable. Therefore, novel approaches are required to improve the therapeutic efficacy of melanoma treatment. In our study, gold nanorods conjugated with Arg-Gly-Asp peptides (RGD-GNRs were used as a sensitizer to enhance the response of melanoma cells to 6 mV radiation.Methods and materials: A375 melanoma cells were treated by gold nanorods or RGD-GNRs with or without irradiation. The antiproliferative impact of the treatments was measured by MTT assay. Radiosensitizing effects were determined by colony formation assay. Apoptosis and cell cycle data were measured by flow cytometry. Integrin αvβ3expression was also investigated by flow cytometry.Results: Addition of RGD-GNRs enhanced the radiosensitivity of A375 cells with a dose-modifying factor of 1.35, and enhanced radiation-induced apoptosis. DNA flow cytometric analysis indicated that RGD-GNRs plus irradiation induced significant G2/M phase arrest in A375 cells. Both spontaneous and radiation-induced expressions of integrin αvβ3 were downregulated by RGD-GNRs.Conclusion: Our study indicated that RGD-GNRs could sensitize melanoma A375 cells to irradiation. It was hypothesized that this was mainly through downregulation of radiation-induced αvβ3, in addition to induction of a higher proportion of cells within the G2/M phase. The combination of RGD-GNRs and

  14. In Vitro Bioactivity Study of RGD-Coated Titanium Alloy Prothesis for Revision Total Hip Arthroplasty

    Directory of Open Access Journals (Sweden)

    Zhentao Man

    2016-01-01

    Full Text Available Total hip arthroplasty (THA is a common procedure for the treatment of end-stage hip joint disease, and the demand for revision THA will double by 2026. Ti6Al4V (Titanium, 6% Aluminum, and 4% Vanadium is a kind of alloy commonly used to make hip prothesis. To promote the osseointegration between the prothesis and host bone is very important for the revision THA. The peptide Arg-Gly-Asp (RGD could increase cell attachment and has been used in the vascular tissue engineering. In this study, we combined the RGD with Ti6Al4V alloy using the covalent cross-linking method to fabricate the functional Ti6Al4V alloy (FTA. The distribution of RGD oligopeptide on the FTA was even and homogeneous. The FTA scaffolds could promote mouse osteoblasts adhesion and spreading. Furthermore, the result of RT-qPCR indicated that the FTA scaffolds were more beneficial to osteogenesis, which may be due to the improvement of osteoblast adhesion by the RGD oligopeptide coated on FTA. Overall, the FTA scaffolds developed herein pave the road for designing and building more efficient prothesis for osseointegration between the host bone and prothesis in revision THA.

  15. Preparation and characterization of a hetero functional system of gold nanoparticles labeled with 99mTc and conjugated to the sequence Arg-Gly-Asp for detection in vivo of angio genesis and evaluation of their toxicity in Hyalella aztec

    International Nuclear Information System (INIS)

    Integrin s play critical roles in many physiological processes including angio genesis and also contribute to pathological events such as tumor invasion and metastasis. The αvβ3 integrin is expressed in normal endothelial cells but it is over-expressed in the tumor neo vasculature. Peptides based on the Arginine-Glycine-Aspartic acid (RGD) sequence have been reported as molecules with high affinity and selectivity for the αvβ3 integrin. Recent studies have demonstrated that conjugating peptides to gold nanoparticles (AuNP) produces biocompatible and stable multifunctional systems with target-specific molecular recognition due to multivalent effects produced by multiple simultaneous interactions between peptides and their receptors. The first aim of this research was to prepare a m ultimeric system of 99mTc labeled gold particles conjugated to c[RGDfK(C)] and to evaluate its biological behavior as a potential radiopharmaceutical for molecular imaging of αvβ3 tumor expression. Hidrazinonicotinamide-G GC (HYNIC-G GC) and C[RGDfK(C)] peptides were synthesized and conjugated to AuNP (20 nm) by means of spontaneous reaction of the thiol groups of cysteine. The nano conjugate was characterized by transmission electron microscopy, Fourier transform-infrared, Ultraviolet-vis, X-ray photoelectron spectroscopy and Raman spectroscopy. To obtain 99mTc-HYNIC-G GC-AuNP-c[RGDfK(C)], the 99mTc-HYNIC-G GC radio peptide was first prepared and added to the AuNP solution followed by c[RGDfK(C)]. Radiochemical purity (Rp) was determined by size-exclusion HPLC and I TLC-Sg analyses. In vitro binding studies were carried out in αvβ3 receptor-positive C6 glioma cancer cells. Biodistribution studies were accomplished in athymic mice with C6-induced tumors with blocked and non blocked receptors, and images were obtained using a micro-SPECT/CT. Transmission electron microscopy and spectroscopy techniques demonstrated that AuNP were functionalized with peptides. Rp was 96 ± 2% without

  16. Non-peptidic analogs of the cell adhesion motif RGD prevent experimental liver injury.

    Science.gov (United States)

    Bruck, R; Hershkoviz, R; Lider, O; Shirin, H; Aeed, H; Halpern, Z

    2000-07-01

    In chronic viral hepatitis, autoimmune hepatitis, and some chronic cholestatic liver diseases, T lymphocytes serve as effector cells of the immunostimulatory processes. Cellular interactions of immune cells with extracellular matrix components are regulated primarily via the beta 1 subfamily of integrin receptors. The target epitope of several such integrin receptors is the Arg-Gly-Asp sequence, a cell adhesion motif shared by several matrix-associated adhesive glycoproteins. We review the use of synthetic non-peptidic analogs of RGD in the prevention of immune-mediated, concanavalin A-induced liver damage in mice and in inhibiting the development of liver cirrhosis in rats. The Con A-induced elevation of serum transaminases and tumor necrosis factor-alpha and the infiltration of liver tissue by inflammatory cells were inhibited by pretreatment of the mice with the synthetic RGD mimetics. In rats, the progression of thioacetamide-induced liver cirrhosis was markedly inhibited by the co-administration of the RGD mimetic SF-6,5. The compounds described here may be examined therapeutically for pathological conditions in the liver, manifested as necro-inflammation and fibrosis. PMID:10909422

  17. Molecular nuclear imaging of tumoral angio genesis using a rgd-containing tracer, Raft-RGD, targeted at the neo vessel-specific integrin αvβ3

    International Nuclear Information System (INIS)

    Tumoral neo-angio genesis targeting is currently a major field of research for the diagnostic and treatment of solid tumors. Endothelial cells from neo vessels over express several specific markers such as the αvβ3 integrin, which binds RGD (-Arg-Gly-Asp-)- containing peptides. We evaluated the potential of a novel radiotracer - RAFT-RGD - for the molecular nuclear imaging of neo vessels. In vitro, the coupling of 4 c(RGDfK) to the RAFT platform resulted in an increased cellular uptake of the tracer by αvβ3 positive cells when compared to c(RGDfK). Furthermore, RAFTRGD has a higher affinity than c(RGDfK) and similar properties for angio genesis inhibition. In vivo, both αvβ3 positive and negative tumors were visible by non invasive whole body planar and tomographic imaging from 30 min to 24 h post-injection, using a gamma camera dedicated to small animal imaging. Despite a lack of significant contrast improvement compare with c(RGDfK), RAFT-RGD could represent a promising tracer for tumoral angio genesis since it could provide invaluable information about tumor development and treatment efficacy in Nuclear Medicine departments. Furthermore, thanks to its chemical structure, RAFT-RGD can be labelled with a variety of radioisotopes including γ and β- emitters, allowing interesting therapeutical applications such as internal targeted radiotherapy. (author)

  18. Gold Nanorods Conjugated with Doxorubicin and cRGD for Combined Anticancer Drug Delivery and PET Imaging

    Directory of Open Access Journals (Sweden)

    Yuling Xiao, Hao Hong, Vyara Z. Matson, Alireza Javadi, Wenjin Xu, Yunan Yang, Yin Zhang, Jonathan W. Engle, Robert J. Nickles, Weibo Cai, Douglas A. Steeber, Shaoqin Gong

    2012-01-01

    Full Text Available A multifunctional gold nanorod (GNR-based nanoplatform for targeted anticancer drug delivery and positron emission tomography (PET imaging of tumors was developed and characterized. An anti-cancer drug (i.e., doxorubicin (DOX was covalently conjugated onto PEGylated (PEG: polyethylene glycol GNR nanocarriers via a hydrazone bond to achieve pH-sensitive controlled drug release. Tumor-targeting ligands (i.e., the cyclo(Arg-Gly-Asp-D-Phe-Cys peptides, cRGD and 64Cu-chelators (i.e., 1,4,7-triazacyclononane-N, N', N''-triacetic acid (NOTA were conjugated onto the distal ends of the PEG arms to achieve active tumor-targeting and PET imaging, respectively. Based on flow cytometry analysis, cRGD-conjugated nanocarriers (i.e., GNR-DOX-cRGD exhibited a higher cellular uptake and cytotoxicity than non-targeted ones (i.e., GNR-DOX in vitro. However, GNR-DOX-cRGD and GNR-DOX nanocarriers had similar in vivo biodistribution according to in vivo PET imaging and biodistribution studies. Due to the unique optical properties of GNRs, this multifunctional GNR-based nanoplatform can potentially be optimized for combined cancer therapies (chemotherapy and photothermal therapy and multimodality imaging (PET, optical, X-ray computed tomography (CT, etc..

  19. Combined Use of RGD-peptide Modified PLGA and TGF-β1 Gene Transfected MSCs to Improve Cell Biobehaviors in vitro

    Institute of Scientific and Technical Information of China (English)

    Changwen LI; Qixin ZHENG; Xiaodong GUO; Daping QUAN; Jie ZHAO

    2009-01-01

    In order to improve the surface properties of PLGA polymer for a better material/cell in-terface to modulate the cells behaviors, we prepared a novel three-block copolymer, PLGA-[ASP-PEG],and immobilized an RGD-containing peptide, Gly-Arg-Gly-Asp-Ser-Pro-Cys (GRGDSPC) on the sur-face of it. Transforming growth factor-131 (TGF-β1) was transfected into bone marrow stromal cells (MSCs) employed as seeded cells. Cell adhesion, spreading, proliferation and differentiation on this material were investigated. The results showed that the cell adhesive ratio on RGD-modified materials was higher than on un-modified materials (P<0.05). The extent of cell spreading was also wider on RGD-modified materials than on un-modified materials. Cell proliferation indices of transfected MSCs were increased as compared with the un-transfected MSCs (P<0.05). The ALP activities in the MSCs cultured with RGD-modified materials were higher than on un-modified materials after 14 days (P<0.05), and those in transfected MSCs were higher than in un-transfected MSCs (P<0.05). It was suggested that the combined use of RGD-modification and TGF-β gene transfection could improve the interaction of biomaterial and cells.

  20. Molecular nuclear imaging of tumoral angio genesis using a rgd-containing tracer, Raft-RGD, targeted at the neo vessel-specific integrin {alpha}{sub v}{beta}{sub 3}; Evaluation d'un radioligand de l'integrine {alpha}{sub v}{beta}{sub 3} (RAFT-RGD) pour l'imagerie moleculaire de l'angiogenese tumorale

    Energy Technology Data Exchange (ETDEWEB)

    Sancey, L

    2006-06-15

    Tumoral neo-angio genesis targeting is currently a major field of research for the diagnostic and treatment of solid tumors. Endothelial cells from neo vessels over express several specific markers such as the {alpha}{sub v}{beta}{sub 3} integrin, which binds RGD (-Arg-Gly-Asp-)- containing peptides. We evaluated the potential of a novel radiotracer - RAFT-RGD - for the molecular nuclear imaging of neo vessels. In vitro, the coupling of 4 c(RGDfK) to the RAFT platform resulted in an increased cellular uptake of the tracer by {alpha}{sub v}{beta}{sub 3} positive cells when compared to c(RGDfK). Furthermore, RAFTRGD has a higher affinity than c(RGDfK) and similar properties for angio genesis inhibition. In vivo, both {alpha}{sub v}{beta}{sub 3} positive and negative tumors were visible by non invasive whole body planar and tomographic imaging from 30 min to 24 h post-injection, using a gamma camera dedicated to small animal imaging. Despite a lack of significant contrast improvement compare with c(RGDfK), RAFT-RGD could represent a promising tracer for tumoral angio genesis since it could provide invaluable information about tumor development and treatment efficacy in Nuclear Medicine departments. Furthermore, thanks to its chemical structure, RAFT-RGD can be labelled with a variety of radioisotopes including {gamma} and {beta}{sup -} emitters, allowing interesting therapeutical applications such as internal targeted radiotherapy. (author)

  1. Radiolabelled of c-DOTA-RGD and c-DOTA-RGDf with 177Lu and evaluation in vitro and in vivo stability

    International Nuclear Information System (INIS)

    Integrin αvβ3 has a critical role in tumor angio genesis and metastasis. Radiolabelled peptides based on the Arg-Gly-Asp (RGD) sequence have been reported as radiopharmaceuticals with high affinity and selectivity for the αvβ3 integrin. The aim of this study was to label c-DOTA-RGD and c-DOTA-RGDf peptides with 177Lu and to evaluate their in vitro and in vivo stability as potential specific therapeutic radiopharmaceuticals. Labelled was carried out by direct reaction of 177LuCl3 with c-DOTA-RGD peptides in 1 M acetate buffer ph 5.5 at 90o C for 30 min. Radiochemical purity and stability studies were realized by reversed phase HPLC and I TLC-Sg analyses in human serum and saline solution. Biological recognition was performed using MCF7 tumor cells (positive αvβ3) and in athymic mice with induced MCF7 tumors. Molecular mechanics and quantum mechanics calculations were performed to explain experimental results associated with the molecular recognition. 177Lu-DOTA-RGD and 177Lu-DOTA-RGDf were obtained with radiochemical purities > 95%, showing adequate in vitro and in vivo stability and specific binding to □v□3 receptors. (Author)

  2. Fetal muscle gene transfer is not enhanced by an RGD capsid modification to high-capacity adenoviral vectors.

    Science.gov (United States)

    Bilbao, R; Reay, D P; Hughes, T; Biermann, V; Volpers, C; Goldberg, L; Bergelson, J; Kochanek, S; Clemens, P R

    2003-10-01

    High levels of alpha(v) integrin expression by fetal muscle suggested that vector re-targeting to integrins could enhance adenoviral vector-mediated transduction, thereby increasing safety and efficacy of muscle gene transfer in utero. High-capacity adenoviral (HC-Ad) vectors modified by an Arg-Gly-Asp (RGD) peptide motif in the HI loop of the adenoviral fiber (RGD-HC-Ad) have demonstrated efficient gene transfer through binding to alpha(v) integrins. To test integrin targeting of HC-Ad vectors for fetal muscle gene transfer, we compared unmodified and RGD-modified HC-Ad vectors. In vivo, unmodified HC-Ad vector transduced fetal mouse muscle with four-fold higher efficiency compared to RGD-HC-Ad vector. Confirming that the difference was due to muscle cell autonomous factors and not mechanical barriers, transduction of primary myogenic cells isolated from murine fetal muscle in vitro demonstrated a three-fold better transduction by HC-Ad vector than by RGD-HC-Ad vector. We hypothesized that the high expression level of coxsackievirus and adenovirus receptor (CAR), demonstrated in fetal muscle cells both in vitro and in vivo, was the crucial variable influencing the relative transduction efficiencies of HC-Ad and RGD-HC-Ad vectors. To explore this further, we studied transduction by HC-Ad and RGD-HC-Ad vectors in paired cell lines that expressed alpha(v) integrins and differed only by the presence or absence of CAR expression. The results increase our understanding of factors that will be important for retargeting HC-Ad vectors to enhance gene transfer to fetal muscle.

  3. Correlative Light-Electron Microscopy Shows RGD-Targeted ZnO Nanoparticles Dissolve in the Intracellular Environment of Triple Negative Breast Cancer Cells and Cause Apoptosis with Intratumor Heterogeneity

    KAUST Repository

    Othman, Basmah A.

    2016-04-01

    ZnO nanoparticles (NPs) are reported to show a high degree of cancer cell selectivity with potential use in cancer imaging and therapy. Questions remain about the mode by which the ZnO NPs cause cell death, whether they exert an intra- or extracellular effect, and the resistance among different cancer cell types to ZnO NP exposure. The present study quantifies the variability between the cellular toxicity, dynamics of cellular uptake, and dissolution of bare and RGD (Arg-Gly-Asp)-targeted ZnO NPs by MDA-MB-231 cells. Compared to bare ZnO NPs, RGD-targeting of the ZnO NPs to integrin αvβ3 receptors expressed on MDA-MB-231 cells appears to increase the toxicity of the ZnO NPs to breast cancer cells at lower doses. Confocal microscopy of live MDA-MB-231 cells confirms uptake of both classes of ZnO NPs with a commensurate rise in intracellular Zn2+ concentration prior to cell death. The response of the cells within the population to intracellular Zn2+ is highly heterogeneous. In addition, the results emphasize the utility of dynamic and quantitative imaging in understanding cell uptake and processing of targeted therapeutic ZnO NPs at the cellular level by heterogeneous cancer cell populations, which can be crucial for the development of optimized treatment strategies.

  4. Noninvasive imaging of tumor integrin expression using {sup 18}F-labeled RGD dimer peptide with PEG{sub 4} linkers

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Zhaofei [Stanford University School of Medicine, Molecular Imaging Program at Stanford (MIPS), Department of Radiology, Biophysics, and Bio-X Program, Stanford, CA (United States); Peking University, Medical Isotopes Research Center, Beijing (China); Liu, Shuanglong [Stanford University School of Medicine, Molecular Imaging Program at Stanford (MIPS), Department of Radiology, Biophysics, and Bio-X Program, Stanford, CA (United States); Wang, Fan [Peking University, Medical Isotopes Research Center, Beijing (China); Liu, Shuang [Purdue University, School of Health Sciences, West Lafayette, IN (United States); Chen, Xiaoyuan [Stanford University School of Medicine, Molecular Imaging Program at Stanford (MIPS), Department of Radiology, Biophysics, and Bio-X Program, Stanford, CA (United States); Stanford University School of Medicine, Molecular Imaging Program at Stanford (MIPS), Department of Radiology and Bio-X Program, Stanford, CA (United States)

    2009-08-15

    Various radiolabeled Arg-Gly-Asp (RGD) peptides have been previously investigated for tumor integrin {alpha}{sub v}{beta}{sub 3} imaging. To further develop RGD radiotracers with enhanced tumor-targeting efficacy and improved in vivo pharmacokinetics, we designed a new RGD homodimeric peptide with two PEG{sub 4} spacers (PEG{sub 4} = 15-amino-4,7,10,13-tetraoxapentadecanoic acid) between the two monomeric RGD motifs and one PEG{sub 4} linker on the glutamate {alpha}-amino group ({sup 18}F-labeled PEG{sub 4}-E[PEG{sub 4}-c(RGDfK)]{sub 2}, P-PRGD2), as a promising agent for noninvasive imaging of integrin expression in mouse models. P-PRGD2 was labeled with {sup 18}F via 4-nitrophenyl 2-{sup 18}F-fluoropropionate ({sup 18}F-FP) prosthetic group. In vitro and in vivo characteristics of the new dimeric RGD peptide tracer {sup 18}F-FP-P-PRGD2 were investigated and compared with those of {sup 18}F-FP-P-RGD2 ({sup 18}F-labeled RGD dimer without two PEG{sub 4} spacers between the two RGD motifs). The ability of {sup 18}F-FP-P-PRGD2 to image tumor vascular integrin expression was evaluated in a 4T1 murine breast tumor model. With the insertion of two PEG{sub 4} spacers between the two RGD motifs, {sup 18}F-FP-P-PRGD2 showed enhanced integrin {alpha}{sub v}{beta}{sub 3}-binding affinity, increased tumor uptake and tumor-to-nontumor background ratios compared with {sup 18}F-FP-P-RGD2 in U87MG tumors. MicroPET imaging with {sup 18}F-FP-P-PRGD2 revealed high tumor contrast and low background in tumor-bearing nude mice. Biodistribution studies confirmed the in vivo integrin {alpha}{sub v}{beta}{sub 3}-binding specificity of {sup 18}F-FP-P-RGD2. {sup 18}F-FP-P-PRGD2 can specifically image integrin {alpha}{sub v}{beta}{sub 3} on the activated endothelial cells of tumor neovasculature. {sup 18}F-FP-P-PRGD2 can provide important information on integrin expression on the tumor vasculature. The high integrin binding affinity and specificity, excellent pharmacokinetic properties and

  5. Human Bone Marrow Mesenchymal Stem Cell Behaviors on PCL/Gelatin Nanofibrous Scaffolds Modified with A Collagen IV-Derived RGD-Containing Peptide

    Directory of Open Access Journals (Sweden)

    Ali Mota

    2014-03-01

    Full Text Available Objective: We introduce an RGD (Arg-Gly-Asp-containing peptide of collagen IV origin that possesses potent cell adhesion and proliferation properties. Materials and Methods: In this experimental study, the peptide was immobilized on an electrospun nanofibrous polycaprolactone/gelatin (PCL/Gel hybrid scaffold by a chemical bonding approach to improve cell adhesion properties of the scaffold. An iodine-modified phenylalanine was introduced in the peptide to track the immobilization process. Native and modified scaffolds were characterized with scanning electron microscopy (SEM and fourier transform infrared spectroscopy (FTIR. We studied the osteogenic and adipogenic differentiation potential of human bone marrow-derived mesenchymal stem cells (hBMSCs. In addition, cell adhesion and proliferation behaviors of hBMSCs on native and peptide modified scaffolds were evaluated by 3-(4,5-dimethylthiazol-2-yl-2,5-diphenyltetrazolium bromide (MTT assay and 4',6-diamidino-2-phenylindole (DAPI staining, and the results compared with tissue culture plate, as the control. Results: FTIR results showed that the peptide successfully immobilized on the scaffold. MTT assay and DAPI staining results indicated that peptide immobilization had a dramatic effect on cell adhesion and proliferation. Conclusion: This peptide modified nanofibrous scaffold can be a promising biomaterial for tissue engineering and regenerative medicine with the use of hBMSCs.

  6. Specific targeting of angiogenesis in lung cancer with RGD-conjugated ultrasmall superparamagnetic iron oxide particles using a 4.7T magnetic resonance scanner

    Institute of Scientific and Technical Information of China (English)

    LIU Can; LIU Dong-bo; LONG Guo-xian; WANG Jun-feng; MEI Qi; HU Guang-yuan; QIU Hong

    2013-01-01

    Background Angiogenesis is an essential step for tumor development and metastasis.The cell adhesion molecule αvβ3 integrin plays an important role in angiogenesis and is a specific marker of tumor angiogenesis.A novel αvβ3 integrintargeted magnetic resonance (MR) imaging contrast agent utilizing Arg-Gly-Asp (RGD) and ultrasmall superparamagnetic iron oxide particles (USPIO) (referred to as RGD-USPIO) was designed and its uptake by endothelial cells was assessed both in vitro and in vivo to evaluate the angiogenic profile of lung cancer.Methods USPIO were coated with-NH3+ and conjugated with RGD peptides.Prussian blue staining was performed to evaluate the specific uptake of RGD-USPIO by human umbilical vein endothelial cells (HUVECs).Targeted uptake and subcellular localization of RGD-USPIO in HUVECs were confirmed by transmission electron microscopy (TEM).The ability of RGD-USPIO to noninvasively assess αvβ3 integrin positive vessels in lung adenocarcinoma A549 tumor xenografts was evaluated with a 4.7T MR scanner.Immunohistochemistry was used to detect αvβ3 integrin expression and vessel distribution in A549 tumor xenografts.Results HUVECs internalized RGD-USPIO significantly more than plain USPIO.The uptake of RGD-USPIO by HUVECs could be competitively inhibited by addition of free RGD.A significant decrease in T2 signal intensity (SI) was observed at the periphery of A549 tumor xenografts at 30 minutes (P <0.05) and 2 hours (P <0.01) after RGD-USPIO was injected via the tail vein.Angiogenic blood vessels were mainly distributed in the periphery of tumor xenografts with positive αvβ3 integrin expression.Conclusions RGD-USPIO could specifically label αvβ3 integrin and be taken up by HUVECs.This molecular MR imaging contrast agent can specifically evaluate the angiogenic profile of lung cancer using a 4.7T MR scanner.

  7. RGD-modified poly(D,L-lactic acid nanoparticles enhance tumor targeting of oridonin

    Directory of Open Access Journals (Sweden)

    Xu J

    2012-01-01

    Full Text Available Jie Xu, Ji-Hui Zhao, Ying Liu, Nian-Ping Feng, Yong-Tai ZhangSchool of Pharmacy, Shanghai University of Traditional Chinese Medicine, Shanghai, People's Republic of ChinaObjective: The purpose of this study was to develop an active targeting strategy to improve the therapeutic antitumor efficacy of oridonin (ORI, the main active ingredient in the medicinal herb Rabdosia rubescens.Methods: A modified spontaneous emulsification solvent diffusion method was used to prepare the ORI-loaded atactic poly(D,L-lactic acid nanoparticles (ORI-PLA-NPs. Surface cross-linking with the peptide Arg-Gly-Asp (RGD further modified the ORI-PLA-NPs, generating ORI-PLA-RGD-NPs. The NPs were characterized and release experiments were performed in vitro. The pharmacokinetics, tissue distribution, and antitumor activity of the NPs were studied in mice bearing hepatocarcinoma 22 (H22-derived tumors.Results: The ORI-PLA-NPs and ORI-PLA-RGD-NPs were smooth, sphere-like, and relatively uniform in size. The RGD surface modification slightly increased the mean particle size (95.8 nm for ORI-PLA-NPs versus 105.2 nm for ORI-PLA-RGD-NPs and considerably altered the surface electrical property (-10.19 mV for ORI-PLA-NPs versus -21.95 mV for ORI-PLA-RGD-NPs, but it had no obvious influence on ORI loading (8.23% ± 0.35% for ORI-PLA-NPs versus 8.02% ± 0.38% for ORI-PLA-RGD-NPs, entrapment efficiency (28.86% ± 0.93% for ORI-PLA-NPs versus 28.24% ± 0.81% for ORI-PLA-RGD-NPs, or the release of ORI. The pharmacokinetic properties of free ORI were improved by encapsulation in NPs, as shown by increased area under the concentration-time curve (11.89 ± 0.35 µg·mL-1 · h for ORI solution versus 22.03 ± 0.01 µg · mL-1 · h for ORI-PLA-RGD-NPs and prolonged mean retention time (2.03 ± 0.09 hours for ORI solution versus 8.68 ± 0.66 hours for ORI-PLA-RGD-NPs. In the tissue distribution study, more ORI targeted tumor tissue in the mice treated with ORI-PLA-RGD-NPs than with ORI

  8. In-vitro and in-vivo phenotype of type Asia 1 foot-and-mouth disease viruses utilizing two non-RGD receptor recognition sites

    Directory of Open Access Journals (Sweden)

    Yin Hong

    2011-06-01

    Full Text Available Abstract Background Foot-and-mouth disease virus (FMDV uses a highly conserved Arg-Gly-Asp (RGD triplet for attachment to host cells and this motif is believed to be essential for virus viability. Previous sequence analyses of the 1D-encoding region of an FMDV field isolate (Asia1/JS/CHA/05 and its two derivatives indicated that two viruses, which contained an Arg-Asp-Asp (RDD or an Arg-Ser-Asp (RSD triplet instead of the RGD integrin recognition motif, were generated serendipitously upon short-term evolution of field isolate in different biological environments. To examine the influence of single amino acid substitutions in the receptor binding site of the RDD-containing FMD viral genome on virus viability and the ability of non-RGD FMDVs to cause disease in susceptible animals, we constructed an RDD-containing FMDV full-length cDNA clone and derived mutant molecules with RGD or RSD receptor recognition motifs. Following transfection of BSR cells with the full-length genome plasmids, the genetically engineered viruses were examined for their infectious potential in cell culture and susceptible animals. Results Amino acid sequence analysis of the 1D-coding region of different derivatives derived from the Asia1/JS/CHA/05 field isolate revealed that the RDD mutants became dominant or achieved population equilibrium with coexistence of the RGD and RSD subpopulations at an early phase of type Asia1 FMDV quasispecies evolution. Furthermore, the RDD and RSD sequences remained genetically stable for at least 20 passages. Using reverse genetics, the RDD-, RSD-, and RGD-containing FMD viruses were rescued from full-length cDNA clones, and single amino acid substitution in RDD-containing FMD viral genome did not affect virus viability. The genetically engineered viruses replicated stably in BHK-21 cells and had similar growth properties to the parental virus. The RDD parental virus and two non-RGD recombinant viruses were virulent to pigs and bovines that

  9. Monomeric, dimeric and multimeric system of RGD peptides radiolabeled with 177Lu for tumors therapy that expressing αβ integrin s

    International Nuclear Information System (INIS)

    The conjugation of peptides to gold nanoparticles (AuNPs) produces biocompatible and stable multimeric systems with target-specific molecular recognition. Peptides based on the cyclic Arg-Gly-Asp (RGD) sequence have been reported as high affinity agents for the α(v)β(3) and α(v)β(5) integrin. The aim of this research was to prepare a multimeric system of 177Lu-labeled gold nanoparticles conjugated to c[RGDfK(C)] [cyclo(Arg-Gly-Asp-Phe-Lys(Cys)] peptides and to compare the radiation absorbed dose with that of 177Lu-labeled monomeric and dimeric RGD peptides to α(v)β(3) integrin-positive U87MG tumors in mice, as well as, evaluate the in vitro potential 177Lu-AuNP-c[RGDfK(C)] as a plasmonic photothermal therapy and targeted radiotherapy system in MCF7 breast cancer cells. DOTA-GGC (1,4,7,10-tetraaza cyclododecane-N,N,N-tetraacetic-Gly-Gly-Cys) and c[RGDfK(C)] peptides were synthesized and conjugated to AuNPs by the spontaneous reaction of the thiol groups. Tem, UV-Vis, XP S, Raman and Far-IR spectroscopy techniques demonstrated that AuNPs were functionalized with the peptides. To obtain 177Lu-AuNP-c[RGDfK(C)], the 177Lu-DOTA-GGC radio peptide was first prepared and added to a solution of AuNPs followed by c[RGDfK(C)] (25 μL, 5 μM) at 18 grades C for 15 min. 177Lu-DOTA-GGC, 177Lu- DOTA-cRGDfK and 177Lu-DOTA-E-c(RGDfK)2 were prepared by adding 177LuCl3 (370 MBq) to 5 μL (1 mg/ml) of the DOTA derivative diluted with 50 μL of 1 M acetate buffer at ph 5. The mixture was incubated at 90 grades C in a block heater for 30 min. Radiochemical purity was determined by ultrafiltration and HPLC analyses. After laser irradiation, the presence of c[RGDfK(C)]-AuNP in cells caused a significant increase in the temperature of the medium (50.5 grades C, compared to 40.3 grades C without AuNPs) resulting in a significant decrease in MCF7 cell viability down to 9 %. After treatment with 177Lu-AuNP-c[RGDfK(C)], the MCF7 cell proliferation was significantly inhibited. Biokinetic

  10. Monomeric, dimeric and multimeric system of RGD peptides radiolabeled with {sup 177}Lu for tumors therapy that expressing αβ integrin s; Sistema monomerico, dimerico y multimerico de peptidos de RGD radiomarcados con {sup 177}Lu para terapia de tumores que expresan integrinas αβ

    Energy Technology Data Exchange (ETDEWEB)

    Luna G, M. A.

    2014-07-01

    The conjugation of peptides to gold nanoparticles (AuNPs) produces biocompatible and stable multimeric systems with target-specific molecular recognition. Peptides based on the cyclic Arg-Gly-Asp (RGD) sequence have been reported as high affinity agents for the α(v)β(3) and α(v)β(5) integrin. The aim of this research was to prepare a multimeric system of {sup 177}Lu-labeled gold nanoparticles conjugated to c[RGDfK(C)] [cyclo(Arg-Gly-Asp-Phe-Lys(Cys)] peptides and to compare the radiation absorbed dose with that of {sup 177}Lu-labeled monomeric and dimeric RGD peptides to α(v)β(3) integrin-positive U87MG tumors in mice, as well as, evaluate the in vitro potential {sup 177}Lu-AuNP-c[RGDfK(C)] as a plasmonic photothermal therapy and targeted radiotherapy system in MCF7 breast cancer cells. DOTA-GGC (1,4,7,10-tetraaza cyclododecane-N,N,N-tetraacetic-Gly-Gly-Cys) and c[RGDfK(C)] peptides were synthesized and conjugated to AuNPs by the spontaneous reaction of the thiol groups. Tem, UV-Vis, XP S, Raman and Far-IR spectroscopy techniques demonstrated that AuNPs were functionalized with the peptides. To obtain {sup 177}Lu-AuNP-c[RGDfK(C)], the {sup 177}Lu-DOTA-GGC radio peptide was first prepared and added to a solution of AuNPs followed by c[RGDfK(C)] (25 μL, 5 μM) at 18 grades C for 15 min. {sup 177}Lu-DOTA-GGC, {sup 177}Lu- DOTA-cRGDfK and {sup 177}Lu-DOTA-E-c(RGDfK){sub 2} were prepared by adding {sup 177}LuCl{sub 3} (370 MBq) to 5 μL (1 mg/ml) of the DOTA derivative diluted with 50 μL of 1 M acetate buffer at ph 5. The mixture was incubated at 90 grades C in a block heater for 30 min. Radiochemical purity was determined by ultrafiltration and HPLC analyses. After laser irradiation, the presence of c[RGDfK(C)]-AuNP in cells caused a significant increase in the temperature of the medium (50.5 grades C, compared to 40.3 grades C without AuNPs) resulting in a significant decrease in MCF7 cell viability down to 9 %. After treatment with {sup 177}Lu

  11. RGD肽介导的脑胶质瘤靶向治疗和显像的研究进展%Progress in RGD Peptide-mediated Targeted Therapy and Imaging for Glioma

    Institute of Scientific and Technical Information of China (English)

    陈中亚; 赵雁; 陶涛

    2013-01-01

    RGD peptides are small peptides containing arginine-glycine-aspartic acid triple peptide motif and can bind specifically to integrin receptors on cell surface. Integrin receptors, especially αvβ3 are highly expressed on tumor cell (glioma for example) surface while showing low expression on mature endothelial cells. Therefore, in the studies of targeted therapy and imaging for glioma, the competitive binding between ectogenic RGD peptides and integrin receptors on the tumor cell surface has been widely studied. In this paper, the typical methods and progress in RGD peptide-mediated targeted therapy and imaging for glioma in recent years are reviewed.%RGD肽是含有精氨酸-甘氨酸-门冬氨酸(Arg-Gly-Asp)序列的一类短肽,能和细胞表面的整合素受体特异性结合.整合素受体,尤其是αvβ3高表达于脑胶质瘤等肿瘤细胞表面,而在成熟血管内皮细胞呈低表达.因此,在脑胶质瘤的靶向治疗和显像研究中外源性RGD肽与肿瘤细胞表面的整合素受体的竞争性结合得到广泛研究.本文综述了RGD肽介导的脑胶质瘤靶向治疗及显像的典型方法及近年来的研究进展.

  12. Differential diagnosis of solitary pulmonary nodules using {sup 99m}Tc-3P{sub 4}-RGD{sub 2} scintigraphy

    Energy Technology Data Exchange (ETDEWEB)

    Ma, Qingjie; Ji, Bin; Gao, Shi; Ji, Tiefeng; Zhao, Guoqing [China-Japan Union Hospital of Jilin University, Department of Nuclear Medicine, Changchun (China); Jia, Bing [Peking University, Medical Isotopes Research Center, Beijing (China); Wang, Xueju [China-Japan Union Hospital of Jilin University, Department of Pathology, Changchun (China); Han, Zhenguo [China-Japan Union Hospital of Jilin University, Department of Thoracic Surgery, Changchun (China)

    2011-12-15

    Targeting of integrin {alpha}{sub {nu}}{beta}{sub 3} with molecular imaging agents offers great potential in early detection and monitoring of tumour angiogenesis. Recently, an RGD (Arg-Gly-Asp) tracer, {sup 99m}Tc-3P{sub 4}-RGD{sub 2}, with high affinity to integrin {alpha}{sub {nu}}{beta}{sub 3} and in vivo tumour uptake was developed. In this study, we evaluate the feasibility of this novel radiotracer in the noninvasive differentiation of solitary pulmonary nodules (SPNs). Twenty-one patients with SPNs on CT were studied scintigraphically after administration of {sup 99m}Tc-3P{sub 4}-RGD{sub 2} with a dose of 939 {+-} 118 MBq. Image interpretation using a 5-point scale was performed by one thoracic radiologist for CT and three nuclear medicine radiologists for single photon emission computed tomography (SPECT). Scintigraphic images were also analysed semiquantitatively by calculating tumour to normal tissue ratio (T/N). The ''gold standard'' was based on the histopathological diagnosis of the surgical samples from all recruited patients. A fraction of the samples were analysed immunohistochemically for integrin {alpha}{sub v}{beta}{sub 3} expression. Among the 21 SPNs, 15 (71%) were diagnosed as malignant and 6 (29%) were benign. The mean size for SPNs was 2.2 {+-} 0.6 cm. The sensitivity and specificity for CT interpretation, SPECT visual and semiquantitative analysis were 80/67%, 100/67% and 100/67%, respectively. All SPNs classified as indeterminate by CT were correctly diagnosed by {sup 99m}Tc-3P{sub 4}-RGD{sub 2} scintigraphy. The empirical receiver-operating characteristic (ROC) areas were 0.811 [95% confidence interval (CI) 58-95%] for CT, 0.833 (95% CI 61-96%) for SPECT and 0.844 (95% CI 62-96%) for T/N ratios, respectively. Immunohistochemistry confirmed {alpha}{sub {nu}}{beta}{sub 3} expression in malignant and benign nodules with uptake in {sup 99m}Tc-3P{sub 4}-RGD{sub 2} scintigraphy. In this first-in-human study, we demonstrated

  13. RGD(F/S/V-Dex: towards the development of novel, effective, and safe glucocorticoids

    Directory of Open Access Journals (Sweden)

    Jiang X

    2016-03-01

    Full Text Available Xueyun Jiang,1 Ming Zhao,1,2 Yuji Wang,1 Haimei Zhu,1 Shurui Zhao,1 Jianhui Wu,1 Yuanbo Song,3 Shiqi Peng1 1Beijing Area Major Laboratory of Peptide and Small Molecular Drugs, Engineering Research Center of Endogenous Prophylactic of Ministry of Education of China, Beijing Laboratory of Biomedical Materials, College of Pharmaceutical Sciences, Capital Medical University, Beijing, People’s Republic of China; 2Faculty of Biomedical Science and Environmental Biology, Kaohsiung Medical University, Kaohsiung, Taiwan; 3Guangxi Pusen Biotechnology Co. Ltd., Nanning, Guangxi, People’s Republic of China Abstract: Dexamethasone (Dex is an effective glucocorticoid in treating inflammation and preventing rejection reaction. However, the side effects limit its clinical application. To improve its druggable profile, the conjugates of RGD-peptide-modified Dex were presented and their enhanced anti-inflammation activity, minimized osteoporotic action, and nanoscaled assembly were explored. (RGD stands for Arg-Gly-Asp. Standard single letter biochemical abbreviations for amino acids have been used throughout this paper. In respect of the rejection reaction, the survival time of the implanted myocardium of the mice treated with 1.43 µmol/kg/d of the conjugates for 15 consecutive days was significantly longer than that of the mice treated with 2.5 µmol/kg/d of Dex, and the conjugates, but not Dex, exhibited no toxic action. At a single dose of 14.3 µmol/kg (100 times minimal effective dose, 0.143 µmol/kg, the conjugates induced no liver, kidney, or systemic toxicity. At the dose of 1.43 µmol/kg, the conjugates, but not Dex, prolonged the bleeding time of the mice, and inhibited the thrombosis of the rats. In water and rat plasma, the conjugates formed nanoparticles of 14–250 and 101–166 nm in diameter, respectively. Since the nanoparticles of ~100 nm in size cannot be entrapped by macrophages in the circulation, RGDF-Dex would particularly be worthy

  14. Libraries of RGD analogs, labeled through ReO3+ or TcO3+ coordination, targeting αVβ3 integrin: development of tracers for the early detection of tumor neo-angiogenesis

    International Nuclear Information System (INIS)

    Integrins form a family of hetero-dimeric integral glycoproteins which play a central role in cell-cell adhesion and cell-matrix interactions. In particular, they are over expressed during tumor neo-angiogenesis. About 10 of them recognize a structured RGD (Arg-Gly-Asp) sequence. Analogs of this sequence can be used for the early detection of tumors and metastases. We developed new tracers, labeled with 99mTc, for the molecular imaging of αVβ3 integrin. Until recently, there was no reliable ab initio structure prediction of complex molecules containing Re and Tc chelates. Therefore, we preferred a combinatorial approach to develop potential ligands of αVβ3 integrin and we attempted to identify efficient tracers by in vivo screening. This method would account for biodistribution and pharmacokinetics properties in the early steps of the study. Tracers were obtained according two strategies: i) cyclization of linear RGD analogs; ii) combinatorial assembling of independent modules through metal core coordination by the well-known NS2+S motif. After synthesis and labeling, the stability of the tracers was investigated in presence of glutathione and in murine plasma. In vitro screening on purified integrin showed that a cyclic rhenium coordinate binds specifically αVβ3. A tumor model (U87-MG tumor on nude mice) was validated in the laboratory and a method was developed to analyze in vivo experiments. Biodistribution data and percentage of activity found in tumors are encouraging for cyclic compounds though identification of efficient tracers is difficult due to their instability in the conditions of analyses. (author)

  15. An RGD helper sequence in CagL of Helicobacter pylori assists in interactions with integrins and injection of CagA

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    Jens eConradi

    2012-05-01

    Full Text Available Helicobacter pylori is a specific gastric pathogen that colonizes the stomach in more than 50% of the world’s human population. Infection with this bacterium can induce several types of gastric pathology, ranging from chronic gastritis to peptic ulcers and even adenocarcinoma. Virulent H. pylori isolates encode components of a type IV secretion system (T4SS, which form a pilus for the injection of virulence proteins such as CagA into host target cells. This is accomplished by a specialized adhesin on the pilus surface, the protein CagL, a putative VirB5 ortholog, which binds to host cell β1 integrin, triggering subsequent delivery of CagA across the host cell membrane. Like the human extracellular matrix protein fibronectin, CagL contains an RGD (Arg-Gly-Asp motif and is able to trigger intracellular signaling pathways by RGD-dependent binding to integrins. While CagL binding to host cells is mediated primarily by the RGD motif, we identified an auxiliary binding motif for CagL-integrin interaction. Here, we report on a surface-exposed FEANE (Phe-Glu-Ala-Asn-Glu interaction motif in spatial proximity to the RGD sequence, which enhances the interactions of CagL with integrins. It will be referred to as RGD helper sequence (RHS. Competitive cell adhesion assays with recombinant wild type CagL and point mutants, competition experiments with synthetic cyclic and linear peptides, and peptide array experiments revealed amino acids essential for the interaction of the RHS motif with integrins. Infection experiments indicate that the RHS motif plays a role in the early interaction of H. pylori T4SS with integrin, to trigger signaling and to inject CagA into host cells. We thus postulate that CagL is a versatile T4SS surface protein equipped with at least two motifs to promote binding to integrins, thereby causing aberrant signaling within host cells and facilitating translocation of CagA into host cells, thus contributing directly to H. pylori

  16. RGD-tagged helical rosette nanotubes aggravate acute lipopolysaccharide-induced lung inflammation

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    Suri SS

    2011-12-01

    Full Text Available Sarabjeet Singh Suri1, Steven Mills1, Gurpreet Kaur Aulakh1, Felaniaina Rakotondradany2, Hicham Fenniri2, Baljit Singh11Department of Veterinary Biomedical Sciences, University of Saskatchewan, Saskatoon; 2National Institute for Nanotechnology and Department of Chemistry, Edmonton, CanadaAbstract: Rosette nanotubes (RNT are a novel class of self-assembled biocompatible nanotubes that offer a built-in strategy for engineering structure and function through covalent tagging of synthetic self-assembling modules (G∧C motif. In this report, the G∧C motif was tagged with peptide Arg-Gly-Asp-Ser-Lys (RGDSK-G∧C and amino acid Lys (K-G∧C which, upon co-assembly, generate RNTs featuring RGDSK and K on their surface in predefined molar ratios. These hybrid RNTs, referred to as Kx/RGDSKy-RNT, where x and y refer to the molar ratios of K-G∧C and RGDSK–G∧C, were designed to target neutrophil integrins. A mouse model was used to investigate the effects of intravenous Kx/RGDSKy-RNT on acute lipopolysaccharide (LPS-induced lung inflammation. Healthy male C57BL/6 mice were treated intranasally with Escherichia coli LPS 80 µg and/or intravenously with K90/RGDSK10-RNT. Here we provide the first evidence that intravenous administration of K90/RGDSK10-RNT aggravates the proinflammatory effect of LPS in the mouse. LPS and K90/RGDSK10-RNT treatment groups showed significantly increased infiltration of polymorphonuclear cells in bronchoalveolar lavage fluid at all time points compared with the saline control. The combined effect of LPS and K90/RGDSK10-RNT was more pronounced than LPS alone, as shown by a significant increase in the expression of interleukin-1ß, MCP-1, MIP-1, and KC-1 in the bronchoalveolar lavage fluid and myeloperoxidase activity in the lung tissues. We conclude that K90/RGDSK10-RNT promotes acute lung inflammation, and when used along with LPS, leads to exaggerated immune response in the lung.Keywords: RGD peptide, helical rosette

  17. Libraries of RGD analogs, labeled through ReO{sup 3+} or TcO{sup 3+} coordination, targeting {alpha}V{beta}3 integrin: development of tracers for the early detection of tumor neo-angiogenesis; Chimiotheques de complexes du technetium et du rhenium ciblant l'integrine {alpha}V{beta}3: developpement de traceurs pour la detection precoce de la neoangiogenese tumorale

    Energy Technology Data Exchange (ETDEWEB)

    Aufort, M.

    2008-11-15

    Integrins form a family of hetero-dimeric integral glycoproteins which play a central role in cell-cell adhesion and cell-matrix interactions. In particular, they are over expressed during tumor neo-angiogenesis. About 10 of them recognize a structured RGD (Arg-Gly-Asp) sequence. Analogs of this sequence can be used for the early detection of tumors and metastases. We developed new tracers, labeled with {sup 99m}Tc, for the molecular imaging of {alpha}{sub V{beta}3} integrin. Until recently, there was no reliable ab initio structure prediction of complex molecules containing Re and Tc chelates. Therefore, we preferred a combinatorial approach to develop potential ligands of {alpha}{sub V{beta}3} integrin and we attempted to identify efficient tracers by in vivo screening. This method would account for biodistribution and pharmacokinetics properties in the early steps of the study. Tracers were obtained according two strategies: i) cyclization of linear RGD analogs; ii) combinatorial assembling of independent modules through metal core coordination by the well-known NS{sub 2}+S motif. After synthesis and labeling, the stability of the tracers was investigated in presence of glutathione and in murine plasma. In vitro screening on purified integrin showed that a cyclic rhenium coordinate binds specifically {alpha}{sub V{beta}3}. A tumor model (U87-MG tumor on nude mice) was validated in the laboratory and a method was developed to analyze in vivo experiments. Biodistribution data and percentage of activity found in tumors are encouraging for cyclic compounds though identification of efficient tracers is difficult due to their instability in the conditions of analyses. (author)

  18. Design and Activity Determination of Cyclic RGD Peptide and Preparation of 99Tcm Labeled Cyclic RGD Dimer%RGD多肽类药物设计、活性测定及99Tcm-cRGD二聚体的制备

    Institute of Scientific and Technical Information of China (English)

    张丽; 张春丽; 王荣福; 闫平; 康磊; 郭凤琴; 魏海亮; 崔永刚; 卢霞

    2011-01-01

    通过V-life计算机模拟软件建立cRGD(cyclic Arg-Gly-Asp,cRGD)多肽类分子库,利用V-life软件中的DOCK功能对分子库内cRGD肽结构进行筛选评分,挑选出能与整合素αvβ3受体高特异性结合的cRGD 结构.将该结构进行改造后制备成二聚体,用99Tcm对该结构进行标记,制备成肿瘤分子探针.并对其标记条件、稳定性、水溶性和亲和力进行评价.结果表明,DOCK功能计算出评分最佳的cRGD分子结构为Cys-Arg-Gly-Asp-(D)Ser-Cys.将该结构进行改造制备成二聚体后,室温下、ρ(SnCl2·2H2O)=1 g/L、反应时间为30 min时,标记率可达(87.42±3.21)%,经Sephadex G10层析柱纯化后,其放化纯大于95%;在室温和37℃条件下,99Tcm-cRGD于生理盐水和正常人新鲜血清中均保持良好的标记稳定性;其脂水分配系数对数值lg P(正辛醇/生理盐水)为-1.96±0.01;与U87人神经胶质瘤细胞进行受体的放射性配基结合分析(radioligand binding assay of receptors,RBA)实验,其平衡解离常数(equilibrium dissociation constant,Kd)为(0.089±0.052)×10-9 L/mol.这表明,通过计算机模拟系统筛选出的cRGD肽可与整合素αvβ3特异性结合,是一种有前景的整合素αvβ3受体阳性肿瘤显像剂.%This paper was to design a cyclic RGD peptide tumor inhibitor with high affinity to integrin αvβ3 receptor by molecular docking technique. cRGD molecular library was built and an optimal structure of cRGD peptide with the lowest score that was Cys-Arg-Gly-Asp-(D)Ser-Cy s was screened out using the function of DOCK procedure of the V-life software. Based on the moiety a dimer linked by Tyr-(D)Ser-Lys-(D)Ser-Ser and with a side chain Gly-Gly-(D)Ala-Gly on Lys was synthesisized and 99Tcm-cRGD dimer was prepared. Its radiolabeled efficiency,stability, water-soluble and affinity in vitro were evaluated. Under the reaction condition of room temperature, 1 g/L SnCl2 · 2H2O and the 30 min of reaction time, labeling efficiency reachs (87

  19. Examination of soluble integrin resistant mutants of foot-and-mouth disease virus (FMDV)

    Science.gov (United States)

    Foot-and-mouth disease virus (FMDV) initiates infection in vitro via recognition of at least four cell-surface integrin molecules avb1, avb3, avb6 or avb8 through the interaction of a highly conserved Arg-Gly-Asp (RGD) amino acid sequence motif located in the GH loop of VP1. In this work, soluble i...

  20. cDNA cloning, mRNA distribution and heterogeneity, chromosomal location, and RFLP analysis of human osteopontin (OPN)

    DEFF Research Database (Denmark)

    Young, M F; Kerr, J M; Termine, J D;

    1990-01-01

    of the Arg-Gly-Asp (RGD) cell attachment site. Chromosomal mapping of the osteopontin gene (OPN) using human-rodent cell hybrids demonstrated a location on chromosome 4 in the human genome. In situ hybridization of metaphase chromosomes using radiolabeled OP1a as a probe indicated that the gene is located...

  1. The practicality of nanoceria-PAN-based (68)Ge/(68)Ga generator toward preparation of (68)Ga-labeled cyclic RGD dimer as a potential PET radiotracer for tumor imaging.

    Science.gov (United States)

    Chakraborty, Sudipta; Chakravarty, Rubel; Sarma, Haladhar D; Dash, Ashutosh; Pillai, M R A

    2013-02-01

    Cyclic RGD (Arg-Gly-Asp) peptides radiolabeled with (68)Ga have great potential for the early tumor detection and noninvasive monitoring of tumor metastasis and therapeutic response. Herein, the preparation of (68)Ga-labeled DOTA-E[c(RGDfK)](2) (DOTA=1,4,7,10-tetraazacylododecane-1,4,7,10-tetracetic acid; E=Glutamic acid; R=Arginine; G=Glycine; D=Aspartic acid; f=phenyl alanine; K=lysine) using (68)Ga directly eluted from a nanoceria-polyacrylonitrile (CeO(2)-PAN)-based (68)Ge/(68)Ga generator developed in-house was reported. The (68)Ga complex of DOTA-E[c(RGDfK)](2) was synthesized with >98% radiochemical purity by incubating 20 μg of the conjugate with (68)GaCl(3) (74-111 MBq) in acetate buffer (pH 3.5-4.0) at 90°C for 10 minutes. The complex exhibited excellent in vitro stability in 0.1 M EDTA solution at room temperature upto 1 hour studied (radiochemical purity: 98.0%). The biological efficacy of the radiolabeled conjugate was studied in C57/BL6 mice bearing melanoma tumors. The results of the biodistribution studies revealed significant tumor uptake (4.14±0.54%ID/g) within 10 minutes postinjection (p.i.), which increased further to 4.61±0.31%ID/g at 30 minutes p.i. The tumor-to-blood ratio was found to increase from 1.75±0.42 at 10 minutes p.i. to 2.25±0.20 at 60 minutes p.i., whereas the tumor-to-liver and tumor-to-muscle ratio between the same time points increased from 2.71±0.76 to 3.31±0.84 and 5.37±1.08 to 8.97±1.32, respectively. The study successfully demonstrated the preparation of (68)Ga-DOTA-E[c(RGDfK)](2) as a potential positron-emission tomography radiotracer for possible use in tumor imaging by using (68)Ga eluted from a reliable, easy-to-handle (68)Ge/(68)Ga generator developed in-house, without any postelution purification of (68)Ga.

  2. Cu2+-RGDFRGDS: exploring the mechanism and high efficacy of the nanoparticle in antithrombotic therapy

    Directory of Open Access Journals (Sweden)

    Wu J

    2015-04-01

    Full Text Available Jianhui Wu,1 Yuji Wang,1 Yaonan Wang,1 Ming Zhao,1,2 Xiaoyi Zhang,1 Lin Gui,1 Shurui Zhao,1 Haimei Zhu,1 Jinghua Zhao,1 Shiqi Peng11Beijing Area Major Laboratory of Peptide and Small Molecular Drugs, Engineering Research Center of Endogenous Prophylactic of Ministry of Education of China, Beijing Laboratory of Biomedical Materials, College of Pharmaceutical Sciences, Capital Medical University, Beijing, People’s Republic of China; 2Faculty of Biomedical Science and Environmental Biology, Kaohsiung Medical University, Kaohsiung, TaiwanAbstract: Thrombosis disease has been the leading cause of morbidity and mortality worldwide. In the discovery of antithrombotic agents, three complexes of Cu2+ and repetitive arginine-glycine-aspartic acid (RGD sequences, Cu(II-Arg-Gly-Asp-Ser-Arg-Gly-Asp-Ser (Cu[II]-4a, Cu(II-Arg-Gly-Asp-Val-Arg-Gly-Asp-Val (Cu[II]-4b, and Cu(II-Arg-Gly-Asp-Phe-Arg-Gly-Asp-Phe (Cu[II]-4c, were previously reported, of which Cu(II-4a and Cu(II-4c possessed the highest in vitro and in vivo activity, respectively. Transmission electron microscopy (TEM images visualized that Cu(II-4a and Cu(II-4c formed nanoaggregates and nanoparticles, respectively. However, the details of the formation of the nanospecies complexes and of the mechanism for inhibiting thrombosis remain to be clarified. For this purpose, this study designed a novel complex of Cu(II and the RGD octapeptide, Arg-Gly-Asp-Phe-Arg-Gly-Asp-Ser (RGDFRGDS, consisting of Arg-Gly-Asp-Phe of Cu(II-4c and Arg-Gly-Asp-Ser of Cu(II-4a, to colligate their biological and nanostructural benefits. In contrast with Cu(II-4a, -4b, and -4c, Cu(II-RGDFRGDS (Cu2+-FS had high antiplatelet and antithrombotic activities, with the formed nanoparticles having a porous surface. Additionally, this paper evidenced the dimer had the basic structural unit of Cu2+-FS in water, theoretically simulated the formation of Cu2+-FS nanoparticles, and identified that Cu2+-FS activity in decreasing

  3. Rat Genome Database (RGD)

    Data.gov (United States)

    U.S. Department of Health & Human Services — The Rat Genome Database (RGD) is a collaborative effort between leading research institutions involved in rat genetic and genomic research to collect, consolidate,...

  4. Induction of mesenchymal stem cell chondrogenesis by polyacrylate substrates

    OpenAIRE

    Glennon-Alty, Laurence; Williams, Rachel; Dixon, Simon; Murray, Patricia

    2013-01-01

    Mesenchymal stem cells (MSCs) can generate chondrocytes in vitro, but typically need to be cultured as aggregates in the presence of transforming growth factor beta (TGF-β), which makes scale-up difficult. Here we investigated if polyacrylate substrates modelled on the functional group composition and distribution of the Arg-Gly-Asp (RGD) integrin-binding site could induce MSCs to undergo chondrogenesis in the absence of exogenous TGF-β. Within a few days of culture on the biomimetic polyacry...

  5. A β-integrin from sea cucumber Apostichopus japonicus exhibits LPS binding activity and negatively regulates coelomocyte apoptosis.

    Science.gov (United States)

    Wang, Zhenhui; Shao, Yina; Li, Chenghua; Lv, Zhimeng; Wang, Haihong; Zhang, Weiwei; Zhao, Xuelin

    2016-05-01

    Integrins are a family of membrane glycoproteins, which are the major receptors for extracellular matrix and cell-cell adhesion molecules. In this study, a 1038 bp sequence representing the full-length cDNA of a novel β-integrin subunit (designated as AjITGB) was cloned from Apostichopus japonicusby using combined transcriptome sequencing and RACE approaches. The deduced amino acid sequence of AjITGB shared a conserved tripeptide Arg-Gly-Asp (RGD) binding domain with an S-diglyceridecysteine or N-Palm cysteine residue (C(31)), a transmembrane domain, and a β-integrin cytoplasmic domain. Spatial distribution analysis showed that AjITGB was constitutively expressed in all tested tissues with dominant expression in the muscles and weak expression in the respiratory tree. The pathogen Vibrio splendidus challenge and LPS stimulation could both significantly down-regulate the mRNA expression of AjITGB. Functional investigation revealed that recombinant AjITGB displayed higher LPS binding activity but lower binding activity to PGN and MAN. More importantly, knockdown of AjITGB by specific siRNA resulted in the significant promotion of coelomocyte apoptosis in vitro. Results indicated that AjITGB may serve as an apoptosis inhibitor with LPS binding activity during host-pathogen interaction in sea cucumber. PMID:26994670

  6. 99Tcm标记RGD环肽四聚体在神经胶质瘤裸鼠模型中的显像研究%Imaging of 99Tcm-cycllc RGD tetramer in nude mice bearing U87MG human glioma xenografts

    Institute of Scientific and Technical Information of China (English)

    余子璘; 贾兵; 刘昭飞; 史继云; 赵慧云; 杨志; 王凡

    2009-01-01

    Objective Multimeric cyclic RGD (Arg-Gly-Asp) peptides are capable of improving the integrin αvβ3-binding affinity due to the polyvalence effect.In this study,the authors prepare 99Tcm-la-bearing cyclic RGD tetramer E{E[c(RGDfK)]2}2,and evaluate its biodistribution and imaging in nude mice beating U87 MG human glioma xenografts with integrinαvβ3-positive.Methods 99Tcm-hydrazino-nictinamide (HYNIC)-E{E[c(RGDfK)]2}2 was prepared by two-step method,while HYNIC wag chosen as bifunctional chelator,and tricine and trisodium triphenylphosphine-3,3,3-trisuifonate (TPPTS) as coligands.The af-finity of c (RGDyK) monomer,HYNIC-E[c(RGDfK)]2 dimer and HYNIC-E{E[c(RGDfK)]2}2 tetramer to integrin αvβ3 was compared by in vitro competitive assay against binding of 125I-c(RGDyK)to integrin αvβ3.positive U87 MG human glioma cells.The biodistribution [the percentage of injection dose per gram of tissue(%ID/g)] and imaging were performed in nude mice bearing UB7MG human glioma xenografts.Re-suits The labeling yield of 99Tcm-HYNIC-E{E[c(RGDfK)2}2 was over 95%,and the radiochemical purity was more than 99%after purification with Sop-Pak C18 cartridge.The 50%inhibiting concentration (IC30) val-ues of c(RGDyk),HYNIC-E[c(RGDfK)]2 and HYNIC-E{E[c(RGDfK)]2}2 were 85.9,9.5 and 4.5 nmol/L, respectively.The result indicated that RGD tetramer possessed a significantly higher affinity to in-tegrinαvβ3.The biodistribution data showed that 99Tcm-HYNIC-E{E[c(RGDfK)]2}2 was excreted mainly through kidneys.The tumor uptake of 99Tcm-HYNIC-E{E[c(RGDfK)]2}2 was two times higher than 99Tcm- HYNIC-E[c(RGDfK)]2,at 1h postinjection,with the uptake of(10.32±0.07)%ID/g and(5.15±0.52)%ID/g,respectively,which was consistent with the in vitro competitive binding data.The tumor up-tale of 99Tcm-HYNIC.E{E[c(RGDfK)]2}2 was still as higher as(9.35±1.35)%ID/g at 4 h postinjec-tion, which demonstrated that the retention time of radiotracer in tumor was long enough.The imaging showed that tumor was clearly

  7. Bioengineered quantum dot/chitosan-tripeptide nanoconjugates for targeting the receptors of cancer cells.

    Science.gov (United States)

    Mansur, Alexandra A P; de Carvalho, Sandhra M; Mansur, Herman S

    2016-01-01

    Nanobiomaterials can be engineered to recognize cancer-specific receptors at the cellular level for diagnostic and therapeutic purposes. In this work, we report the synthesis of novel multifunctional nanoconjugates composed of fluorescent inorganic semiconductor quantum dot (QD) cores and tripeptide-modified polysaccharide organic shells. These structures were designed for targeting and imaging the αvβ3 integrin receptors of cancer cells. Initially, chitosan was covalently bound with the RGD peptide using a crosslinker to form bioconjugates (RGD-chitosan), which were later utilized as capping ligands for the production of surface-functionalized CdS QDs via a single-step process in aqueous media at room temperature. These core-shell nanostructures were extensively characterized by UV-vis spectroscopy, photoluminescence (PL) spectroscopy, Fourier transform infrared spectroscopy (FTIR), transmission electron microscopy (TEM), zeta potential (ZP) and dynamic light scattering (DLS). The TEM images and the UV-vis absorption results indicated the formation of ultra-small CdS QD nanocrystals with average diameters between 2.0 and 3.0 nm. In addition, the PL results demonstrated that the nanobioconjugates exhibited intense green fluorescence under excitation. The CdS-RGD-chitosan systems were effective at specific targeting integrin when assayed in vitro using two model cell cultures, HEK 293 (non-cancerous human embryonic kidney cell) and SAOS (cancerous sarcoma osteogenic-derived cells) imaged using fluorescence microscopy. PMID:26499085

  8. In-vivo study of adhesion and bone growth around implanted laser groove/RGD-functionalized Ti-6Al-4V pins in rabbit femurs

    International Nuclear Information System (INIS)

    Titanium surfaces were designed, produced, and evaluated for levels of osseointegration into the femurs of rabbits. A total of 36 Ti-6Al-4V pins (15 mm length, 1.64 mm diameter) were prepared into three experimental groups. These were designed to test the effects of osseointegration on laser grooved, RGD coated, and polished control surfaces, as well as combined effects. Circumferential laser grooves were introduced onto pin surfaces (40 μm spacing) using a UV laser (λ = 355 nm). The tripeptide sequence, Arginine-Glycine-Aspartic acid (RGD), was functionalized onto laser grooved surfaces. Of the prepared samples, surface morphology and chemistry were analyzed using scanning electron microscopy (SEM) and Immunoflourescence (IF) spectroscopy, respectively. The experimental pin surfaces were surgically implanted into rabbit femurs. The samples were then harvested and evaluated histologically. Sections of the sample were preserved in a methylmethacralate mold, sliced via a hard microtome, and polished systematically. In the case of the RGD coated and laser grooved surfaces, histological results showed accelerated bone growth into the implant, pull-out tests were also used to compare the adhesion between bone and the titanium pins with/without laser textures and/or RGD coatings. - Research highlights: → Circumferential laser grooves were introduced onto pin surfaces using a UV laser. → The tripeptide sequence, RGD, was functionalized onto laser grooved surfaces. → The experimental pin surfaces were surgically implanted into rabbit femurs. → RGD coated laser groove surfaces accelerated bone growth into the implant. → RGD coated laser grooved surfaces enhanced the adhesion between the bone and implant.

  9. In-vivo study of adhesion and bone growth around implanted laser groove/RGD-functionalized Ti-6Al-4V pins in rabbit femurs

    Energy Technology Data Exchange (ETDEWEB)

    Chen, J., E-mail: jianboc@gmail.com [Princeton Institute of Science and Technology of Materials and Department of Mechanical and Aerospace Engineering, Princeton University, Princeton, NJ 08544 (United States); Bly, R.A. [Princeton Institute of Science and Technology of Materials and Department of Mechanical and Aerospace Engineering, Princeton University, Princeton, NJ 08544 (United States); Saad, M.M.; AlKhodary, M.A.; El-Backly, R.M. [Tissue Engineering laboratories, Faculty of Dentistry, Alexandria University, Alexandria (Egypt); Cohen, D.J.; Kattamis, N. [Princeton Institute of Science and Technology of Materials and Department of Mechanical and Aerospace Engineering, Princeton University, Princeton, NJ 08544 (United States); Fatta, M.M.; Moore, W.A. [Tissue Engineering laboratories, Faculty of Dentistry, Alexandria University, Alexandria (Egypt); Arnold, C.B. [Princeton Institute of Science and Technology of Materials and Department of Mechanical and Aerospace Engineering, Princeton University, Princeton, NJ 08544 (United States); Marei, M.K. [Tissue Engineering laboratories, Faculty of Dentistry, Alexandria University, Alexandria (Egypt); Soboyejo, W.O. [Princeton Institute of Science and Technology of Materials and Department of Mechanical and Aerospace Engineering, Princeton University, Princeton, NJ 08544 (United States)

    2011-07-20

    Titanium surfaces were designed, produced, and evaluated for levels of osseointegration into the femurs of rabbits. A total of 36 Ti-6Al-4V pins (15 mm length, 1.64 mm diameter) were prepared into three experimental groups. These were designed to test the effects of osseointegration on laser grooved, RGD coated, and polished control surfaces, as well as combined effects. Circumferential laser grooves were introduced onto pin surfaces (40 {mu}m spacing) using a UV laser ({lambda} = 355 nm). The tripeptide sequence, Arginine-Glycine-Aspartic acid (RGD), was functionalized onto laser grooved surfaces. Of the prepared samples, surface morphology and chemistry were analyzed using scanning electron microscopy (SEM) and Immunoflourescence (IF) spectroscopy, respectively. The experimental pin surfaces were surgically implanted into rabbit femurs. The samples were then harvested and evaluated histologically. Sections of the sample were preserved in a methylmethacralate mold, sliced via a hard microtome, and polished systematically. In the case of the RGD coated and laser grooved surfaces, histological results showed accelerated bone growth into the implant, pull-out tests were also used to compare the adhesion between bone and the titanium pins with/without laser textures and/or RGD coatings. - Research highlights: {yields} Circumferential laser grooves were introduced onto pin surfaces using a UV laser. {yields} The tripeptide sequence, RGD, was functionalized onto laser grooved surfaces. {yields} The experimental pin surfaces were surgically implanted into rabbit femurs. {yields} RGD coated laser groove surfaces accelerated bone growth into the implant. {yields} RGD coated laser grooved surfaces enhanced the adhesion between the bone and implant.

  10. Recombinant expression of mutants of the Frankenstein disintegrin, RTS-ocellatusin. Evidence for the independent origin of RGD and KTS/RTS disintegrins.

    Science.gov (United States)

    Sanz-Soler, Raquel; Lorente, Carolina; Company, Beatriz; Sanz, Libia; Juárez, Paula; Pérez, Alicia; Zhang, Yun; Jin, Yang; Chen, Runqiang; Eble, Johannes A; Calvete, Juan J; Bolás, Gema

    2012-09-15

    The requirements to transform a short disintegrin of the RGD clade into an RTS disintegrin, were investigated through the generation of recombinant mutants of ocellatusin in which the RGD tripeptide was substituted for RTS in different positions along the integrin-specificity loop. Any attempt to create an active integrin α(1)β(1) inhibitory motif within the specificity loop of ocellatusin was unsuccessful. Replacing the whole RGD-loop of ocellatusin by the RTS-loop of jerdostatin was neither sufficient for confering α(1)β(1) binding specificity to this ocellatusin-RTS Frankenstein(2) mutant. Factors other than the integrin-binding loop sequence per se are thus required to transform a disintegrin scaffold from the RGD clade into another scaffold from the RTS/KTS clade. Moreover, our results provide evidences, that the RTS/KTS short disintegrins have potentially been recruited into the venom gland of Eurasian vipers independently from the canonical neofunctionalization pathway of the RGD disintegrins. PCR-amplifications of jerdostatin-like sequences from a number of taxa across reptiles, including snakes (Crotalinae, Viperinae, and Elapidae taxa) and lizards (Lacertidae and Iguanidae) clearly showed that genes coding for RTS/KTS disintegrins existed long before the split of Lacertidae and Iguania, thus predating the recruitment of the SVMP precursors of disintegrins, providing strong support for the view of an independent evolutionary history of the RTS/KTS and the RGD clades of short disintegrins.

  11. Comparative study on the cellular activities of osteoblast-like cells and new bone formation of anorganic bone mineral coated with tetra-cell adhesion molecules and synthetic cell binding peptide

    OpenAIRE

    Yu, Hyeon-Seok; Noh, Woo-Chang; Park, Jin-Woo; Lee, Jae-Mok; Yang, Dong-Jun; Park, Kwang-Bum; Suh, Jo-Young

    2011-01-01

    Purpose We have previously reported that tetra-cell adhesion molecule (T-CAM) markedly enhanced the differentiation of osteoblast-like cells grown on anorganic bone mineral (ABM). T-CAM comprises recombinant peptides containing the Arg-Gly-Asp (RGD) sequence in the tenth type III domain, Pro-His-Ser-Arg-Asn (PHSRN) sequence in the ninth type III domain of fibronectin (FN), and the Glu-Pro-Asp-Ilu-Met (EPDIM) and Tyr-His (YH) sequence in the fourth fas-1 domain of βig-h3. Therefore, the purpos...

  12. Effect of enamel matrix derivative and parathyroid hormone on bone formation in standardized osseous defects: an experimental study in minipigs

    DEFF Research Database (Denmark)

    Jensen, Simon S; Chen, B; Bornstein, Michael M;

    2011-01-01

    or directly on the bone-forming cells. In addition, it is not known if the presentation of PTH by adding the amino acid sequence Arg-Gly-Asp (RGD) is essential for its osteopromotive effect. Local delivery of a bioactive substance at the right time and in the right concentration often constitutes a major...... challenge. Polyethylene glycol-based hydrogel (PEG) is a degradable vehicle developed for delivery of bioactive proteins. To enhance the mechanical stability of the PEG-bioactive substance complex, an osteoconductive bone substitute material is often needed....

  13. Biomedical applications of dipeptides and tripeptides.

    Science.gov (United States)

    Santos, Sara; Torcato, Inês; Castanho, Miguel A R B

    2012-01-01

    Peptides regulate many physiological processes, acting at some sites as endocrine or paracrine signals and at others as neurotransmitters or growth factors, for instance. These molecules represent a major evolution in medical and industrial fields, as it is becoming mandatory to design and exploit molecules that do not necessarily fit the description of classical drug classes. The list of peptides with potential biomedical applications is huge and is growing each year. These biomedical applications range from uses as drugs to flavor-active peptides as ingredients in natural health products, nutraceuticals and functional foods. Among the peptide family, dipeptides and tripeptides are very appealing for drug discovery and development because of their cost-effectiveness, possibility of oral administration, and simplicity to perform molecular structural and quantitative structure-activity studies. Our objective is to review different actual and future uses of dipeptides and tripeptides as well as the major advances and obstacles in this growing area. PMID:23193593

  14. Development of a QSAR model for binding of tripeptides and tripeptidomimetics to the human intestinal di-/tripeptide transporter hPEPT1

    DEFF Research Database (Denmark)

    Andersen, Rikke; Jørgensen, Flemming Steen; Olsen, Lars;

    2006-01-01

    The aim of this study was to develop a three-dimensional quantitative structure-activity relationship (QSAR) model for binding of tripeptides and tripeptidomimetics to hPEPT1 based on a series of 25 diverse tripeptides.......The aim of this study was to develop a three-dimensional quantitative structure-activity relationship (QSAR) model for binding of tripeptides and tripeptidomimetics to hPEPT1 based on a series of 25 diverse tripeptides....

  15. Enzymatic Synthesis of a CCK-8 Tripeptide Derivative

    Institute of Scientific and Technical Information of China (English)

    2003-01-01

    The enzymatic synthesis of CCK-8 tripeptide derivative Phac-Met-Asp(OMe)-Phe-NH2 is reported. Starting with Phac-Met-OCam, we have successfully synthesized the target tripeptide with three free or immobilized enzymes, α-chymotrypsin, papain and thermolysin in reasonable yields. The key steps in this synthesis were the coupling of Phac-Met-OCam and H-Asp(OMe)2 to form Met-Asp peptide bond catalyzed by α-chymotrypsin and the selective hydrolysis of α-ester of Phac-Met-Asp(OMe)2 catalyzed by papain.

  16. Advantages of RGD peptides for directing cell association with biomaterials

    OpenAIRE

    Bellis, Susan L.

    2011-01-01

    Despite many years of in vitro research confirming the effectiveness of RGD in promoting cell attachment to a wide variety of biomaterials, animal studies evaluating tissue responses to implanted RGD-functionalized substrates have yielded more variable results The goals of this report are to present some of the reasons why cell culture studies may not always reliably predict in vivo responses, and more importantly, to highlight potential applications that may benefit from the use of RGD pepti...

  17. Fabrication of multi-biofunctional gelatin-based electrospun fibrous scaffolds for enhancement of osteogenesis of mesenchymal stem cells.

    Science.gov (United States)

    Lin, Wei-Han; Yu, Jiashing; Chen, Guoping; Tsai, Wei-Bor

    2016-02-01

    Biofunctional scaffolds that support the adhesion, proliferation, and osteo-differentiation of mesenchymal stem cells (MSCs) are critical for bone tissue engineering. In this study, a simple in situ UV-crosslinking strategy was utilized to fabricate gelatin electrospun fibrous (GEF) scaffolds with multiple biosignals, including cell adhesive Arg-Gly-Asp (RGD) peptide, osteo-conductive hydroxyapatite (HAp) nanoparticles, and osteo-inductive bone morphogenic protein-2 (BMP-2). The adhesion and proliferation of MSCs on the GEF scaffolds were improved by the incorporation of RGD. Meanwhile, the incorporation of HAp and BMP-2 enhanced osteo-differentiation of MSCs. The three incorporated bio-factors exert a synergistic effect on osteogenesis of MSCs in the GEF scaffolds. This strategy of incorporating multiple biomolecules could be used to fabricate crosslinked electrospun scaffolds of natural polymers for tissue-engineering applications. PMID:26642073

  18. Asociación de índices infecciosos del virus de la fiebre aftosa a receptores celulares en el ganado criollo bon

    OpenAIRE

    Ariza Botero Manuel Fernando; Rodríguez Morales Alejandra

    2006-01-01

    Los miembros de la familia de receptores celulares, integrinas (·V‚1, ·V‚3 ·V‚6), han sido identificados como factores de adhesión de diferentes virus a las células del hospedero. Para el caso del virus de la Fiebre Aftosa, estas integrinas se unen al sitio de reconocimiento celular en la secuencia tripéptidica Arg-Gly-Asp (RGD) e igualmente están localizadas en la proteína VP1 del virus. Evidencia genética de esta interacción ha sido obtenida mediante la mutación de la secuencia RGD en clone...

  19. Does ligand-receptor mediated competitive effect or penetrating effect of iRGD peptide when co-administration with iRGD-modified SSL?

    Science.gov (United States)

    Zhang, Wei-Qiang; Yu, Ke-Fu; Zhong, Ting; Luo, Li-Min; Du, Ruo; Ren, Wei; Huang, Dan; Song, Ping; Li, Dan; Zhao, Yang; Wang, Chao; Zhang, Xuan

    2015-12-01

    Ligand-mediated targeting of anticancer therapeutic agents is a useful strategy for improving anti-tumor efficacy. It has been reported that co-administration of a tumor-penetrating peptide iRGD (CRGDK/RGPD/EC) enhances the efficacy of anticancer drugs. Here, we designed an experiment involving co-administration of iRGD-SSL-DOX with free iRGD to B16-F10 tumor bearing mice to examine the action of free iRGD. We also designed an experiment to investigate the location of iRGD-modified SSL when co-administered with free iRGD or free RGD to B16-F10 tumor bearing nude mice. Considering the sequence of iRGD, we selected the GPDC, RGD and CRGDK as targeting ligands to investigate the targeting effect of these peptides compared with iRGD on B16-F10 and MCF-7 cells, with or without enzymatic degradation. Finally, we selected free RGD, free CRGDK and free iRGD as ligand to investigate the inhibitory effect on RGD-, CRGDK- or iRGD-modified SSL on B16-F10 or MCF-7 cells. Our results indicated that iRGD targeting to tumor cells was ligand-receptor mediated involving RGD to αv-integrin receptor and CRGDK to NRP-1 receptor. Being competitive effect, the administration of free iRGD would not be able to further enhance the anti-tumor activity of iRGD-modified SSL. There is no need to co-administrate of free iRGD with the iRGD-modified nanoparticles for further therapeutic benefit.

  20. The tripeptide feG inhibits leukocyte adhesion

    OpenAIRE

    Davison Joseph S; Christie Emily; Mathison Ronald D

    2008-01-01

    Abstract Background The tripeptide feG (D-Phe-D-Glu-Gly) is a potent anti-inflammatory peptide that reduces the severity of type I immediate hypersensitivity reactions, and inhibits neutrophil chemotaxis and adhesion to tissues. feG also reduces the expression of β1-integrin on circulating neutrophils, but the counter ligands involved in the anti-adhesive actions of the peptide are not known. In this study the effects of feG on the adhesion of rat peritoneal leukocytes and extravasated neutro...

  1. Construction and Characterization of Novel Staphylokinase Variants with Antiplatelet Aggregation Activity and Reduced Immunogenecity

    Institute of Scientific and Technical Information of China (English)

    Hua-Bo SU; Yu-Gao ZHANG; Jin-Tian HE; Wei MO; Yan-Ling ZHANG; Xian-Mei TAO; Hou-Yan SONG

    2004-01-01

    To develop target thrombolytic agents with fibrinolytic activity, antiplatelet aggregation activity and reduced immunogenicity, two staphylokinase variants containing Arg-Gly-Asp (RGD) motif were constructed. Gene expression was induced in E. coli JF1125 and the variants, designated DGR and RL1, were purified with gel filtration and ion-exchange chromatography and the purity was over 95%. The fibrinolytic activity and kinetic constants of the two variants were comparable to those of recombinant wild-type staphylokinase. Both the variants can inhibit the platelet aggregation at a final concentration of 2 μM. Thetiters of antibodies against variants were much lower than those against recombinant staphylokinase in guineapigs, which indicated that the immunogenicity of the variants was greatly reduced. These results confirm thatit is possible to design and produce a bifunctional protein that possesses fibrinolytic and antiplatelet aggregation activities.

  2. Enzymatic Synthesis of a CCK-8 Tripeptide Fragment

    Institute of Scientific and Technical Information of China (English)

    XIANG,Hua(向华); ECKSTEIN,Heiner

    2004-01-01

    A process for the synthesis of CCK-8 tripeptide H-Gly-Trp-Met-OH catalyzed by immobilized enzyme was reported. Enzymes were used for the formation of peptide bonds and the removal of protecting group. Starting with phenylacetyl (PhAc) glycin, N-protected dipeptide PhAc-Gly-Trp-OMe was obtained by coupling PhAc-protected glycine carboxamidomethyl ester (OCam) with Trp-OMe catalyzed by immobilized papain in buffered ethyl acetate.motrypsin catalysis in solvent free system. Basic hydrolysis was followed getting PhAc-Gly-Trp-Met-OH. The PhAc-group was removed with penicillin G amidase and H-Gly-Trp-Met-OH was obtained in an overall yield of 43.9%. The reaction conversion of tripeptide in solvent free system was strongly affected by the system of basic salts added. The influence of the support materials used to deposit enzymes and structures of acyl donor and nucleophile on the reaction was also investigated.

  3. Cooperative assembly of Zn cross-linked artificial tripeptides with pendant hydroxyquinoline ligands.

    Science.gov (United States)

    Zhang, Meng; Gallagher, Joy A; Coppock, Matthew B; Pantzar, Lisa M; Williams, Mary Elizabeth

    2012-11-01

    An artificial peptide with three pendant hydroxyquinoline (hq) ligands on a palindromic backbone was designed and used to form multimetallic assemblies. Reaction of the tripeptide with zinc acetate led to a highly fluorescent tripeptide duplex with three Zn(II) coordinative cross-links. The binding process was monitored using spectrophotometric absorbance and emission titrations; NMR spectroscopy and mass spectrometry confirmed the identity and stoichiometry of the product structure. Titrations monitoring duplex formation of the zinc-tripeptide structure had a sigmoidal shape, equilibrium constant larger than the monomeric analogue, and a Hill coefficient >1, all of which indicate positive cooperativity. Photophysical characterization of the quantum yield, excited state lifetime, and polarization anisotropy are compared with the monometallic zinc-hq analogue. A higher than expected quantum yield for the trimetallic complex suggests a structure in which the central chromophore is shielded from solvent by π-stacking with neighboring Zn(II) complexes.

  4. Evaluation of 111In-Labeled Cyclic RGD Peptides: Tetrameric Not Tetravalent

    OpenAIRE

    Chakraborty, Sudipta; Shi, Jiyun; Kim, Young-Seung; Zhou, Yang; Jia, Bing; Wang, Fan; Liu, Shuang

    2010-01-01

    This report presents the synthesis and evaluation of 111In(DOTA-6G-RGD4) (DOTA = 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetracetic acid; 6G-RGD4 = E{G3-E[G3-c(RGDfK)]2}2 and G3 = Gly-Gly-Gly), 111In(DOTA-RGD4) (RGD4 = E{E[c(RGDfK)]2}2) and 111In(DOTA-3G-RGD2) (3G-RGD2 = G3-E[G3-c(RGDfK)]2) as new radiotracers for imaging integrin αvβ3–positive tumors. The IC50 values of DOTA-6G-RGD4, DOTA-RGD4 and DOTA-3G-RGD2 were determined to be 0.4 ± 0.1, 1.5 ± 0.2 and 1.3 ± 0.2 nM against 125I-c(RGDyK) ...

  5. RGD functionalized polymeric nanoparticles targeting periodontitis epithelial cells for the enhanced treatment of periodontitis in dogs.

    Science.gov (United States)

    Yao, Wenxin; Xu, Peicheng; Zhao, Jingjing; Ling, Li; Li, Xiaoxia; Zhang, Bo; Cheng, Nengneng; Pang, Zhiqing

    2015-11-15

    Long term retention of antimicrobials with effective drug concentration in gingival crevicular fluid (GCF) is of vital importance for the treatment of chronic periodontitis. In this study, a novel epithelial cell-targeting nanoparticle drug delivery system by conjugating minocycline-loaded poly(ethylene glycol)-poly(lactic acid) (PEG-PLA) nanoparticles (NP-MIN) with RGD peptide were developed and administrated locally for targeting periodontitis epithelial cells and enhancing the treatment of periodontitis in dogs. Biodegradable NP-MIN was made with an emulsion/solvent evaporation technique. RGD peptide was conjugated to the surface of nanoparticles via Maleimide group reaction with hydrosulfide in RGD peptide (RGD-NP-MIN). Transmission electron microscopy examination and dynamic light scattering results revealed that RGD-NP-MIN had a sphere shape, with a mean diameter around 106nm. In vitro release of minocycline from RGD-NP-MIN showed that RGD modification did not change the remarkable sustained releasing characteristic of NP-MIN. To elucidate the interaction of RGD-NP and epithelial cells, RGD-NP binding, uptake and cellular internalization mechanisms by calu-3 cells were investigated. It was shown RGD modification significantly enhanced nanoparticles binding and uptake by Calu-3 cells, and RGD-NP uptake was an energy-dependent process through receptor-mediated endocytosis. Both clathrin-associated endocytosis and caveolae-dependent endocytosis pathway were involved in the RGD-NP uptake, and the intracellular transport of RGD-NP was related to lysosome and Golgi apparatus. Finally, in vivo pharmacokinetics of minocycline in the periodontal pockets and anti-periodontitis effects of RGD-NP-MIN on periodontitis-bearing dogs were evaluated. After local administration of RGD-NP-MIN, minocycline concentration in gingival crevicular fluid decreased slowly and maintained an effective drug concentration for a longer time than that of NP-MIN. Anti-periodontitis effects

  6. Comparison of 90Y/177Lu labeled DOTA-Bz-RGD tetramer and DOTA-RGD tetramer

    International Nuclear Information System (INIS)

    90Y/177Lu labeled DOTA-Bz-RGD tetramer and DOTA-RGD tetramer were prepared, and the effect of Bz-DOTA and DOTA on labeling conditions and in vitro stability of radiolabeled compounds was compared. The labeling conditions, including reaction pH, reaction temperature and reaction time, were investigated. ITLC and HPLC show that the labeling yields of four radiolabeled compounds are more than 95% under optimal conditions (pH=6.0, reacting at 100 degree C for 15-20 min); the four radiolabeled compounds show pretty good stability in saline and fetal bovine serum. Although introducing of Bz has no effect on labeling conditions and in vitro stability of radiolabeled compounds, it brings a little change on molecule polarity. HPLC analysis and lg P values reveal that introducing of Bz increases the lipophilicity of radiolabeled compounds. (authors)

  7. The rescue and evaluation of FLAG and HIS epitope-tagged Asia 1 type foot-and-mouth disease viruses.

    Science.gov (United States)

    Yang, Bo; Yang, Fan; Zhang, Yan; Liu, Huanan; Jin, Ye; Cao, Weijun; Zhu, Zixiang; Zheng, Haixue; Yin, Hong

    2016-02-01

    The VP1 G-H loop of the foot-and-mouth disease virus (FMDV) contains the primary antigenic site, as well as an Arg-Gly-Asp (RGD) binding motif for the αv-integrin family of cell surface receptors. We anticipated that introducing a foreign epitope tag sequence downstream of the RGD motif would be tolerated by the viral capsid and would not destroy the antigenic site of FMDV. In this study, we have designed, generated, and characterized two recombinant FMDVs with a FLAG tag or histidine (HIS) inserted in the VP1 G-H loop downstream of the RGD motif +9 position. The tagged viruses were genetically stable and exhibited similar growth properties with their parental virus. What is more, the recombinant viruses rFMDV-FLAG and rFMDV-HIS showed neutralization sensitivity to FMDV type Asia1-specific mAbs, as well as to polyclonal antibodies. Additionally, the r1 values of the recombinant viruses were similar to that of the parental virus, indicating that the insertion of FLAG or HIS tag sequences downstream of the RGD motif +9 position do not eradicate the antigenic site of FMDV and do not affect its antigenicity. These results indicated that the G-H loop of Asia1 FMDV is able to effectively display the foreign epitopes, making this a potential approach for novel FMDV vaccines development. PMID:26732485

  8. Construction of hydroxypropyl-{beta}-cyclodextrin copolymer nanoparticles and targeting delivery of paclitaxel

    Energy Technology Data Exchange (ETDEWEB)

    Miao Qinghua; Li Suping; Han Siyuan [National Center for Nanoscience and Technology of China, CAS Key Laboratory for Biological Effects of Nanomaterials and Nanosafety (China); Wang Zhi, E-mail: wangzhi@jlu.edu.cn [Jilin University, Key Laboratory for Molecular Enzymology and Engineering, Ministry of Education (China); Wu Yan, E-mail: wuy@nanoctr.cn; Nie Guangjun, E-mail: niegj@nanoctr.cn [National Center for Nanoscience and Technology of China, CAS Key Laboratory for Biological Effects of Nanomaterials and Nanosafety (China)

    2012-08-15

    A novel amphiphilic copolymer with p-maleimidophenyl isocyanate-hydroxypropyl-{beta}-cyclodextrin-polylactide-1, 2-dipalmitoyl-sn-glycero-3-phosphoethanolamine to generate copolymer nanoparticles (NPs) has been designed. In order to develop an active targeting system, integrin {alpha}{sub v}{beta}{sub 3}-specific targeting peptide cyclo(Arg-Gly-Asp-D-Phe-Cys), cRGD, was conjugated to the surface of NPs (NPs-RGD). These NPs were used to encapsulate anti-tumor drug, paclitaxel. The resulting NPs exhibited high drug-loading capacity and controlled drug release in vitro at acidic pH. In vitro cytotoxicity assay demonstrates that paclitaxel-loaded NPs-RGD significantly inhibited B16 tumor cell (high {alpha}{sub v}{beta}{sub 3}) proliferation relative to free paclitaxel and paclitaxel-loaded NPs at high concentrations. Paclitaxel-loaded NPs-RGD localized mainly in lysosomes in B16 cells as revealed by confocal microscopy. These results suggest a novel strategy for fabrication-functionalizing hydroxypropyl-{beta}-cyclodextrin copolymer nanoparticles for targeting delivery of paclitaxel to integrin {alpha}{sub v}{beta}{sub 3}-rich tumor cells. These nanocarriers can be readily extended to couple other bioactive molecules for active targeting and delivery of various chemotherapeutic drugs.

  9. Engineering Factor Xa Inhibitor with Multiple Platelet-Binding Sites Facilitates its Platelet Targeting

    Science.gov (United States)

    Zhu, Yuanjun; Li, Ruyi; Lin, Yuan; Shui, Mengyang; Liu, Xiaoyan; Chen, Huan; Wang, Yinye

    2016-01-01

    Targeted delivery of antithrombotic drugs centralizes the effects in the thrombosis site and reduces the hemorrhage side effects in uninjured vessels. We have recently reported that the platelet-targeting factor Xa (FXa) inhibitors, constructed by engineering one Arg-Gly-Asp (RGD) motif into Ancylostoma caninum anticoagulant peptide 5 (AcAP5), can reduce the risk of systemic bleeding than non-targeted AcAP5 in mouse arterial injury model. Increasing the number of platelet-binding sites of FXa inhibitors may facilitate their adhesion to activated platelets, and further lower the bleeding risks. For this purpose, we introduced three RGD motifs into AcAP5 to generate a variant NR4 containing three platelet-binding sites. NR4 reserved its inherent anti-FXa activity. Protein-protein docking showed that all three RGD motifs were capable of binding to platelet receptor αIIbβ3. Molecular dynamics simulation demonstrated that NR4 has more opportunities to interact with αIIbβ3 than single-RGD-containing NR3. Flow cytometry analysis and rat arterial thrombosis model further confirmed that NR4 possesses enhanced platelet targeting activity. Moreover, NR4-treated mice showed a trend toward less tail bleeding time than NR3-treated mice in carotid artery endothelium injury model. Therefore, our data suggest that engineering multiple binding sites in one recombinant protein is a useful tool to improve its platelet-targeting efficiency. PMID:27432161

  10. Engineering Factor Xa Inhibitor with Multiple Platelet-Binding Sites Facilitates its Platelet Targeting.

    Science.gov (United States)

    Zhu, Yuanjun; Li, Ruyi; Lin, Yuan; Shui, Mengyang; Liu, Xiaoyan; Chen, Huan; Wang, Yinye

    2016-01-01

    Targeted delivery of antithrombotic drugs centralizes the effects in the thrombosis site and reduces the hemorrhage side effects in uninjured vessels. We have recently reported that the platelet-targeting factor Xa (FXa) inhibitors, constructed by engineering one Arg-Gly-Asp (RGD) motif into Ancylostoma caninum anticoagulant peptide 5 (AcAP5), can reduce the risk of systemic bleeding than non-targeted AcAP5 in mouse arterial injury model. Increasing the number of platelet-binding sites of FXa inhibitors may facilitate their adhesion to activated platelets, and further lower the bleeding risks. For this purpose, we introduced three RGD motifs into AcAP5 to generate a variant NR4 containing three platelet-binding sites. NR4 reserved its inherent anti-FXa activity. Protein-protein docking showed that all three RGD motifs were capable of binding to platelet receptor αIIbβ3. Molecular dynamics simulation demonstrated that NR4 has more opportunities to interact with αIIbβ3 than single-RGD-containing NR3. Flow cytometry analysis and rat arterial thrombosis model further confirmed that NR4 possesses enhanced platelet targeting activity. Moreover, NR4-treated mice showed a trend toward less tail bleeding time than NR3-treated mice in carotid artery endothelium injury model. Therefore, our data suggest that engineering multiple binding sites in one recombinant protein is a useful tool to improve its platelet-targeting efficiency. PMID:27432161

  11. Integrin activation and focal complex formation in cardiac hypertrophy

    Science.gov (United States)

    Laser, M.; Willey, C. D.; Jiang, W.; Cooper, G. 4th; Menick, D. R.; Zile, M. R.; Kuppuswamy, D.

    2000-01-01

    Cardiac hypertrophy is characterized by both remodeling of the extracellular matrix (ECM) and hypertrophic growth of the cardiocytes. Here we show increased expression and cytoskeletal association of the ECM proteins fibronectin and vitronectin in pressure-overloaded feline myocardium. These changes are accompanied by cytoskeletal binding and phosphorylation of focal adhesion kinase (FAK) at Tyr-397 and Tyr-925, c-Src at Tyr-416, recruitment of the adapter proteins p130(Cas), Shc, and Nck, and activation of the extracellular-regulated kinases ERK1/2. A synthetic peptide containing the Arg-Gly-Asp (RGD) motif of fibronectin and vitronectin was used to stimulate adult feline cardiomyocytes cultured on laminin or within a type-I collagen matrix. Whereas cardiocytes under both conditions showed RGD-stimulated ERK1/2 activation, only collagen-embedded cells exhibited cytoskeletal assembly of FAK, c-Src, Nck, and Shc. In RGD-stimulated collagen-embedded cells, FAK was phosphorylated only at Tyr-397 and c-Src association occurred without Tyr-416 phosphorylation and p130(Cas) association. Therefore, c-Src activation is not required for its cytoskeletal binding but may be important for additional phosphorylation of FAK. Overall, our study suggests that multiple signaling pathways originate in pressure-overloaded heart following integrin engagement with ECM proteins, including focal complex formation and ERK1/2 activation, and many of these pathways can be activated in cardiomyocytes via RGD-stimulated integrin activation.

  12. Therapeutic efficacy of an oncolytic adenovirus containing RGD ligand in minor capsid protein IX and Fiber, Δ24DoubleRGD, in an ovarian cancer model

    Directory of Open Access Journals (Sweden)

    Anton V Borovjagin

    2012-02-01

    Full Text Available Ovarian cancer is the leading cause of gynecological disease death despite advances in medicine. Therefore, novel strategies are required for ovarian cancer therapy. Conditionally replicative adenoviruses (CRAds, genetically modified as anti-cancer therapeutics, are one of the most attractive candidate agents for cancer therapy. However, a paucity of coxsackie B virus and adenovirus receptor (CAR expression on the surface of ovarian cancer cells has impeded treatment of ovarian cancer using this approach.This study sought to engineer a CRAd with enhanced oncolytic ability in ovarian cancer cells, “Δ24DoubleRGD.” Δ24DoubleRGD carries an arginine-glycine-aspartate (RGD motif incorporated into both fiber and capsid protein IX (pIX and its oncolytic efficacy was evaluated in ovarian cancer. In vitro analysis of cell viability showed that infection of ovarian cancer cells with Δ24DoubleRGD leads to increased cell killing relative to the control CRAds. Data from this study suggested that not only an increase in number of RGD motifs on the CRAd capsid, but also a change in the repertoir of targeted integrins could lead to enhanced oncolytic potency of Δ24DoubleRGD in ovarian cancer cells in vitro. In an intraperitoneal model of ovarian cancer, mice injected with Δ24DoubleRGD showed, however, a similar survival rate as mice treated with control CRAds.

  13. Is the structural diversity of tripeptides sufficient for developing functional food additives with satisfactory multiple bioactivities?

    Science.gov (United States)

    Wang, Jian-Hui; Liu, Yong-Le; Ning, Jing-Heng; Yu, Jian; Li, Xiang-Hong; Wang, Fa-Xiang

    2013-05-01

    Multifunctional peptides have attracted increasing attention in the food science community because of their therapeutic potential, low toxicity and rapid intestinal absorption. However, previous study demonstrated that the limited structural variations make it difficult to optimize dipeptide molecules in a good balance between desirable and undesirable properties (F. Tian, P. Zhou, F. Lv, R. Song, Z. Li, J. Pept. Sci. 13 (2007) 549-566). In the present work, we attempt to answer whether the structural diversity is sufficient for a tripeptide to have satisfactory multiple bioactivities. Statistical test, structural examination and energetic analysis confirm that peptides of three amino acids long can bind tightly to human angiotensin converting enzyme (ACE) and thus exert significant antihypertensive efficacy. Further quantitative structure-activity relationship (QSAR) modeling and prediction of all 8000 possible tripeptides reveal that their ACE-inhibitory potency exhibits a good (positive) relationship to antioxidative activity, but has only a quite modest correlation with bitterness. This means that it is possible to find certain tripeptide entities possessing the optimal combination of strong ACE-inhibitory potency, high antioxidative activity and weak bitter taste, which are the promising candidates for developing multifunctional food additives with satisfactory multiple bioactivities. The marked difference between dipeptide and tripeptide can be attributed to the fact that the structural diversity of peptides increases dramatically with a slight change in sequence length.

  14. Oxymatrine liposome attenuates hepatic fibrosis via targeting hepatic stellate cells

    Institute of Scientific and Technical Information of China (English)

    Ning-Li Chai; Qiang Fu; Hui Shi; Chang-Hao Cai; Jun Wan; Shi-Ping Xu; Ben-Yan Wu

    2012-01-01

    AIM:To investigate the potential mechanism of ArgGly-Asp (RGD) peptide-labeled liposome loading oxymatrine (OM) therapy in CCl4-induced hepatic fibrosis in METHODS:We constructed a rat model of CCl4-induced hepatic fibrosis and treated the rats with different formulations of OM.To evaluate the antifibrotic effect of OM,we detected levels of alkaline phosphatase,hepatic histopathology (hematoxylin and eosin stain and Masson staining) and fibrosis-related gene expression of matrix metallopeptidase (MMP)-2,tissue inhibitor of metalloproteinase (TIMP)-1 as well as type Ⅰ procollagen via quantitative real-time polymerase chain reaction.To detect cell viability and apoptosis of hepatic stellate cells (HSCs),we performed 3-(4,5)-dimethylthiahiazo(-z-y1)-3,5-diphenytetrazoliumromide assay and flow cytometry.To reinforce the combination of oxymatrine with HSCs,we constructed fluorescein-isothiocyanate-conjugated Arg-Gly-Asp peptide-labeled liposomes loading OM,and its targeting of HSCs was examined by fluorescent microscopy.RESULTS:OM attenuated CCl4-induced hepatic fibrosis,as defined by reducing serum alkaline phosphatase (344.47 ± 27.52 U/L vs 550.69 ± 43.78 U/L,P < 0.05),attenuating liver injury and improving collagen deposits (2.36% ± 0.09% vs 7.70% ± 0.60%,P < 0.05) and downregulating fibrosis-related gene expression,that is,MMP-2,TIMP-1 and type Ⅰ procollagen (P < 0.05).OM inhibited cell viability and induced apoptosis of HSCs in vitro.RGD promoted OM targeting of HSCs and enhanced the therapeutic effect of OM in terms of serum alkaline phosphatase (272.51 ± 19.55 U/L vs 344.47 ± 27.52 U/L,P < 0.05),liver injury,collagen deposits (0.26% ± 0.09% vs 2.36% ± 0.09%,P < 0.05) and downregulating fibrosis-related gene expression,that is,MMP-2,TIMP-1 and type Ⅰ procollagen (P < 0.05).Moreover,in vitro assay demonstrated that RGD enhanced the effect of OM on HSC viability and apoptosis.CONCLUSION:OM attenuated hepatic fibrosis by

  15. Evaluation of the therapeutic efficacy and radiotoxicity of the conjugates 177Lu-DOTA-E-c(RGDfK)2 and 177Lu-DOTA-GGC-AuNP-c[RGDfk(C)] in a murine model and their relationship with the inhibition of the angiogenic factors VEGF and HIF-1α

    International Nuclear Information System (INIS)

    Molecular targeting therapy has become a relevant therapeutic strategy for cancer. The principle that peptide receptors can be used successfully for in vivo targeting of human cancers has been proven, and radiolabeled peptides have been demonstrated to be effective in patients with malignant tumors. Peptides based on the cyclic Arg-Gly-Asp (RGD) sequence have been designed to antagonize the function of α(v)β(3) integrin, thereby inhibiting angio genesis. The conjugation of RGD peptides to radiolabeled gold nanoparticles (AuNP) produces biocompatible and stable m ultimeric systems with target-specific molecular recognition. The aim of this research was to evaluate the therapeutic response of 177Lu-AuNP-RGD in athymic mice bearing α(v)β(3)-integrin-positive C6 gliomas and compare with that of 177Lu-AuNP or 177Lu-RGD. The radiation absorbed dose, metabolic activity (SUV, [18F]fluor-deoxy-glucose-micro PET/CT), renal radiotoxicity, renal and tumoral histological characteristics as well as tumoral VEGF and HIF-1? gene expression (by realtime polymerase chain reaction) following treatment with 177Lu-AuNP-RGD, 177Lu-AuNP or 177Lu-RGD were assessed. Of the radiopharmaceuticals evaluated, 177Lu-AuNP-RGD delivered the highest tumor radiation absorbed dose (63.8 ± 7.9 Gy) vs other treatments. These results correlated with the observed therapeutic response, in which 177Lu-AuNP-RGD significantly (p177Lu). There was a low uptake in non-target organs and no induction of renal toxicity. 177Lu-AuNP-RGD demonstrates properties suitable for use as an agent for molecular targeting radiotherapy. (Author)

  16. Synthesis of RGD-aPEG-lactoside, a Potential Anti-metastasis Glycoconjugate

    Institute of Scientific and Technical Information of China (English)

    Shu Chun LI; Li Min NIU; Hui LI; Zhong Jun LI; Qing LI

    2004-01-01

    The adhesive interaction between tumor cells and host cells or the extra cellular matrix plays a crucial role in metastasis. Due to the anti-metastasis effects of RGD (arginyl-glycyl -aspartic acid) and some oligosaccharides, RGD-aPEG-Lactoside was prepared which will be used on anti-metastasis.

  17. Synthesis and Characterization of a Biodegradable Copolymer: RGD Peptide Modification of Poly (lactic acid-co-lysine)

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    The poly ( lactic acid- co-lysine ) was synthesized using IR and 1 H NMR to characterize the copolymer. And then the RGD modification copolymer RGD-PLAL was prepared. The contact angles were used to see the RGD modification occurrence. Also high molecular weight polymer was controlled to the reaction of polymerization of copolymer.

  18. Biofunctionalization of polycaprolactone scaffolds with RGD peptides for the better cells integration

    Science.gov (United States)

    Matveeva, V. G.; Seifalian, A. M.; Antonova, L. V.; Velikanova, E. A.; Sergeeva, E. A.; Krivkina, E. O.; Glushkova, T. V.; Kudryavtseva, Yu. A.; Barbarash, O. L.; Barbarash, L. S.

    2016-08-01

    Here we tested in vitro electrospun polycaprolactone (PCL) scaffolds carbodiimide linkage with RGD peptides and their unconjugated counterparts. The scaffolds possessed highly porous structure and were formed by randomly distributed fibers. Orange II staining and ninhydrin test confirmed successful amination of the PCL. For the assessment of cell adhesion, we colonized scaffolds with primary human fibroblasts and counted the number of alive and dead cells. After 6 days of incubation, the number of fibroblasts on the scaffolds modified by RGD peptides significantly exceeded the number on unmodified scaffolds; however, the distribution of the cells on functionalized scaffolds was uneven, possibly due to uneven distribution of RGD peptides. The percentage of dead cells on the scaffolds with RGD peptides was significantly lower compared to their unmodified counterparts. Therefore, conjugation of PCL scaffolds with RGD peptides improves their integration with cells. This can be used in regenerative medicine.

  19. Targeting In-Stent-Stenosis with RGD- and CXCL1-Coated Mini-Stents in Mice

    Science.gov (United States)

    Weinandy, Stefan; Schreiber, Fabian; Megens, Remco T. A.; Theelen, Wendy; Smeets, Ralf; Jockenhövel, Stefan; Gries, Thomas; Möller, Martin; Klee, Doris; Weber, Christian; Zernecke, Alma

    2016-01-01

    Atherosclerotic lesions that critically narrow the artery can necessitate an angioplasty and stent implantation. Long-term therapeutic effects, however, are limited by excessive arterial remodeling. We here employed a miniaturized nitinol-stent coated with star-shaped polyethylenglycole (star-PEG), and evaluated its bio-functionalization with RGD and CXCL1 for improving in-stent stenosis after implantation into carotid arteries of mice. Nitinol foils or stents (bare metal) were coated with star-PEG, and bio-functionalized with RGD, or RGD/CXCL1. Cell adhesion to star-PEG-coated nitinol foils was unaltered or reduced, whereas bio-functionalization with RGD but foremost RGD/CXCL1 increased adhesion of early angiogenic outgrowth cells (EOCs) and endothelial cells but not smooth muscle cells when compared with bare metal foils. Stimulation of cells with RGD/CXCL1 furthermore increased the proliferation of EOCs. In vivo, bio-functionalization with RGD/CXCL1 significantly reduced neointima formation and thrombus formation, and increased re-endothelialization in apoE-/- carotid arteries compared with bare-metal nitinol stents, star-PEG-coated stents, and stents bio-functionalized with RGD only. Bio-functionalization of star-PEG-coated nitinol-stents with RGD/CXCL1 reduced in-stent neointima formation. By supporting the adhesion and proliferation of endothelial progenitor cells, RGD/CXCL1 coating of stents may help to accelerate endothelial repair after stent implantation, and thus may harbor the potential to limit the complication of in-stent restenosis in clinical approaches. PMID:27192172

  20. Targeting In-Stent-Stenosis with RGD- and CXCL1-Coated Mini-Stents in Mice.

    Directory of Open Access Journals (Sweden)

    Sakine Simsekyilmaz

    Full Text Available Atherosclerotic lesions that critically narrow the artery can necessitate an angioplasty and stent implantation. Long-term therapeutic effects, however, are limited by excessive arterial remodeling. We here employed a miniaturized nitinol-stent coated with star-shaped polyethylenglycole (star-PEG, and evaluated its bio-functionalization with RGD and CXCL1 for improving in-stent stenosis after implantation into carotid arteries of mice. Nitinol foils or stents (bare metal were coated with star-PEG, and bio-functionalized with RGD, or RGD/CXCL1. Cell adhesion to star-PEG-coated nitinol foils was unaltered or reduced, whereas bio-functionalization with RGD but foremost RGD/CXCL1 increased adhesion of early angiogenic outgrowth cells (EOCs and endothelial cells but not smooth muscle cells when compared with bare metal foils. Stimulation of cells with RGD/CXCL1 furthermore increased the proliferation of EOCs. In vivo, bio-functionalization with RGD/CXCL1 significantly reduced neointima formation and thrombus formation, and increased re-endothelialization in apoE-/- carotid arteries compared with bare-metal nitinol stents, star-PEG-coated stents, and stents bio-functionalized with RGD only. Bio-functionalization of star-PEG-coated nitinol-stents with RGD/CXCL1 reduced in-stent neointima formation. By supporting the adhesion and proliferation of endothelial progenitor cells, RGD/CXCL1 coating of stents may help to accelerate endothelial repair after stent implantation, and thus may harbor the potential to limit the complication of in-stent restenosis in clinical approaches.

  1. Targeting In-Stent-Stenosis with RGD- and CXCL1-Coated Mini-Stents in Mice.

    Science.gov (United States)

    Simsekyilmaz, Sakine; Liehn, Elisa A; Weinandy, Stefan; Schreiber, Fabian; Megens, Remco T A; Theelen, Wendy; Smeets, Ralf; Jockenhövel, Stefan; Gries, Thomas; Möller, Martin; Klee, Doris; Weber, Christian; Zernecke, Alma

    2016-01-01

    Atherosclerotic lesions that critically narrow the artery can necessitate an angioplasty and stent implantation. Long-term therapeutic effects, however, are limited by excessive arterial remodeling. We here employed a miniaturized nitinol-stent coated with star-shaped polyethylenglycole (star-PEG), and evaluated its bio-functionalization with RGD and CXCL1 for improving in-stent stenosis after implantation into carotid arteries of mice. Nitinol foils or stents (bare metal) were coated with star-PEG, and bio-functionalized with RGD, or RGD/CXCL1. Cell adhesion to star-PEG-coated nitinol foils was unaltered or reduced, whereas bio-functionalization with RGD but foremost RGD/CXCL1 increased adhesion of early angiogenic outgrowth cells (EOCs) and endothelial cells but not smooth muscle cells when compared with bare metal foils. Stimulation of cells with RGD/CXCL1 furthermore increased the proliferation of EOCs. In vivo, bio-functionalization with RGD/CXCL1 significantly reduced neointima formation and thrombus formation, and increased re-endothelialization in apoE-/- carotid arteries compared with bare-metal nitinol stents, star-PEG-coated stents, and stents bio-functionalized with RGD only. Bio-functionalization of star-PEG-coated nitinol-stents with RGD/CXCL1 reduced in-stent neointima formation. By supporting the adhesion and proliferation of endothelial progenitor cells, RGD/CXCL1 coating of stents may help to accelerate endothelial repair after stent implantation, and thus may harbor the potential to limit the complication of in-stent restenosis in clinical approaches.

  2. Closed headpiece of integrin [alpah]IIb[beta]3 and its complex with an [alpha]IIb[beta]3-specific antagonist that does not induce opening

    Energy Technology Data Exchange (ETDEWEB)

    Zhu, Jieqing; Zhu, Jianghai; Negri, Ana; Provasi, Davide; Filizola, Marta; Coller, Barry S.; Springer, Timothy A. (Sinai); (Rockefeller); (CH-Boston)

    2011-08-24

    The platelet integrin {alpha}{sub IIb}{beta}{sub 3} is essential for hemostasis and thrombosis through its binding of adhesive plasma proteins. We have determined crystal structures of the {alpha}{sub IIb}{beta}{sub 3} headpiece in the absence of ligand and after soaking in RUC-1, a novel small molecule antagonist. In the absence of ligand, the {alpha}{sub IIb}{beta}{sub 3} headpiece is in a closed conformation, distinct from the open conformation visualized in presence of Arg-Gly-Asp (RGD) antagonists. In contrast to RGD antagonists, RUC-1 binds only to the {alpha}{sub IIb} subunit. Molecular dynamics revealed nearly identical binding. Two species-specific residues, {alpha}{sub IIb} Y190 and {alpha}{sub aIIb} D232, in the RUC-1 binding site were confirmed as important by mutagenesis. In sharp contrast to RGD-based antagonists, RUC-1 did not induce {alpha}{sub IIb}{beta}{sub 3} to adopt an open conformation, as determined by gel filtration and dynamic light scattering. These studies provide insights into the factors that regulate integrin headpiece opening, and demonstrate the molecular basis for a novel mechanism of integrin antagonism.

  3. Vitronectin-Based, Biomimetic Encapsulating Hydrogel Scaffolds Support Adipogenesis of Adipose Stem Cells.

    Science.gov (United States)

    Clevenger, Tracy N; Hinman, Cassidy R; Ashley Rubin, Rebekah K; Smither, Kate; Burke, Daniel J; Hawker, Craig J; Messina, Darin; Van Epps, Dennis; Clegg, Dennis O

    2016-04-01

    Soft tissue defects are relatively common, yet currently used reconstructive treatments have varying success rates, and serious potential complications such as unpredictable volume loss and reabsorption. Human adipose-derived stem cells (ASCs), isolated from liposuction aspirate have great potential for use in soft tissue regeneration, especially when combined with a supportive scaffold. To design scaffolds that promote differentiation of these cells down an adipogenic lineage, we characterized changes in the surrounding extracellular environment during adipogenic differentiation. We found expression changes in both extracellular matrix proteins, including increases in expression of collagen-IV and vitronectin, as well as changes in the integrin expression profile, with an increase in expression of integrins such as αVβ5 and α1β1. These integrins are known to specifically interact with vitronectin and collagen-IV, respectively, through binding to an Arg-Gly-Asp (RGD) sequence. When three different short RGD-containing peptides were incorporated into three-dimensional (3D) hydrogel cultures, it was found that an RGD-containing peptide derived from vitronectin provided strong initial attachment, maintained the desired morphology, and created optimal conditions for in vitro 3D adipogenic differentiation of ASCs. These results describe a simple, nontoxic encapsulating scaffold, capable of supporting the survival and desired differentiation of ASCs for the treatment of soft tissue defects. PMID:26956095

  4. Efficient preparation of 99mTc(III) '4+1' mixed-ligand complexes for peptide labeling with high specific activity

    International Nuclear Information System (INIS)

    An improved labeling procedure for peptides attached to organometallic 99mTc(III) '4+1' mixed-ligand complexes in which the radiometal is coordinated by a tripodal tetradentate chelator 2,2',2''-nitrilotriethanethiol (NS3) and a monodentate isocyanide ligand is presented. The labeling procedure was evaluated by the synthesis of [99mTc(NS3)(L2-RGD)]. The containing radiopharmaceutically interesting RGD-peptide cyclo[Arg-Gly-Asp-D-Tyr-Lys] was modified with 4-isocyanobutanoic acid (L2) as linker conjugated to N6-Lys to get the monodentate ligand L2-RGD. The structural identity of the 99mTc-conjugate was confirmed by comparison to a Re reference compound. The Tc- and Re-conjugates had matching retention times under identical HPLC conditions. The 99mTc-labeling was performed in a novel one-step procedure using the eluate of a 99Mo/99mTc generator, NS3, the isocyanide modified peptide, SnCl2, Na2EDTA, mannitol and ascorbic acid in the reaction mixture. Using optimized reagents it is possible to label 50 nmol peptide with 99mTc within 60 min at room temperature with a radiochemical yield higher than 95% and a specific activity of ∼20 GBq/μmol.

  5. Tobacco Mosaic Virus-Based 1D Nanorod-Drug Carrier via the Integrin-Mediated Endocytosis Pathway.

    Science.gov (United States)

    Tian, Ye; Gao, Sijia; Wu, Man; Liu, Xiangxiang; Qiao, Jing; Zhou, Quan; Jiang, Shidong; Niu, Zhongwei

    2016-05-01

    For cancer therapy, viruses have been utilized as excellent delivery vehicles because of their facile transfection efficiency in their host cells. However, their inherent immunogenicity has become the major obstacle for their translation into approved pharmaceuticals. Herein, we utilized rodlike plant virus, tobacco mosaic virus (TMV), which is nontoxic to mammals and mainly infects tobacco species, as anticancer nanorod-drug vector for cancer therapy study. Doxorubicin (DOX) was installed in the inner cavity of TMV by hydrazone bond, which enabled the pH-sensitive drug release property. Conjugation of cyclic Arg-Gly-Asp (cRGD) on the surface of TMV can enhance HeLa cell uptake of the carrier via the integrin-mediated endocytosis pathway. Comparing with free DOX, the cRGD-TMV-hydra-DOX vector had similar cell growth inhibition and much higher apoptosis efficiency on HeLa cells. Moreover, the in vivo assay assumed that cRGD-TMV-hydra-DOX behaved similar antitumor efficiency but much lower side effect on HeLa bearing Balb/c-nu mice. Our work provides novel insights into potentially cancer therapy based on rodlike plant viral nanocarriers. PMID:27062971

  6. Treatment of Klebsiella pneumoniae septicemia in normal and leukopenic mice by liposome-encapsulated muramyl tripeptide phosphatidylethanolamide

    NARCIS (Netherlands)

    P.M. Melissen (Petronella Maria Bernadette); W. van Vianen (Wim); I.A.J.M. Bakker-Woudenberg (Irma)

    1994-01-01

    textabstractThe effect of free muramyl tripeptide phosphatidylethanolamide (MTPPE) and liposome-encapsulated MTPPE (LE-MTPPE) on Klebsiella pneumoniae septicemia resulting from intraperitoneal bacterial inoculation was investigated in mice. When administering a single p

  7. Novel Approach to Prepare {sup 99m}Tc-Based Multivalent RGD Peptides

    Energy Technology Data Exchange (ETDEWEB)

    Shuang Liu

    2012-10-24

    This project presents a novel approach to prepare the {sup 99m}Tc-bridged multivalent RGD (arginine-glycine-aspartate) peptides. This project will focus on fundamentals of {sup 99m}Tc radiochemistry. The main objective of this project is to demonstrate the proof-of-principle for the proposed radiotracers. Once a kit formulation is developed for preparation of the {sup 99m}Tc-bridged multivalent RGD peptides, various tumor-bearing animal models will be used to evaluate their potential for SPECT (single photon-emission computed tomography) imaging of cancer. We have demonstrated that (1) multimerization of cyclic RGD peptides enhances the integrin {alpha}{sub v}{beta}{sub 3} bonding affinity and radiotracer tumor uptake; (2) addition of G{sub 3} or PEG{sub 4} linkers makes it possible for two RGD motifs in 3P-RGD{sub 2} and 3G-RGD{sub 2} to achieve simultaneous integrin {alpha}{sub v}{beta}{sub 3} binding; and (3) multimers are actually bivalent (not multivalent), the presence of extra RGD motifs can enhance the tumor retention time of the radiotracer.

  8. A High-Adhesive Lysine-Cyclic RGD Peptide Designed for Selective Cell Retention Technology.

    Science.gov (United States)

    Luo, Keyu; Mei, Tieniu; Li, Zhiqiang; Deng, Moyuan; Zhang, Zehua; Hou, Tianyong; Dong, Shiwu; Xie, Zhao; Xu, Jianzhong; Luo, Fei

    2016-06-01

    Cell adhesion is an important property of biomaterials used in selective cell retention (SCR) technology, which fabricates bone grafts rapidly in clinical settings. This could be improved by physical and biologic manipulations. To facilitate retention of the cells on the scaffold, especially osteoprogenitors from bone marrow in the convenient SCR procedure, a lysine-cyclic RGD (LcRGD) peptide was here designed to coordinate positively charged amino acids and the RGD sequence to enhance the adhesion performance of the scaffold. Demineralized bone matrix (DBM) is an important therapeutic resource, but its cell adhesion ability and osteoinductive capacity are low because of its processing. These capabilities can be increased to enhance the performance of DBM when used in SCR technology. Here, LcRGD peptide was used to modify DBM and produce a DBM/LcRGD composite. This composite exhibited enhanced adhesion performance on cultured human bone marrow-derived mesenchymal stem cells and retained more osteoprogenitors from bone marrow than other materials did. The DBM/LcRGD composite displayed a preferable osteoinduction in vitro and osteogenic capacity in vivo. Thus, LcRGD peptide as a commendable modifier of DBM applied in SCR technology can improve bone transplantation. PMID:27154386

  9. In Silico and in Vitro Study of Binding Affinity of Tripeptides to Amyloid β Fibrils: Implications for Alzheimer's Disease.

    Science.gov (United States)

    Viet, Man Hoang; Siposova, Katarina; Bednarikova, Zuzana; Antosova, Andrea; Nguyen, Truc Trang; Gazova, Zuzana; Li, Mai Suan

    2015-04-23

    Self-assembly of Aβ peptides into amyloid aggregates has been suggested as the major cause of Alzheimer's disease (AD). Nowadays, there is no medication for AD, but experimental data indicate that reversion of the process of amyloid aggregation reduces the symptoms of disease. In this paper, all 8000 tripeptides were studied for their ability to destroy Aβ fibrils. The docking method and the more sophisticated MM-PBSA (molecular mechanics Poisson-Boltzmann surface area) method were employed to calculate the binding affinity and mode of tripeptides to Aβ fibrils. The ability of these peptides to depolymerize Aβ fibrils was also investigated experimentally using atomic force microscopy and fluorescence spectroscopy (Thioflavin T assay). It was shown that tripeptides prefer to bind to hydrophobic regions of 6Aβ9-40 fibrils. Tripeptides WWW, WWP, WPW and PWW were found to be the most potent binders. In vitro experiments showed that tight-binding tripeptides have significant depolymerizing activities and their DC50 values determined from dose-response curves were in micromolar range. The ability of nonbinding (GAM, AAM) and weak-binding (IVL and VLA) tripeptides to destroy Aβ fibrils was negligible. In vitro data of tripeptide depolymerizing activities support the predictions obtained by molecular docking and all-atom simulation methods. Our results suggest that presence of multiple complexes of heterocycles forming by tryptophan and proline residues in tripeptides is crucial for their tight binding to Aβ fibrils as well as for extensive fibril depolymerization. We recommend PWW for further studies as it has the lowest experimental binding constant. PMID:25815792

  10. Cellular compatibility of RGD-modified chitosan nanofibers with aligned or random orientation

    Energy Technology Data Exchange (ETDEWEB)

    Wang Yanyan; Lue Lanxin; Feng Zhangqi; Xiao Zhongdang; Huang Ningping, E-mail: nphuang@seu.edu.c, E-mail: zdxiao@seu.edu.c [State Key Laboratory of Bioelectronics, School of Biological Science and Medical Engineering, Southeast University, Nanjing 210096 (China)

    2010-10-01

    Aligned and randomly oriented chitosan nanofibers were prepared by electrospinning. The fibers were modified with the RGD cell-adhesive peptide through a heterobifunctional crosslinker containing a segment of poly(ethylene glycol) (PEG). PEG rendered the surface hydrophilic and provided flexible spacers, allowing the preservation of the bioactivity of further captured RGD peptides. NIH 3T3 cells were used to test the cellular compatibility of these chitosan nanofibrous scaffolds. Cell morphology and viability were investigated by SEM, fluorescent staining and cell counting. The results indicate that RGD-modified surfaces significantly improve the cellular compatibility of chitosan nanofibers and suggest a good candidate as a scaffold employed in tissue engineering.

  11. Interruptions in the collagen repeating tripeptide pattern can promote supramolecular association

    OpenAIRE

    Hwang, Eileen S; Thiagarajan, Geetha; Parmar, Avanish S.; Brodsky, Barbara

    2010-01-01

    The standard collagen triple-helix requires a perfect (Gly-Xaa-Yaa)n sequence, yet all nonfibrillar collagens contain interruptions in this tripeptide repeating pattern. Defining the structural consequences of disruptions in the sequence pattern may shed light on the biological role of sequence interruptions, which have been suggested to play a role in molecular flexibility, collagen degradation, and ligand binding. Previous studies on model peptides with 1- and 4-residue interruptions showed...

  12. Biomaterial arrays with defined adhesion ligand densities and matrix stiffness identify distinct phenotypes for tumorigenic and nontumorigenic human mesenchymal cell types.

    Science.gov (United States)

    Hansen, Tyler D; Koepsel, Justin T; Le, Ngoc Nhi; Nguyen, Eric H; Zorn, Stefan; Parlato, Matthew; Loveland, Samuel G; Schwartz, Michael P; Murphy, William L

    2014-05-01

    Here, we aimed to investigate migration of a model tumor cell line (HT-1080 fibrosarcoma cells, HT-1080s) using synthetic biomaterials to systematically vary peptide ligand density and substrate stiffness. A range of substrate elastic moduli were investigated by using poly(ethylene glycol) (PEG) hydrogel arrays (0.34 - 17 kPa) and self-assembled monolayer (SAM) arrays (~0.1-1 GPa), while cell adhesion was tuned by varying the presentation of Arg-Gly-Asp (RGD)-containing peptides. HT-1080 motility was insensitive to cell adhesion ligand density on RGD-SAMs, as they migrated with similar speed and directionality for a wide range of RGD densities (0.2-5% mol fraction RGD). Similarly, HT-1080 migration speed was weakly dependent on adhesion on 0.34 kPa PEG surfaces. On 13 kPa surfaces, a sharp initial increase in cell speed was observed at low RGD concentration, with no further changes observed as RGD concentration was increased further. An increase in cell speed ~ two-fold for the 13 kPa relative to the 0.34 kPa PEG surface suggested an important role for substrate stiffness in mediating motility, which was confirmed for HT-1080s migrating on variable modulus PEG hydrogels with constant RGD concentration. Notably, despite ~ two-fold changes in cell speed over a wide range of moduli, HT-1080s adopted rounded morphologies on all surfaces investigated, which contrasted with well spread primary human mesenchymal stem cells (hMSCs). Taken together, our results demonstrate that HT-1080s are morphologically distinct from primary mesenchymal cells (hMSCs) and migrate with minimal dependence on cell adhesion for surfaces within a wide range of moduli, whereas motility is strongly influenced by matrix mechanical properties.

  13. PROTEIN ADSORPTION AND CELL ADHESION ON RGD-FUNCTIONALIZED SILICON SUBSTRATE SURFACES

    Institute of Scientific and Technical Information of China (English)

    Wei-fang Tong; Xiao-li Liu; Fei Pan; Zhao-qiang Wu; Wen-wen Jiang

    2013-01-01

    A method was developed to modify silicon surfaces with good protein resistance and specific cell attachment.A silicon surface was initially deposited using a block copolymer of N-vinylpyrrolidone (NVP) and 2-hydroxyethyl methacrylate (HEMA) (PVP-b-PHEMA) film through surface-initiated atom transfer radical polymerization and then further immobilized using a short arginine-glycine-aspartate (RGD) peptide.Our results demonstrate that the RGD-modified surfaces (Si-RGD) can suppress non-specific adsorption of proteins and induce the adhesion of L929 cells.The Si-RGD surface exhibited higher cell proliferation rates than the unmodified silicon surface.This research established a simple method for the fabrication of dual-functional silicon surface that combines antifouling and cell attachment promotion.

  14. Synthesis and Cell Adhesive Properties of Linear and Cyclic RGD Functionalized Polynorbornene Thin Films

    OpenAIRE

    Patel, Paresma R.; Kiser, Rosemary Conrad; Lu, Ying Y.; Fong, Eileen; Ho, Wilson C.; Tirrell, David A.; Grubbs, Robert H.

    2012-01-01

    Described herein is the efficient synthesis and evaluation of bioactive arginine-glycine-aspartic acid (RGD) functionalized polynorbornene-based materials for cell adhesion and spreading. Polynorbornenes containing either linear or cyclic RGD peptides were synthesized by ring-opening metathesis polymerization (ROMP) using the well-defined ruthenium initiator [(H_(2)IMes)(pyr)_(2)(Cl)_(2)Ru═CHPh]. The random copolymerization of three separate norbornene monomers allowed for the incorporation o...

  15. RGD Gifted PDLLA-PRGD Conduits Promotes the Sciatic Nerve Regeneration

    Institute of Scientific and Technical Information of China (English)

    FANG Xiaoqing; QIU Tong; XIE Lijuan; YIN Yixia; LI Binbin; YAN Qiongjiao; DAI Honglian; WANG Xinyu; LI Shipu

    2014-01-01

    Schwann cells play a key role in peripheral nerve growth and regeneration. The aim of this study was to evaluate the effects of RGD peptides on Schwann cell behavior, and to identify the effects of the modified PDLLA films with RGD in vivo. The results revealed that RGD coating with the concentration of 100-500 ug/mL promoted the cell proliferation and boosted the cell migration. Molecularly, RGD coating also enhanced the expression of the proliferation related genes (c-fos and c-jun) and the cell behavior related genes (actin, tublin, tau and MAP1) at first stages of the seeding, which is similar to the effects from laminin coating. In vivo, RGD addition improved the recovery efficiency of the transected nerve in regard of the more survived Schwann cells in vivo and the formation of more mature myelin sheath. Taken together, RGD peptides are good candidates to enhance the biocompatibility of the biomaterials and facilitate the peripheral nerve regeneration by prompting responses in Schwann cells.

  16. Low-Molecular Weight Polyethylenimine Modified with Pluronic 123 and RGD- or Chimeric RGD-NLS Peptide: Characteristics and Transfection Efficacy of Their Complexes with Plasmid DNA

    OpenAIRE

    Jing Hu; Wenfang Zhao; Kehai Liu; Qian Yu; Yuan Mao; Zeyu Lu; Yaguang Zhang; Manman Zhu

    2016-01-01

    To solve the problem of transfection efficiency vs. cytotoxicity and tumor-targeting ability when polyethylenimine (PEI) was used as a nonviral gene delivery vector, new degradable PEI polymers were synthesized via cross-linking low-molecular-weight PEI with Pluronic P123 and then further coupled with a targeting peptide R4 (RGD) and a bifunctional R11 (RGD-NLS), which were termed as P123-PEI-R4 and P123-PEI-R11, respectively. Agarose gel electrophoresis showed that both P123-PEI-R4 and P123-...

  17. Hypertonicity-induced transmitter release at Drosophila neuromuscular junctions is partly mediated by integrins and cAMP/protein kinase A

    Science.gov (United States)

    Suzuki, Kazuhiro; Grinnell, Alan D.; Kidokoro, Yoshiaki

    2002-01-01

    The frequency of quantal transmitter release increases upon application of hypertonic solutions. This effect bypasses the Ca(2+) triggering step, but requires the presence of key molecules involved in vesicle fusion, and hence could be a useful tool for dissecting the molecular process of vesicle fusion. We have examined the hypertonicity response at neuromuscular junctions of Drosophila embryos in Ca(2+)-free saline. Relative to wild-type, the response induced by puff application of hypertonic solution was enhanced in a mutant, dunce, in which the cAMP level is elevated, or in wild-type embryos treated with forskolin, an activator of adenylyl cyclase, while protein kinase A (PKA) inhibitors decreased it. The response was also smaller in a mutant, DC0, which lacks the major subunit of PKA. Thus the cAMP/PKA cascade is involved in the hypertonicity response. Peptides containing the sequence Arg-Gly-Asp (RGD), which inhibit binding of integrins to natural ligands, reduced the response, whereas a peptide containing the non-binding sequence Arg-Gly-Glu (RGE) did not. A reduced response persisted in a mutant, myospheroid, which expresses no integrins, and the response in DC0 was unaffected by RGD peptides. These data indicate that there are at lease two components in the hypertonicity response: one that is integrin mediated and involves the cAMP/PKA cascade, and another that is not integrin mediated and does not involve the cAMP/PKA cascade.

  18. Synthesis of Zn-Cu-In-S/ZnS Core/Shell Quantum Dots with Inhibited Blue-Shift Photoluminescence and Applications for Tumor Targeted Bioimaging

    Directory of Open Access Journals (Sweden)

    Weisheng Guo, Na chen, Yu Tu, Chunhong Dong, Bingbo Zhang, Chunhong Hu, Jin Chang

    2013-01-01

    Full Text Available A facile strategy is reported here for synthesis of Zn-Cu-In-S/ZnS (ZCIS/ZnS core/shell QDs to address the synthetic issues that the unexpected blue-shift of CuInS2-based nanocrystals. In this strategy, Zn2+ ions are intentionally employed for the synthesis of alloyed ZCIS core QDs before ZnS shell coating, which contributes to the reduced blue-shift in photoluminescence (PL emission. The experimental results demonstrate this elaborate facile strategy is effective for the reduction of blue-shift during shell growth. Particularly, a hypothesis is proposed and proved for explanation of this effective strategy. Namely, both cation exchange inhibition and ions accumulation are involved during the synthesis of ZCIS/ZnS QDs. Furthermore, the obtained near infrared (NIR ZCIS/ZnS QDs are transferred into aqueous phase by a polymer coating technique and coupled with cyclic Arg-Gly-Asp peptide (cRGD peptides. After confirmation of biocompability by cytotoxicity test on normal 3T3 cells, these QDs are injected via tail vein into nude mice bearing U87 MG tumor. The result indicates that the signals detected in the tumor region are much more distinguishing injected with ZCIS/ZnS-cRGD QDs than that injected with ZCIS/ZnS QDs.

  19. Ca2+-dependent cessation of cytoplasmic streaming induced by hypertonic treatment in Vallisneria mesophyll cells: possible role of cell wall-plasma membrane adhesion.

    Science.gov (United States)

    Hayashi, Teruyuki; Takagi, Shingo

    2003-10-01

    In mesophyll cells of the aquatic angiosperm Vallisneria gigantea Graebner, a rapid and transient inhibition of cytoplasmic streaming was induced by hypertonic treatment with sorbitol. Higher concentrations of sorbitol induced the response more rapidly and in more cells. The response to hypertonic treatment was strictly dependent on the presence of extracellular Ca2+ and was sensitive to Ca2+-channel blockers, including the stretch-activated Ca2+-channel blocker Gd3+. Deplasmolyzed cells never responded to a second hypertonic treatment administered immediately after plasmolysis and subsequent deplasmolysis. Responsiveness was gradually recovered during 24 h of incubation; however, cycloheximide, cordycepin, and trypsin completely suppressed the recovery. Although an Arg-Gly-Asp (RGD) hexapeptide markedly disturbed the pattern of cytoplasmic streaming, it exhibited no specific effects on the response to hypertonic treatment or on the recovery of responsiveness. Taken together, our results demonstrate that leaf mesophyll cells in a multicellular plant can respond to mechanical stimuli and that a Ca2+ influx through stretch-activated Ca2+ channels plays an indispensable role in the response. Furthermore, the possible involvement of RGD-insensitive but trypsin-sensitive protein factor(s), whose function is impaired by detachment of the plasma membrane from the cell wall, is suggested. PMID:14581627

  20. Design of a Vitronectin-Based Recombinant Protein as a Defined Substrate for Differentiation of Human Pluripotent Stem Cells into Hepatocyte-Like Cells.

    Directory of Open Access Journals (Sweden)

    Masato Nagaoka

    Full Text Available Maintenance and differentiation of human pluripotent stem cells (hPSCs usually requires culture on a substrate for cell adhesion. A commonly used substratum is Matrigel purified from Engelbreth-Holm-Swarm sarcoma cells, and consists of a complex mixture of extracellular matrix proteins, proteoglycans, and growth factors. Several studies have successfully induced differentiation of hepatocyte-like cells from hPSCs. However, most of these studies have used Matrigel as a cell adhesion substrate, which is not a defined culture condition. In an attempt to generate a substratum that supports undifferentiated properties and differentiation into hepatic lineage cells, we designed novel substrates consisting of vitronectin fragments fused to the IgG Fc domain. hPSCs adhered to these substrates via interactions between integrins and the RGD (Arg-Gly-Asp motif, and the cells maintained their undifferentiated phenotypes. Using a previously established differentiation protocol, hPSCs were efficiently differentiated into mesendodermal and hepatic lineage cells on a vitronectin fragment-containing substrate. We found that full-length vitronectin did not support stable cell adhesion during the specification stage. Furthermore, the vitronectin fragment with the minimal RGD-containing domain was sufficient for differentiation of human induced pluripotent stem cells into hepatic lineage cells under completely defined conditions that facilitate the clinical application of cells differentiated from hPSCs.

  1. Interactions between chitosan and cells measured by AFM

    Energy Technology Data Exchange (ETDEWEB)

    Hsiao, Sheng-Wen; Thien, Doan Van Hong; Ho, Ming-Hua [Department of Chemical Engineering, National Taiwan University of Science and Technology, Taipei 10617, Taiwan (China); Hsieh, Hsyue-Jen [Department of Chemical Engineering, National Taiwan University, Taipei 10617, Taiwan (China); Li, Chung-Hsing [Division of Orthodontics and Pediatric Dentistry, Department of Dentistry, Tri-Service General Hospital, Taipei, Taiwan (China); Hung, Chang-Hsiang [Department of Dentistry, Kinmen Hospital Department of Health, Taiwan (China); Li, Hsi-Hsin, E-mail: mhho@mail.ntust.edu.t [Deputy Superintendent, Kinmen Hospital Department of Health, Taiwan (China)

    2010-10-01

    Chitosan, a biocompatible material that has been widely used in bone tissue engineering, is believed to have a high affinity to osteoblastic cells. This research is the first to prove this hypothesis. By using atomic force microscopy (AFM) with a chitosan-modified cantilever, quantitative evaluation of the interforce between chitosan and cells was carried out. A chitosan tip functionalized with Arg-Gly-Asp (RGD) was also used to measure the interforce between RGD-chitosan and osteoblastic cells. This research concluded by examining cell adhesion and spreading of chitosan substrates as further characterization of the interactions between cells and chitosan. The force measured by AFM showed that the interforce between chitosan and osteoblasts was the highest (209 nN). The smallest adhesion force (61.8 nN) appeared between chitosan and muscle fibroblasts, which did not demonstrate any osteoblastic properties. This result proved that there was a significant interaction between chitosan and bone cells, and correlated with the observations of cell attachment and spreading. The technique developed in this research directly quantified the adhesion between chitosan and cells. This is the first study to demonstrate that specific interaction exists between chitosan and osteoblasts.

  2. Interactions between chitosan and cells measured by AFM

    International Nuclear Information System (INIS)

    Chitosan, a biocompatible material that has been widely used in bone tissue engineering, is believed to have a high affinity to osteoblastic cells. This research is the first to prove this hypothesis. By using atomic force microscopy (AFM) with a chitosan-modified cantilever, quantitative evaluation of the interforce between chitosan and cells was carried out. A chitosan tip functionalized with Arg-Gly-Asp (RGD) was also used to measure the interforce between RGD-chitosan and osteoblastic cells. This research concluded by examining cell adhesion and spreading of chitosan substrates as further characterization of the interactions between cells and chitosan. The force measured by AFM showed that the interforce between chitosan and osteoblasts was the highest (209 nN). The smallest adhesion force (61.8 nN) appeared between chitosan and muscle fibroblasts, which did not demonstrate any osteoblastic properties. This result proved that there was a significant interaction between chitosan and bone cells, and correlated with the observations of cell attachment and spreading. The technique developed in this research directly quantified the adhesion between chitosan and cells. This is the first study to demonstrate that specific interaction exists between chitosan and osteoblasts.

  3. Improving Drug Penetrability with iRGD Leverages the Therapeutic Response to Sorafenib and Doxorubicin in Hepatocellular Carcinoma.

    Science.gov (United States)

    Schmithals, Christian; Köberle, Verena; Korkusuz, Hüdayi; Pleli, Thomas; Kakoschky, Bianca; Augusto, Eduardo Alonso; Ibrahim, Ahmed Atef; Arencibia, Jose M; Vafaizadeh, Vida; Groner, Bernd; Korf, Horst-Werner; Kronenberger, Bernd; Zeuzem, Stefan; Vogl, Thomas J; Waidmann, Oliver; Piiper, Albrecht

    2015-08-01

    iRGD is a derivative of the integrin-binding peptide RGD, which selectively increases the penetrability of tumor tissue to various coadministered substances in several preclinical models. In this study, we investigated the ability of iRGD to improve the delivery of sorafenib and doxorubicin therapy in hepatocellular carcinoma (HCC) using established mouse models of the disease. A contrast-enhanced MRI method was developed in parallel to assess the in vivo effects of iRGD in this setting. We found that iRGD improved the delivery of marker substances to the tumors of HCC-bearing mice about three-fold without a parallel increase in normal tissues. Control peptides lacking the critical CendR motif had no effect. Similarly, iRGD also selectively increased the signal intensity from tumors in Gd-DTPA-enhanced MRI. In terms of antitumor efficacy, iRGD coadministration significantly augmented the individual inhibitory effects of sorafenib and doxorubicin without increasing systemic toxicity. Overall, our results offered a preclinical proof of concept for the use of iRGD coadministration as a strategy to widen the therapeutic window for HCC chemotherapy, as monitored by Gd-DTPA-enhanced MRI as a noninvasive, clinically applicable method to identify iRGD-reactive tumors. PMID:26239478

  4. Preparation of 177Lu-DOTA/DTPA-Bz-Cys-RGD dimer and biodistribution evaluation in normal mice

    International Nuclear Information System (INIS)

    177Lu-DOTA-Bz-Cys-RGD dimer and 177Lu-DTPA-Bz-Cys-RGD dimer were prepared, and the in vitro and in vivo properties were compared. TLC and HPLC show that the labeling yields of two radiolabeled compounds are more than 95% under optimal conditions (pH=5.0, reacting at 100 degree C for 15-20 min), and the two radiolabeled compounds show pretty good in vitro stability. HPLC analyses and lg P values reveal that lipophilicity of 177Lu-DOTA-Bz-Cys- RGD dimer is higher than 177Lu-DTPA-Bz-Cys-RGD dimer. The uptake of 177Lu-DTPA-Bz-Cys- RGD dimer in other tissues is significantly higher than that of 177Lu-DOTA-Bz-Cys-RGD dimer at 4 h postinjection, except for blood and spleen. The in vivo stability of 177Lu-DOTA-Bz-Cys-RGD dimer is much better than 177Lu-DTPA-Bz-Cys-RGD dimer. Bz-DOTA is an ideal bifunctional chelator for 177Lu labeling of RGD dimer. (authors)

  5. Engineered peptide-based nanobiomaterials for electrochemical cell chip

    Science.gov (United States)

    Kafi, Md. Abdul; Cho, Hyeon-Yeol; Choi, Jeong-Woo

    2016-07-01

    Biomaterials having cell adhesion ability are considered to be integral part of a cell chip. A number of researches have been carried out to search for a suitable material for effective immobilization of cell on substrate. Engineered ECM materials or their components like collagen, Poly- l-Lysine (PLL), Arg-Gly-Asp (RGD) peptide have been extensively used for mammalian cell adhesion and proliferation with the aim of tissue regeneration or cell based sensing application. This review focuses on the various approaches for two- and three-dimensionally patterned nanostructures of a short peptide i.e. RGD peptide on chip surfaces together with their effects on cell behaviors and electrochemical measurements. Most of the study concluded with positive remarks on the well-oriented engineered RGD peptide over their homogenous thin film. The engineered RGD peptide not only influences cell adhesion, spreading and proliferation but also their periodic nano-arrays directly influence electrochemical measurements of the chips. The electrochemical signals found to be enhanced when RGD peptides were used in well-defined two-dimensional nano-arrays. The topographic alteration of three-dimensional structure of engineered RGD peptide was reported to be suitably contacted with the integrin receptors of cellular membrane which results indicated the enhanced cell-electrode adhesion and efficient electron exchange phenomenon. This enhanced electrochemical signal increases the sensitivity of the chip against the target analytes. Therefore, development of engineered cellular recognizable peptides and its 3D topological design for fabrication of cell chip will provide the synergetic effect on bio-affinity, sensitivity and accuracy for the in situ real-time monitoring of analytes.

  6. Evaluation of a Fiber-Modified Adenovirus Vector Vaccine against Foot-and-Mouth Disease in Cattle

    Science.gov (United States)

    Medina, Gisselle N.; Montiel, Nestor; Diaz-San Segundo, Fayna; Sturza, Diego; Ramirez-Medina, Elizabeth; Grubman, Marvin J.

    2015-01-01

    Novel vaccination approaches against foot-and-mouth disease (FMD) include the use of replication-defective human adenovirus type 5 (Ad5) vectors that contain the capsid-encoding regions of FMD virus (FMDV). Ad5 containing serotype A24 capsid sequences (Ad5.A24) has proved to be effective as a vaccine against FMD in livestock species. However, Ad5-vectored FMDV serotype O1 Campos vaccine (Ad5.O1C.2B) provides only partial protection of cattle against homologous challenge. It has been reported that a fiber-modified Ad5 vector expressing Arg-Gly-Asp (RGD) enhances transduction of antigen-presenting cells (APC) in mice. In the current study, we assessed the efficacy of a fiber-modified Ad5 (Adt.O1C.2B.RGD) in cattle. Expression of FMDV capsid proteins was superior in cultured cells infected with the RGD-modified vector. Furthermore, transgene expression of Adt.O1C.2B.RGD was enhanced in cell lines that constitutively express integrin αvβ6, a known receptor for FMDV. In contrast, capsid expression in cattle-derived enriched APC populations was not enhanced by infection with this vector. Our data showed that vaccination with the two vectors yielded similar levels of protection against FMD in cattle. Although none of the vaccinated animals had detectable viremia, FMDV RNA was detected in serum samples from animals with clinical signs. Interestingly, CD4+ and CD8+ gamma interferon (IFN-γ)+ cell responses were detected at significantly higher levels in animals vaccinated with Adt.O1C.2B.RGD than in animals vaccinated with Ad5.O1C.2B. Our results suggest that inclusion of an RGD motif in the fiber of Ad5-vectored FMD vaccine improves transgene delivery and cell-mediated immunity but does not significantly enhance vaccine performance in cattle. PMID:26607309

  7. Evaluation of a Fiber-Modified Adenovirus Vector Vaccine against Foot-and-Mouth Disease in Cattle.

    Science.gov (United States)

    Medina, Gisselle N; Montiel, Nestor; Diaz-San Segundo, Fayna; Sturza, Diego; Ramirez-Medina, Elizabeth; Grubman, Marvin J; de los Santos, Teresa

    2015-11-25

    Novel vaccination approaches against foot-and-mouth disease (FMD) include the use of replication-defective human adenovirus type 5 (Ad5) vectors that contain the capsid-encoding regions of FMD virus (FMDV). Ad5 containing serotype A24 capsid sequences (Ad5.A24) has proved to be effective as a vaccine against FMD in livestock species. However, Ad5-vectored FMDV serotype O1 Campos vaccine (Ad5.O1C.2B) provides only partial protection of cattle against homologous challenge. It has been reported that a fiber-modified Ad5 vector expressing Arg-Gly-Asp (RGD) enhances transduction of antigen-presenting cells (APC) in mice. In the current study, we assessed the efficacy of a fiber-modified Ad5 (Adt.O1C.2B.RGD) in cattle. Expression of FMDV capsid proteins was superior in cultured cells infected with the RGD-modified vector. Furthermore, transgene expression of Adt.O1C.2B.RGD was enhanced in cell lines that constitutively express integrin αvβ6, a known receptor for FMDV. In contrast, capsid expression in cattle-derived enriched APC populations was not enhanced by infection with this vector. Our data showed that vaccination with the two vectors yielded similar levels of protection against FMD in cattle. Although none of the vaccinated animals had detectable viremia, FMDV RNA was detected in serum samples from animals with clinical signs. Interestingly, CD4(+) and CD8(+) gamma interferon (IFN-γ)(+) cell responses were detected at significantly higher levels in animals vaccinated with Adt.O1C.2B.RGD than in animals vaccinated with Ad5.O1C.2B. Our results suggest that inclusion of an RGD motif in the fiber of Ad5-vectored FMD vaccine improves transgene delivery and cell-mediated immunity but does not significantly enhance vaccine performance in cattle.

  8. Efficacy evaluation of two synthetic lysine lipidated tripeptides as vaccine adjuvants against HBsAg.

    Science.gov (United States)

    Sidiq, Tabasum; Khajuria, Anamika; Shafi, Syed; Ismail, Tabasum; Sampath Kumar, Halmathur; Kannappa Srinivas, Vellimedu; Krishna, Ella; Kamal Johri, Rakesh

    2013-04-01

    In the present investigation, adjuvant potential of two novel lipidated tripeptide lysine derivatives (KKSM and KKSMB) was evaluated using various in vitro and animal-derived models of humoral and cell-mediated immune events in response to hepatitis B surface antigen (HBsAg). The results were compared with alum adjuvanted with HBsAg. Both these molecules were found to stimulate anti-HBsAg IgG and neutralizing (IgG1 and IgG2a) antibody titres in mice sera. The two molecules stimulated the proliferation of T-lymphocyte sub-sets (CD4/CD8) as well as the production of soluble mediators of Th1 (IL-2 and IFN-γ) and Th2 response (IL-4) in spleen cell culture supernatant. Furthermore, the two lipidated tripeptides enhanced the CD4, CD8, CD3 and CD19 cell populations as well as CD4/CD8 derived IL-2, IL-4, IFN-γ and TNF-α in whole blood of treated mice. There was found to be the significant enhancement in the release of IL-12, IFN-γ and nitrite content in macrophage supernatant. Moreover, the two lipidated tripeptides enhanced the population of CD80 and CD86 in spleen-derived macrophages and did not show any hemolytic effect on rabbit RBCs. Taken together, these results suggest that both these molecules are the potent enhancers of anti-HBsAg immune response via augmenting Th1/Th2 response in a dose dependent manner. PMID:23474022

  9. Osteoblastic potency of bone marrow cells cultivated on functionalized biometals with cyclic RGD-peptide.

    Science.gov (United States)

    Jäger, M; Böge, C; Janissen, R; Rohrbeck, D; Hülsen, T; Lensing-Höhn, S; Krauspe, R; Herten, M

    2013-10-01

    The fixation of cementless endoprostheses requires excellent fixation at the bone implant interface. Although the surface structures of these implants are designed to promote osteoblastic differentiation, poor bone quality may prevent or delay osseointegration. There is evidence that RGD peptides known as recognition motifs for various integrins, promote cellular adhesion, influence cellular proliferation, and differentiation of local cells. In this study, five different metal surfaces were analyzed: Sandblasted (TiSa) and polished (TiPol) Ti6Al4V, porocoated (CCPor) and polished (CCPol) cobalt chrome and polished stainless steel (SS) were coated by ethanol amine and poly(ethylene glycol) to attach covalently RGD peptides. Human mesenchymal stromal cells of healthy donors were cultivated onto prior functionalized metal surfaces for 14 days without osteogenic stimulation. Cell proliferation and differentiation were quantitatively evaluated for native (I), NaOH pre-activated (II), NaOH pre-activated, and PEG-coated (III) as well as for RGD (IV) coated surfaces. The RGD immobilization efficiency was analyzed by epi-fluorescence spectroscopy, cell morphology was documented by light and scanning electron microscopy. The RGD-binding efficiency was TiSa > TiPol > SS > CCPor > CCPol. RGD coated surfaces showed the highest average cell proliferation on CCPol > SS > CCPor > TiSa ≥ TiPol, whereas cellular differentiation mostly correlated with the observed proliferation results, such as CCPol > TiSa > SS > CCPor > TiPol. Considering statistical analyses (significance level of α = 0.05), the RGD-coating of all biometals in comparison and in respect of their particular controls showed no significant improvement in cellular proliferation and osteoblastic differentiation. PMID:23529934

  10. 68Ga-DOTA-RGD peptide: biodistribution and binding into atherosclerotic plaques in mice

    International Nuclear Information System (INIS)

    Increased expression of αvβ3/αvβ5 integrin is involved in angiogenesis and the inflammatory process in atherosclerotic plaques. The novel 68Ga-DOTA-RGD peptide binds with high affinity to αvβ3/αvβ5 integrin. The aim of this study was to investigate the uptake of the 68Ga-DOTA-RGD peptide in atherosclerotic plaques. Uptake of intravenously administered 68Ga-DOTA-RGD peptide was studied ex vivo in excised tissue samples and aortic sections of LDLR-/-ApoB100/100 atherosclerotic mice. The uptake of the tracer in aortic cryosections was examined by using digital autoradiography. Subsequently, the autoradiographs were combined with histological and immunohistological analysis of the sections. DOTA-RGD peptide was successfully labelled with the generator-produced 68Ga. The tracer had reasonably good specific radioactivity (8.7 ± 1.1 GBq/μmol) and was quite stable in vivo. According to ex vivo biodistribution results, 68Ga-DOTA-RGD was cleared rapidly from the blood circulation and excreted through the kidneys to the urine with high radioactivity in the intestine, lungs, spleen and liver. Autoradiography results showed significantly higher uptake of 68Ga-DOTA-RGD peptide in the atherosclerotic plaques compared to healthy vessel wall (mean ratio ± SD 1.4 ± 0.1, p = 0.0004). We observed that 68Ga-DOTA-RGD is accumulated into the plaques of atherosclerotic mice. However, this data only shows the feasibility of the approach, while the clinical significance still remains to be proven. Further studies are warranted to assess the uptake of this tracer into human atherosclerotic plaques. (orig.)

  11. Phe-Gly dipeptidomimetics designed for the di-/tripeptide transporters PEPT1 and PEPT2

    DEFF Research Database (Denmark)

    Våbenø, Jon; Lejon, Tore; Nielsen, Carsten Uhd;

    2004-01-01

    A series of five Phe-Gly dipeptidomimetics containing different amide bond replacements have been synthesized in a facile way from the readily available unsaturated ketoester 1, and their affinities for the di-/tripeptide transporters hPEPT1 (Caco-2 cells) and rPEPT2 (SKPT cells) were tested. The...... inhibitor Gly-Pro, indicating contribution from an active transport component. No active uptake of 3a and 4a was observed. Transepithelial transport studies also indicated active transport of 2a across Caco-2 monolayers....

  12. Synthesis of octadecyl esters of histidine-containing tripeptides as potential regulators of plant growth

    Energy Technology Data Exchange (ETDEWEB)

    Ogrel, A.A.; Zvonkova, E.N. [Lomonosov Academy of Fine Chemical Technology, Moscow (Russian Federation); Gafurov, R.G. [Institute of Physiologically Active Substances, Chernogolovka (Russian Federation)

    1995-08-01

    Octadecyl esters of dipeptides and tripeptides of the type Phe-His, Val-His, Phe-Val-His and Val-Phe-His were synthesized using different methods. The minimum energy conformations of these peptides were calculated with computer minimization programs and compared with those of paclobutrazol, a well-known regulator of plant growth. It was demonstrated that the elongation of the peptide chain leads to a higher topochemical correspondence between paclobutrazol and the peptide derivatives than between paclobutrazol and amino acid derivatives. 9 refs., 2 figs., 3 tabs.

  13. Genetic characterization and molecular cloning of the tripeptide permease (tpp) genes of Salmonella typhimurium.

    OpenAIRE

    Gibson, M M; Price, M; Higgins, C F

    1984-01-01

    Of the three bacterial peptide transport systems only one, the oligopeptide permease, has been characterized in any detail. We have now isolated Salmonella typhimurium mutants deficient in a second transport system, the tripeptide permease (Tpp), using the toxic peptide alafosfalin. Alafosfalin resistance mutations map at three loci, the gene encoding peptidase A (pepA) and two transport-defective loci, tppA and tppB. Locus tppA has been mapped to 74 min on the S. typhimurium chromosome, cotr...

  14. Sequence Determination of a Novel Tripeptide Isolated from the Young Leaves of Azadirachta indica A. Juss

    Directory of Open Access Journals (Sweden)

    M. Rajeswari Prabha

    2013-01-01

    Full Text Available The neem tree has long been recognized for its unique properties, both against insects and in improving human health. Every part of the tree has been used as a traditional medicine for household remedy against various human ailments, from antiquity. Although the occurrence of various phytochemicals in neem has been studied, we have identified the presence of a novel tripeptide in the young leaves of neem using a simple and inexpensive paper chromatographic method, detected by Cu(II-ninhydrin reagent. The peptide nature of the isolated compound is confirmed by spectral studies. The sequence of the peptide is determined using de novo sequencing by tandem MS after purification.

  15. Development of a new anti-cancer agent for targeted radionuclide therapy: β- radiolabeled RAFT-RGD

    International Nuclear Information System (INIS)

    β-emitters radiolabeled RAFT-RGD as new agents for internal targeted radiotherapy. The αvβ3 integrin is known to play an important role in tumor-induced angiogenesis, tumor proliferation, survival and metastasis. Because of its overexpression on neo-endothelial cells such as those present in growing tumors, as well as on tumor cells of various origins, αvβ3 integrin is an attractive molecular target for diagnosis and therapy of the rapidly growing and metastatic tumors. A tetrameric RGD-based peptide, regioselectively addressable functionalized template-(cyclo-[RGDfK])4 (RAFT-RGD), specifically targets integrin αvβ3 in vitro and in vivo. RAFT-RGD has been used for tumor imaging and drug targeting. This study is the first to evaluate the therapeutic potential of the β-emitters radiolabeled tetrameric RGD peptide RAFT-RGD in a Nude mouse model of αvβ3 -expressing tumors. An injection of 37 MBq of 90Y-RAFT-RGD or 177Lu-RAFT-RGD in mice with αvβ3 -positive tumors caused a significant growth delay as compared with mice treated with 37 MBq of 90Y-RAFT-RAD or 177Lu-RAFT-RAD or untreated mice. In comparison, an injection of 30 MBq of 90Y-RAFT-RGD had no efficacy for the treatment of αvβ3 -negative tumors. 90Y-RAFT-RGD and 177Lu-RAFT-RGD are potent αvβ3 -expressing tumor targeting agents for internal targeted radiotherapy. (author)

  16. RGD-peptide conjugated inulin-ibuprofen nanoparticles for targeted delivery of Epirubicin.

    Science.gov (United States)

    Zhang, Luzhong; Li, Guicai; Gao, Ming; Liu, Xin; Ji, Bing; Hua, Ruheng; Zhou, Youlang; Yang, Yumin

    2016-08-01

    Recently, chemotherapy-based polymeric nanoparticles have been extensively investigated for solid tumor treatment. Tumor targeted nanoparticles demonstrated great potential for improved accumulation in the tumor tissue, superior anticancer activity and reduced side effects. Thus, inulin-ibuprofen polymer was synthesized by esterification between inulin and ibuprofen, and RGD targeted epirubicin (EPB) loaded nanoparticles were prepared by the self-assembly of inulin-ibuprofen polymer and in situ encapsulation of EPB. RGD conjugated EPB loaded nanoparticles were characterized by dynamic light scattering (DLS) and transmission electron microscope (TEM). The EPB release from the nanoparticles showed pH-dependent profile and accelerated by the decreased pH value, which would favor the effective drug delivery in vivo. Intracellular uptake analysis suggested that RGD conjugated nanoparticles could be easily internalized by the cancer cells. In vitro cytotoxicity revealed that RGD conjugated EPB loaded nanoparticles exhibited the better antitumor efficacy compared with non-conjugated nanoparticles. More importantly, RGD conjugated EPB loaded nanoparticles showed superior anticancer effects and reduced toxicity than free EPB and non-conjugated nanoparticles by in vivo antitumor activity, EPB biodistribution and histology analysis.

  17. Self-assembly of ciprofloxacin and a tripeptide into an antimicrobial nanostructured hydrogel.

    Science.gov (United States)

    Marchesan, Silvia; Qu, Yue; Waddington, Lynne J; Easton, Christopher D; Glattauer, Veronica; Lithgow, Trevor J; McLean, Keith M; Forsythe, John S; Hartley, Patrick G

    2013-05-01

    This work reports the self-assembly of a sparingly soluble antibiotic (ciprofloxacin) and a hydrophobic tripeptide ((D)Leu-Phe-Phe) into supramolecular nanostructures that yield a macroscopic hydrogel at physiological pH. Drug incorporation results in modified morphology and rheological properties of the self-assembled hydrogel. These changes can be correlated with intermolecular interactions between the drug and the peptide, as confirmed by spectroscopic analysis (fluorescence, circular dichroism, IR). The drug appears bound within the hydrogel by non-covalent interactions, and retains its activity over a prolonged release timescale. Antimicrobial activity of the ciprofloxacin-peptide self-assembled hydrogel was evaluated against Staphylococcus aureus, Escherichia coli, and a clinical strain of Klebsiella pneumoniae. Interestingly, the peptide hydrogel alone exhibited a mild anti-bacterial activity against Gram-negative bacteria. While toxic to bacteria, no major cytotoxicity was seen in haemolysis assays of human red blood cells or in mouse fibroblast cell cultures. This new approach of drug incorporation into the nanostructure of a simple tripeptide hydrogel by self-assembly may have important applications for cost-effective wound dressings and novel antimicrobial formulations. PMID:23422591

  18. The influence of hydrogen bonding on the diffusion behaviour of diastereoisomeric tripeptide derivatives

    Science.gov (United States)

    Gröbel, Angela; Plass, Monika

    1999-05-01

    The diffusion behaviour of the diastereoisomers of Z-Ala-Phe-Val-OMe and Z-Ala-Leu-Val-OMe was studied in solutions of carbon tetrachloride and toluene. The capillary method according to Anderson was used for the diffusion experiment. The loss of the concentration of the tripeptide derivatives in the course of the time was monitored by infrared spectroscopy using their NH stretching vibrations. In general, the diffusion rate of the substances in toluene was 50-100 times larger than in carbon tetrachloride. Also the diastereoisomers differ in their diffusion properties. In carbon tetrachloride this effect is very small but still significant. It can be explained by the strong intramolecular hydrogen bonding of the peptides which leads to C 5 and C 7 rings. In toluene the different configuration of the compounds whose changes are connected with the change in their polar properties are responsible for the observed diffusion rates. The diffusion rate will be discussed in terms of equilibrium constants describing the intramolecular association behaviour and molecular descriptors of the tripeptide derivatives obtained from HPLC measurements in polar solvents. It will be shown that the diffusion rate correlates with the McGowan volume Vx and in part with the effective hydrogen bond acidity ∑ α2H.

  19. Unzipping the role of chirality in nanoscale self-assembly of tripeptide hydrogels.

    Science.gov (United States)

    Marchesan, Silvia; Waddington, Lynne; Easton, Christopher D; Winkler, David A; Goodall, Liz; Forsythe, John; Hartley, Patrick G

    2012-11-01

    Change of chirality is a useful tool to manipulate the aqueous self-assembly behaviour of uncapped, hydrophobic tripeptides. In contrast with other short peptides, these tripeptides form hydrogels at a physiological pH without the aid of organic solvents or end-capping groups (e.g. Fmoc). The novel hydrogel forming peptide (D)Leu-Phe-Phe ((D)LFF) and its epimer Leu-Phe-Phe (LFF) exemplify dramatic supramolecular effects induced by subtle changes to stereochemistry. Only the d-amino acid-containing peptide instantly forms a hydrogel in aqueous solution following a pH switch, generating long fibres (>100 μm) that entangle into a 3D network. However, unexpected nanostructures are observed for both peptides and they are particularly heterogeneous for LFF. Structural analyses using CD, FT-IR and fluorescent amyloid staining reveal anti-parallel beta-sheets for both peptides. XRD analysis also identifies key distances consistent with beta-sheet formation in both peptides, but suggests additional high molecular order and extended molecular length for (D)LFF only. Molecular modelling of the two peptides highlights the key interactions responsible for self-assembly; in particular, rapid self-assembly of (D)LFF is promoted by a phenylalanine zipper, which is not possible because of steric factors for LFF. In conclusion, this study elucidates for the first time the molecular basis for how chirality can dramatically influence supramolecular organisation in very short peptide sequences. PMID:22955637

  20. Unzipping the role of chirality in nanoscale self-assembly of tripeptide hydrogels

    Science.gov (United States)

    Marchesan, Silvia; Waddington, Lynne; Easton, Christopher D.; Winkler, David A.; Goodall, Liz; Forsythe, John; Hartley, Patrick G.

    2012-10-01

    Change of chirality is a useful tool to manipulate the aqueous self-assembly behaviour of uncapped, hydrophobic tripeptides. In contrast with other short peptides, these tripeptides form hydrogels at a physiological pH without the aid of organic solvents or end-capping groups (e.g. Fmoc). The novel hydrogel forming peptide DLeu-Phe-Phe (DLFF) and its epimer Leu-Phe-Phe (LFF) exemplify dramatic supramolecular effects induced by subtle changes to stereochemistry. Only the d-amino acid-containing peptide instantly forms a hydrogel in aqueous solution following a pH switch, generating long fibres (>100 μm) that entangle into a 3D network. However, unexpected nanostructures are observed for both peptides and they are particularly heterogeneous for LFF. Structural analyses using CD, FT-IR and fluorescent amyloid staining reveal anti-parallel beta-sheets for both peptides. XRD analysis also identifies key distances consistent with beta-sheet formation in both peptides, but suggests additional high molecular order and extended molecular length for DLFF only. Molecular modelling of the two peptides highlights the key interactions responsible for self-assembly; in particular, rapid self-assembly of DLFF is promoted by a phenylalanine zipper, which is not possible because of steric factors for LFF. In conclusion, this study elucidates for the first time the molecular basis for how chirality can dramatically influence supramolecular organisation in very short peptide sequences.Change of chirality is a useful tool to manipulate the aqueous self-assembly behaviour of uncapped, hydrophobic tripeptides. In contrast with other short peptides, these tripeptides form hydrogels at a physiological pH without the aid of organic solvents or end-capping groups (e.g. Fmoc). The novel hydrogel forming peptide DLeu-Phe-Phe (DLFF) and its epimer Leu-Phe-Phe (LFF) exemplify dramatic supramolecular effects induced by subtle changes to stereochemistry. Only the d-amino acid-containing peptide

  1. Pokemon siRNA Delivery Mediated by RGD-Modified HBV Core Protein Suppressed the Growth of Hepatocellular Carcinoma.

    Science.gov (United States)

    Kong, Jing; Liu, Xiaoping; Jia, Jianbo; Wu, Jinsheng; Wu, Ning; Chen, Jun; Fang, Fang

    2015-10-01

    Hepatocellular carcinoma (HCC) is a deadly human malignant tumor that is among the most common cancers in the world, especially in Asia. Hepatitis B virus (HBV) infection has been well established as a high risk factor for hepatic malignance. Studies have shown that Pokemon is a master oncogene for HCC growth, suggesting it as an ideal therapeutic target. However, efficient delivery system is still lacking for Pokemon targeting treatment. In this study, we used core proteins of HBV, which is modified with RGD peptides, to construct a biomimetic vector for the delivery of Pokemon siRNAs (namely, RGD-HBc-Pokemon siRNA). Quantitative PCR and Western blot assays revealed that RGD-HBc-Pokemon siRNA possessed the highest efficiency of Pokemon suppression in HCC cells. In vitro experiments further indicated that RGD-HBc-Pokemon-siRNA exerted a higher tumor suppressor activity on HCC cell lines, evidenced by reduced proliferation and attenuated invasiveness, than Pokemon-siRNA or RGD-HBc alone. Finally, animal studies demonstrated that RGD-HBc-Pokemon siRNA suppressed the growth of HCC xenografts in mice by a greater extent than Pokemon-siRNA or RGD-HBc alone. Based on the above results, Pokemon siRNA delivery mediated by RGD-modified HBV core protein was shown to be an effective strategy of HCC gene therapy. PMID:26356810

  2. Tumor-targeted inhibition by a novel strategy - mimoretrovirus expressing siRNA targeting the Pokemon gene.

    Science.gov (United States)

    Tian, Zhiqiang; Wang, Huaizhi; Jia, Zhengcai; Shi, Jinglei; Tang, Jun; Mao, Liwei; Liu, Hongli; Deng, Yijing; He, Yangdong; Ruan, Zhihua; Li, Jintao; Wu, Yuzhang; Ni, Bing

    2010-12-01

    Pokemon gene has crucial but versatile functions in cell differentiation, proliferation and tumorigenesis. It is a master regulator of the ARF-HDM2-p53 and Rb-E2F pathways. The facts that the expression of Pokemon is essential for tumor formation and many kinds of tumors over-express the Pokemon gene make it an attractive target for therapeutic intervention for cancer treatment. In this study, we used an RNAi strategy to silence the Pokemon gene in a cervical cancer model. To address the issues involving tumor specific delivery and durable expression of siRNA, we applied the Arg-Gly-Asp (RGD) peptide ligand and polylysine (K(18)) fusion peptide to encapsulate a recombinant retrovirus plasmid expressing a siRNA targeting the Pokemon gene and produced the 'mimoretrovirus'. At charge ratio 2.0 of fusion peptide/plasmid, the mimoretrovirus formed stable and homogenous nanoparticles, and provided complete DNase I protection and complete gel retardation. This nanoparticle inhibited SiHa cell proliferation and invasion, while it promoted SiHa cell apoptosis. The binding of the nanoparticle to SiHa cells was mediated via the RGD-integrin α(v)β(3) interaction, as evidenced by the finding that unconjugated RGD peptide inhibited this binding significantly. This tumor-targeting mimoretrovirus exhibited excellent anti-tumor capacity in vivo in a nude mouse model. Moreover, the mimoretrovirus inhibited tumor growth with a much higher efficiency than recombinant retrovirus expressing siRNA or the K(18)/P4 nanoparticle lacking the RGD peptide. Results suggest that the RNAi/RGD-based mimoretrovirus developed in this study represents a novel anti-tumor strategy that may be applicable to most research involving cancer therapy and, thus, has promising potential as a cervical cancer treatment. PMID:20879980

  3. Immobilization of RGD Peptide onto the Surface of Apatite-Wollastonite Ceramic for Enhanced Osteoblast Adhesion and Bone Regeneration

    Institute of Scientific and Technical Information of China (English)

    ZHANG Xiang; GU Jianwen; ZHANG Yue; TAN Yanfei; ZHOU Jiabei; ZHOU Dali

    2014-01-01

    The arginine-glycine-aspartic (RGD) acid peptide was grafted to the surface of apatite-wollastonite (AW) ceramic in an effort to improve its cell adhesion, proliferation and osteoinduction. RGD peptide was covalently immobilized onto the surface of AW ceramic via the synthetic cross linker AAPTS-E and 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide (EDC). The modified surfaces were characterized by attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR) and X-ray photoelectron spectroscopy (XPS). The chemical analysis indicated that RGD peptide had been immobilized onto the AW surface successfully. The growth of osteoblast-like cells (MG63) showed that modifying the AW surface with RGD peptide enhanced the cell adhesion and proliferation. And the histological evaluation of RGD-AW showed that the bone regeneration and remodeling process were significantly enhanced compared to the original AW ceramics after 2, 4 and 8 weeks implantation in rabbit’s femoral condyles.

  4. Improved tumor-targeting MRI contrast agents: Gd(DOTA) conjugates of a cycloalkane-based RGD peptide

    Energy Technology Data Exchange (ETDEWEB)

    Park, Ji-Ae, E-mail: jpark@kirams.re.kr [Molecular Imaging Research Center, Korea Institute of Radiological and Medical Sciences, Seoul (Korea, Republic of); Lee, Yong Jin; Ko, In Ok [Molecular Imaging Research Center, Korea Institute of Radiological and Medical Sciences, Seoul (Korea, Republic of); Kim, Tae-Jeong; Chang, Yongmin [Institute of Biomedical Engineering, Kyungpook National University, Daegu (Korea, Republic of); Lim, Sang Moo [Department of Nuclear Medicine, Korea Institute of Radiological and Medical Sciences, Seoul (Korea, Republic of); Kim, Kyeong Min [Molecular Imaging Research Center, Korea Institute of Radiological and Medical Sciences, Seoul (Korea, Republic of); Kim, Jung Young, E-mail: jykim@kirams.re.kr [Molecular Imaging Research Center, Korea Institute of Radiological and Medical Sciences, Seoul (Korea, Republic of)

    2014-12-12

    Highlights: • Development of improved tumor-targeting MRI contrast agents. • To increase the targeting ability of RGD, we developed cycloalkane-based RGD peptides. • Gd(DOTA) conjugates of cycloalkane-based RGD peptide show improved tumor signal enhancement in vivo MR images. - Abstract: Two new MRI contrast agents, Gd-DOTA-c(RGD-ACP-K) (1) and Gd-DOTA-c(RGD-ACH-K) (2), which were designed by incorporating aminocyclopentane (ACP)- or aminocyclohexane (ACH)-carboxylic acid into Gd-DOTA (gadolinium-tetraazacyclo dodecanetetraacetic acid) and cyclic RGDK peptides, were synthesized and evaluated for tumor-targeting ability in vitro and in vivo. Binding affinity studies showed that both 1 and 2 exhibited higher affinity for integrin receptors than cyclic RGDyK peptides, which were used as a reference. These complexes showed high relaxivity and good stability in human serum and have the potential to improve target-specific signal enhancement in vivo MR images.

  5. {sup 18}F-labeled RGD peptide: initial evaluation for imaging brain tumor angiogenesis

    Energy Technology Data Exchange (ETDEWEB)

    Chen Xiaoyuan; Park, Ryan; Shahinian, Anthony H.; Tohme, Michel; Khankaldyyan, Vazgen; Bozorgzadeh, Mohammed H.; Bading, James R.; Moats, Rex; Laug, Walter E.; Conti, Peter S. E-mail: pconti@usc.edu

    2004-02-01

    Brain tumors are highly angiogenesis dependent. The cell adhesion receptor integrin {alpha}{sub v}{beta}{sub 3} is overexpressed in glioma and activated endothelial cells and plays an important role in brain tumor growth, spread and angiogenesis. Suitably labeled {alpha}{sub v}{beta}{sub 3}-integrin antagonists may therefore be useful for imaging brain tumor associated angiogenesis. Cyclic RGD peptide c(RGDyK) was labeled with {sup 18}F via N-succinimidyl-4-[{sup 18}F]fluorobenzoate through the side-chain {epsilon}-amino group of the lysine residue. The radiotracer was evaluated in vivo for its tumor targeting efficacy and pharmacokinetics in subcutaneously implanted U87MG and orthotopically implanted U251T glioblastoma nude mouse models by means of microPET, quantitative autoradiography and direct tissue sampling. The N-4-[{sup 18}F]fluorobenzoyl-RGD ([{sup 18}F]FB-RGD) was produced in less than 2 h with 20-25% decay-corrected yields and specific activity of 230 GBq/{mu}mol at end of synthesis. The tracer showed very rapid blood clearance and both hepatobiliary and renal excretion. Tumor-to-muscle uptake ratio at 30 min was approximately 5 in the subcutaneous U87MG tumor model. MicroPET imaging with the orthotopic U251T brain tumor model revealed very high tumor-to-brain ratio, with virtually no uptake in the normal brain. Successful blocking of tumor uptake of [{sup 18}F]FB-RGD in the presence of excess amount of c(RGDyK) revealed receptor specific activity accumulation. Hence, N-4-[{sup 18}F]fluorobenzoyl labeled cyclic RGD peptide [{sup 18}F]FB-RGD is a potential tracer for imaging {alpha}{sub v}{beta}{sub 3}-integrin positive tumors in brain and other anatomic locations.

  6. [18F]Galacto-RGD: synthesis, radiolabeling, metabolic stability, and radiation dose estimates.

    Science.gov (United States)

    Haubner, Roland; Kuhnast, Bertrand; Mang, Christian; Weber, Wolfgang A; Kessler, Horst; Wester, Hans-Jürgen; Schwaiger, Markus

    2004-01-01

    It has been demonstrated in various murine tumor models that radiolabeled RGD-peptides can be used for noninvasive determination of alphavbeta3 integrin expression. Introduction of sugar moieties improved the pharmacokinetic properties of these peptides and led to tracer with good tumor-to-background ratios. Here we describe the synthesis, radiolabeling, and the metabolic stability of a glycosylated RGD-peptide ([18F]Galacto-RGD) and give first radiation dose estimates for this tracer. The peptide was assembled on a solid support using Fmoc-protocols and cyclized under high dilution conditions. It was conjugated with a sugar amino acid, which can be synthesized via a four-step synthesis starting from pentaacetyl-protected galactose. For radiolabeling of the glycopeptide, 4-nitrophenyl-2-[18F]fluoropropionate was used. This prosthetic group allowed synthesis of [18F]Galacto-RGD with a maximum decay-corrected radiochemical yield of up to 85% and radiochemical purity >98%. The overall radiochemical yield was 29 +/- 5% with a total reaction time including final HPLC preparation of 200 +/- 18 min. The metabolic stability of [18F]Galacto-RGD was determined in mouse blood and liver, kidney, and tumor homogenates 2 h after tracer injection. The average fraction of intact tracer in these organs was approximately 87%, 76%, 69%, and 87%, respectively, indicating high in vivo stability of the radiolabeled glycopeptide. The expected radiation dose to humans after injection of [18F]Galacto-RGD has been estimated on the basis of dynamic PET studies with New Zealand white rabbits. According to the residence times in these animals the effective dose was calculated using the MIRDOSE 3.0 program as 2.2 x 10(-2) mGy/MBq. In conclusion, [18F]Galacto-RGD can be synthesized in high radiochemical yields and radiochemical purity. Despite the time-consuming synthesis of the prosthetic group 185 MBq of [18F]Galacto-RGD, a sufficient dose for patient studies, can be produced starting with

  7. Identification and characterization of di- and tripeptide transporter DtpT of Listeria monocytogenes EGD-e

    NARCIS (Netherlands)

    Wouters, J.A.; Hain, T.; Darji, A.; Hüfner, E.; Wemekamp-Kamphuis, H.H.; Chakraborty, T.; Abee, T.

    2005-01-01

    Listeria monocytogenes is a gram-positive intracellular pathogen responsible for opportunistic infections in humans and animals. Here we identified and characterized the dtpT gene (lmo0555) of L. monocytogenes EGD-e, encoding the di- and tripeptide transporter, and assessed its role in growth under

  8. Peptide length, steric effects, and ion solvation govern zwitterion stabilization in barium-chelated di- and tripeptides

    NARCIS (Netherlands)

    R.C. Dunbar; J.D. Steill; N.C. Polfer; J. Oomens

    2009-01-01

    Infrared multiple-photon dissociation (IRMPD) spectroscopy has given infrared spectra of complexes of di- and tripeptides (AlaAla, AlaAlaAla, AlaPhe, PheAla) with singly and doubly charged metal ions (K+, Ca2+, Sr2+, and Ba2+). The switch between charge-solvated (CS) and salt-bridged zwitterion (SB)

  9. Peptide Length, Steric Effects, and Ion Solvation Govern Zwitterion Stabilization in Barium-Chelated Di- and Tripeptides

    NARCIS (Netherlands)

    Dunbar, R. C.; Steill, J. D.; Polfer, N. C.; Oomens, J.

    2009-01-01

    Infrared multiple-photon dissociation (IRMPD) spectroscopy has given infrared spectra of complexes of di- and tripeptides (AlaAla, AlaAlaAla, AlaPhe, PheAla) with singly and doubly charged metal ions (K+, Ca2+, Sr2+, and Ba2+). The switch between charge-solvated (CS) and salt-bridged zwitterion (SB)

  10. Cloning and functional expression in Escherichia coli of the gene encoding the di- and tripeptide transport protein of Lactobacillus helveticus

    NARCIS (Netherlands)

    Nakajima, H.; Hagting, A; Kunji, E.R S; Poolman, B.; Konings, W.N

    1997-01-01

    The gene encoding the di- and tripeptide transport protein (DtpT) of Lactobacillus helveticus (DtpT(LH)) was cloned with the aid of the inverse PCR technique and used to complement the dipeptide transport-deficient and proline-auxotrophic Escherichia coil E1772. Functional expression of the peptide

  11. alpha-MSH tripeptide analogs activate the melanocortin 1 receptor and reduce UV-induced DNA damage in human melanocytes.

    Science.gov (United States)

    Abdel-Malek, Zalfa A; Ruwe, Andrew; Kavanagh-Starner, Renny; Kadekaro, Ana Luisa; Swope, Viki; Haskell-Luevano, Carrie; Koikov, Leonid; Knittel, James J

    2009-10-01

    One skin cancer prevention strategy that we are developing is based on synthesizing and testing melanocortin analogs that reduce and repair DNA damage resulting from exposure to solar ultraviolet (UV) radiation, in addition to stimulating pigmentation. Previously, we reported the effects of tetrapeptide analogs of alpha-melanocortin (alpha-MSH) that were more potent and stable than the physiological alpha-MSH, and mimicked its photoprotective effects against UV-induced DNA damage in human melanocytes. Here, we report on a panel of tripeptide analogs consisting of a modified alpha-MSH core His(6)-d-Phe(7)-Arg(8), which contained different N-capping groups, C-terminal modifications, or arginine mimics. The most potent tripeptides in activating cAMP formation and tyrosinase of human melanocytes were three analogs with C-terminal modifications. The most effective C-terminal tripeptide mimicked alpha-MSH in reducing hydrogen peroxide generation and enhancing nucleotide excision repair following UV irradiation. The effects of these three analogs required functional MC1R, as they were absent in human melanocytes that expressed non-functional receptor. These results demonstrate activation of the MC1R by tripeptide melanocortin analogs. Designing small analogs for topical delivery should prove practical and efficacious for skin cancer prevention.

  12. Preparation and evaluation of 177Lu-DOTA-Bz-RGD dimer and 177Lu-DOTA-Bz-PEG4-RGD dimer

    International Nuclear Information System (INIS)

    177Lu-DOTA-Bz-RGD dimer and 177Lu-DOTA-Bz-PEG4-RGD dimer were pre- pared, and the effect of PEG4 on labeling conditions and in vitro stability as well as pharma-cokinetic properties and biodistribution in normal mice for the radiolebeled compounds was compared. The results of TLC and HPLC show that the labeling yield of two radiolabeled compounds was more than 95% under optimal conditions (pH 4.0 and pH 6.0, respectively, reacting at 100 degree C for 15-20 min). The two radiolabeled compounds show pretty good stability in saline. HPLC analyses and lgPow values revealed that introducing of PEG4 increased the lipophilic character of radiolabeled compounds, but had no significant changes on pharmacokinetic properties and biodistribution in normal mice. (authors)

  13. Radiolabeled antibodies and RGD-peptides for the treatment of ovarian cancer

    NARCIS (Netherlands)

    Janssen, M.L.H.

    2004-01-01

    In this thesis, preclinical studies on new treatment modalities for ovarian cancer are descibed, applying radiolabeled antibodies and radiolabeled RGD-peptides. In chapter 2 a study is described comparing the therapeutic efficacy of the antibody HMFG1 radiolabeled with several beta-emitting radionuc

  14. Helicoidal multi-lamellar features of RGD-functionalized silk biomaterials for corneal tissue engineering.

    Science.gov (United States)

    Gil, Eun Seok; Mandal, Biman B; Park, Sang-Hyug; Marchant, Jeffrey K; Omenetto, Fiorenzo G; Kaplan, David L

    2010-12-01

    RGD-coupled silk protein-biomaterial lamellar systems were prepared and studied with human cornea fibroblasts (hCFs) to match functional requirements. A strategy for corneal tissue engineering was pursued to replicate the structural hierarchy of human corneal stroma within thin stacks of lamellae-like tissues, in this case constructed from scaffolds constructed with RGD-coupled, patterned, porous, mechanically robust and transparent silk films. The influence of RGD-coupling on the orientation, proliferation, ECM organization, and gene expression of hCFs was assessed. RGD surface modification enhanced cell attachment, proliferation, alignment and expression of both collagens (type I and V) and proteoglycans (decorin and biglycan). Confocal and histological images of the lamellar systems revealed that the bio-functionalized silk human cornea 3D constructs exhibited integrated corneal stroma tissue with helicoidal multi-lamellar alignment of collagen-rich and proteoglycan-rich extracellular matrix, with transparency of the construct. This biomimetic approach to replicate corneal stromal tissue structural hierarchy and architecture demonstrates a useful strategy for engineering human cornea. Further, this approach can be exploited for other tissue systems due to the pervasive nature of such helicoids in most human tissues. PMID:20801503

  15. New Glucocyclic RGD Dimers for Positron Emission Tomography Imaging of Tumor Integrin Receptors.

    Science.gov (United States)

    Lee, Ji Woong; Park, Ji-Ae; Lee, Yong Jin; Shin, Un Chol; Kim, Suhng Wook; Kim, Byung Il; Lim, Sang Moo; An, Gwang Il; Kim, Jung Young; Lee, Kyo Chul

    2016-08-01

    Most studies of radiolabeled arginine-glycine-aspartic acid (RGD) peptides have shown in vitro affinity for integrin ανβ3, allowing for the targeting of receptor-positive tumors in vivo. However, major differences have been found in the pharmacokinetic profiles of different radiolabeled RGD peptide analogs. The purposes of this study were to prepare (64)Cu-DOTA-gluco-E[c(RGDfK)]2 (R8), (64)Cu-NOTA-gluco-E[c(RGDfK)]2 (R9), and (64)Cu-NODAGA-gluco-E[c(RGDfK)]2 (R10) and compare their pharmacokinetics and tumor imaging properties using small-animal positron emission tomography (PET). All three compounds were produced with high specific activity within 10 minutes. The IC50 values were similar for all the substances, and their affinities were greater than that of c(RGDyK). R8, R9, and R10 were stable for 24 hours in human and mouse serums and showed high uptake in U87MG tumors with high tumor-to-blood ratios. Compared to the control, a cyclic RGD peptide dimer without glucosamine, R10, showed low uptake in the liver. Because of their good imaging qualities and improved pharmacokinetics, (64)Cu-labeled dimer RGD conjugates (R8, R9, and R10) may have potential applications as PET radiotracers. R9 (NOTA) with highly in vivo stability consequentially showed an improved PET tumor uptake than R8 (DOTA) or R10 (NODAGA). PMID:27403677

  16. Improved tumor targeting of radiolabeled RGD peptides using rapid dose fractionation.

    NARCIS (Netherlands)

    Janssen, M.; Frielink, C.; Dijkgraaf, I.; Oyen, W.J.G.; Edwards, D.S.; Liu, S.; Rajopadhye, M.; Massuger, L.F.A.G.; Corstens, F.H.M.; Boerman, O.C.

    2004-01-01

    Arginine-glycine-aspartic acid (RGD) peptides preferentially bind to alphavbeta3 integrin, an integrin expressed on newly formed endothelial cells and on various tumor cells. When labeled with beta-emitting radionuclides, these peptides can be used for peptide-receptor radionuclide therapy of malign

  17. Preparation of RGD-modified Long Circulating Liposome Loading Matrine, and its in vitro Anti-cancer Effects

    Directory of Open Access Journals (Sweden)

    Xiao-yan Liu, Li-ming Ruan, Wei-wei Mao, Jin-Qiang Wang, You-qing Shen, Mei-hua Sui

    2010-01-01

    Full Text Available Aim: To prepare RGD-modified long circulating liposome (LCL loading matrine (RGD-M-LCL to improve the tumor-targeting and efficacy of matrine. Methods: LCL which was prepared with HSPC, cholesterol, DSPE-PEG2000 and DSPE-PEG-MAL was modified with an RGD motif confirmed by high performance liquid chromatography (HPLC. The encapsulation efficiency of RGD-M-LCL was also detected by HPLC. MTT assay was used to examine the effects of RGD-M-LCL on the proliferation of Bcap-37, HT-29 and A375 cells. The percentage of apoptotic cells and morphological changes in Bcap-37 cells treated with RGD-M-LCL were detected by Annexin-V-FITC/PI affinity assay and observed under light microscope, respectively. Results: Spherical or oval single-chamber particles of uniform sizes with little agglutination or adhesion were observed under transmission electronic microscope. The RGD motif was successfully coupled to the DSPE-PEG-MAL on liposomes, as confirmed by HPLC. An encapsulation efficiency of 83.13% was obtained when the drug-lipid molar ratio was 0.1, and the encapsulation efficiency was negatively related to the drug-lipid ratio in the range of 0.1~0.4, and to the duration of storage. We found that, compared with free matrine, RGD-M-LCL had much stronger in vitro activity, leading to anti-proliferative and pro-apoptotic effects against cancer cells (P<0.01. Conclusion: RGD-M-LCL, a novel delivery system for anti-cancer drugs, was successfully prepared, and we demonstrated that the use of this material could augment the effects of matrine on cancer cells in vitro.

  18. Density-tunable conjugation of cyclic RGD ligands with polyion complex vesicles for the neovascular imaging of orthotopic glioblastomas

    Science.gov (United States)

    Kawamura, Wataru; Miura, Yutaka; Kokuryo, Daisuke; Toh, Kazuko; Yamada, Naoki; Nomoto, Takahiro; Matsumoto, Yu; Sueyoshi, Daiki; Liu, Xueying; Aoki, Ichio; Kano, Mitsunobu R.; Nishiyama, Nobuhiro; Saga, Tsuneo; Kishimura, Akihiro; Kataoka, Kazunori

    2015-06-01

    Introduction of ligands into 100 nm scaled hollow capsules has great potential for diagnostic and therapeutic applications in drug delivery systems. Polyethylene glycol-conjugated (PEGylated) polyion complex vesicles (PICsomes) are promising hollow nano-capsules that can survive for long periods in the blood circulation and can be used to deliver water-soluble macromolecules to target tissues. In this study, cyclic RGD (cRGD) peptide, which is specifically recognized by αVβ3 and αvβ5 integrins that are expressed at high levels in the neovascular system, was conjugated onto the distal end of PEG strands on PICsomes for active neovascular targeting. Density-tunable cRGD-conjugation was achieved using PICsomes with definite fraction of end-functionalized PEG, to substitute 20, 40, and 100% of PEG distal end of the PICsomes to cRGD moieties. Compared with control-PICsomes without cRGD, cRGD-PICsomes exhibited increased uptake into human umbilical vein endothelial cells. Intravital confocal laser scanning microscopy revealed that the 40%-cRGD-PICsomes accumulated mainly in the tumor neovasculature and remained in the perivascular region even after 24 h. Furthermore, we prepared superparamagnetic iron oxide (SPIO)-loaded cRGD-PICsomes for magnetic resonance imaging (MRI) and successfully visualized the neovasculature in an orthotopic glioblastoma model, which suggests that SPIO-loaded cRGD-PICsomes might be useful as a MRI contrast reagent for imaging of the tumor microenvironment, including neovascular regions that overexpress αVβ3 integrins.

  19. Induction of mesenchymal stem cell chondrogenesis by polyacrylate substrates.

    Science.gov (United States)

    Glennon-Alty, Laurence; Williams, Rachel; Dixon, Simon; Murray, Patricia

    2013-04-01

    Mesenchymal stem cells (MSCs) can generate chondrocytes in vitro, but typically need to be cultured as aggregates in the presence of transforming growth factor beta (TGF-β), which makes scale-up difficult. Here we investigated if polyacrylate substrates modelled on the functional group composition and distribution of the Arg-Gly-Asp (RGD) integrin-binding site could induce MSCs to undergo chondrogenesis in the absence of exogenous TGF-β. Within a few days of culture on the biomimetic polyacrylates, both mouse and human MSCs, and a mesenchymal-like mouse-kidney-derived stem cell line, began to form multi-layered aggregates and started to express the chondrocyte-specific markers, Sox9, collagen II and aggrecan. Moreover, collagen II tended to be expressed in the centre of the aggregates, similarly to developing limb buds in vivo. Surface analysis of the substrates indicated that those with the highest surface amine content were most effective at promoting MSC chondrogenesis. These results highlight the importance of surface group functionality and the distribution of those groups in the design of substrates to induce MSC chondrogenesis. PMID:23237986

  20. Selective activation of mechanosensitive ion channels using magnetic particles.

    Science.gov (United States)

    Hughes, Steven; McBain, Stuart; Dobson, Jon; El Haj, Alicia J

    2008-08-01

    This study reports the preliminary development of a novel magnetic particle-based technique that permits the application of highly localized mechanical forces directly to specific regions of an ion-channel structure. We demonstrate that this approach can be used to directly and selectively activate a mechanosensitive ion channel of interest, namely TREK-1. It is shown that manipulation of particles targeted against the extended extracellular loop region of TREK-1 leads to changes in whole-cell currents consistent with changes in TREK-1 activity. Responses were absent when particles were coated with RGD (Arg-Gly-Asp) peptide or when magnetic fields were applied in the absence of magnetic particles. It is concluded that changes in whole-cell current are the result of direct force application to the extracellular loop region of TREK-1 and thus these results implicate this region of the channel structure in mechano-gating. It is hypothesized that the extended loop region of TREK-1 may act as a tension spring that acts to regulate sensitivity to mechanical forces, in a nature similar to that described for MscL. The development of a technique that permits the direct manipulation of mechanosensitive ion channels in real time without the need for pharmacological drugs has huge potential benefits not only for basic biological research of ion-channel gating mechanisms, but also potentially as a tool for the treatment of human diseases caused by ion-channel dysfunction.

  1. Disintegrins: integrin selective ligands which activate integrin-coupled signaling and modulate leukocyte functions

    Directory of Open Access Journals (Sweden)

    Barja-Fidalgo C.

    2005-01-01

    Full Text Available Extracellular matrix proteins and cell adhesion receptors (integrins play essential roles in the regulation of cell adhesion and migration. Interactions of integrins with the extracellular matrix proteins lead to phosphorylation of several intracellular proteins such as focal adhesion kinase, activating different signaling pathways responsible for the regulation of a variety of cell functions, including cytoskeleton mobilization. Once leukocytes are guided to sites of infection, inflammation, or antigen presentation, integrins can participate in the initiation, maintenance, or termination of the immune and inflammatory responses. The modulation of neutrophil activation through integrin-mediated pathways is important in the homeostatic control of the resolution of inflammatory states. In addition, during recirculation, T lymphocyte movement through distinct microenvironments is mediated by integrins, which are critical for cell cycle, differentiation and gene expression. Disintegrins are a family of low-molecular weight, cysteine-rich peptides first identified in snake venom, usually containing an RGD (Arg-Gly-Asp motif, which confers the ability to selectively bind to integrins, inhibiting integrin-related functions in different cell systems. In this review we show that, depending on the cell type and the microenvironment, disintegrins are able to antagonize the effects of integrins or to act agonistically by activating integrin-mediated signaling. Disintegrins have proven useful as tools to improve the understanding of the molecular events regulated by integrin signaling in leukocytes and prototypes in order to design therapies able to interfere with integrin-mediated effects.

  2. Integrin-like Protein Is Involved in the Osmotic Stress-induced Abscisic Acid Biosynthesis in Arabidopsis thaliana

    Institute of Scientific and Technical Information of China (English)

    Bing Lü; Feng Chen; Zhong-Hua Gong; Hong Xie; Jian-Sheng Liang

    2007-01-01

    We studied the perception of plant cells to osmotic stress that leads to the accumulation of abscisic acid (ABA) in stressed Arabidopsis thaliana L. cells. A significant difference was found between protoplasts and cells in terms of their responses to osmotic stress and ABA biosynthesis, implying that cell wall and/or cell wall-plasma membrane interaction are essential in identifying osmotic stress. Western blotting and immunofluorescence localization experiments, using polyclonal antibody against human integrin β1, revealed the existence of a protein similar to the integrin protein of animals in the suspension-cultured cells located in the plasma membrane fraction.Treatment with a synthetic pentapeptide, Gly-Arg-Gly-Asp-Ser (GRGDS), which contains an RGD domain and interacts specifically with integrin protein and thus blocks the cell wall-plasma membrane interaction, significantly inhibited osmotic stress-induced ABA biosynthesis in cells, but not in protoplasts. These results demonstrate that cell wall and/or cell wall-plasma membrane interaction mediated by integrin-like proteins played important roles in osmotic stress-induced ABA biosynthesis in Arabidopsis thaliana.

  3. Molecular cloning and characterisation of a pattern recognition molecule, lipopolysaccharide- and beta-1,3-glucan binding protein (LGBP) from the white shrimp Litopenaeus vannamei.

    Science.gov (United States)

    Cheng, Winton; Liu, Chun-Hung; Tsai, Chiung-Hui; Chen, Jiann-Chu

    2005-04-01

    A lipopolysaccharide- and beta-1,3-glucan binding protein (LGBP) cDNA was cloned from the haemocyte and hepatopancreas of white shrimp Litopenaeus vannamei using oligonucleotide primers and RT-PCR. Both 3'- and 5'-regions were isolated by rapid amplification of cDNA end RACE method. Analysis of nucleotide sequence revealed that the cDNA clone has an open reading frame of 1101 bp encoding a protein of 367 amino acids including a 17 amino acid signal peptide. The calculated molecular mass of the mature proteins (350 amino acids) is 39.92 kDa with an estimated pI of 4.37. Two putative integrin binding motifs (cell adhesion site), RGD (Arg-Gly-Asp) and a potential recognition motif for beta- (1-->3) linkage of polysaccharides were observed in the LGBP. Sequence comparison showed that LGBP deduced amino acid of L. vannamei has an overall similarity of 95%, 92% and 61% to that of blue shrimp Litopenaeus stylirostris LGBP, tiger shrimp Penaeus monodon BGBP and crayfish Pacifastacus leniusculus LGBP, respectively. Quantitative real-time RT-PCR analysis showed that LGBP transcript in haemocyte of L. vannamei increased in 3- and 6-h post Vibrio alginolyticus injection. PMID:15561560

  4. Shrimp arginine kinase being a binding protein of WSSV envelope protein VP31

    Science.gov (United States)

    Ma, Cuiyan; Gao, Qiang; Liang, Yan; Li, Chen; Liu, Chao; Huang, Jie

    2016-03-01

    Viral entry into the host is the earliest stage of infection in the viral life cycle in which attachment proteins play a key role. VP31 (WSV340/WSSV396), an envelope protein of white spot syndrome virus (WSSV), contains an Arg-Gly-Asp (RGD) peptide domain known as a cellular attachment site. At present, the process of VP31 interacting with shrimp host cells has not been explored. Therefore, the VP31 gene was cloned into pET30a (+), expressed in Escherichia coli strain BL21 and purified with immobilized metal ion affinity chromatography. Four gill cellular proteins of shrimp (Fenneropenaeus chinensis) were pulled down by an affinity column coupled with recombinant VP31 (rVP31), and the amino acid sequences were identified with MALDI-TOF/TOF mass spectrometry. Hemocyanin, beta-actin, arginine kinase (AK), and an unknown protein were suggested as the putative VP31 receptor proteins. SDS-PAGE showed that AK is the predominant binding protein of VP31. An i n vitro binding activity experiment indicated that recombinant AK's (rAK) binding activity with rVP31 is comparable to that with the same amount of WSSV. These results suggested that AK, as a member of the phosphagen kinase family, plays a role in WSSV infection. This is the first evidence showing that AK is a binding protein of VP31. Further studies on this topic will elucidate WSSV infection mechanism in the future.

  5. Molecular cloning and characterisation of a pattern recognition protein, lipopolysaccharide and beta-1,3-glucan binding protein (LGBP) from Chinese shrimp Fenneropenaeus chinensis.

    Science.gov (United States)

    Liu, Fengsong; Li, Fuhua; Dong, Bo; Wang, Xiaomei; Xiang, Jianhai

    2009-03-01

    A pattern recognition protein (PRP), lipopolysaccharide and beta-1,3-glucan binding protein (LGBP) cDNA was cloned from the haemocyte of Chinese shrimp Fenneropenaeus chinensis by the techniques of homology cloning and RACE. Analysis of nucleotide sequence revealed that the full-length cDNA of 1,275 bp has an open reading frame of 1,098 bp encoding a protein of 366 amino acids including a 17 amino acid signal peptide. Sequence comparison of the deduced amino acid sequence of F. chinensis LGBP showed a high identity of 94%, 90%, 87%, 72% and 63% with Penaeus monodon BGBP, Litopenaeus stylirostris LGBP, Marsupenaeu japonicus BGBP, Homarus gammarus BGBP and Pacifastacus leniusculus LGBP, respectively. The calculated molecular mass of the mature protein is 39,857 Da with a deduced pI of 4.39. Two putative integrin binding motifs, RGD (Arg-Gly-Asp) and a potential recognition motif for beta-1,3-linkage of polysaccharides were observed in LGBP sequence. RT-PCR analysis showed that LGBP gene expresses in haemocyte and hepatopancreas only, but not in other tissues. Capillary electrophoresis RT-PCR method was used to quantify the variation of mRNA transcription level during artificial infection with heat-killed Vibrio anguillarum and Staphylococcus aureusin. A significant enhancement of LGBP transcription was appeared at 6 h post-injection in response to bacterial infection. These results have provided useful information to understand the function of LGBP in shrimp. PMID:18163220

  6. Direct influence of culture dimensionality on human mesenchymal stem cell differentiation at various matrix stiffnesses using a fibrous self-assembling peptide hydrogel.

    Science.gov (United States)

    Hogrebe, Nathaniel J; Gooch, Keith J

    2016-09-01

    Much is unknown about the effects of culture dimensionality on cell behavior due to the lack of biomimetic substrates that are suitable for directly comparing cells grown on two-dimensional (2D) and encapsulated within three-dimensional (3D) matrices of the same stiffness and biochemistry. To overcome this limitation, we used a self-assembling peptide hydrogel system that has tunable stiffness and cell-binding site density as well as a fibrous microarchitecture resembling the structure of collagen. We investigated the effect of culture dimensionality on human mesenchymal stem cell differentiation at different values of matrix stiffness (G' = 0.25, 1.25, 5, and 10 kPa) and a constant RGD (Arg-Gly-Asp) binding site concentration. In the presence of the same soluble induction factors, culture on top of stiff gels facilitated the most efficient osteogenesis, while encapsulation within the same stiff gels resulted in a switch to predominantly terminal chondrogenesis. Adipogenesis dominated at soft conditions, and 3D culture induced better adipogenic differentiation than 2D culture at a given stiffness. Interestingly, initial matrix-induced cell morphology was predictive of these end phenotypes. Furthermore, optimal culture conditions corresponded to each cell type's natural niche within the body, highlighting the importance of incorporating native matrix dimensionality and stiffness into tissue engineering strategies. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 2356-2368, 2016. PMID:27163888

  7. Effects of fused hirudin on activity of thrombin and function of platelets

    Institute of Scientific and Technical Information of China (English)

    SHEN Li; CHEN Shao-ping; CAI Zai-long; YANG Sheng-sheng; QIN Yong-wen

    2005-01-01

    Objective: To investigate whether fused hirudin peptide has both antithrombin and antiplatelet functions. Methods: The core region of fused hirudin was the C-terminal tail of hirudin(hirudin53-64),which could bind to the anion binding exosite (ABE) of thrombin.Arg-Pro-Pro-Gly-Phe(RPPGF) amino acid sequence,a metabolite of bradykinin,was added to the N-terminus of hirudin53-64.It bound to the active site of thrombin.Additionally,Arg-Gly-Asp(RGD)amino acid sequence,an inibitor of glycoprotein Ⅱb/Ⅲa( GP Ⅱb/Ⅲa) receptor,was linked to C-terminus of hirudin53-64.This 26-animo acid-fused hirudin peptide was artificially synthesized,purified and analysed. Results: Fused hirudin peptide significantly lengthened the activated partial thromboplastin time(APTT),thrombin time(TT)and prothrombin time(PT) and inhibited the amidolytic activity of thrombin.The ADP-induced platelet aggregation was markedly inhibited by fused hirudin peptide. Conclusion: Fused hirudin peptide has activity of antithrombin as well as antiplatelet.Therefore bifunctional anticoagulation peptide has capacity to target various components of haemostatic process and may become more powerful antithrombosis agent.

  8. Direct influence of culture dimensionality on human mesenchymal stem cell differentiation at various matrix stiffnesses using a fibrous self-assembling peptide hydrogel.

    Science.gov (United States)

    Hogrebe, Nathaniel J; Gooch, Keith J

    2016-09-01

    Much is unknown about the effects of culture dimensionality on cell behavior due to the lack of biomimetic substrates that are suitable for directly comparing cells grown on two-dimensional (2D) and encapsulated within three-dimensional (3D) matrices of the same stiffness and biochemistry. To overcome this limitation, we used a self-assembling peptide hydrogel system that has tunable stiffness and cell-binding site density as well as a fibrous microarchitecture resembling the structure of collagen. We investigated the effect of culture dimensionality on human mesenchymal stem cell differentiation at different values of matrix stiffness (G' = 0.25, 1.25, 5, and 10 kPa) and a constant RGD (Arg-Gly-Asp) binding site concentration. In the presence of the same soluble induction factors, culture on top of stiff gels facilitated the most efficient osteogenesis, while encapsulation within the same stiff gels resulted in a switch to predominantly terminal chondrogenesis. Adipogenesis dominated at soft conditions, and 3D culture induced better adipogenic differentiation than 2D culture at a given stiffness. Interestingly, initial matrix-induced cell morphology was predictive of these end phenotypes. Furthermore, optimal culture conditions corresponded to each cell type's natural niche within the body, highlighting the importance of incorporating native matrix dimensionality and stiffness into tissue engineering strategies. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 2356-2368, 2016.

  9. Atomic basis for the species-specific inhibition of αV integrins by monoclonal antibody 17E6 is revealed by the crystal structure of αVβ3 ectodomain-17E6 Fab complex.

    Science.gov (United States)

    Mahalingam, Bhuvaneshwari; Van Agthoven, Johannes F; Xiong, Jian-Ping; Alonso, José Luis; Adair, Brian D; Rui, Xianliang; Anand, Saurabh; Mehrbod, Mehrdad; Mofrad, Mohammad R K; Burger, Christa; Goodman, Simon L; Arnaout, M Amin

    2014-05-16

    The function-blocking, non-RGD-containing, and primate-specific mouse monoclonal antibody 17E6 binds the αV subfamily of integrins. 17E6 is currently in phase II clinical trials for treating cancer. To elucidate the structural basis of recognition and the molecular mechanism of inhibition, we crystallized αVβ3 ectodomain in complex with the Fab fragment of 17E6. Protein crystals grew in presence of the activating cation Mn(2+). The integrin in the complex and in solution assumed the genuflected conformation. 17E6 Fab bound exclusively to the Propeller domain of the αV subunit. At the core of αV-Fab interface were interactions involving Propeller residues Lys-203 and Gln-145, with the latter accounting for primate specificity. The Propeller residue Asp-150, which normally coordinates Arg of the ligand Arg-Gly-Asp motif, formed contacts with Arg-54 of the Fab that were expected to reduce soluble FN10 binding to cellular αVβ3 complexed with 17E6. This was confirmed in direct binding studies, suggesting that 17E6 is an allosteric inhibitor of αV integrins.

  10. Mapping of amino acid residues responsible for adhesion of cell culture-adapted foot-and-mouth disease SAT type viruses.

    Science.gov (United States)

    Maree, Francois F; Blignaut, Belinda; de Beer, Tjaart A P; Visser, Nico; Rieder, Elizabeth A

    2010-10-01

    Foot-and-mouth disease virus (FMDV) infects host cells by adhering to the alpha(V) subgroup of the integrin family of cellular receptors in a Arg-Gly-Asp (RGD) dependent manner. FMD viruses, propagated in non-host cell cultures are reported to acquire the ability to enter cells via alternative cell surface molecules. Sequencing analysis of SAT1 and SAT2 cell culture-adapted variants showed acquisition of positively charged amino acid residues within surface-exposed loops of the outer capsid structural proteins. The fixation of positively charged residues at position 110-112 in the beta F-beta G loop of VP1 of SAT1 isolates is thought to correlate with the acquisition of the ability to utilise alternative glycosaminoglycan (GAG) molecules for cell entry. Similarly, two SAT2 viruses that adapted readily to BHK-21 cells accumulated positively charged residues at positions 83 and 85 of the beta D-beta E loop of VP1. Both regions surround the fivefold axis of the virion. Recombinant viruses containing positively charged residues at position 110 and 112 of VP1 were able to infect CHO-K1 cells (that expresses GAG) and demonstrated increased infectivity in BHK-21 cells. Therefore, recombinant SAT viruses engineered to express substitutions that induce GAG-binding could be exploited in the rational design of vaccine seed stocks with improved growth properties in cell cultures. PMID:20637812

  11. Use of snake venom fractions in the coagulation laboratory.

    Science.gov (United States)

    Marsh, N A

    1998-07-01

    Snake venom toxins are now regularly used in the coagulation laboratory for assaying haemostatic parameters and as coagulation reagents. Snake venom thrombin-like enzymes (SVTLE) are used for fibrinogen and fibrinogen breakdown product assay as well as detecting dysfibrinogenaemias. Significantly, because SVTLE are not inhibited by heparin, they can be used for defibrinating samples that contain the anticoagulant before assay of haemostatic variables. Prothrombin activators are found in many snake venoms and are used in prothrombin assays, for studying dysprothrombinaemias and preparing meizothrombin and non-enzymic prothrombin. Russell's viper (Daboia russelli) venom (RVV) contains a number of compounds useful in the assay of factors V, VII, X, platelet factor 3 and lupus anticoagulants. Activators from the taipan, Australian brown snake and saw-scaled viper have been used to assay lupus anticoagulants. Protein C and activated protein C resistance can be measured by means of RVV and Protac, a fast acting inhibitor from Southern copperhead snake venom and von Willebrand factor can be studied with Botrocetin from Bothrops jararaca venom. Finally, phospholipase A2 enzymes and the disintegrins, a family of Arg-Gly-Asp (RGD)-containing proteins found in snake venoms, show great potential for the study of haemostasis including, notably, platelet glycoprotein receptors GPIIb/IIIa and Ib. PMID:9712287

  12. Expression cloning and chromosomal mapping of the leukocyte activation antigen CD97, a new seven-span transmembrane molecule of the secretin receptor superfamily with an unusual extracellular domain

    Energy Technology Data Exchange (ETDEWEB)

    Hamann, J. [Univ. of Amsterdam (Netherlands)]|[Max Planck Society, Berlin-Buch (Germany); Hamann, D.; Lier, R.A.W. [Univ. of Amsterdam (Netherlands)] [and others

    1995-08-15

    CD97 is a monomeric glycoprotein of 75 to 85 kDa that is induced rapidly on the surface of most leukocytes upon activation. We herein report the isolation of a cDNA encoding human CD97 by expression cloning in COS cells. The 3-kb cDNA clone encodes a mature polypeptide chain of 722 amino acids with a predicted molecular mass of 79 kDa. Within the C-terminal part of the protein, a region with seven hydrophobic segments was identified, suggesting that CD97 is a seven-span transmembrane molecule. Sequence comparison indicates that CD97 is the first leukocyte Ag in a recently described superfamily that includes the receptors for secretin, calcitonin, and other mammalian and insect peptide hormones. Different from these receptors, CD97 has an extended extracellular region of 433 amino acids that possesses three N-terminal epidermal growth factor-like domains, two of them with a calcium-binding site, and single Arg-Gly-Asp (RGD) motif. The existence of structural elements characteristic for extracellular matrix proteins in a seven-span transmembrane molecule makes CD97 a receptor potentially involved in both adhesion and signaling processes early after leukocyte activation. The gene encoding CD97 is localized on chromosome 19 (19p13.12-13.2).

  13. Mesangial cell αvβ8-integrin regulates glomerular capillary integrity and repair.

    Science.gov (United States)

    Lakhe-Reddy, Sujata; Li, Vincent; Arnold, Thomas D; Khan, Shenaz; Schelling, Jeffrey R

    2014-06-15

    αvβ8-Integrin is most abundantly expressed in the kidney, brain, and female reproductive organs, and its cognate ligand is latent transforming growth factor (LTGF)-β. Kidney αvβ8-integrin localizes to mesangial cells, and global β8-integrin gene (Itgb8) deletion results in embryonic lethality due to impaired placentation and cerebral hemorrhage. To circumvent the lethality and better define kidney αvβ8-integrin function, Cre-lox technology was used to generate mesangial-specific Itgb8-null mice. Platelet-derived growth factor-β receptor (PDGFBR)-Cre mice crossed with a reporter strain revealed functional Cre recombinase activity in a predicted mesangial pattern. However, mating between two different PDGFBR-Cre or Ren1(d)-Cre strains with Itgb8 (flox/-) mice consistently resulted in incomplete recombination, with no renal phenotype in mosaic offspring. Induction of a renal phenotype with Habu snake venom, a reversible mesangiolytic agent, caused exaggerated glomerular capillary microaneurysms and delayed recovery in Cre(+/-) PDGFRB (flox/-) mice compared with Cre(+/-) PDGFRB (flox/+) control mice. To establish the mechanism, in vitro experiments were conducted in Itgb8-null versus Itgb8-expressing mesangial cells and fibroblasts, which revealed β8-integrin-regulated adhesion to Arg-Gly-Asp (RGD) peptides within a mesangial-conditioned matrix as well as β8-integrin-dependent migration on RGD-containing LTGF-β or vitronectin matrices. We speculate that kidney αvβ8-integrin indirectly controls glomerular capillary integrity through mechanical tension generated by binding RGD peptides in the mesangial matrix, and healing after glomerular injury may be facilitated by mesangial cell migration, which is guided by transient β8-integrin interactions with RGD ligands.

  14. Nanoprobes for two-photon excitation time-resolved imaging of living animals: In situ analysis of tumor-targeting dynamics of nanocarriers.

    Science.gov (United States)

    Yang, Wen; Fu, Li-Min; Wen, Xue; Liu, Ying; Tian, Ye; Liu, Yu-Chen; Han, Rong-Cheng; Gao, Zhi-Yue; Wang, Tian-En; Sha, Yin-Lin; Jiang, Yu-Qiang; Wang, Yuan; Zhang, Jian-Ping

    2016-09-01

    Great challenges remain in the noninvasive luminescence imaging analysis of tumor-targeting dynamics of nanocarriers in living animals which is of significance for the development of anti-cancer nanomedicine. In this work, luminescent nanoparticles Eu(tta)3bpt@SMA (dav = 15 nm), which exhibited good water dispersion stability and high yields of red Eu-luminescence under near-infrared two-photon excitation, were prepared by a modified microfluidic mixing method in the absence of surfactants. Tumor-targeting agents, Arg-Gly-Asp-D-Phe-Lys (cRGD) polypeptide or transferrin (Tf), were then anchored on the nanoparticle surfaces to form the desired nanocarriers Eu@SMA-RGD or Eu@SMA-Tf. The tumor-targeting processes of the prepared nanocarriers in intact living mice were analyzed on a home-built two-photon excitation time-resolved (TPE-TR) imaging apparatus having a wide view filed. The TPE-TR strategy could effectively suppress the interference from biological autofluorescence, which allowed the targeted domains to be visualized with a high signal-to-noise ratio. It was found that the tumor-tissue trapping efficacy of Eu@SMA-RGD was much higher than that of Eu@SMA-Tf, and the desorption process from the tumor tissues of Eu@SMA-RGD was slower than that of Eu@SMA-Tf. The methods developed in this work pave a way to investigate the in vivo tumor-targeting dynamics of nanocarriers by noninvasive luminescence imaging of living animals. PMID:27258485

  15. Utilizing cell-matrix interactions to modulate gene transfer to stem cells inside hyaluronic acid hydrogels.

    Science.gov (United States)

    Gojgini, Shiva; Tokatlian, Talar; Segura, Tatiana

    2011-10-01

    The effective delivery of DNA locally would increase the applicability of gene therapy in tissue regeneration, where diseased tissue is to be repaired in situ. One promising approach is to use hydrogel scaffolds to encapsulate and deliver plasmid DNA in the form of nanoparticles to the diseased tissue, so that cells infiltrating the scaffold are transfected to induce regeneration. This study focuses on the design of a DNA nanoparticle-loaded hydrogel scaffold. In particular, this study focuses on understanding how cell-matrix interactions affect gene transfer to adult stem cells cultured inside matrix metalloproteinase (MMP) degradable hyaluronic acid (HA) hydrogel scaffolds. HA was cross-linked to form a hydrogel material using a MMP degradable peptide and Michael addition chemistry. Gene transfer inside these hydrogel materials was assessed as a function of polyplex nitrogen to phosphate ratio (N/P = 5 to 12), matrix stiffness (100-1700 Pa), RGD (Arg-Gly-Asp) concentration (10-400 μM), and RGD presentation (0.2-4.7 RGDs per HA molecule). All variables were found to affect gene transfer to mouse mensenchymal stem cells culture inside the DNA loaded hydrogels. As expected, higher N/P ratios lead to higher gene transfer efficiency but also higher toxicity; softer hydrogels resulted in higher transgene expression than stiffer hydrogels, and an intermediate RGD concentration and RGD clustering resulted in higher transgene expression. We believe that the knowledge gained through this in vitro model can be utilized to design better scaffold-mediated gene delivery for local gene therapy.

  16. X-ray structure of engineered human Aortic Preferentially Expressed Protein-1 (APEG-1

    Directory of Open Access Journals (Sweden)

    Scheich Christoph

    2005-12-01

    Full Text Available Abstract Background Human Aortic Preferentially Expressed Protein-1 (APEG-1 is a novel specific smooth muscle differentiation marker thought to play a role in the growth and differentiation of arterial smooth muscle cells (SMCs. Results Good quality crystals that were suitable for X-ray crystallographic studies were obtained following the truncation of the 14 N-terminal amino acids of APEG-1, a region predicted to be disordered. The truncated protein (termed ΔAPEG-1 consists of a single immunoglobulin (Ig like domain which includes an Arg-Gly-Asp (RGD adhesion recognition motif. The RGD motif is crucial for the interaction of extracellular proteins and plays a role in cell adhesion. The X-ray structure of ΔAPEG-1 was determined and was refined to sub-atomic resolution (0.96 Å. This is the best resolution for an immunoglobulin domain structure so far. The structure adopts a Greek-key β-sandwich fold and belongs to the I (intermediate set of the immunoglobulin superfamily. The residues lying between the β-sheets form a hydrophobic core. The RGD motif folds into a 310 helix that is involved in the formation of a homodimer in the crystal which is mainly stabilized by salt bridges. Analytical ultracentrifugation studies revealed a moderate dissociation constant of 20 μM at physiological ionic strength, suggesting that APEG-1 dimerisation is only transient in the cell. The binding constant is strongly dependent on ionic strength. Conclusion Our data suggests that the RGD motif might play a role not only in the adhesion of extracellular proteins but also in intracellular protein-protein interactions. However, it remains to be established whether the rather weak dimerisation of APEG-1 involving this motif is physiogically relevant.

  17. Development of novel AAV serotype 6 based vectors with selective tropism for human cancer cells.

    Science.gov (United States)

    Sayroo, R; Nolasco, D; Yin, Z; Colon-Cortes, Y; Pandya, M; Ling, C; Aslanidi, G

    2016-01-01

    Viral vectors-based gene therapy is an attractive alternative to common anti-cancer treatments. In the present studies, AAV serotype 6 vectors were identified to be particularly effective in the transduction of human prostate (PC3), breast (T47D) and liver (Huh7) cancer cells. Next, we developed chimeric AAV vectors with Arg-Gly-Asp (RGD) peptide incorporated into the viral capsid to enable specific targeting of integrin-overexpressing malignant cells. These AAV6-RGD vectors improved transduction efficiency approximately 3-fold compared with wild-type AAV6 vectors by enhancing the viral entry into the cells. We also observed that transduction efficiency significantly improved, up to approximately 5-fold, by the mutagenesis of surface-exposed tyrosine and threonine residues involved in the intracellular trafficking of AAV vectors. Therefore, in our study, the AAV6-Y705-731F+T492V vector was identified as the most efficient. The combination of RGD peptide, tyrosine and threonine mutations on the same AAV6 capsid further increased the transduction efficiency, approximately 8-fold in vitro. In addition, we mutated lysine (K531E) to impair the affinity of AAV6 vectors to heparan sulfate proteoglycan. Finally, we showed a significant increase in both specificity and efficiency of AAV6-RGD-Y705-731F+T492V+K531E vectors in a xenograft animal model in vivo. In summary, the approach described here can lead to the development of AAV vectors with selective tropism to human cancer cells.

  18. Bioabsorbable bypass grafts biofunctionalised with RGD have enhanced biophysical properties and endothelialisation tested in vivo

    Directory of Open Access Journals (Sweden)

    Larisa V Antonova

    2016-05-01

    Full Text Available Small diameter arterial bypass grafts are considered as unmet clinical need since the current grafts have poor patency of 25% within 5 years. We have developed a 3D scaffold manufactured from natural and synthetic biodegradable polymers, poly(3-hydroxybutyrate-co-3-hydroxyvalerate (PHBV and poly(ε-caprolactone (PCL, respectively. Further to improve the biophysical properties as well as endothelialisation, the grafts were covalently conjugated with arginine-glycine-aspartic acid (RGD bioactive peptides. The biophysical properties as well as endothelialisation of PHBV/PCL and PCL 2 mm diameter bypass grafts were assessed with and without biofunctionalisation with RGD peptides in vitro and in vivo. Morphology of the grafts was assessed by scanning electron microscopy, whereas physico-mechanical properties were evaluated using a physiological circulating system equipped with a state of art ultrasound vascular wall tracking system. Endothelialisation of the grafts in vitro and in vivo were assessed using a cell viability assay and rat abdominal aorta replacement model, respectively. The biofunctionalisation with RGD bioactive peptides decreased mean fiber diameter and mean pore area in PHBV/PCL grafts; however, this was not the case for PCL grafts. Both PHBV/PCL and PCL grafts with RGD peptides had lower durability compared to those without; these durability values were similar to those of internal mammary artery. Modification of PHBV/PCL and PCL grafts with RGD peptides increased endothelial cell viability in vitro by a factor of 8 and enhanced the formation of an endothelial cell monolayer in vivo one month postimplantation. In conclusion, PHBV/PCL small-caliber graft can be a suitable 3D scaffold for the development of a tissue engineering arterial bypass graft.

  19. RGD-modified pH-sensitive liposomes for docetaxel tumor targeting.

    Science.gov (United States)

    Chang, Minglu; Lu, Shanshan; Zhang, Fang; Zuo, Tiantian; Guan, Yuanyuan; Wei, Ting; Shao, Wei; Lin, Guimei

    2015-05-01

    Phosphatidylethanolamine-based pH-sensitive liposomes of various compositions have been described as efficient systems for delivery of therapeutic molecules into tumor cells. The aim of this work was to develop a drug delivery system based on pH-sensitive liposomes (PLPs) that were modified with arginine-glycine-aspartic acid (RGD) peptide to enhance the effectiveness of docetaxel treatment. Docetaxel/coumarin-6 loaded PLPs were prepared by the thin-film dispersion method and characterized in detail, including by particle size, polydispersity, zeta potential and drug encapsulation efficiency. In vitro studies using MCF-7, HepG2and A549 cells were employed to investigate cytotoxicity and cellular uptake of the drug solution or docetaxel/coumarin-6 loaded PLPs. The accumulation of 7-nitro-2-1,3-benzoxadiazol-4-yl (NBD)-labeled liposomes in vivo was studied through tumor section imaging of xenograft mouse models of MCF-7 24h after intravenous administration. The particle size of the non-coated or RGD modified PLPs ranged between 146 and 129nm. Drug release in vitro was modestly prolonged and had good pH sensitivity. In the in vitro study, RGD-coated PLPs showed higher cytotoxicity and cellular uptake relative to non-coated ones. The results of the in vivo study showed that RGD-coated PLPs had higher fluorescence, which suggested a more efficient accumulation than normal PLPs in tumors. In conclusion, these results confirmed RGD-modified PLPs as a potential drug delivery system to achieve controlled release and tumor targeting. PMID:25851582

  20. Molecular mechanics of the cooperative adsorption of a Pro-Hyp-Gly tripeptide on a hydroxylated rutile TiO2(110) surface mediated by calcium ions.

    Science.gov (United States)

    Zheng, Ting; Wu, Chunya; Chen, Mingjun; Zhang, Yu; Cummings, Peter T

    2016-07-20

    The interaction of amino acids with inorganic materials at interfaces plays an important role in enhancing the biocompatibility of titanium-based alloys. The adsorption of a tripeptide, i.e. Pro-Hyp-Gly, on the hydroxylated rutile TiO2(110) surface was investigated by the MD simulations. The changes in free energy during the adsorption of both the tripeptide and calcium ions were calculated by using the PMF method in order to obtain the adsorption strength. The results suggested that the adsorption of the tripeptide on the TiO2 surface through the carboxyl groups in glycine residues can be more stable compared with other binding conformations. Special attention was focused on the cooperative adsorption of the tripeptide with the assistance of calcium ions. Calcium ions preferred to absorb at the tetradentate or monodentate sites on the negatively charged TiO2 surface. As a result of the strong attraction between the carboxyl group and calcium ions, the tripeptide can be pulled down to the surface by following the trajectory of the calcium ions, forming an indirect interaction with a sandwich structure of peptide-cation-TiO2. However, this indirect interaction could eventually transform to the direct adsorption of the tripeptide on the TiO2 surface with higher binding energy. The results may help to interpret the adsorption of peptides on inorganic materials in aqueous solution with ions. PMID:27383367

  1. The Effect of Superparamagnetic Iron Oxide with iRGD Peptide on the Labeling of Pancreatic Cancer Cells In Vitro: A Preliminary Study

    Directory of Open Access Journals (Sweden)

    Hou Dong Zuo

    2014-01-01

    Full Text Available The iRGD peptide loaded with iron oxide nanoparticles for tumor targeting and tissue penetration was developed for targeted tumor therapy and ultrasensitive MR imaging. Binding of iRGD, a tumor homing peptide, is mediated by integrins, which are widely expressed on the surface of cells. Several types of small molecular drugs and nanoparticles can be transfected into cells with the help of iRGD peptide. Thus, we postulate that SPIO nanoparticles, which have good biocompatibility, can also be transfected into cells using iRGD. Despite the many kinds of cell labeling studies that have been performed with SPIO nanoparticles and RGD peptide or its analogues, only a few have applied SPIO nanoparticles with iRGD peptide in pancreatic cancer cells. This paper reports our preliminary findings regarding the effect of iRGD peptide (CRGDK/RGPD/EC combined with SPIO on the labeling of pancreatic cancer cells. The results suggest that SPIO with iRGD peptide can enhance the positive labeling rate of cells and the uptake of SPIO. Optimal functionalization was achieved with the appropriate concentration or concentration range of SPIO and iRGD peptide. This study describes a simple and economical protocol to label panc-1 cells using SPIO in combination with iRGD peptide and may provide a useful method to improve the sensitivity of pancreatic cancer imaging.

  2. Ligand Conformation Dictates Membrane and Endosomal Trafficking of Arginine-Glycine-Aspartate (RGD)-Functionalized Mesoporous Silica Nanoparticles

    Energy Technology Data Exchange (ETDEWEB)

    Fang, I-Ju; Slowing, Igor I; Wu, Kevin C.W.; Lin, Victor S.Y.; Trewyn, Brian

    2012-05-15

    Recent breakthrough research on mesoporous silica nanoparticle (MSN) materials has illustrated their significant potential in biological applications due to their excellent drug delivery and endocytotic behavior. We set out to determine if MSN, covalently functionalized with conformation specific bioactive molecules (either linear or cyclic RGD ligands), behave towards mammalian cells in a similar manner as the free ligands. We discovered that RGD immobilized on the MSN surface did not influence the integrity of the porous matrix and improved the endocytosis efficiency of the MSN materials. Through competition experiments with free RGD ligands, we also discovered a conformation specific receptor–integrin association. The interaction between RGD immobilized on the MSN surface and integrins plays an important role in endosome trafficking, specifically dictating the kinetics of endosomal escape. Thus, covalent functionalization of biomolecules on MSN assists in the design of a system for controlling the interface with cancer cells.

  3. MicroPET imaging of brain tumor angiogenesis with {sup 18}F-labeled PEGylated RGD peptide

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Xiaoyuan; Park, Ryan; Hou, Yingping; Tohme, Michel; Bading, James R.; Conti, Peter S. [PET Imaging Science Center, Department of Radiology, University of Southern California Keck School of Medicine, 1510 San Pablo St., Suite 350, CA 90033, Los Angeles (United States); Khankaldyyan, Vazgen; Gonzales-Gomez, Ignacio; Laug, Walter E. [Department of Pediatrics, Childrens Hospital Los Angeles, CA 90027, Los Angeles (United States)

    2004-08-01

    We have previously labeled cyclic RGD peptide c(RGDyK) with fluorine-18 through conjugation labeling via a prosthetic 4-[{sup 18}F]fluorobenzoyl moiety and applied this [{sup 18}F]FB-RGD radiotracer for {alpha}{sub v}-integrin expression imaging in different preclinical tumor models with good tumor-to-background contrast. However, the unfavorable hepatobiliary excretion and rapid tumor washout rate of this tracer limit its potential clinical applications. The aims of this study were to modify the [{sup 18}F]FB-RGD tracer by inserting a heterobifunctional poly(ethylene glycol) (PEG, M.W. =3,400) between the {sup 18}F radiolabel and the RGD moiety and to test this [{sup 18}F]FB-PEG-RGD tracer for brain tumor targeting and in vivo kinetics. [{sup 18}F]FB-PEG-RGD was prepared by coupling the RGD-PEG conjugate with N-succinimidyl 4-[{sup 18}F]fluorobenzoate ([{sup 18}F]SFB) under slightly basic conditions (pH=8.5). The radiochemical yield was about 20-30% based on the active ester [{sup 18}F]SFB, and specific activity was over 100 GBq/{mu}mol. This tracer had fast blood clearance, rapid and high tumor uptake in the subcutaneous U87MG glioblastoma model (5.2{+-}0.5%ID/g at 30 min p.i.). Moderately rapid tumor washout was observed, with the activity accumulation decreased to 2.2{+-}0.4%ID/g at 4 h p.i. MicroPET and autoradiography imaging showed a very high tumor-to-background ratio and limited activity accumulation in the liver, kidneys and intestinal tracts. U87MG tumor implanted into the mouse forebrain was well visualized with [{sup 18}F]FB-PEG-RGD. Although uptake in the orthotopic tumor was significantly lower (P<0.01) than in the subcutaneous tumor, the maximum tumor-to-brain ratio still reached 5.0{+-}0.6 due to low normal brain background. The results of H and E staining post mortem agreed with the anatomical information obtained from non-invasive microPET imaging. In conclusion, PEGylation suitably modifies the physiological behavior of the RGD peptide. [{sup 18

  4. An RGD-restricted substrate interface is sufficient for the adhesion, growth and cartilage forming capacity of human chondrocytes

    Directory of Open Access Journals (Sweden)

    D Vonwil

    2010-11-01

    Full Text Available This study aimed at testing whether an RGD-restricted substrate interface is sufficient for adhesion and growth of human articular chondrocytes (HAC, and whether it enhances their post expansion chondrogenic capacity. HAC/substrate interaction was restricted to RGD by modifying tissue culture polystyrene (TCPS with a poly(ethylene glycol (PEG based copolymer system that renders the surface resistant to protein adsorption while at the same time presenting the bioactive RGD-containing peptide GCRGYGRGDSPG (RGD. As compared to TCPS, HAC cultured on RGD spread faster (1.9-fold, maintained higher type II collagen mRNA expression (4.9-fold and displayed a 19% lower spreading area. On RGD, HAC attachment efficiency (66±10% and proliferation rate (0.56±0.04 doublings/day, as well as type II collagen mRNA expression in the subsequent chondrogenic differentiation phase, were similar to those of cells cultured on TCPS. In contrast, cartilaginous matrix deposition by HAC expanded on RGD was slightly but consistently higher (15% higher glycosaminoglycan-to-DNA ratio. RDG (bioinactive peptide and PEG (no peptide ligand controls yielded drastically reduced attachment efficiency (lower than 11% and proliferation (lower than 0.20 doublings/day. Collectively, these data indicate that restriction of HAC interaction with a substrate through RGD peptides is sufficient to support their adhesion, growth and maintenance of cartilage forming capacity. The concept could thus be implemented in materials for cartilage repair, whereby in situ recruited/infiltrated chondroprogenitor cells would proliferate while maintaining their ability to differentiate and generate cartilage tissue.

  5. Low-Molecular Weight Polyethylenimine Modified with Pluronic 123 and RGD- or Chimeric RGD-NLS Peptide: Characteristics and Transfection Efficacy of Their Complexes with Plasmid DNA

    Directory of Open Access Journals (Sweden)

    Jing Hu

    2016-05-01

    Full Text Available To solve the problem of transfection efficiency vs. cytotoxicity and tumor-targeting ability when polyethylenimine (PEI was used as a nonviral gene delivery vector, new degradable PEI polymers were synthesized via cross-linking low-molecular-weight PEI with Pluronic P123 and then further coupled with a targeting peptide R4 (RGD and a bifunctional R11 (RGD-NLS, which were termed as P123-PEI-R4 and P123-PEI-R11, respectively. Agarose gel electrophoresis showed that both P123-PEI-R4 and P123-PEI-R11 efficaciously condense plasmid DNA at a polymer-to-pDNA w/w ratio of 3.0 and 0.4, respectively. The polyplexes were stable in the presence of serum and could protect plasmid DNA against DNaseI. They had uniform spherical nanoparticles with appropriate sizes around 100–280 nm and zeta-potentials about +40 mV. Furthermore, in vitro experiments showed that these polyplexes had lower cytotoxicity at any concentration compared with PEI 25 kDa, thus giving promise to high transfection efficiency as compared with another P123-PEI derivate conjugated with trifunctional peptide RGD-TAT-NLS (P123-PEI-R18. More importantly, compared with the other polymers, P123-PEI-R11 showed the highest transfection efficiency with relatively lower cytotoxicity at any concentration, indicating that the new synthetic polymer P123-PEI-R11 could be used as a safe and efficient gene deliver vector.

  6. Low-Molecular Weight Polyethylenimine Modified with Pluronic 123 and RGD- or Chimeric RGD-NLS Peptide: Characteristics and Transfection Efficacy of Their Complexes with Plasmid DNA.

    Science.gov (United States)

    Hu, Jing; Zhao, Wenfang; Liu, Kehai; Yu, Qian; Mao, Yuan; Lu, Zeyu; Zhang, Yaguang; Zhu, Manman

    2016-01-01

    To solve the problem of transfection efficiency vs. cytotoxicity and tumor-targeting ability when polyethylenimine (PEI) was used as a nonviral gene delivery vector, new degradable PEI polymers were synthesized via cross-linking low-molecular-weight PEI with Pluronic P123 and then further coupled with a targeting peptide R4 (RGD) and a bifunctional R11 (RGD-NLS), which were termed as P123-PEI-R4 and P123-PEI-R11, respectively. Agarose gel electrophoresis showed that both P123-PEI-R4 and P123-PEI-R11 efficaciously condense plasmid DNA at a polymer-to-pDNA w/w ratio of 3.0 and 0.4, respectively. The polyplexes were stable in the presence of serum and could protect plasmid DNA against DNaseI. They had uniform spherical nanoparticles with appropriate sizes around 100-280 nm and zeta-potentials about +40 mV. Furthermore, in vitro experiments showed that these polyplexes had lower cytotoxicity at any concentration compared with PEI 25 kDa, thus giving promise to high transfection efficiency as compared with another P123-PEI derivate conjugated with trifunctional peptide RGD-TAT-NLS (P123-PEI-R18). More importantly, compared with the other polymers, P123-PEI-R11 showed the highest transfection efficiency with relatively lower cytotoxicity at any concentration, indicating that the new synthetic polymer P123-PEI-R11 could be used as a safe and efficient gene deliver vector. PMID:27213305

  7. Evaluation of the therapeutic efficacy and radiotoxicity of the conjugates {sup 177}Lu-DOTA-E-c(RGDfK){sub 2} and {sup 177}Lu-DOTA-GGC-AuNP-c[RGDfk(C)] in a murine model and their relationship with the inhibition of the angiogenic factors VEGF and HIF-1α; Evaluacion de la eficacia terapeutica y radiotoxicidad de los conjugados {sup 177}Lu-DOTA-E-c(RGDfK){sub 2} y {sup 177}Lu-DOTA-GGC-AuNP-c[RGDfK(C)] en un modelo murino y su relacion con la inhibicion de los factores angiogenicos VEGF y HIF-1α

    Energy Technology Data Exchange (ETDEWEB)

    Vilchis J, A.

    2013-07-01

    Molecular targeting therapy has become a relevant therapeutic strategy for cancer. The principle that peptide receptors can be used successfully for in vivo targeting of human cancers has been proven, and radiolabeled peptides have been demonstrated to be effective in patients with malignant tumors. Peptides based on the cyclic Arg-Gly-Asp (RGD) sequence have been designed to antagonize the function of α(v)β(3) integrin, thereby inhibiting angio genesis. The conjugation of RGD peptides to radiolabeled gold nanoparticles (AuNP) produces biocompatible and stable m ultimeric systems with target-specific molecular recognition. The aim of this research was to evaluate the therapeutic response of {sup 177}Lu-AuNP-RGD in athymic mice bearing α(v)β(3)-integrin-positive C6 gliomas and compare with that of {sup 177}Lu-AuNP or {sup 177}Lu-RGD. The radiation absorbed dose, metabolic activity (SUV, [18F]fluor-deoxy-glucose-micro PET/CT), renal radiotoxicity, renal and tumoral histological characteristics as well as tumoral VEGF and HIF-1? gene expression (by realtime polymerase chain reaction) following treatment with {sup 177}Lu-AuNP-RGD, {sup 177}Lu-AuNP or {sup 177}Lu-RGD were assessed. Of the radiopharmaceuticals evaluated, {sup 177}Lu-AuNP-RGD delivered the highest tumor radiation absorbed dose (63.8 ± 7.9 Gy) vs other treatments. These results correlated with the observed therapeutic response, in which {sup 177}Lu-AuNP-RGD significantly (p<0.05) reduced tumor progression, tumor metabolic activity, intratumoral vessels and VEGF gene expression compared to the other radiopharmaceuticals. This was consequence of high tumor retention and a combination of molecular targeting therapy (m ultimeric RGD system) and radiotherapy ({sup 177}Lu). There was a low uptake in non-target organs and no induction of renal toxicity. {sup 177}Lu-AuNP-RGD demonstrates properties suitable for use as an agent for molecular targeting radiotherapy. (Author)

  8. Evaluation of 111In-Labeled Cyclic RGD Peptides: Effects of Peptide and Linker Multiplicity on Their Tumor Uptake, Excretion Kinetics and Metabolic Stability

    Directory of Open Access Journals (Sweden)

    Jiyun Shi, Yang Zhou, Sudipta Chakraborty, Young-Seung Kim, Bing Jia, Fan Wang, Shuang Liu

    2011-01-01

    Full Text Available Purpose: The purpose of this study was to demonstrate the valence of cyclic RGD peptides, P-RGD (PEG4-c(RGDfK: PEG4 = 15-amino-4,710,13-tetraoxapentadecanoic acid, P-RGD2 (PEG4-E[c(RGDfK]2, 2P-RGD4 (E{PEG4-E[c(RGDfK]2}2, 2P4G-RGD4 (E{PEG4-E[G3-c(RGDfK]2}2: G3 = Gly-Gly-Gly and 6P-RGD4 (E{PEG4-E[PEG4-c(RGDfK]2}2 in binding to integrin αvβ3, and to assess the impact of peptide and linker multiplicity on biodistribution properties, excretion kinetics and metabolic stability of their corresponding 111In radiotracers.Methods: Five new RGD peptide conjugates (DOTA-P-RGD (DOTA =1,4,7,10-tetraazacyclododecane-1,4,7,10-tetracetic acid, DOTA-P-RGD2, DOTA-2P-RGD4, DOTA-2P4G-RGD4, DOTA-6P-RGD4, and their 111In complexes were prepared. The integrin αvβ3 binding affinity of cyclic RGD conjugates were determined by a competitive displacement assay against 125I-c(RGDyK bound to U87MG human glioma cells. Biodistribution, planar imaging and metabolism studies were performed in athymic nude mice bearing U87MG human glioma xenografts.Results: The integrin αvβ3 binding affinity of RGD conjugates follows the order of: DOTA-6P-RGD4 (IC50 = 0.3 ± 0.1 nM ~ DOTA-2P4G-RGD4 (IC50 = 0.2 ± 0.1 nM ~ DOTA-2P-RGD4 (IC50 = 0.5 ± 0.1 nM > DOTA-3P-RGD2 (DOTA-PEG4-E[PEG4-c(RGDfK]2: IC50 = 1.5 ± 0.2 nM > DOTA-P-RGD2 (IC50 = 5.0 ± 1.0 nM >> DOTA-P-RGD (IC50 = 44.3 ± 3.5 nM ~ c(RGDfK (IC50 = 49.9 ± 5.5 nM >> DOTA-6P-RGK4 (IC50 = 437 ± 35 nM. The fact that DOTA-6P-RGK4 had much lower integrin αvβ3 binding affinity than DOTA-6P-RGD4 suggests that the binding of DOTA-6P-RGD4 to integrin αvβ3 is RGD-specific. This conclusion is consistent with the lower tumor uptake for 111In(DOTA-6P-RGK4 than that for 111In(DOTA-6P-RGD4. It was also found that the G3 and PEG4 linkers between RGD motifs have a significant impact on the integrin αvβ3-targeting capability, biodistribution characteristics, excretion kinetics and metabolic stability of 111In-labeled cyclic RGD peptides

  9. Liposomal muramyl tripeptide phosphatidylethanolamine (MTP-PE) promotes haemopoietic recovery in irradiated mouse

    International Nuclear Information System (INIS)

    Pretreatment of C57B1/6 mouse with the macrophage activator muramyl tripeptide phosphatidylethanolamine encapsulated in liposomes (MTP-PE/MLV) induced haemopoietic recovery in subsequently irradiated mouse. An optimal endoCFU-S survival was observed when 200 μg MTP-PE/MLV was administered i.p. 24 h before irradiation. MTP-PE/MLV did not affect the day 8 exogenous CFU-S survival in the bone marrow immediately after irradiation. However, 3, 6, 9 and 14 days after irradiation the number of day 8 CFU-S was almost 2 to 4-fold higher in the bone marrow of the MTP-PE/MLV injected mouse. Also, recovery of the GM-CFC pools in femoral bone marrow after irradiation proceeded at a faster rate in the MTP-PE/MLV-treated animal than in control groups. Pretreatment with MTP-PE/MLV protected the C57B1/6 mouse in a dose-dependent manner from the lethal effects of ionizing radiation. (Author)

  10. Design, Synthesis, and Evaluation of New Tripeptides as COX-2 Inhibitors

    Directory of Open Access Journals (Sweden)

    Ermelinda Vernieri

    2013-01-01

    Full Text Available Cyclooxygenase (COX is a key enzyme in the biosynthetic pathway leading to the formation of prostaglandins, which are mediators of inflammation. It exists mainly in two isoforms COX-1 and COX-2. The conventional nonsteroidal anti-inflammatory drugs (NSAIDs have gastrointestinal side effects because they inhibit both isoforms. Recent data demonstrate that the overexpression of these enzymes, and in particular of cyclooxygenases-2, promotes multiple events involved in tumorigenesis; in addition, numerous studies show that the inhibition of cyclooxygenases-2 can delay or prevent certain forms of cancer. Agents that inhibit COX-2 while sparing COX-1 represent a new attractive therapeutic development and offer a new perspective for a further use of COX-2 inhibitors. The present study extends the evaluation of the COX activity to all 203 possible natural tripeptide sequences following a rational approach consisting in molecular modeling, synthesis, and biological tests. Based on data obtained from virtual screening, only those peptides with better profile of affinity have been selected and classified into two groups called S and E. Our results suggest that these novel compounds may have potential as structural templates for the design and subsequent development of the new selective COX-2 inhibitors drugs.

  11. A nickel tripeptide as a metallodithiolate ligand anchor for resin-bound organometallics.

    Science.gov (United States)

    Green, Kayla N; Jeffery, Stephen P; Reibenspies, Joseph H; Darensbourg, Marcetta Y

    2006-05-17

    The molecular structure of the acetyl CoA synthase enzyme has clarified the role of individual nickel atoms in the dinickel active site which mediates C-C and C-S coupling reactions. The NiN2S2 portion of the biocatalyst (N2S2 = a cysteine-glycine-cysteine or CGC4- tripeptide ligand) serves as an S-donor ligand comparable to classical bidentate ligands operative in organometallic chemistry, ligating the second nickel which is redox and catalytically active. Inspired by this biological catalyst, the synthesis of NiN2S2 metalloligands, including the solid-phase synthesis of resin-bound Ni(CGC)2-, and sulfur-based derivatization with W(CO)5 and Rh(CO)2+ have been carried out. Through comparison to analogous well-characterized, solution-phase complexes, Attenuated Total Reflectance FTIR spectroscopy establishes the presence of unique heterobimetallic complexes, of the form [Ni(CGC)]M(CO)x, both in solution and immobilized on resin beads. This work provides the initial step toward exploitation of such an evolutionarily optimized nickel peptide as a solid support anchor for hybrid bioinorganic-organometallic catalysts.

  12. Egg-derived tri-peptide IRW exerts antihypertensive effects in spontaneously hypertensive rats.

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    Kaustav Majumder

    Full Text Available BACKGROUND: There is a growing interest in using functional food components as therapy for cardiovascular diseases such as hypertension. We have previously characterized a tri-peptide IRW (Ile-Arg-Trp from egg white protein ovotransferrin; this peptide showed anti-inflammatory, anti-oxidant and angiotensin converting enzyme (ACE inhibitor properties in vitro. Given the pathogenic roles played by angiotensin, oxidative stress and inflammation in the spontaneously hypertensive rat (SHR, we tested the therapeutic potential of IRW in this well-established model of hypertension. METHODS AND RESULTS: 16-17 week old male SHRs were orally administered IRW at either a low dose (3 mg/Kg BW or a high dose (15 mg/Kg BW daily for 18 days. Blood pressure (BP and heart rate were measured by telemetry. Animals were sacrificed at the end of the treatment for vascular function studies and measuring markers of inflammation. IRW treatment attenuated mean BP by ~10 mmHg and ~40 mmHg at the low- and high-dose groups respectively compared to untreated SHRs. Heart rate was not affected. Reduction in BP was accompanied by the restoration of diurnal variations in BP, preservation of nitric oxide dependent vasorelaxation, as well as reduction of plasma angiotensin II, other inflammatory markers and tissue fibrosis. CONCLUSION: Our results demonstrate anti-hypertensive effects of IRW in vivo likely mediated through ACE inhibition, endothelial nitric oxide synthase and anti-inflammatory properties.

  13. Bond cleavage reactions in the tripeptide tri-alanine upon free electron capture

    International Nuclear Information System (INIS)

    In the present study we performed dissociative electron attachment (DEA) measurements with the tripeptide tri-alanine, C9H17N3O4, utilizing a crossed electron-molecular beam experiment with high electron energy resolution (about 100 meV). Anion efficiency yields as a function of the incident electron energy are obtained for the most abundant anions up to electron energies of ∼ eV. Quantum chemical calculations are performed to determine the thermochemical thresholds for the anions observed in the measurements. There is no evidence of a molecular anion with lifetime of mass spectrometric timescales. The dehydrogenated closed shell anion (M-H)- is one of the fragment anions observed for which the calculations show that H-loss is energetically possible from carboxyl, as well as amide groups. In contrast to the dipeptide di-alanine and monomer alanine the cleavage of the N-Cα bond in the peptide chain is already possible by attachment of electrons at ∼ 0 eV. (authors)

  14. Evaluation of 111In-Labeled Cyclic RGD Peptides: Effects of Peptide and Linker Multiplicity on Their Tumor Uptake, Excretion Kinetics and Metabolic Stability

    OpenAIRE

    Jiyun Shi, Yang Zhou, Sudipta Chakraborty, Young-Seung Kim, Bing Jia, Fan Wang, Shuang Liu

    2011-01-01

    Purpose: The purpose of this study was to demonstrate the valence of cyclic RGD peptides, P-RGD (PEG4-c(RGDfK): PEG4 = 15-amino-4,710,13-tetraoxapentadecanoic acid), P-RGD2 (PEG4-E[c(RGDfK)]2, 2P-RGD4 (E{PEG4-E[c(RGDfK)]2}2, 2P4G-RGD4 (E{PEG4-E[G3-c(RGDfK)]2}2: G3 = Gly-Gly-Gly) and 6P-RGD4 (E{PEG4-E[PEG4-c(RGDfK)]2}2) in binding to integrin αvβ3, and to assess the impact of peptide and linker multiplicity on biodistribution properties, excretion kinetics and metabolic stability of ...

  15. Evaluation of 111In-Labeled Cyclic RGD Peptides: Effects of Peptide and Linker Multiplicity on Their Tumor Uptake, Excretion Kinetics and Metabolic Stability

    OpenAIRE

    Shi, Jiyun; Zhou, Yang; Chakraborty, Sudipta; Kim, Young-Seung; Jia, Bing; Wang, Fan; LIU, SHUANG

    2011-01-01

    Purpose: The purpose of this study was to demonstrate the valence of cyclic RGD peptides, P-RGD (PEG4-c(RGDfK): PEG4 = 15-amino-4,710,13-tetraoxapentadecanoic acid), P-RGD2 (PEG4-E[c(RGDfK)]2, 2P-RGD4 (E{PEG4-E[c(RGDfK)]2}2, 2P4G-RGD4 (E{PEG4-E[G3-c(RGDfK)]2}2: G3 = Gly-Gly-Gly) and 6P-RGD4 (E{PEG4-E[PEG4-c(RGDfK)]2}2) in binding to integrin αvβ3, and to assess the impact of peptide and linker multiplicity on biodistribution properties, excretion kinetics and metabolic stability of their corr...

  16. Immobilization of RGD Peptides onto Decellularized Valve Scaffolds to Promote Cell Adhesion

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    Porcine aortic valves were decellularized with trypsinase/EDTA and Triton-100. With the help of a coupling reagent Sulfo-LC-SPDP, the biological valve scaffolds were immobilized with one of RGD(arginine-glycine-aspartic acid) containing peptides, called GRGDSPC peptide. Myofibroblasts harvested from rats were seeded onto them. Based on the spectra of X-ray photoelectron spectroscopy, we could find conjugation of GRGDSPC peptide and the scaffolds. Cell count by both microscopy and MTT assay showed that myofibroblasts were easier to adhere to the modified scaffolds. It is proved that it is feasible to immobilize RGD peptides onto decellularized valve scaffolds, and effective to promote cell adhesion, which is beneficial for constructing tissue engineering heart valves in vitro.

  17. The Adenovirus Type 3 Dodecahedron's RGD Loop Comprises an HSPG Binding Site That Influences Integrin Binding

    Directory of Open Access Journals (Sweden)

    E. Gout

    2010-01-01

    Full Text Available Human type 3 adenovirus dodecahedron (a virus like particle made of twelve penton bases features the ability to enter cells through Heparan Sulphate Proteoglycans (HSPGs and integrins interaction and is used as a versatile vector to deliver DNA or proteins. Cryo-EM reconstruction of the pseudoviral particle with Heparan Sulphate (HS oligosaccharide shows an extradensity on the RGD loop. A set of mutants was designed to study the respective roles of the RGD sequence (RGE mutant and of a basic sequence located just downstream. Results showed that the RGE mutant binding to the HS deficient CHO-2241 cells was abolished and unexpectedly, mutation of the basic sequence (KQKR to AQAS dramatically decreased integrin recognition by the viral pseudoparticle. This basic sequence is thus involved in integrin docking, showing a close interplay between HSPGs and integrin receptors.

  18. Rational Design of Cancer-Targeted Benzoselenadiazole by RGD Peptide Functionalization for Cancer Theranostics.

    Science.gov (United States)

    Yang, Liye; Li, Wenying; Huang, Yanyu; Zhou, Yangliang; Chen, Tianfeng

    2015-09-01

    A cancer-targeted conjugate of the selenadiazole derivative BSeC (benzo[1,2,5] selenadiazole-5-carboxylic acid) with RGD peptide as targeting molecule and PEI (polyethylenimine) as a linker is rationally designed and synthesized in the present study. The results show that RGD-PEI-BSeC forms nanoparticles in aqueous solution with a core-shell nanostructure and high stability under physiological conditions. This rational design effectively enhances the selective cellular uptake and cellular retention of BSeC in human glioma cells, and increases its selectivity between cancer and normal cells. The nanoparticles enter the cells through receptor-mediated endocytosis via clathrin-mediated and nystatin-dependent lipid raft-mediated pathways. Internalized nanoparticles trigger glioma cell apoptosis by activation of ROS-mediated p53 phosphorylation. Therefore, this study provides a strategy for the rational design of selenium-containing cancer-targeted theranostics.

  19. An iRGD Based Strategy to Study Electrochemically the Species Inside a Cell

    Directory of Open Access Journals (Sweden)

    Genxi Li

    2012-08-01

    Full Text Available This paper reports a method for electrical communication between the inner part of cells and an electrode with the help of iRGD peptide. Due to the enhancement of the cell penetration caused by iRGD peptide, DNA molecules, previously modified on a gold electrode surface, can be easily transfected into the cells. At the same time, doxorubicin, an anticancer drug, can also be transfected into cells with high penetration. Consequently, doxorubicin binds to DNA chains through electrostatic interaction, and the redox reaction is transferred out of the cell across the cell membrane. As a result, this work may provide a novel way to get information from inside of cells.

  20. Radiolabeled antibodies and RGD-peptides for the treatment of ovarian cancer

    OpenAIRE

    Janssen, M.L.H.

    2004-01-01

    In this thesis, preclinical studies on new treatment modalities for ovarian cancer are descibed, applying radiolabeled antibodies and radiolabeled RGD-peptides. In chapter 2 a study is described comparing the therapeutic efficacy of the antibody HMFG1 radiolabeled with several beta-emitting radionuclides in mice with intraperitoneal ovarian carcinoma. Mice were injected with the radiopharmaceuticals 90Y-HMFG1, 186Re-HMFG1 or 131I-HMFG1. Each of the i.p. administered radiolabeled antibody prep...

  1. Measurement of interaction force between RGD-peptide and Hela cell surface by optical tweezers

    Institute of Scientific and Technical Information of China (English)

    Mincheng Zhong; Guosheng Xue; Jinhua Zhou; Ziqiang Wang; Yinmei Li

    2012-01-01

    Since RGD peptides (R:arginine; G:glycine; D:aspartic acid) are found to promote cell adhesion,they are modified at numerous materials surface for medical applications such as drug delivery and regenerative medicine.Peptide-cell surface interactions play a key role in the above applications.In this letter,we study the adhesion force between the RGD-coated bead and Hela cell surface by optical tweezes.The adhesion is dominated by the binding of α5β1 and RGD-peptide with higher adhesion probability and stronger adhesion strength compared with the adhesion of bare bead and cell surface.The binding force for a single α5β1-GRGDSP pair is determined to be 16.8 pN at a loading rate of 1.5 nN/s.The unstressed off-rate is 1.65 × 10-2 s-1 and the distance of transition state for the rigid binding model is 3.0 nm.

  2. Synthesis of dimeric cyclic RGD based near-infrared probe for in vivo tumor diagnosis

    Science.gov (United States)

    Cao, Jie; Wan, Shunan; Tian, Junmei; Chi, Xuemei; Du, Changli; Deng, Dawei; Chen, Wei R.; Gu, Yueqing

    2012-03-01

    Cell adhesion molecule integrin αvβ3 is an excellent target for tumor interventions because of its unique expression on the surface of several types of solid tumor cells and on almost all sprouting tumor vasculatures. In this manuscript, we describe the synthesis of near-infrared (NIR) fluorochrome ICG-Der-02-labeled dimeric cyclic RGD peptides (ICG-Der-02-c(RGDyK)2) for in vivo tumor integrin targeting. The optical properties and structure of the probe were intensively characterized. Afterwards, the integrin specificity of the fluorescent probe was tested in vitro for receptor binding assay and fluorescence microscopy and in vivo for subcutaneous MDA-MB-231 and U87MG tumor targeting. The results indicated that after labeling RGD peptide, the optical properties of ICG-Der-02 showed no obvious change. Besides, in vitro and in vivo tumor targeting experiment indicated that the ICG-Der-02-c(RGDyK)2 probe with high integrin affinity showed excellent tumor activity accumulation. Noninvasive NIR fluorescence imaging is able to detect tumor integrin expression based upon the highly potent RGD peptide probe.

  3. Locally-Delivered T-Cell-Derived Cellular Vehicles Efficiently Track and Deliver Adenovirus Delta24-RGD to Infiltrating Glioma

    Directory of Open Access Journals (Sweden)

    Rutger K. Balvers

    2014-08-01

    Full Text Available Oncolytic adenoviral vectors are a promising alternative for the treatment of glioblastoma. Recent publications have demonstrated the advantages of shielding viral particles within cellular vehicles (CVs, which can be targeted towards the tumor microenvironment. Here, we studied T-cells, often having a natural capacity to target tumors, for their feasibility as a CV to deliver the oncolytic adenovirus, Delta24-RGD, to glioblastoma. The Jurkat T-cell line was assessed in co-culture with the glioblastoma stem cell (GSC line, MGG8, for the optimal transfer conditions of Delta24-RGD in vitro. The effect of intraparenchymal and tail vein injections on intratumoral virus distribution and overall survival was addressed in an orthotopic glioma stem cell (GSC-based xenograft model. Jurkat T-cells were demonstrated to facilitate the amplification and transfer of Delta24-RGD onto GSCs. Delta24-RGD dosing and incubation time were found to influence the migratory ability of T-cells towards GSCs. Injection of Delta24-RGD-loaded T-cells into the brains of GSC-bearing mice led to migration towards the tumor and dispersion of the virus within the tumor core and infiltrative zones. This occurred after injection into the ipsilateral hemisphere, as well as into the non-tumor-bearing hemisphere. We found that T-cell-mediated delivery of Delta24-RGD led to the inhibition of tumor growth compared to non-treated controls, resulting in prolonged survival (p = 0.007. Systemic administration of virus-loaded T-cells resulted in intratumoral viral delivery, albeit at low levels. Based on these findings, we conclude that T-cell-based CVs are a feasible approach to local Delta24-RGD delivery in glioblastoma, although efficient systemic targeting requires further improvement.

  4. Distinct specificities of Mycobacterium tuberculosis and mammalian proteasomes for N-acetyl tripeptide substrates.

    Science.gov (United States)

    Lin, Gang; Tsu, Christopher; Dick, Lawrence; Zhou, Xi K; Nathan, Carl

    2008-12-01

    The proteasome of Mycobacterium tuberculosis (Mtb) is a validated and drug-treatable target for therapeutics. To lay ground-work for developing peptide-based inhibitors with a useful degree of selectivity for the Mtb proteasome over those of the host, we used a library of 5,920 N-acetyl tripeptide-aminomethylcoumarins to contrast the substrate preferences of the recombinant Mtb proteasome wild type and open gate mutant, the Rhodococcus erythropolis proteasome, and the bovine proteasome with activator PA28. The Mtb proteasome was distinctive in strictly preferring P1 = tryptophan, particularly in combination with P3 = glycine, proline, lysine or arginine. Screening results were validated with Michalis-Menten kinetic analyses of 21 oligopeptide aminomethyl-coumarin substrates. Bortezomib, a proteasome inhibitor in clinical use, and 17 analogs varying only at P1 were used to examine the differential impact of inhibitors on human and Mtb proteasomes. The results with the inhibitor panel confirmed those with the substrate panel in demonstrating differential preferences of Mtb and mammalian proteasomes at the P1 amino acid. Changing P1 in bortezomib from Leu to m-CF(3)-Phe led to a 220-fold increase in IC(50) against the human proteasome, whereas changing a P1 Ala to m-F-Phe decreased the IC(50) 400-fold against the Mtb proteasome. The change of a P1 Ala to m-Cl-Phe led to an 8000-fold shift in inhibitory potency in favor of the Mtb proteasome, resulting in 8-fold selectivity. Combinations of preferred amino acids at different sites may thus improve the species selectivity of peptide-based inhibitors that target the Mtb proteasome. PMID:18829465

  5. Potentiometric and spectroscopic study of the complexation of copper(II) ions by tripeptides containing aromatic side-chains

    Science.gov (United States)

    Ghalem, S.; Fan, B.-T.; Xiao, L.

    1998-01-01

    The complexation of copper(II) ions with L,L-Gly-Phe-Phe, L,L-Phe-Gly-Phe and L,L-Phe-Phe-Gly was studied by potentiometric and spectroscopic measurements. Only four complexes have been found for each copper(II)-tripeptide system, and no species with two ligand molecules was observed. The results show influences of aromatic side-chains. These influences are dependent upon the location of two aromatic rings in studied tripeptides. The stabilization or destabilization of a given complex is probably the result of several different effects, including steric hindrance, hydrophobic effect, electrodonor effect and π-d interaction. The spectroscopic measurements, e.s.r and electronic absorption, are useful to determine the complex structures. La complexation du cuivre(II) par Gly-Phe-Phe-L,L, Phe-Gly-Phe-L,L et Phe-Phe-Gly-L,L a été étudiée par potentiométrie et par spectroscopies. Seulement quatre espèces ont été mises en évidence pour chaque système Cu(II)-tripeptide. Aucun complexe contenant deux molécules de ligand n'a été observé. Les résultats obtenus montrent des influences évidentes liées aux chaînes latérales aromatiques. Ces influences dépendent des positions des résidus phénylalanines. La stabilisation ou déstabilisation d'un complexe est probablement le résultat d'un ensemble de différents effets : effet stérique, effet hydrophobe, électrodonneur et l'interaction π-d. Les spectroscopies RPE et visible ont été utilisées pour la détermination structurale des complexes.

  6. Formation of Peptide Bound Pyrraline in the Maillard Model Systems with Different Lys-Containing Dipeptides and Tripeptides

    Directory of Open Access Journals (Sweden)

    Zhili Liang

    2016-04-01

    Full Text Available Peptide-bound advanced glycation end-products (peptide-bound AGEs can be formed when peptides are heated with reducing saccharides. Pyrraline is the one of most commonly studied AGEs in foods, but the relative importance of the precursor peptide structure is uncertain. In the present study, model systems were prepared by heating peptides with glucose from 60 °C to 220 °C for up to 65 min, and the amounts of peptide-bound pyrraline formed were monitored to evaluate the effect of the neighboring amino acids on the peptide-bound pyrraline formation. The physico-chemical properties were introduced to explore the quantitative structure-reactivity relationships between physicochemical properties and peptide bound formation. 3-DG content in dipeptide-glucose model system was higher than that in the corresponding tripeptide-glucose model systems. Dipeptides produced higher amounts of peptide-bound pyrraline than the corresponding tripeptides. The peptide-bound pyrraline and 3-DG production were influenced by the physico-chemical properties of the side chain of amino acids adjacent to Lys in the following order: Lys-Leu/glucose > Lys-Ile/glucose > Lys-Val/ glucose > Lys-Thr/glucose > Lys-Ser/glucose > Lys-Ala/ glucose > Lys-Gly/glucose; Lys-Leu-Gly/glucose > Lys-Ile-Gly/glucose > Lys-Val-Gly/glucose > Lys-Thr-Gly/glucose > Lys-Ser-Gly/glucose > Lys-Ala-Gly/glucose > Lys-Gly-Gly/glucose. For the side chain of amino acids adjacent to Lys in dipeptides, residue volume, polarizability, molecular volume and localized electrical effect were positively related to the yield of peptide bound pyrraline, while hydrophobicity and pKb were negatively related to the yield of peptide bound pyrraline. In terms of side chain of amino acid adjacent to Lys in tripeptides, a similar result was observed, except hydrophobicity was positively related to the yield of peptide bound pyrraline.

  7. Intravenous administration of the conditionally replicative adenovirus Ad5-Δ24RGD induces regression of osteosarcoma lung metastases

    Directory of Open Access Journals (Sweden)

    Gerritsen Winald R

    2008-01-01

    Full Text Available Abstract Metastatic osteosarcoma (OS has a very poor prognosis. New treatments are therefore wanted. The conditionally replicative adenovirus Ad5-Δ24RGD has shown promising anti-tumor effects on local cancers, including OS. The purpose of this study was to determine whether intravenous administration of Ad5-Δ24RGD could suppress growth of human OS lung metastases. Mice bearing SaOs-lm7 OS lung metastases were treated with Ad5-Δ24RGD at weeks 1, 2 and 3 or weeks 5, 6 and 7 after tumor cell injection. Virus treatment at weeks 1–3 did not cause a statistically significant effect on lung weight and total body weight. However, the number of macroscopic lung tumor nodules was reduced from a median of >158 in PBS-treated control mice to 58 in Ad5-Δ24RGD-treated mice (p = 0.15. Moreover, mice treated at weeks 5–7 showed a significantly reduced lung weight (decrease of tumor mass, p 149, p = 0.12 compared to PBS treated control animals. Adenovirus hexon expression was detected in lung tumor nodules at sacrifice three weeks after the last intravenous adenovirus administration, suggesting ongoing viral infection. These findings suggest that systemic administration of Ad5-Δ24RGD might be a promising new treatment strategy for metastatic osteosarcoma.

  8. Synthesis, Characterization, and Evaluation of a Novel Amphiphilic Polymer RGD-PEG-Chol for Target Drug Delivery System

    Directory of Open Access Journals (Sweden)

    Shi Zeng

    2014-01-01

    Full Text Available An amphiphilic polymer RGD-PEG-Chol which can be produced in large scale at a very low cost has been synthesized successfully. The synthesized intermediates and final products were characterized and confirmed by 1H nuclear magnetic resonance spectrum (1H NMR and Fourier transform infrared spectrum (FT-IR. The paclitaxel- (PTX- loaded liposomes based on RGD-PEG-Chol were then prepared by film formation method. The liposomes had a size within 100 nm and significantly enhanced the cytotoxicity of paclitaxel to B16F10 cell as demonstrated by MTT test (IC50 = 0.079 μg/mL of RGD-modified PTX-loaded liposomes compared to 9.57 μg/mL of free PTX. Flow cytometry analysis revealed that the cellular uptake of coumarin encapsulated in the RGD-PEG-Chol modified liposome was increased for HUVEC cells. This work provides a reasonable, facile, and economic approach to prepare peptide-modified liposome materials with controllable performances and the obtained linear RGD-modified PTX-loaded liposomes might be attractive as a drug delivery system.

  9. Identification and Characterization of Di- and Tripeptide Transporter DtpT of Listeria monocytogenes EGD-e

    OpenAIRE

    Wouters, Jeroen A.; Hain, Torsten; Darji, Ajub; Hüfner, Eric; Wemekamp-Kamphuis, Henrike; Chakraborty, Trinad; Abee, Tjakko

    2005-01-01

    Listeria monocytogenes is a gram-positive intracellular pathogen responsible for opportunistic infections in humans and animals. Here we identified and characterized the dtpT gene (lmo0555) of L. monocytogenes EGD-e, encoding the di- and tripeptide transporter, and assessed its role in growth under various environmental conditions as well as in the virulence of L. monocytogenes. Uptake of the dipeptide Pro-[14C]Ala was mediated by the DtpT transporter and was abrogated in a ΔdtpT isogenic del...

  10. Identification and characterization of di- and tripeptide transporter DtpT of Listeria monocytogenes EGD-e

    OpenAIRE

    Wouters, J.A.; Hain, T.; Darji, A; Hüfner, E.; Wemekamp-Kamphuis, H.H.; Chakraborty, T.; Abee, T

    2005-01-01

    Listeria monocytogenes is a gram-positive intracellular pathogen responsible for opportunistic infections in humans and animals. Here we identified and characterized the dtpT gene (lmo0555) of L. monocytogenes EGD-e, encoding the di- and tripeptide transporter, and assessed its role in growth under various environmental conditions as well as in the virulence of L. monocytogenes. Uptake of the dipeptide Pro-[14C]Ala was mediated by the DtpT transporter and was abrogated in a dtpT isogenic dele...

  11. Nanosolvation by acetonitrile and 18-crown-6 ether induce strongly different effects on the electron-capture induced dissociation of aromatic tripeptide cations in the gas phase

    DEFF Research Database (Denmark)

    MacLot, S.; Rangama, J.; Nielsen, Steen Brøndsted;

    2013-01-01

    Experimental gas-phase Electron Capture-Induced Dissociation (ECID) coupled to mass-spectrometry has been performed on the doubly-protonated tripeptides Lys-Trp-Lys (KWK) and Lys-Tyr-Lys (KYK). In this report, we focus on the influence of non-covalent binding of two different molecules......, acetonitrile and 18-crown-6 ether (CE), to these tripeptide cations on the relative probabilities of their main fragmentation channels (H loss, NH3 loss and N — Cα bond cleavage) after electron capture from sodium atoms. First, we recorded the spectra of bare peptide ions, and found that N — Cα bond cleavage...... leads to fragments containing the aromatic amino acid. The structures and energies of the low-lying conformers of the tripeptide dications and radical monocations obtained from our DFT and MP2 calculations are in line with this observation. Second, the ECID spectra of KWK and KYK dications nanosolvated...

  12. The HDAC inhibitors scriptaid and LBH589 combined with the oncolytic virus Delta24-RGD exert enhanced anti-tumor efficacy in patient-derived glioblastoma cells

    NARCIS (Netherlands)

    Pont, L.M.E.B. (Lotte M. E. Berghauser); A. Kleijn (Anne); J.J. Kloezeman (Jenneke); Van Bossche, W.D. (Wouter Den); Kaufmann, J.K. (Johanna K.); J. Vrij (Jeroen); S. Leenstra (Sieger); C.M.F. Dirven (Clemens); M.L.M. Lamfers (Martine)

    2015-01-01

    textabstractBackground: A phase I/II trial for glioblastoma with the oncolytic adenovirus Delta24-RGD was recently completed. Delta24-RGD conditionally replicates in cells with a disrupted retinoblastomapathway and enters cells via αvβ3/5 integrins. Glioblastomas are differentially sensitive to Delt

  13. Standard Thermodynamic Functions of Tripeptides N-Formyl-l-methionyl-l-leucyl-l-phenylalaninol and N-Formyl-l-methionyl-l-leucyl-l-phenylalanine Methyl Ester

    OpenAIRE

    Markin, Alexey V.; Markhasin, Evgeny; Sologubov, Semen S.; Smirnova, Natalia N.; Griffin, Robert G.

    2014-01-01

    The heat capacities of tripeptides N-formyl-l-methionyl-l-leucyl-l-phenylalaninol (N-f-MLF-OH) and N-formyl-l-methionyl-l-leucyl-l-phenylalanine methyl ester (N-f-MLF-OMe) were measured by precision adiabatic vacuum calorimetry over the temperature range from T = (6 to 350) K. The tripeptides were stable over this temperature range, and no phase change, transformation, association, or thermal decomposition was observed. The standard thermodynamic functions: molar heat capacity C p,m, enthalpy...

  14. Non-canonical peroxisome targeting signals: identification of novel PTS1 tripeptides and characterization of enhancer elements by computational permutation analysis

    Directory of Open Access Journals (Sweden)

    Chowdhary Gopal

    2012-08-01

    Full Text Available Abstract Background High-accuracy prediction tools are essential in the post-genomic era to define organellar proteomes in their full complexity. We recently applied a discriminative machine learning approach to predict plant proteins carrying peroxisome targeting signals (PTS type 1 from genome sequences. For Arabidopsis thaliana 392 gene models were predicted to be peroxisome-targeted. The predictions were extensively tested in vivo, resulting in a high experimental verification rate of Arabidopsis proteins previously not known to be peroxisomal. Results In this study, we experimentally validated the predictions in greater depth by focusing on the most challenging Arabidopsis proteins with unknown non-canonical PTS1 tripeptides and prediction scores close to the threshold. By in vivo subcellular targeting analysis, three novel PTS1 tripeptides (QRL>, SQM>, and SDL> and two novel tripeptide residues (Q at position −3 and D at pos. -2 were identified. To understand why, among many Arabidopsis proteins carrying the same C-terminal tripeptides, these proteins were specifically predicted as peroxisomal, the residues upstream of the PTS1 tripeptide were computationally permuted and the changes in prediction scores were analyzed. The newly identified Arabidopsis proteins were found to contain four to five amino acid residues of high predicted targeting enhancing properties at position −4 to −12 in front of the non-canonical PTS1 tripeptide. The identity of the predicted targeting enhancing residues was unexpectedly diverse, comprising besides basic residues also proline, hydroxylated (Ser, Thr, hydrophobic (Ala, Val, and even acidic residues. Conclusions Our computational and experimental analyses demonstrate that the plant PTS1 tripeptide motif is more diverse than previously thought, including an increasing number of non-canonical sequences and allowed residues. Specific targeting enhancing elements can be predicted for particular sequences

  15. Measurement and modeling of acid dissociation constants of tri-peptides containing Glu, Gly, and His using potentiometry and generalized multiplicative analysis of variance.

    Science.gov (United States)

    Khoury, Rima Raffoul; Sutton, Gordon J; Hibbert, D Brynn; Ebrahimi, Diako

    2013-02-28

    We report pK(a) values with measurement uncertainties for all labile protons of the 27 tri-peptides prepared from the amino acids glutamic acid (E), glycine (G) and histidine (H). Each tri-peptide (GGG, GGE, GGH, …, HHH) was subjected to alkali titration and pK(a) values were calculated from triplicate potentiometric titrations data using HyperQuad 2008 software. A generalized multiplicative analysis of variance (GEMANOVA) of pK(a) values for the most acidic proton gave the optimum model having two terms, an interaction between the end amino acids plus an isolated main effect of the central amino acid.

  16. Computational study of the RGD-peptide interactions with perovskite-type BFO-(1 1 1) membranes under aqueous conditions

    Science.gov (United States)

    Li, Hai-long; Bian, Liang; Hou, Wen-ping; Dong, Fa-Qin; Song, Mian-Xin; Zhang, Xiao-yan; Wang, Li-sheng

    2016-07-01

    We elucidated a number of facets regarding arginine-glycine-aspartate (RGD)-bismuth ferrite (BFO)-(1 1 1) membrane interactions and reactivity that have previously remained unexplored on a molecular level. Results demonstrate the intra-molecular interaction facilitates a "horseshoe" structure of RGD adsorbed onto the BFO-(1 1 1) membrane, through the electrostatic (Asp-cation-Fe) and water-bridge (Osbnd H2O and H2Osbnd NH2) interactions. The effect of structural and electron-transfer interactions is attributed to the cation-valences, indicating that the divalent cations are electron-acceptors and the monovalent cations as electron-donors. Notably, the strongly bound Ca2+ ion exerts a "gluing" effect on the Asp-side-chain, indicating a tightly packed RGD-BFO configuration. Thus, modulating the biological response of BFO-(1 1 1) membrane will allow us to design more appropriate interfaces for implantable diagnostic and therapeutic perovskite-type micro-devices.

  17. 68Ga- and 111In-labelled DOTA-RGD peptides for imaging of αvβ3 integrin expression

    International Nuclear Information System (INIS)

    αvβ3 integrins are important cell adhesion receptors involved in angiogenic processes. Recently, we demonstrated using [18F]Galacto-RGD that monitoring of αvβ3 expression is feasible. Here, we introduce 68Ga- and 111In-labelled derivatives and compare them with [18F]Galacto-RGD. For radiolabelling, cyclo(RGDfK(DOTA)) was synthesised using SPPS. For in vitro characterisation determination of partition coefficients, protein binding, metabolic stability, αvβ3 affinity and cell uptake and for in vivo characterization, biodistribution studies and micro positron emission tomography (PET) imaging were carried out. For in vivo and in vitro studies, human melanoma M21 (αvβ3 positive) and M21-L (αvβ3 negative) cells were used. Both tracers can be synthesised straightforward. The compounds showed hydrophilic properties and high metabolic stability. Up to 23% protein-bound activity for [68Ga]DOTA-RGD and only up to 1.4% for [111In]DOTA-RGD was found. Cell uptake studies indicate receptor-specific accumulation. This is confirmed by the biodistribution data. One hour p.i. accumulation in αvβ3-positive tumours was 2.9 ± 0.3%ID/g and in αvβ3-negative tumours 0.8 ± 0.1%ID/g for [68Ga]DOTA-RGD ([111In]DOTA-RGD: 1.9 ± 0.3%ID/g and 0.5 ± 0.2%ID/g; [18F]Galacto-RGD: 1.6 ± 0.2%ID/g and 0.4 ± 0.1%ID/g). Thus, tumour uptake ratios were comparable. Due to approx. 3-fold higher blood pool activities for [68Ga]DOTA-RGD, tumour/blood ratios were higher for [111In]DOTA-RGD and [18F]Galacto-RGD. However, microPET studies demonstrated that visualisation of αvβ3-positive tumours using [68Ga]DOTA-RGD is possible. Our data indicate that [68Ga]DOTA-RGD allows monitoring of αvβ3 expression. Especially, the much easier radiosynthesis compared to [18F]Galacto-RGD would make it an attractive alternative. However, due to higher blood pool activity, [18F]Galacto-RGD remains superior for imaging αvβ3 expression. Introduction of alternative chelator systems may overcome the

  18. The vitronectin RGD motif regulates TGF-β-induced alveolar epithelial cell apoptosis.

    Science.gov (United States)

    Wheaton, Amanda K; Velikoff, Miranda; Agarwal, Manisha; Loo, Tiffany T; Horowitz, Jeffrey C; Sisson, Thomas H; Kim, Kevin K

    2016-06-01

    Transforming growth factor-β (TGF-β) is a critical driver of acute lung injury and fibrosis. Injury leads to activation of TGF-β, which regulates changes in the cellular and matrix makeup of the lung during the repair and fibrosis phase. TGF-β can also initiate alveolar epithelial cell (AEC) apoptosis. Injury leads to destruction of the laminin-rich basement membrane, which is replaced by a provisional matrix composed of arginine-glycine-aspartate (RGD) motif-containing plasma matrix proteins, including vitronectin and fibronectin. To determine the role of specific matrix proteins on TGF-β-induced apoptosis, we studied primary AECs cultured on different matrix conditions and utilized mice with deletion of vitronectin (Vtn(-/-)) or mice in which the vitronectin RGD motif is mutated to nonintegrin-binding arginine-glycine-glutamate (RGE) (Vtn(RGE/RGE)). We found that AECs cultured on fibronectin and vitronectin or in wild-type mouse serum are resistant to TGF-β-induced apoptosis. In contrast, AECs cultured on laminin or in serum from Vtn(-/-) or Vtn(RGE/RGE) mice undergo robust TGF-β-induced apoptosis. Plasminogen activator inhibitor-1 (PAI-1) sensitizes AECs to greater apoptosis by disrupting AEC engagement to vitronectin. Inhibition of integrin-associated signaling proteins augments AEC apoptosis. Mice with transgenic deletion of PAI-1 have less apoptosis after bleomycin, but deletion of vitronectin or disruption of the vitronectin RGD motif reverses this protection, suggesting that the proapoptotic function of PAI-1 is mediated through vitronectin inhibition. Collectively, these data suggest that integrin-matrix signaling is an important regulator of TGF-β-mediated AEC apoptosis and that PAI-1 functions as a natural regulator of this interaction. PMID:27106291

  19. Expression and purification of rhIL-10-RGD from Escherichia coli as a potential wound healing agent.

    Science.gov (United States)

    Yang, Fangfang; Wan, Yi; Liu, Jiaqi; Yang, Xuekang; Wang, Hongtao; Tao, Ke; Han, Juntao; Shi, Jihong; Hu, Dahai

    2016-08-01

    Various protocols for recombinant Interleukin-10 (IL-10) purification in wound healing have been reported previously. However, the therapeutic effect was not obvious. Thus, it is of great importance to find new and effective approaches for therapy. In this study, we propose that IL-10 and Arginine-Glycine-Aspartic (RGD) peptide would be a valuable therapeutic for wound healing. To explore a high-efficiency and cost-effective approach for the production of IL-10 and RGD peptide with bioactivity, a synthetic gene was cloned into a recombinant pTWIN1 vector. As a consequence, rhIL-10-RGD and the pH-induced self-cleavable Ssp DnaB mini-intein as a fusion protein was highly expressed by IPTG induction in Escherichia coli Rosetta without extra residues in a bioreactor. After Ni affinity chromatographic purification, rhIL-10-RGD was released by the Ssp DnaB intein-mediated self-cleavage that is triggered by pH shift. SDS-PAGE and silver staining showed a major band with an estimated molecular mass of 19.3kDa. Cell proliferation assay confirmed its potent proliferation activity on MC/9 murine mast cells. In conclusion, we report a novel strategy to produce rhIL-10-RGD mediated by the pH-induced self-cleavable Ssp DnaB mini-intein, and show that rhIL-10-RGD could play an effective role in wound healing of BALB/c mice. PMID:27241829

  20. {sup 99m}Tc-HYNIC-RGD analog for monitoring of integrin expression in ischemic animal model

    Energy Technology Data Exchange (ETDEWEB)

    Queiroz, Rodrigo G.; Faintuch, Bluma L.; Teodorao, Rodrigo; Muramoto, Emiko [Instituto de Pesquisas Energeticas e Nucleares (IPEN/CNEN-SP), Sao Paulo, SP (Brazil)]. E-mail: teodoro_rodrigo@yahoo.com.br; blfaintuch@hotmail.com

    2007-07-01

    Introduction: The development of new blood vessels, from existing network of angiogenesis, occurs during embryonic development, cancer, arthritis and in response to ischemia. Ischemia in muscle tissue increases expression of integrin in the form of proliferating endothelial cells, including the {alpha}{sub v}{beta}{sub 3} receptor. Cyclic peptides containing RGD sequences have high affinity and selectivity for {alpha}{sub v}{beta}{sub 3} integrin, and may thus be used for angiogenesis monitoring. Aim: The aim of this study was the investigation of neovascularization in an ischemic hindlimb model using RGD conjugated with HYNIC and labeled with technetium-99m. Methods: {sup 99m}Tc-HYNIC-RGD analog was prepared using tricine and EDDA as coligands. Unilateral hindlimb ischemia was induced in Wistar rats by surgical ligation of the femoral artery. Radiolabeled RGD was injected in rats after 2 h, as well as 1, 3, 5, 7, 10 and 14 days of ischemic induction. Evaluation of uptake by ischemic limb and normal contralateral limb was done by planar imaging and biodistribution studies. Pertechnetate distribution, which has no affinity for regeneration tissue and in the circumstances indicates just blood flow, was also assessed for confirmation. Results: The highest uptake ratio between ischemic hindlimb and control side was achieved at the seventh day (2.62 {+-} 0.95), with substantial decreased by the fourteenth day. For pertechnetate the seventh day ratio was 0.87 {+-} 0.23, consistent with reduced perfusion following arterial ligation. Scintigraphic imaging confirmed the RGD results. Both renal and hepatic elimination was observed, with noticeable uptake also by intestine, spleen and bone. Conclusions: {sup 99m}Tc-HYNIC-RGD analog corresponded to the expectations in this animal model, displaying elevated uptake in ischemic tissue by the time of widespread angiogenesis. Utilization in other ischemic models can be recommended. (author)

  1. Facile synthesis of RGD peptide-modified iron oxide nanoparticles with ultrahigh relaxivity for targeted MR imaging of tumors.

    Science.gov (United States)

    Hu, Yong; Li, Jingchao; Yang, Jia; Wei, Ping; Luo, Yu; Ding, Ling; Sun, Wenjie; Zhang, Guixiang; Shi, Xiangyang; Shen, Mingwu

    2015-05-01

    We report the facile synthesis of arginine-glycine-aspartic acid (RGD) peptide-targeted iron oxide (Fe3O4) nanoparticles (NPs) with ultrahigh relaxivity for in vivo tumor magnetic resonance (MR) imaging. In this study, stable polyethyleneimine (PEI)-coated Fe3O4 NPs were first prepared by a mild reduction route. The formed aminated Fe3O4 NPs with PEI coating were sequentially conjugated with fluorescein isothiocyanate (FI) and polyethylene glycol (PEG)-RGD segment, followed by acetylation of the remaining PEI surface amines. The thus-formed Fe3O4@PEI·NHAc-FI-PEG-RGD NPs were characterized via different techniques. We show that the multifunctional RGD-targeted Fe3O4 NPs with a mean size of 9.1 nm are water-dispersible, colloidally stable, and hemocompatible and cytocompatible in the given concentration range. With the displayed ultrahigh r2 relaxivity (550.04 mM(-1) s(-1)) and RGD-mediated targeting specificity to αvβ3 integrin-overexpressing cancer cells as confirmed by flow cytometry and confocal microscopy, the developed multifunctional Fe3O4@PEI·NHAc-FI-PEG-RGD NPs are able to be used as a highly efficient nanoprobe for targeted MR imaging of αvβ3 integrin-overexpressing cancer cells in vitro and the xenografted tumor model in vivo. Given the versatile PEI amine-enabled conjugation chemistry, the developed PEI-coated Fe3O4 NPs may be functionalized with other biological ligands or drugs for various biomedical applications, in particular, the diagnosis and therapy of different types of cancer. PMID:26222591

  2. The Human Tripeptide GHK-Cu in Prevention of Oxidative Stress and Degenerative Conditions of Aging: Implications for Cognitive Health

    Directory of Open Access Journals (Sweden)

    Loren Pickart

    2012-01-01

    Full Text Available Oxidative stress, disrupted copper homeostasis, and neuroinflammation due to overproduction of proinflammatory cytokines are considered leading causative factors in development of age-associated neurodegenerative conditions. Recently, a new mechanism of aging—detrimental epigenetic modifications—has emerged. Thus, compounds that possess antioxidant, anti-inflammatory activity as well as compounds capable of restoring copper balance and proper gene functioning may be able to prevent age-associated cognitive decline and ward off many common neurodegenerative conditions. The aim of this paper is to bring attention to a compound with a long history of safe use in wound healing and antiaging skin care. The human tripeptide GHK was discovered in 1973 as an activity in human albumin that caused old human liver tissue to synthesize proteins like younger tissue. It has high affinity for copper ions and easily forms a copper complex or GHK-Cu. In addition, GHK possesses a plethora of other regenerative and protective actions including antioxidant, anti-inflammatory, and wound healing properties. Recent studies revealed its ability to up- and downregulate a large number of human genes including those that are critical for neuronal development and maintenance. We propose GHK tripeptide as a possible therapeutic agent against age-associated neurodegeneration and cognitive decline.

  3. The human tripeptide GHK-Cu in prevention of oxidative stress and degenerative conditions of aging: implications for cognitive health.

    Science.gov (United States)

    Pickart, Loren; Vasquez-Soltero, Jessica Michelle; Margolina, Anna

    2012-01-01

    Oxidative stress, disrupted copper homeostasis, and neuroinflammation due to overproduction of proinflammatory cytokines are considered leading causative factors in development of age-associated neurodegenerative conditions. Recently, a new mechanism of aging-detrimental epigenetic modifications-has emerged. Thus, compounds that possess antioxidant, anti-inflammatory activity as well as compounds capable of restoring copper balance and proper gene functioning may be able to prevent age-associated cognitive decline and ward off many common neurodegenerative conditions. The aim of this paper is to bring attention to a compound with a long history of safe use in wound healing and antiaging skin care. The human tripeptide GHK was discovered in 1973 as an activity in human albumin that caused old human liver tissue to synthesize proteins like younger tissue. It has high affinity for copper ions and easily forms a copper complex or GHK-Cu. In addition, GHK possesses a plethora of other regenerative and protective actions including antioxidant, anti-inflammatory, and wound healing properties. Recent studies revealed its ability to up- and downregulate a large number of human genes including those that are critical for neuronal development and maintenance. We propose GHK tripeptide as a possible therapeutic agent against age-associated neurodegeneration and cognitive decline. PMID:22666519

  4. Synchrotron X-ray fluorescence studies of a bromine-labelled cyclic RGD peptide interacting with individual tumor cells

    International Nuclear Information System (INIS)

    The first example of synchrotron X-ray fluorescence imaging of cultured mammalian cells in cyclic peptide research is reported. The study reports the first quantitative analysis of the incorporation of a bromine-labelled cyclic RGD peptide and its effects on the biodistribution of endogenous elements (for example, K and Cl) within individual tumor cells. The first example of synchrotron X-ray fluorescence imaging of cultured mammalian cells in cyclic peptide research is reported. The study reports the first quantitative analysis of the incorporation of a bromine-labelled cyclic RGD peptide and its effects on the biodistribution of endogenous elements (for example, K and Cl) within individual tumor cells

  5. Synthesis, Characterization, and Evaluation of a Novel Amphiphilic Polymer RGD-PEG-Chol for Target Drug Delivery System

    OpenAIRE

    Shi Zeng; Fengbo Wu; Bo Li; Xiangrong Song; Yu Zheng; Gu He; Cheng Peng; Wei Huang

    2014-01-01

    An amphiphilic polymer RGD-PEG-Chol which can be produced in large scale at a very low cost has been synthesized successfully. The synthesized intermediates and final products were characterized and confirmed by 1H nuclear magnetic resonance spectrum (1H NMR) and Fourier transform infrared spectrum (FT-IR). The paclitaxel- (PTX-) loaded liposomes based on RGD-PEG-Chol were then prepared by film formation method. The liposomes had a size within 100 nm and significantly enhanced the cytotoxicit...

  6. Development of laminated fiber-reinforced nanocomposites for bone regeneration

    Science.gov (United States)

    Xu, Weijie

    There have been numerous efforts to develop synthetic and/or natural tissue engineering scaffolds that are suitable for bone regeneration applications to replace autograft and allograft bones. Current biomaterials as a scaffold for bone regeneration are limited by the extent of degradation concurrent with bone formation, mechanical strength, and the extent of osteogenic differentiation of marrow stromal cells migrating from the surrounding tissues. In this project, a novel laminated nanocomposite scaffold is fabricated, consisting of poly (L-lactide ethylene oxide fumarate) (PLEOF) hydrogel reinforced with poly (L-lactic acid) (PLLA) electrospun nanofibers and hydroxyapatite (HA) nanoparticles. PLEOF is a novel in situ crosslinkable macromer synthesized from biocompatible building units which can be functionalized with bioactive peptides like the cell-adhesive Arg--Gly--Asp (RGD) amino acid sequence. The hydrophilicity and degradation rate of the macromer can be tailored to a particular application by controlling the ratio of PEG to PLA blocks in the macromer and the unsaturated fumarate units can be used for in-situ crosslinking. The PLLA nanofibers were electrospun from high molecular weight PLLA. The laminated nanocomposites were fabricated by dry-hand lay up technique followed by compression molding and thermal crosslinking. The laminated nanocomposites were evaluated with respect to degradation, water uptake, mechanical strength, and the extent of osteogenic differentiation of bone marrow stromal (BMS) cells. Laminates with or without HA nanoparticles showed modulus values much higher than that of trabecular bone (50-100 MPa). The effect of laminated nanocomposites on osteogenic differentiation of BMS cells was determined in terms of cell number, ALPase activity and calcium content. Our results demonstrate that grafting RGD peptide and HA nanoparticles to a PLEOF hydrogel reinforced with PLLA nanofibers synergistically enhance osteogenic differentiation of BMS

  7. The VP1 S154D mutation of type Asia1 foot-and-mouth disease virus enhances viral replication and pathogenicity.

    Science.gov (United States)

    Lian, Kaiqi; Yang, Fan; Zhu, Zixiang; Cao, Weijun; Jin, Ye; Liu, Huanan; Li, Dan; Zhang, Keshan; Guo, Jianhong; Liu, Xiangtao; Zheng, Haixue

    2016-04-01

    One of the proteins encoded by the foot-and-mouth disease virus (FMDV), the VP1 protein, a capsid protein, plays an important role in integrin receptor attachment and humoral immunity-mediated host responses. The integrin receptor recognition motif and an important antigenic epitope exist within the G-H loop, which is comprised of amino acids 134-160 of the VP1 protein. FMDV strain, Asia1/HN/CHA/06, isolated from a pig, was passaged four times in suckling mice and sequenced. Sequencing analyses showed that there was a mutation of the integrin receptor recognition motif Arg-Gly-Asp/Arg-Asp-Asp (RGD/RDD, VP1 143-145) and a VP1 154 serine/Asp (VP1 S154D) mutation in the G-H loop of the VP1 protein. The influence of the RGD/RDD mutation on Asia1 FMDV disease phenotype has been previously studied. In this study, to determine the influence of the VP1 S154D mutation on FMDV Asia1 replication and pathogenicity, two recombinant FMDVs with different residues only at the VP1 154 site were rescued by reverse genetics techniques and their infectious potential in host cells and pathogenicity in pigs were compared. Our data indicates that the VP1 S154D mutation increases the replication level of FMDV Asia1/HN/CHA/06 in BHK-21, IB-RS-2, and PK-15 cells and enhances pathogenicity in pigs. Through the transient transfection-infection assay to compare integrin receptor usage of two recombinant viruses, the result shows that the VP1 S154D mutation markedly increases the ability of type Asia1 FMDV to use the integrin receptors αυβ6 and αυβ8 from pig. This study identifies a key research target for illuminating the role of residues located at G-H loop in FMDV pathogenicity. PMID:26792712

  8. ‘Living’ PEGylation on gold nanoparticles to optimize cancer cell uptake by controlling targeting ligand and charge densities

    Science.gov (United States)

    Chen, Hongwei; Paholak, Hayley; Ito, Masayuki; Sansanaphongpricha, Kanokwan; Qian, Wei; Che, Yong; Sun, Duxin

    2013-09-01

    We report and demonstrate biomedical applications of a new technique—‘living’ PEGylation—that allows control of the density and composition of heterobifunctional PEG (HS-PEG-R; thiol-terminated poly(ethylene glycol)) on gold nanoparticles (AuNPs). We first establish ‘living’ PEGylation by incubating HS-PEG5000-COOH with AuNPs (˜20 nm) at increasing molar ratios from zero to 2000. This causes the hydrodynamic layer thickness to differentially increase up to 26 nm. The controlled, gradual increase in PEG-COOH density is revealed after centrifugation, based on the ability to re-suspend the pellet and increase the AuNP absorption. Using a fluorescamine-based assay we quantify differential HS-PEG5000-NH2 binding to AuNPs, revealing that it is highly efficient until AuNP saturation is reached. Furthermore, the zeta potential incrementally changes from -44.9 to +52.2 mV and becomes constant upon saturation. Using ‘living’ PEGylation we prepare AuNPs with different ratios of HS-PEG-RGD (RGD: Arg-Gly-Asp) and incubate them with U-87 MG (malignant glioblastoma) and non-target cells, demonstrating that targeting ligand density is critical to maximizing the efficiency of targeting of AuNPs to cancer cells. We also sequentially control the HS-PEG-R density to develop multifunctional nanoparticles, conjugating positively charged HS-PEG-NH2 at increasing ratios to AuNPs containing negatively charged HS-PEG-COOH to reduce uptake by macrophage cells. This ability to minimize non-specific binding/uptake by healthy cells could further improve targeted nanoparticle efficacy.

  9. Structure-guided creation of AcAP5-derived and platelet targeted factor Xa inhibitors.

    Science.gov (United States)

    Zhu, Yuanjun; Lin, Yuan; Liu, Aihua; Shui, Mengyang; Li, Ruyi; Liu, Xiaoyan; Hu, Wenhui; Wang, Yinye

    2015-06-15

    Anticoagulants and anti-platelet agents are simultaneously administrated in clinical practice (i.e. percutaneous coronary intervention), which cause significant risk of systemic bleeding. Targeted delivery of anticoagulants to the activated platelets at sites of vascular injuries may condense the site-specific anticoagulant effect and reduce the hemorrhage side effects in uninjured vessels. To this end, we prepared three ancylostoma caninum anticoagulant peptide 5 (AcAP5) variants NR1, NR2 and NR3 engineered with a platelet-binding Arg-Gly-Asp (RGD) motif and evaluated their anti-Factor Xa (FXa) and platelet-binding effects. These RGD-containing AcAP5 variants were capable of interacting with platelet receptor αIIbβ3 as shown in computational analysis. All variants, especially NR2 and NR3, retained entirely the anti-FXa function of parent AcAP5. Moreover, they prevented the formation of occlusive thrombi in rat carotid artery injury model, suggesting that they inhibit platelet aggregation in vivo. Further functional investigation of NR3 demonstrated that NR3 inhibited platelet aggregation in vitro and FXa activity in vivo, and prolonged the coagulation time, all in a dose-dependent manner. Through flow cytometry assay, we confirmed the binding of NR3 to αIIbβ3 receptor. In mouse model of carotid artery endothelium injury, NR3-treated mice showed less tail bleeding time than AcAP5-treated mice, and aspirin plus NR3 treatment exhibited moderate reduction of blood loss compared with aspirin plus AcAP5 treatment. These results indicate the feasibility to engineer a novel FXa inhibitor specifically targeting the activated platelets, which centralizes its anticoagulation efficacy in the injured vascular endothelium and reduces the risk of systemic bleeding. PMID:25887920

  10. Induced Pluripotent Stem Cell-derived Mesenchymal Stem Cell Seeding on Biofunctionalized Calcium Phosphate Cements

    Institute of Scientific and Technical Information of China (English)

    WahWah TheinHan; Jun Liu; Minghui Tang; Wenchuan Chen; Linzhao Cheng; Hockin H. K. Xu

    2013-01-01

    Induced pluripotent stem cells (iPSCs) have great potential due to their proliferation and differentiation capability. The objectives of this study were to generate iPSC-derived mesenchymal stem cells (iPSC-MSCs), and investigate iPSC-MSC proliferation and osteogenic differentiation on calcium phosphate cement (CPC) containing biofunctional agents for the first time. Human iPSCs were derived from marrow CD34+ cells which were reprogrammed by a single episomal vector. iPSCs were cultured to form embryoid bodies (EBs), and MSCs migrated out of EBs. Five biofunctional agents were incorporated into CPC:RGD (Arg-Gly-Asp) peptides, fibronectin (Fn), fibronectin-like engineered polymer protein (FEPP), extracellular matrix Geltrex, and platelet concentrate. iPSC-MSCs were seeded on five biofunctionalized CPCs:CPC-RGD, CPC-Fn, CPC-FEPP, CPC-Geltrex, and CPC-Platelets. iPSC-MSCs on biofunctional CPCs had enhanced proliferation, actin fiber expression, osteogenic differentiation and mineralization, compared to control. Cell proliferation was greatly increased on biofunctional CPCs. iPSC-MSCs underwent osteogenic differentiation with increased alkaline phosphatase, Runx2 and collagen-I expressions. Mineral synthesis by iPSC-MSCs on CPC-Platelets was 3-fold that of CPC control. In conclusion, iPSCs showed high potential for bone engineering. iPSC-MSCs on biofunctionalized CPCs had cell proliferation and bone mineralization that were much better than traditional CPC. iPSC-MSC-CPC constructs are promising to promote bone regeneration in craniofacial/orthopedic repairs.

  11. RGD-Binding Integrins in Prostate Cancer: Expression Patterns and Therapeutic Prospects against Bone Metastasis

    International Nuclear Information System (INIS)

    Prostate cancer is the third leading cause of male cancer deaths in the developed world. The current lack of highly specific detection methods and efficient therapeutic agents for advanced disease have been identified as problems requiring further research. The integrins play a vital role in the cross-talk between the cell and extracellular matrix, enhancing the growth, migration, invasion and metastasis of cancer cells. Progression and metastasis of prostate adenocarcinoma is strongly associated with changes in integrin expression, notably abnormal expression and activation of the β3 integrins in tumour cells, which promotes haematogenous spread and tumour growth in bone. As such, influencing integrin cell expression and function using targeted therapeutics represents a potential treatment for bone metastasis, the most common and debilitating complication of advanced prostate cancer. In this review, we highlight the multiple ways in which RGD-binding integrins contribute to prostate cancer progression and metastasis, and identify the rationale for development of multi-integrin antagonists targeting the RGD-binding subfamily as molecularly targeted agents for its treatment

  12. Cyclic isoDGR and RGD peptidomimetics containing bifunctional diketopiperazine scaffolds are integrin antagonists.

    Science.gov (United States)

    Panzeri, Silvia; Zanella, Simone; Arosio, Daniela; Vahdati, Leila; Dal Corso, Alberto; Pignataro, Luca; Paolillo, Mayra; Schinelli, Sergio; Belvisi, Laura; Gennari, Cesare; Piarulli, Umberto

    2015-04-13

    The cyclo[DKP-isoDGR] peptidomimetics 2-5, containing bifunctional diketopiperazine (DKP) scaffolds that differ in the configuration of the two DKP stereocenters and in the substitution at the DKP nitrogen atoms, were prepared and examined in vitro in competitive binding assays with purified αv β3 and αv β5 integrin receptors. IC50 values ranged from low nanomolar (ligand 3) to submicromolar with αv β3 integrin. The biological activities of ligands cyclo[DKP3-RGD] 1 and cyclo[DKP3-isoDGR] 3, bearing the same bifunctional DKP scaffold and showing similar αV β3 integrin binding values, were compared in terms of their cellular effects in human U373 glioblastoma cells. Compounds 1 and 3 displayed overlapping inhibitory effects on the FAK/Akt integrin activated transduction pathway and on integrin-mediated cell infiltration processes, and qualify therefore, despite the different RGD and isoDGR sequences, as integrin antagonists. Both compounds induced apoptosis in glioma cells after 72 hour treatment.

  13. Installing multifunctionality on titanium with RGD-decorated polyurethane-polyurea roxithromycin loaded nanoparticles: toward new osseointegrative therapies.

    Science.gov (United States)

    Rocas, Pau; Hoyos-Nogués, Mireia; Rocas, Josep; Manero, José M; Gil, Javier; Albericio, Fernando; Mas-Moruno, Carlos

    2015-09-16

    A novel class of polyurethane-polyurea nanoparticles (PUUa NPs) to install multifunctionality on biomaterials is presented. Biofunctionalization of titanium with roxithromycin loaded RGD-decorated PUUa NPs results in an outstanding improvement of osteoblast adhesion and strong suppression of bacterial attachment. This strategy represents a powerful approach to enhance the osseointegration of implant materials. PMID:26274361

  14. PET imaging of alphavbeta integrin expression in tumours with Ga-labelled mono-, di- and tetrameric RGD peptides

    NARCIS (Netherlands)

    Dijkgraaf, I.; Yim, C.B.; Franssen, G.M.; Schuit, R.C.; Luurtsema, G.; Liu, S.; Oyen, W.J.G.; Boerman, O.C.

    2011-01-01

    PURPOSE: Due to the restricted expression of alpha(v)beta(3) in tumours, alpha(v)beta(3) is considered a suitable receptor for tumour targeting. In this study the alpha(v)beta(3)-binding characteristics of (68)Ga-labelled monomeric, dimeric and tetrameric RGD peptides were determined and compared wi

  15. Effects of linker variation on the in vitro and in vivo characteristics of an 111In-labeled RGD peptide.

    NARCIS (Netherlands)

    Dijkgraaf, I.; Liu, S.; Kruijtzer, J.A.; Soede, A.C.; Oyen, W.J.G.; Liskamp, R.M.; Corstens, F.H.M.; Boerman, O.C.

    2007-01-01

    INTRODUCTION: Due to the selective expression of the alpha(v)beta3 integrin in tumors, radiolabeled arginine-glycine-aspartic acid (RGD) peptides are attractive candidates for tumor targeting. Minor modifications of these peptides could have a major impact on in vivo characteristics. In this study,

  16. Towards a biocompatible artificial lung: Covalent functionalization of poly(4-methylpent-1-ene (TPX with cRGD pentapeptide

    Directory of Open Access Journals (Sweden)

    Lena Möller

    2013-02-01

    Full Text Available Covalent multistep coating of poly(methylpentene, the membrane material in lung ventilators, by using a copper-free “click” approach with a modified cyclic RGD peptide, leads to a highly biocompatible poly(methylpentene surface. The resulting modified membrane preserves the required excellent gas-flow properties while being densely seeded with lung endothelial cells.

  17. cRGD-directed, NIR-responsive and robust AuNR/PEG-PCL hybrid nanoparticles for targeted chemotherapy of glioblastoma in vivo.

    Science.gov (United States)

    Zhong, Yinan; Wang, Chao; Cheng, Ru; Cheng, Liang; Meng, Fenghua; Liu, Zhuang; Zhong, Zhiyuan

    2014-12-10

    cRGD-directed, NIR-responsive and robust AuNR/PEG-PCL hybrid nanoparticles (cRGD-HNs) were designed and developed for targeted chemotherapy of human glioma xenografts in mice. As expected, cRGD-HNs had excellent colloidal stability. The in vitro release studies showed that drug release from DOX-loaded cRGD-HNs (cRGD-HN-DOX) was minimal under physiological conditions but markedly accelerated upon NIR irradiation at a low power density of 0.2 W/cm2, due to photothermally induced phase transition of PCL regime. MTT assays showed that the antitumor activity of cRGD-HN-DOX in αvβ3 integrin over-expressed human glioblastoma U87MG cells was greatly boosted by mild NIR irradiation, which was significantly more potent than non-targeting HN-DOX counterpart under otherwise the same conditions and was comparable or superior to free DOX, supporting receptor-mediated endocytosis mechanism. The in vivo pharmacokinetics studies showed that cRGD-HN-DOX had much longer circulation time than free DOX. The in vivo imaging and biodistribution studies revealed that cRGD-HN-DOX could actively target human U87MG glioma xenograft in nude mice. The therapeutic studies in human U87MG glioma xenografts exhibited that cRGD-HN-DOX in combination with NIR irradiation completely inhibited tumor growth and possessed much lower side effects than free DOX. The Kaplan-Meier survival curves showed that all mice treated with cRGD-HN-DOX plus NIR irradiation survived over an experimental period of 48 days while control groups treated with PBS, cRGD-HN-DOX, cRGD-HNs with NIR irradiation, free DOX, or HN-DOX with NIR irradiation (non-targeting control) had short life spans of 15-40 days. Ligand-directed AuNR/PEG-PCL hybrid nanoparticles with evident tumor-targetability as well as superior spatiotemporal and rate control over drug release have emerged as an appealing platform for cancer chemotherapy in vivo.

  18. Adhesive and migratory effects of phosphophoryn are modulated by flanking peptides of the integrin binding motif.

    Directory of Open Access Journals (Sweden)

    Shigeki Suzuki

    Full Text Available Phosphophoryn (PP is generated from the proteolytic cleavage of dentin sialophosphoprotein (DSPP. Gene duplications in the ancestor dentin matrix protein-1 (DMP-1 genomic sequence created the DSPP gene in toothed animals. PP and DMP-1 are phosphorylated extracellular matrix proteins that belong to the family of small integrin-binding ligand N-linked glycoproteins (SIBLINGs. Many SIBLING members have been shown to evoke various cell responses through the integrin-binding Arg-Gly-Asp (RGD domain; however, the RGD-dependent function of PP is not yet fully understood. We demonstrated that recombinant PP did not exhibit any obvious cell adhesion ability, whereas the simultaneously purified recombinant DMP-1 did. A cell adhesion inhibitory analysis was performed by pre-incubating human osteosarcoma MG63 cells with various PP peptides before seeding onto vitronectin. The results obtained revealed that the incorporation of more than one amino acid on both sides of the PP-RGD domain was unable to inhibit the adhesion of MG63 cells onto vitronectin. Furthermore, the inhibitory activity of a peptide containing the PP-RGD domain with an open carboxyl-terminal side (H-463SDESDTNSESANESGSRGDA482-OH was more potent than that of a peptide containing the RGD domain with an open amino-terminal side (H-478SRGDASYTSDESSDDDNDSDSH499-OH. This phenomenon was supported by the potent cell adhesion and migration abilities of the recombinant truncated PP, which terminated with Ala482. Furthermore, various point mutations in Ala482 and/or Ser483 converted recombinant PP into cell-adhesive proteins. Therefore, we concluded that the Ala482-Ser483 flanking sequence, which was detected in primates and mice, was the key peptide bond that allowed the PP-RGD domain to be sequestered. The differential abilities of PP and DMP-1 to act on integrin imply that DSPP was duplicated from DMP-1 to serve as a crucial extracellular protein for tooth development rather than as an integrin

  19. Construction of expressing vectors including melanoma differentiation-associated gene-7 (mda-7 fused with the RGD sequences for better tumor targeting

    Directory of Open Access Journals (Sweden)

    Mahboobeh Khodadad

    2015-08-01

    Conclusion: Theoretically RGD tagged mda-7 would be able to induce apoptosis with more specificity and stronger than the standard one, therefore, these new constructs may have the potential for further researches.

  20. Diversity of RGD radiotracers in monitoring antiangiogenesis of flavopiridol and paclitaxel in ovarian cancer xenograft-bearing mice

    International Nuclear Information System (INIS)

    Introduction: Although encouraging results had been shown in antiangiogenesis therapy monitoring, the underlying mechanism of RGD radiotracer accumulation needs to be further illustrated. This study was aimed to investigate the diversity of RGD radiotracers in monitoring antiangiogenic agent's effects and the underlying mechanism in ovarian cancer-bearing mice with a new agent flavopiridol compared with paclitaxel. Methods: Ovarian cancer SKOV-3 xenograft-bearing mice were established and divided into three groups, flavopiridol, paclitaxel and control. Flavopiridol (5 mg/kg body weight) and paclitaxel (20 mg/kg body weight) were administered every 3 days for 16 days. Tumor growth and proliferation were monitored by caliper measurements and immunofluorescence staining. Antiangiogenic effects were determined by tumor microvessel density (MVD) in vivo and by endothelial cell tube formation assay in vitro, respectively. 99mTc-3P-RGD2 was prepared, and its biodistribution studies were carried out. The effect of antiangiogenesis therapy on integrin αvβ3 expression was studied by immunohistochemical staining and flow cytometry. Results: Both paclitaxel and flavopiridol therapy could apparently inhibit tumor growth and proliferation, and antiangiogenic effects of therapy were validated in vivo and in vitro. However, compared with the control group, ID%/g tumor uptake of 99mTc-3P-RGD2 showed a significant decrease at 2 hours (by 39.96% ± 8.23%, P = 0.044) and at 4 hours (by 35.76% ± 11.42%, P = 0.024) post injection in the paclitaxel-treated group, but a slight increase of tumor uptake in the flavopiridol-treated group at 2 hours (by 4.42% ± 0.24%, p = 0.898) and at 4 hours (by 12.2% ± 1.84%, P = 0.702). The further studies indicated flavopiridol therapy has a dual-effect, reducing integrin αvβ3 expression on endothelial cells due to the reduction of tumor MVD and up-regulating the integrin αvβ3 expression on tumor cells. Conclusions: There is diversity in

  1. Standard Thermodynamic Functions of Tripeptides N-Formyl-l-methionyl-l-leucyl-l-phenylalaninol and N-Formyl-l-methionyl-l-leucyl-l-phenylalanine Methyl Ester.

    Science.gov (United States)

    Markin, Alexey V; Markhasin, Evgeny; Sologubov, Semen S; Smirnova, Natalia N; Griffin, Robert G

    2014-04-10

    The heat capacities of tripeptides N-formyl-l-methionyl-l-leucyl-l-phenylalaninol (N-f-MLF-OH) and N-formyl-l-methionyl-l-leucyl-l-phenylalanine methyl ester (N-f-MLF-OMe) were measured by precision adiabatic vacuum calorimetry over the temperature range from T = (6 to 350) K. The tripeptides were stable over this temperature range, and no phase change, transformation, association, or thermal decomposition was observed. The standard thermodynamic functions: molar heat capacity Cp,m, enthalpy H(T) - H(0), entropy S(T), and Gibbs energy G(T) - H(0) of peptides were calculated over the range from T = (0 to 350) K. The low-temperature (T ≤ 50 K) heat capacities dependencies were analyzed using the Debye's and the multifractal theories. The standard entropies of formation of peptides at T = 298.15 K were calculated.

  2. Stimulation of collagen synthesis in fibroblast cultures by the tripeptide-copper complex glycyl-L-histidyl-L-lysine-Cu2+.

    Science.gov (United States)

    Maquart, F X; Pickart, L; Laurent, M; Gillery, P; Monboisse, J C; Borel, J P

    1988-10-10

    Glycyl-L-histidyl-L-lysine (GHK) is a tripeptide with affinity for copper(II) ions and was isolated from human plasma. This peptide appears to play a physiological role in wound healing. We report the stimulating effect of GHK-Cu on collagen synthesis by fibroblasts. The stimulation began between 10(-12) and 10(-11) M, maximized at 10(-9) M, and was independent of any change in cell number. The presence of a GHK triplet in the alpha 2(I) chain of type I collagen suggests that the tripeptide might be liberated by proteases at the site of a wound and exert in situ healing effects. PMID:3169264

  3. Standard Thermodynamic Functions of Tripeptides N-Formyl-l-methionyl-l-leucyl-l-phenylalaninol and N-Formyl-l-methionyl-l-leucyl-l-phenylalanine Methyl Ester.

    Science.gov (United States)

    Markin, Alexey V; Markhasin, Evgeny; Sologubov, Semen S; Smirnova, Natalia N; Griffin, Robert G

    2014-04-10

    The heat capacities of tripeptides N-formyl-l-methionyl-l-leucyl-l-phenylalaninol (N-f-MLF-OH) and N-formyl-l-methionyl-l-leucyl-l-phenylalanine methyl ester (N-f-MLF-OMe) were measured by precision adiabatic vacuum calorimetry over the temperature range from T = (6 to 350) K. The tripeptides were stable over this temperature range, and no phase change, transformation, association, or thermal decomposition was observed. The standard thermodynamic functions: molar heat capacity C p,m, enthalpy H(T) - H(0), entropy S(T), and Gibbs energy G(T) - H(0) of peptides were calculated over the range from T = (0 to 350) K. The low-temperature (T ≤ 50 K) heat capacities dependencies were analyzed using the Debye's and the multifractal theories. The standard entropies of formation of peptides at T = 298.15 K were calculated. PMID:24803685

  4. Development and biophysical characterization of HK polymer for siRNA delivery to tumor in a mouse model

    Science.gov (United States)

    Chou, Szu-Ting

    Delivery has been the major obstacle for nucleic acid therapeutics, including the RNA interference (RNAi) approach. Mixson's lab has been focused on the development of a non-viral peptide-based delivery system, HK (histidine-lysine) polymers, which have shown promise as carriers of plasmids and small interference RNA (siRNA) in several cell lines and in tumor-bearing models. In a previous study, a four-branched peptide, denoted H3K(+H)4b, with the predominant repeating -HHHK- sequence in the branch, has been shown to be the most effective and least toxic carrier in vitro and in vivo.. Building on these results, I utilized different approaches including several structure and stability molecular characterization methods to study polyplex and to develop more effective carriers for improved therapy with siRNAs targeting malignancies. To understand the role of histidine in the stability of the H3K(+H)4b/siRNA polyplex, the physicochemical properties were investigated. With the use of isothermal titration calorimetry and heteronuclear single quantum coherence NMR, histidines were shown to form hydrogen bonds with siRNA, which enhanced the stability and biological activity of the polyplexes. In addition, to enhance resistance to nucleases and to target the tumors selectively, H3K(+H)4b was chemically modified with different patterns of polyethylene glycol (PEG) and cyclic RGD (Arg-Gly-Asp, cRGD) peptide conjugates. The luciferase marker gene expressed stably by tumor xenografts in mouse models was targeted in order to evaluate the efficacy of HK carriers of siRNA that differed in location and number of cRGD-PEG attachments. The most effective carrier was (RGD-PEG)4H3K(+H) (RP-HK), which has a cRGD-PEG on each of its four terminal branches. Consistent with its prolonged stability, as observed by pharmacokinetic studies, the RP-HK polyplex down-regulated luciferase activity in tumor xenografts by nearly 70% compared with the untreated group. Subsequently, the RP-HK polyplex

  5. Theranostic cRGD-BioShuttle Constructs Containing Temozolomide- and Cy7 For NIR-Imaging and Therapy

    OpenAIRE

    Wiessler, Manfred; Hennrich, Ute; Pipkorn, Rüdiger; Waldeck, Waldemar; Cao, Liji; Peter, Jörg; Ehemann, Volker; Semmler, Wolfhard; Lammers, Twan; Braun, Klaus

    2011-01-01

    Innovative and personalized therapeutic approaches result from the identification and control of individual aberrantly expressed genes at the transcriptional and post-transcriptional level. Therefore, it is of high interest to establish diagnostic, therapeutic and theranostic strategies at these levels. In the present study, we used the Diels-Alder Reaction with inverse electron demand (DARinv) click chemistry to prepare a series of cyclic RGD-BioShuttle constructs. These constructs carry the...

  6. Biological evaluation of an ornithine-modified 99mTc-labeled RGD peptide as an angiogenesis imaging agent

    International Nuclear Information System (INIS)

    Introduction: Radiolabeled RGD peptides that specifically target integrin ανβ3 have great potential in early tumor detection through noninvasive monitoring of tumor angiogenesis. Based on previous findings of our group on radiopeptides containing positively charged aminoacids, we developed a new cyclic cRGDfK derivative, c(RGDfK)-(Orn)3-CGG. This new peptide availing the polar linker (Orn)3 and the 99mTc-chelating moiety CGG (Cys-Gly-Gly) is appropriately designed for 99mTc-labeling, as well as consequent conjugation onto nanoparticles. Methods: A tumor imaging agent, c(RGDfK)-(Orn)3-[CGG-99mTc], is evaluated with regard to its radiochemical, radiobiological and imaging characteristics. Results: The complex c(RGDfK)-(Orn)3-[CGG-99mTc] was obtained in high radiochemical yield (> 98%) and was stable in vitro and ex vivo. It presented identical to the respective, fully analytically characterized 185/187Re complex retention time in RP-HPLC. In contrary to other RGD derivatives, we showed that the new radiopeptide exhibits kidney uptake and urine excretion due to the ornithine linker. High tumor uptake (3.87 ± 0.48% ID/g at 60 min p.i.) was observed and was maintained relatively high even at 24 h p.i. (1.83 ± 0.05 % ID/g), thus providing well-defined scintigraphic imaging. Accumulation in other organs was negligible. Blocking experiments indicated target specificity for integrin receptors in U87MG glioblastoma cells. Conclusion: Due to its relatively high tumor uptake, renal elimination and negligible abdominal localization, the new 99mTc-RGD peptide is considered promising in the field of imaging ανβ3-positive tumors. However, the preparation of multifunctional SPECT/MRI contrast agents (RGD-conjugated nanoparticles) for dual modality imaging of integrin expressing tumors should be further investigated

  7. Dynamic PET and Optical Imaging and Compartment Modeling using a Dual-labeled Cyclic RGD Peptide Probe

    Directory of Open Access Journals (Sweden)

    Lei Zhu, Ning Guo, Quanzheng Li, Ying Ma, Orit Jacboson, Seulki Lee, Hak Soo Choi, James R. Mansfield, Gang Niu, Xiaoyuan Chen

    2012-01-01

    Full Text Available Purpose: The aim of this study is to determine if dynamic optical imaging could provide comparable kinetic parameters to that of dynamic PET imaging by a near-infrared dye/64Cu dual-labeled cyclic RGD peptide.Methods: The integrin αvβ3 binding RGD peptide was conjugated with a macrocyclic chelator 1,4,7,10-tetraazacyclododecane-1,4,7,10-tetraacetic acid (DOTA for copper labeling and PET imaging and a near-infrared dye ZW-1 for optical imaging. The in vitro biological activity of RGD-C(DOTA-ZW-1 was characterized by cell staining and receptor binding assay. Sixty-min dynamic PET and optical imaging were acquired on a MDA-MB-435 tumor model. Singular value decomposition (SVD method was applied to compute the dynamic optical signal from the two-dimensional optical projection images. Compartment models were used to quantitatively analyze and compare the dynamic optical and PET data.Results: The dual-labeled probe 64Cu-RGD-C(DOTA-ZW-1 showed integrin specific binding in vitro and in vivo. The binding potential (Bp derived from dynamic optical imaging (1.762 ± 0.020 is comparable to that from dynamic PET (1.752 ± 0.026.Conclusion: The signal un-mixing process using SVD improved the accuracy of kinetic modeling of 2D dynamic optical data. Our results demonstrate that 2D dynamic optical imaging with SVD analysis could achieve comparable quantitative results as dynamic PET imaging in preclinical xenograft models.

  8. Optimization of the production process of hybrid and multivalent formulation Bombesin/RGD for the opportune detection of breast cancer

    International Nuclear Information System (INIS)

    The radiopharmaceuticals of third generation are used in nuclear medicine to obtain images of specific molecular targets, and they are unique in their capacity to detect in vivo specific biochemical sites as receptors that are over-expressed in diverse illness. In cancer cells several types of receptors are over-expressed, as the integrin s α(v)β(3) and α(v)β(5) that specifically recognize the sequence RGD (Arginine-Glycin-Ac. Aspartic) and gastrin-releasing peptide that recognizes specifically to the peptide Lys3-Bombesin. The integrin s α(v)β(3) and α(v)β(5) are involved in the tumor angio genesis processes and the gastrin-releasing peptide is over-expressed in breast and prostate cancer. The molecular recognition of the specific receptors is the basis to be utilized as targets of the radiopharmaceuticals 99mTc-HYNIC-Bombesin and 99mTc-HYNIC-RGD. In this work was developed a lyophilized pharmaceutical formulation effective, stable and safe for the simultaneous obtaining of the radiopharmaceuticals 99mTc-HYNIC-Bombesin (99mTc-EDDA/HYNIC-Lys3-Bombesin) and 99mTc-HYNIC-RGD (99mTc EDDA/HYNIC-E-[c(RGDfK)]2). Later on the production process of the product HYNIC-Bombesin/RGD-Sn was optimized using a factorial design and the formulation was transferred to the production plant of radiopharmaceuticals of the Instituto Nacional de Investigaciones Nucleares (ININ). The optimized formulation is described in the following chart: HYNIC-[Lys3]-Bombesin - 12.5 μg; HYNIC-E-c[RGDfK]2 - 12.5 μg; Stannous chloride (SnCl2) - 20 μg; Ethylenediamine diacetic acid (EDDA) - 10 mg; N-tris(hydroxymethyl)methyl glycin (Tricine) - 20 mg; Mannitol - 50 mg. The production process was validated and were carried out the stability studies under refrigeration conditions. (Author)

  9. RGD-functionalisation of PLLA nanofibers by surface coupling using plasma treatment: influence on stem cell differentiation.

    Science.gov (United States)

    Paletta, Jürgen Rudolf Josef; Bockelmann, Sarah; Walz, Andreas; Theisen, Christina; Wendorff, Joachim Heinz; Greiner, Andreas; Fuchs-Winkelmann, Susanne; Schofer, Markus Dietmar

    2010-04-01

    The aim of this study was to functionalize the surface of synthetic poly-(l-lactic) (PLLA) nanofibers with RGD peptide, in order to promote growth and osteogenic differentiation of human mesenchymal stem cells (hMSC) in vitro. The cRGD was coupled onto PLLA nanofibers using oxygen plasma combined with EDC/sulfo-NHS activation. Matrices were seeded with hMSC and cultivated over a period of 22 days under growth conditions and analyzed during the course of cultivation. The plasma activation of PLLA nanofibers resulted in a reduction of hydrophobicity as well as a formation of carboxyl groups on the surface of the fibers. Furthermore, maximum load, but not young's modulus was influenced by the treatment with oxygen plasma. When hMSC were cultured onto the cRGD functionalized scaffolds, cells showed no increased proliferation or cell density but an induction of genes associated with the osteoblast lineage. In brief, this study indicates that functional peptides of the extracellular matrix can be coupled onto PLLA nanofibers using plasma treatment in combination with EDC/sulfo-NHS treatment. These groups are accessible for the growing cell and mediate probably some osteoinductive properties of collagen nanofibers. PMID:19943087

  10. Radiolabeled biomolecules for early cancer detection and therapy via angiogenesis targeting

    Energy Technology Data Exchange (ETDEWEB)

    Bouziotis, P. [Institute of Radioisotopes-Radiodiagnostic Products, N.C.S.R. ' Demokritos' , 153 10 Athens, Hellas (Greece)]. E-mail: pennybil@yahoo.gr; Psimadas, D. [Institute of Radioisotopes-Radiodiagnostic Products, N.C.S.R. ' Demokritos' , 153 10 Athens, Hellas (Greece): Biomedica Life Sciences SA, Athens, Hellas (Greece); Fani, M. [Institute of Radioisotopes-Radiodiagnostic Products, N.C.S.R. ' Demokritos' , 153 10 Athens, Hellas (Greece): Biomedica Life Sciences SA, Athens, Hellas (Greece); Gourni, E. [Institute of Radioisotopes-Radiodiagnostic Products, N.C.S.R. ' Demokritos' , 153 10 Athens, Hellas (Greece); Loudos, G. [Biomedical Simulations and Imaging Laboratory, N.T.U.A., Athens, Hellas (Greece); Xanthopoulos, S. [Institute of Radioisotopes-Radiodiagnostic Products, N.C.S.R. ' Demokritos' , 153 10 Athens, Hellas (Greece); Archimandritis, S.C. [Institute of Radioisotopes-Radiodiagnostic Products, N.C.S.R. ' Demokritos' , 153 10 Athens, Hellas (Greece); Varvarigou, A.D. [Institute of Radioisotopes-Radiodiagnostic Products, N.C.S.R. ' Demokritos' , 153 10 Athens, Hellas (Greece)

    2006-12-20

    Tumors cannot grow or metastasize without the formation of new blood vessels, i.e. without angiogenesis. A variety of anti-angiogenic agents leading to angiogenesis inhibition are in the clinical trial phase, among which are: (i) molecules which inhibit the action of Vascular Endothelial Growth Factors, VEGF and (ii) molecules which obstruct migration, differentiation and proliferation of endothelial cells, via their binding to receptors of the {alpha}{sub {nu}}{beta}{sub 3} integrins. Certain derivatives of the abovementioned categories, labeled with radionuclides, which emit {gamma}-radiation or {beta}-particles or positrons, have been proposed and are being evaluated as possible radiopharmaceuticals, for the detection and/or treatment of primary or metastatic cancer at an early stage. For the study of angiogenesis the following have been described: (a) antibodies targeting VEGF, labeled with radionuclides emitting {beta}- and/or {gamma}-radiation, which can be applied for the diagnosis and, possibly, for the treatment of cancer (b) peptide derivatives which contain the amino-acid sequence RGD (Arg-Gly-Asp) and compete for the {alpha}{sub {nu}}{beta}{sub 3} integrins, with the proteins of the stroma. It has been found that these radiolabeled biomolecules localize in tumors and can be used for the visualization and, possibly, for tumor eradication of primary and metastatic cancer. In our laboratory radiolabeling of biomolecules by beta and/or gamma emitters is a principal research goal. In the present work we are presenting our results on the labeling of monoclonal antibodies and peptides with {beta}- and {gamma}-emitting isotopes, as well as on their in vivo evaluation in experimental animal models, by use of specially dedicated imaging devices.

  11. The Anti-Cancer Potency and Mechanism of a Novel Tumor-Activated Fused Toxin, DLM

    Directory of Open Access Journals (Sweden)

    Dejun Sun

    2015-02-01

    Full Text Available Melittin, which acts as a membrane-disrupting lytic peptide, is not only cytotoxic to tumors, but also vital to normal cells. Melittin had low toxicity when coupled with target peptides. Despite significant research development with the fused toxin, a new fused toxin is needed which has a cleavable linker such that the fused toxin can release melittin after protease cleavage on the tumor cell surface. We describe a novel fused toxin, composed of disintegrin, uPA (urokinase-type plasminogen activator-cleavable linker, and melittin. Disintegrin is a single strand peptide (73 aa isolated from Gloydius Ussuriensis venom. The RGD (Arg-Gly-Asp site of disintegrin dominates its interaction with integrins on the surface of the tumor cells. uPA is over-expressed and plays an important role in tumor cell invasiveness and metastatic progression. The DLM (disintegrin-linker-melittin linker is uPA-cleavable, enabling DLM to release melittin. We compared binding activity of our synthesized disintegrin with native disintegrin and report that DLM had less binding activity than the native form. uPA-cleavage was evaluated in vitro and the uPA-cleavable linker released melittin. Treating tumors expressing uPA with DLM enhanced tumor cell killing as well as reduced toxicity to erythrocytes and other non-cancerous normal cells. The mechanism behind DLM tumor cell killing was tested using a DNA ladder assay, fluorescent microscopy, flow cytometry, and transmission electron microscopy. Data revealed tumor cell necrosis as the mechanism of cell death, and the fused DLM toxin with an uPA-cleavable linker enhanced tumor selectivity and killing ability.

  12. Practical applications of snake venom toxins in haemostasis.

    Science.gov (United States)

    Marsh, Neville; Williams, Vaughan

    2005-06-15

    Snake venom toxins affecting haemostasis have facilitated extensively the routine assays of haemostatic parameters in the coagulation laboratory. Snake venom thrombin-like enzymes (SVTLE) are used for fibrinogen/fibrinogen breakdown product assay and for the detection of fibrinogen dysfunction. SVTLE are not inhibited by heparin and can thus can be used for assaying antithrombin III and other haemostatic variables in heparin-containing samples. Snake venoms are a rich source of prothrombin activators and these are utilised in prothrombin assays, for studying dysprothrombinaemias and for preparing meizothrombin and non-enzymic forms of prothrombin. Russell's viper (Daboia russelli) venom (RVV) contains toxins which have been used to assay blood clotting factors V, VII, X, platelet factor 3 and, importantly, lupus anticoagulants (LA). Other prothrombin activators (from the taipan, Australian brown snake and saw-scaled viper) have now been used to assay LA. Protein C and activated protein C resistance can be measured by means of RVV and Protac, a fast acting inhibitor from Southern copperhead snake venom and von Willebrand factor can be studied with botrocetin from Bothrops jararaca venom. The disintegrins, a large family of Arg-Gly-Asp (RGD)-containing snake venom proteins, show potential for studying platelet glycoprotein receptors, notably, GPIIb/IIIa and Ib. Snake venom toxins affecting haemostasis are also used in the therapeutic setting: Ancrod (from the Malayan pit viper, Calloselasma rhodostoma), in particular, has been used as an anticoagulant to achieve 'therapeutic defibrination'. Other snake venom proteins show promise in the treatment of a range of haemostatic disorders. PMID:15922782

  13. Force via integrins but not E-cadherin decreases Oct3/4 expression in embryonic stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Uda, Yuhei [Department of Mechanical Science and Engineering, University of Illinois at Urbana-Champaign, Urbana, IL 61801 (United States); Department of Biomedical Engineering, Graduate School of Biomedical Engineering, Tohoku University, 6-6-01, Aramaki-aoba, Aoba-ward, Sendai City (Japan); Poh, Yeh-Chuin; Chowdhury, Farhan; Wu, Douglas C. [Department of Mechanical Science and Engineering, University of Illinois at Urbana-Champaign, Urbana, IL 61801 (United States); Tanaka, Tetsuya S. [Department of Animal Sciences, University of Illinois at Urbana-Champaign, Urbana, IL 61801 (United States); Sato, Masaaki [Department of Biomedical Engineering, Graduate School of Biomedical Engineering, Tohoku University, 6-6-01, Aramaki-aoba, Aoba-ward, Sendai City (Japan); Wang, Ning, E-mail: nwangrw@illinois.edu [Department of Mechanical Science and Engineering, University of Illinois at Urbana-Champaign, Urbana, IL 61801 (United States)

    2011-11-18

    Highlights: Black-Right-Pointing-Pointer Force via integrins or cadherins induces similar cell stiffening responses. Black-Right-Pointing-Pointer Force via integrins but not cadherins induces cell spreading. Black-Right-Pointing-Pointer Force via integrins but not cadherins induces differentiation of embryonic stem cells. -- Abstract: Increasing evidence suggests that mechanical factors play a critical role in fate decisions of stem cells. Recently we have demonstrated that a local force applied via Arg-Gly-Asp (RGD) peptides coated magnetic beads to mouse embryonic stem (ES) cells increases cell spreading and cell stiffness and decreases Oct3/4 (Pou5f1) gene expression. However, it is not clear whether the effects of the applied stress on these functions of ES cells can be extended to natural extracellular matrix proteins or cell-cell adhesion molecules. Here we show that a local cyclic shear force applied via fibronectin or laminin to integrin receptors increased cell spreading and stiffness, downregulated Oct3/4 gene expression, and decreased cell proliferation rate. In contrast, the same cyclic force applied via cell-cell adhesion molecule E-cadherin (Cdh1) had no effects on cell spreading, Oct3/4 gene expression, and the self-renewal of mouse ES cells, but induced significant cell stiffening. Our findings demonstrate that biological responses of ES cells to force applied via integrins are different from those to force via E-cadherin, suggesting that mechanical forces might play different roles in different force transduction pathways to shape early embryogenesis.

  14. 3D culture of adult mouse neural stem cells within functionalized self-assembling peptide scaffolds

    Directory of Open Access Journals (Sweden)

    Cunha C

    2011-05-01

    Full Text Available Carla Cunha1,2, Silvia Panseri3,4, Omar Villa1,2, Diego Silva1,2, Fabrizio Gelain1,21Department of Biotechnology and Biosciences, University of Milano-Bicocca; 2Center for Nanomedicine and Tissue Engineering, CNTE – A.O. Ospedale Niguarda Ca' Granda, Milan; 3Laboratory of Biomechanics and Technology Innovation, Rizzoli Orthopaedic Institute, Bologna; 4Laboratory of Nano-Biomagnetism, Institute of Science and Technology for Ceramics, National Research Council, Faenza, ItalyAbstract: Three-dimensional (3D in vitro models of cell culture aim to fill the gap between the standard two-dimensional cell studies and the in vivo environment. Especially for neural tissue regeneration approaches where there is little regenerative capacity, these models are important for mimicking the extracellular matrix in providing support, allowing the natural flow of oxygen, nutrients, and growth factors, and possibly favoring neural cell regrowth. We have previously demonstrated that a new self-assembling nanostructured biomaterial, based on matrigel, was able to support adult neural stem cell (NSC culture. In this study, we developed a new 3D cell culture system that takes advantage of the nano- and microfiber assembling process, under physiologic conditions, of these biomaterials. The assembled scaffold forms an intricate and biologically active matrix that displays specifically designed functional motifs: RGD (Arg-Gly-Asp, BMHP1 (bone marrow homing peptide 1, and BMHP2, for the culture of adult NSCs. These scaffolds were prepared at different concentrations, and microscopic examination of the cell-embedded scaffolds showed that NSCs are viable and they proliferate and differentiate within the nanostructured environment of the scaffold. Such a model has the potential to be tailored to develop ad hoc designed peptides for specific cell lines.Keywords: biomaterials, tissue engineering, 3D in vitro model

  15. Cytoskeletal mechanics in pressure-overload cardiac hypertrophy

    Science.gov (United States)

    Tagawa, H.; Wang, N.; Narishige, T.; Ingber, D. E.; Zile, M. R.; Cooper, G. 4th

    1997-01-01

    We have shown that the cellular contractile dysfunction characteristic of pressure-overload cardiac hypertrophy results not from an abnormality intrinsic to the myofilament portion of the cardiocyte cytoskeleton but rather from an increased density of the microtubule component of the extramyofilament portion of the cardiocyte cytoskeleton. To determine how, in physical terms, this increased microtubule density mechanically overloads the contractile apparatus at the cellular level, we measured cytoskeletal stiffness and apparent viscosity in isolated cardiocytes via magnetic twisting cytometry, a technique by which magnetically induced force is applied directly to the cytoskeleton through integrin-coupled ferromagnetic beads coated with Arg-Gly-Asp (RGD) peptide. Measurements were made in two groups of cardiocytes from cats with right ventricular (RV) hypertrophy induced by pulmonary artery banding: (1) those from the pressure-overloaded RV and (2) those from the normally loaded same-animal control left ventricle (LV). Cytoskeletal stiffness increased almost twofold, from 8.53 +/- 0.77 dyne/cm2 in the normally loaded LV cardiocytes to 16.46 +/- 1.32 dyne/cm2 in the hypertrophied RV cardiocytes. Cytoskeletal apparent viscosity increased almost fourfold, from 20.97 +/- 1.92 poise in the normally loaded LV cardiocytes to 87.85 +/- 6.95 poise in the hypertrophied RV cardiocytes. In addition to these baseline data showing differing stiffness and, especially, apparent viscosity in the two groups of cardiocytes, microtubule depolymerization by colchicine was found to return both the stiffness and the apparent viscosity of the pressure overload-hypertrophied RV cells fully to normal. Conversely, microtubule hyperpolymerization by taxol increased the stiffness and apparent viscosity values of normally loaded LV cardiocytes to the abnormal values given above for pressure-hypertrophied RV cardiocytes. Thus, increased microtubule density constitutes primarily a viscous load on

  16. 皖南尖吻蝮蛇毒中类去整合蛋白/富含半胱氨酸两结构域cDNA的克隆和表达%Cloning,Expression and Characteristics of Disintegrin-like/cysteine-rich Domains cDNA from Agkistrodon acutus Venom

    Institute of Scientific and Technical Information of China (English)

    刘庆都; 张家琳; 承新; 刘爱平; 刘兢

    2000-01-01

    为了探讨出血毒金属蛋白酶结构功能关系,通过RT-PCR方法,从皖南尖吻蝮蛇(Agkistrodon acutus)毒腺总RNA中扩增得到编码P-Ⅲ型出血毒金属蛋白酶的完整类去整合蛋白和富含半胱氨酸两个结构域cDNA(AA/DC).它全长964 bp的cDNA,开放阅读框架编码216个氨基酸残基,序列比较分析表明它同来自Bothrops jararaca的jararhagin-C、来自Crotalus atrox的catrocollastatin-C有很高的同源性.在类去整合蛋白结构域中,Ser-Glu-Cys-Asp(SECD)代替了去整合蛋白中相应部位的Arg-Gly-Asp(RGD)三肽序列.将编码区基因克隆入pGEX-2T栽体中,转化大肠杆菌TG-1,用IPTG诱导表达,表达产物具有抑制胶原诱导的血小板凝集活性,但不抑制ADP诱导的血小板凝集.该研究为进一步阐述蛇毒金属蛋白酶结构功能关系和药物开发奠定了基础.

  17. Preparation and characterization of 99mTc(CO)3-BPy-RGD complex as αvβ3 integrin receptor-targeted imaging agent

    International Nuclear Information System (INIS)

    The aim of this study is to develop a novel arginine-glycine-aspartic acid (RGD) peptide-containing ligand for 99mTc labeling as αvβ3 integrin receptor-targeted imaging agent. BPy-RGD conjugate was successfully synthesized by coupling of 5-carboxylate-2,2'-bipyridine and c(RGDyK) peptide through EDC/SNHS in aqueous solution and was characterized by MADLI-TOF-MS (m/z=802.72, C38H48N11O9). 99mTc(CO)3-BPy-RGD was prepared by exchange reaction between [99mTc(H2O)3(CO)3]+ and BPy-RGD. Final product was purified by HPLC and tested for octanol/water partition coefficient. Cell-binding assays of BPy-RGD and unmodified c(RGDyK) were tested in MDA-MB-435 cells (125I-echistatin as radioligand). Preliminary biodistribution of the 99mTc(I)-labeled radiotracer in orthotopic MDA-MB-435 breast tumor xenograft model was also evaluated. The BPy-RGD conjugate had good integrin-binding affinity (50% inhibitory concentration (IC50)=92.51+/-22.69nM), slightly lower than unmodified c(RGDyK) (IC50=59.07+/-11.03nM). The hydrophilic radiotracer also had receptor-mediated activity accumulation in MDA-MB-435 tumor (1.45+/-0.25 percentage of injected dose per gram (%ID/g) at 1.5h postinjection (p.i.)), which is known to be integrin positive. After blocking with c(RGDyK), the tumor uptake was reduced from 0.71+/-0.01%ID/g to 0.33+/-0.18%ID/g at 4h p.i. 99mTc(I) tricarbonyl complex of cyclic RGD peptide is a promising strategy for integrin targeting. Further modification of the bipyridine-conjugated RGD peptide by using more potent RGD peptides and fine tuning of the tether group between the RGD moiety and 99mTc(CO)3+ core to improve the tumor targeting efficacy and in vivo kinetic profiles is currently in progress

  18. Stimulation of sulfated glycosaminoglycan synthesis by the tripeptide-copper complex glycyl-L-histidyl-L-lysine-Cu2+.

    Science.gov (United States)

    Wegrowski, Y; Maquart, F X; Borel, J P

    1992-01-01

    Glycyl-L-histidyl-L-lysine-copper (II) complex (GHK-Cu) is a naturally occurring tripeptide with potential healing properties. We studied the effect of GHK-Cu on the synthesis of glycosaminoglycans (GAGs) by normal human fibroblasts in culture. Cells were incubated with 3H glucosamine and 35S sulfate and the radioactivity of isolated GAGs was determined. GHK-Cu induced a dose-dependent increase of the synthesis of total GAGs secreted into the culture medium and those associated with the cell layer. The effect of GHK-Cu was biphasic with a maximal stimulation at 10(-9) to 10(-8) M. At higher concentrations, the rate of synthesis returned progressively to that of control cultures. Electrophoretic analysis of the different GAG populations showed that GHK-Cu preferentially stimulated the synthesis of extracellular dermatan sulfate and cell layer associated heparan sulfate. No influence of GHK-Cu on the synthesis of hyaluronic acid was observed. GHK-Cu stimulation of GAG synthesis may be one of the phenomenons implicated in the wound healing properties of the peptide. PMID:1522753

  19. Free-energy calculations of residue mutations in a tripeptide using various methods to overcome inefficient sampling.

    Science.gov (United States)

    Graf, Michael M H; Maurer, Manuela; Oostenbrink, Chris

    2016-11-01

    Previous free-energy calculations have shown that the seemingly simple transformation of the tripeptide KXK to KGK in water holds some unobvious challenges concerning the convergence of the forward and backward thermodynamic integration processes (i.e., hysteresis). In the current study, the central residue X was either alanine, serine, glutamic acid, lysine, phenylalanine, or tyrosine. Interestingly, the transformation from alanine to glycine yielded the highest hysteresis in relation to the extent of the chemical change of the side chain. The reason for that could be attributed to poor sampling of φ2 /ψ2 dihedral angles along the transformation. Altering the nature of alanine's Cβ atom drastically improved the sampling and at the same time led to the identification of high energy barriers as cause for it. Consequently, simple strategies to overcome these barriers are to increase simulation time (computationally expensive) or to use enhanced sampling techniques such as Hamiltonian replica exchange molecular dynamics and one-step perturbation. © 2016 The Authors. Journal of Computational Chemistry Published by Wiley Periodicals, Inc.

  20. Novel Antioxidant Tripeptide "ACQ" Can Prevent UV-Induced Cell Death and Preserve the Number of Epidermal Stem Cells.

    Science.gov (United States)

    Choi, Hye-Ryung; Shin, Jung-Won; Na, Jung-Im; Nam, Kyung-Mi; Lee, Hyun-Sun; Park, Kyoung-Chan

    2015-01-01

    We found that tripeptide "ACQ: alanine-cysteine-glutamine" has significant DPPH scavenging activity compared to that of glutathione. Antioxidant effects of ACQ were tested in in vitro and in vivo models. When treated with H2O2, mock treated fibroblasts and keratinocytes showed strong staining by H2DCFA. But, ACQ showed good protective effects against hydrogen peroxide treatment. When mice were fed for 2 or 4 weeks, similar protective effects were observed. In the control group, epidermis was severely damaged by UV irradiation and apoptotic keratinocytes were observed. There were also numerous TUNEL positive cells. But in the ACQ group, epidermis became thicker and there was no sign of severe damage. Interestingly, the number of p63 cells was also higher in ACQ fed mice. To confirm the stem cell rescuing effects of ACQ, three-dimensional skin samples were constructed. Results showed that ACQ increased the expression of integrin α6 and the number of p63 positive cells. These findings showed that ACQ has good antioxidant activity and may increase stem cell activities by the regulation of integrin α6.

  1. A tripeptide isolated from Bothrops atrox venom has neuroprotective and neurotrophic effects on a cellular model of Parkinson's disease.

    Science.gov (United States)

    Martins, N M; Santos, N A G; Sartim, M A; Cintra, A C O; Sampaio, S V; Santos, A C

    2015-06-25

    Parkinson's disease (PD) is the second most common neurodegenerative disorder; however, there is no treatment able to prevent the loss of dopaminergic neurons or its consequences. Trophic factors such as NGF and BDNF has positive effects on different disorders of the brain, including neurodegeneration. Additionally, studies have suggested the use of venom peptides as a therapeutic strategy for neurological disorders. Therefore, in the present study, we investigated the neuroprotective activity of a peptide isolated from Bothrops atrox venom and its trophic ability by using a cellular model of dopaminergic neurotoxicity induced by 1-methyl-4-phenylpyridinium (MPP(+)) in PC12 cells. We showed that it decreased the activities of the apoptotic proteases caspase-9 (mitochondrial) and caspase-3 (executor) and increased cell viability and proliferation in this model. Additionally, it increased neuritogenesis in non-treated PC12 cells (neuronal model) as well as in PC12 cells treated with the dopaminergic neurotoxin. The amino acid sequence of the peptide was identified as Glutamic acid-Valine-Tryptophan (Glu-Val-Trp). These findings suggest that this tripeptide has the potential to protect against the dopaminergic neurons loss and that trophic stimulation of neuroplasticity might be involved in its mechanism of neuroprotection. PMID:25868679

  2. Dual targeting strategy of magnetic nanoparticle-loaded and RGD peptide-activated stimuli-sensitive polymeric micelles for delivery of paclitaxel

    Energy Technology Data Exchange (ETDEWEB)

    Lin, Meng Meng [Tsinghua University, Department of Chemical Engineering (China); Kang, Yoon Joong [Jungwon University, Department of Biomedical Science (Korea, Republic of); Sohn, Youngjoo [Kyung Hee University, Department of Anatomy, College of Korean Medicine (Korea, Republic of); Kim, Do Kyung, E-mail: eurokorean@gmail.com, E-mail: dokyung@konyang.ac.kr [Konyang University, Industry Cooperation Foundation (Korea, Republic of)

    2015-06-15

    A double targeting strategy of anti-neoplastic agent paclitaxel (PTX) was developed by incorporating magnetic nanoparticles and RGD peptide for enhanced cell cytotoxicity effect at lower dosage. A dual targeting mechanism including magnetic targeting and RGD ligand-specific targeting enhanced the overall cytotoxicity and reduced the effective dosage of PTX to achieve enhanced and sustained release of PTX in vitro. We addressed the issues of water-insolubility of oleic acid (OA)-stabilized SPIONs and low incorporation efficiency of hydrophobic PTX with SPION nanocarriers by using an amphiphilic polymer poly[(N-isopropylacrylamide-r-acrylamide)-b-l-lactic acid] (PNAL) as micelle-forming materials. A targeting moiety, GGGGRGD peptide, a RGD sequence-containing peptide with a short linker, is attached to the surface of PNAL-SPIONs via a homo-crosslinker. Confocal microscopy image analysis revealed that the cellular uptake was increased from (1.5 ± 0.5 % (PNAL) to 11.7 ± 0.8 % (RGD-PNAL-SPIONs) at 6 h incubation, once both RGD peptide and magnetic force attraction were incorporated into the carriers. Such multi-targeting nanocarriers showed promising potential in cancer-oriented diagnosis and therapy.

  3. Modular Small Diameter Vascular Grafts with Bioactive Functionalities.

    Directory of Open Access Journals (Sweden)

    Meik Neufurth

    Full Text Available We report the fabrication of a novel type of artificial small diameter blood vessels, termed biomimetic tissue-engineered blood vessels (bTEBV, with a modular composition. They are composed of a hydrogel scaffold consisting of two negatively charged natural polymers, alginate and a modified chitosan, N,O-carboxymethyl chitosan (N,O-CMC. Into this biologically inert scaffold two biofunctionally active biopolymers are embedded, inorganic polyphosphate (polyP and silica, as well as gelatin which exposes the cell recognition signal, Arg-Gly-Asp (RGD. These materials can be hardened by exposure to Ca(2+ through formation of Ca(2+ bridges between the polyanions, alginate, N,O-CMC, and polyP (alginate-Ca(2+-N,O-CMC-polyP. The bTEBV are formed by pressing the hydrogel through an extruder into a hardening solution, containing Ca(2+. In this universal scaffold of the bTEBV biomaterial, polycations such as poly(L-Lys, poly(D-Lys or a His/Gly-tagged RGD peptide (three RGD units were incorporated, which promote the adhesion of endothelial cells to the vessel surface. The mechanical properties of the biopolymer material (alginate-Ca(2+-N,O-CMC-polyP-silica revealed a hardness (elastic modulus of 475 kPa even after a short incubation period in CaCl2 solution. The material of the artificial vascular grafts (bTEBVs with an outer size 6 mm and 1.8 mm, and an inner diameter 4 mm and 0.8 mm, respectively turned out to be durable in 4-week pulsatile flow experiments at an alternating pressure between 25 and 100 mbar (18.7 and 75.0 mm Hg. The burst pressure of the larger (smaller vessels was 850 mbar (145 mbar. Incorporation of polycationic poly(L-Lys, poly(D-Lys, and especially the His/Gly-tagged RGD peptide, markedly increased the adhesion of human, umbilical vein/vascular endothelial cells, EA.HY926 cells, to the surface of the hydrogel. No significant effect of the polyP samples on the clotting of human plasma is measured. We propose that the metabolically

  4. Tripeptide-copper complex GHK-Cu (II) transiently improved healing outcome in a rat model of ACL reconstruction.

    Science.gov (United States)

    Fu, Sai-Chuen; Cheuk, Yau-Chuk; Chiu, Wai-Yin Vivien; Yung, Shu-Hang; Rolf, Christer G; Chan, Kai-Ming

    2015-07-01

    After anterior cruciate ligament reconstruction (ACLR), the biological healing of the graft is a rate-limiting step which can contribute to graft failure. The tripeptide-copper complex glycyl-L-histidyl-L-lysine-Cu(II) (GHK-Cu) is a well-known activator of tissue remodeling. We investigated whether GHK-Cu can improve graft healing following ACLR. Seventy-two rats underwent unilateral ACLR were randomized to saline, 0.3 or 3 mg/ml GHK-Cu groups (n = 24). Post-operational intra-articular injections were given from week 2, once a week, for 4 weeks. Gait analysis was performed pre-injury and at harvesting time. At 6 or 12 weeks post-operation, knee specimens were harvested for knee laxity test, graft pull-out test, and histology. At 6 weeks post-ACLR, GHK-Cu groups resulted in a smaller side-to-side difference in knee laxity as compared to the saline group (p = 0.009), but there was no significant difference at 12 weeks post-operation. The graft complex in the 0.3 mg/ml GHK-Cu group had higher stiffness than saline group at 6 weeks post-operation (p = 0.026), but there was no significant difference in ultimate load, gait parameters, and histological scores among treatment groups. All grafts failed mid-substance during pull-out test. Intra-articular supplementation with a bioactive small molecule GHK-Cu improved graft healing following ACLR in rat, but the beneficial effects could not last as treatment discontinued. PMID:25731775

  5. A pilot study imaging integrin αvβ3 with RGD PET/CT in suspected lung cancer patients

    Energy Technology Data Exchange (ETDEWEB)

    Gao, Song [Shandong Cancer Hospital and Institute, Department of Radiation Oncology, Jinan, Shandong (China); University of Jinan-Shandong Academy of Medical Sciences, School of Medicine and Life Sciences, Jinan, Shandong (China); Wu, Honghu [Wuyi County People' s Hospital of Hengshui City, Hengshui, Hebei Province (China); Li, Wenwu; Zhao, Shuqiang; Teng, Xuepeng; Lu, Hong [Shandong Cancer Hospital and Institute, Department of Radiology, Jinan, Shandong (China); Hu, Xudong; Wang, Suzhen; Yu, Jinming; Yuan, Shuanghu [Shandong Cancer Hospital and Institute, Department of Radiation Oncology, Jinan, Shandong (China)

    2015-12-15

    Angiogenesis is an essential step in tumour development and metastasis. Integrin αvβ3 plays a major role in angiogenesis, tumour growth and progression. A new tracer, {sup 18}F-AL-NOTA-PRGD2, denoted as {sup 18}F-alfatide, has been developed for positron emission tomography (PET) imaging of integrin αvβ3. This is a pilot study to test the safety and diagnostic value of {sup 18}F- arginine-glycine-aspartic acid (RGD) PET/computed tomography (CT) in suspected lung cancer patients. Twenty-six patients with suspected lung cancer on enhanced CT underwent {sup 18}F-alfatide RGD PET/CT examination before surgery and puncture biopsy. Standard uptake values (SUVs) and the tumour-to-blood ratios were measured, and diagnoses were pathologically confirmed. RGD PET/CT with {sup 18}F-alfatide was performed successfully in all patients and no clinically significant adverse events were observed. The {sup 18}F-alfatide RGD PET/CT analysis correctly recognized 17 patients with lung cancer, 4 patients (hamartoma) as true negative, and 5 patients (4 chronic inflammation and 1 inflammatory pseudotumour) as false positive. The sensitivity, specificity, accuracy, positive predictive value (PPV) and negative predictive value (NPV) of {sup 18}F-alfatide RGD PET/CT for the diagnosis of suspected lung cancer patients was 100, 44.44, 80.77, 77.27, and 100 %, respectively. The area under a receiver operating characteristic (ROC) curve was 0.75 (P = 0.038), and ROC analysis suggested an SUVmax cut-off value of 2.65 to differentiate between malignant lesions and benign lesions. The SUV for malignant lesions was 5.37 ± 2.17, significantly higher than that for hamartomas (1.60 ± 0.11; P < 0.001). The difference between the tumour-to-blood ratio for malignant lesions (4.13 ± 0.91) and tissue of interest-to-blood ratio for hamartomas (1.56 ± 0.24) was also statistically significant (P < 0.001). Neither the SUVmax nor the tumour-to-blood ratio was significantly different between malignant

  6. Reproducibility study of [{sup 18}F]FPP(RGD){sub 2} uptake in murine models of human tumor xenografts

    Energy Technology Data Exchange (ETDEWEB)

    Chang, Edwin; Liu, Shuangdong; Chin, Frederick; Cheng, Zhen [Stanford University, Molecular Imaging Program at Stanford, Department of Radiology, School of Medicine, Stanford, CA (United States); Gowrishankar, Gayatri; Yaghoubi, Shahriar [Stanford University, Molecular Imaging Program at Stanford, Department of Radiology, School of Medicine, Stanford, CA (United States); Stanford University, Molecular Imaging Program at Stanford, Department of Bioengineering, School of Medicine, Stanford, CA (United States); Wedgeworth, James Patrick [Stanford University, Molecular Imaging Program at Stanford, Department of Bioengineering, School of Medicine, Stanford, CA (United States); Berndorff, Dietmar; Gekeler, Volker [Bayer Schering Pharma AG, Global Drug Discovery, Berlin (Germany); Gambhir, Sanjiv S. [Stanford University, Molecular Imaging Program at Stanford, Department of Radiology, School of Medicine, Stanford, CA (United States); Stanford University, Molecular Imaging Program at Stanford, Department of Bioengineering, School of Medicine, Stanford, CA (United States); Canary Center at Stanford for Cancer Early Detection, Nuclear Medicine, Departments of Radiology and Bioengineering, Molecular Imaging Program at Stanford, Stanford, CA (United States)

    2011-04-15

    An {sup 18}F-labeled PEGylated arginine-glycine-aspartic acid (RGD) dimer [{sup 18}F]FPP(RGD){sub 2} has been used to image tumor {alpha}{sub v}{beta}{sub 3} integrin levels in preclinical and clinical studies. Serial positron emission tomography (PET) studies may be useful for monitoring antiangiogenic therapy response or for drug screening; however, the reproducibility of serial scans has not been determined for this PET probe. The purpose of this study was to determine the reproducibility of the integrin {alpha}{sub v}{beta}{sub 3}-targeted PET probe, [{sup 18}F ]FPP(RGD){sub 2} using small animal PET. Human HCT116 colon cancer xenografts were implanted into nude mice (n = 12) in the breast and scapular region and grown to mean diameters of 5-15 mm for approximately 2.5 weeks. A 3-min acquisition was performed on a small animal PET scanner approximately 1 h after administration of [{sup 18}F]FPP(RGD){sub 2} (1.9-3.8 MBq, 50-100 {mu}Ci) via the tail vein. A second small animal PET scan was performed approximately 6 h later after reinjection of the probe to assess for reproducibility. Images were analyzed by drawing an ellipsoidal region of interest (ROI) around the tumor xenograft activity. Percentage injected dose per gram (%ID/g) values were calculated from the mean or maximum activity in the ROIs. Coefficients of variation and differences in %ID/g values between studies from the same day were calculated to determine the reproducibility. The coefficient of variation (mean {+-}SD) for %ID{sub mean}/g and %ID{sub max}/g values between [{sup 18}F]FPP(RGD){sub 2} small animal PET scans performed 6 h apart on the same day were 11.1 {+-} 7.6% and 10.4 {+-} 9.3%, respectively. The corresponding differences in %ID{sub mean}/g and %ID{sub max}/g values between scans were -0.025 {+-} 0.067 and -0.039 {+-} 0.426. Immunofluorescence studies revealed a direct relationship between extent of {alpha}{sub {nu}}{beta}{sub 3} integrin expression in tumors and tumor vasculature

  7. Covalent bonding of YIGSR and RGD to PEDOT/PSS/MWCNT-COOH composite material to improve the neural interface.

    Science.gov (United States)

    Wang, Kun; Tang, Rong-Yu; Zhao, Xiao-Bo; Li, Jun-Jie; Lang, Yi-Ran; Jiang, Xiao-Xia; Sun, Hong-Ji; Lin, Qiu-Xia; Wang, Chang-Yong

    2015-11-28

    The development of coating materials for neural interfaces has been a pursued to improve the electrical, mechanical and biological performances. For these goals, a bioactive coating was developed in this work featuring a poly(3,4-ethylenedioxythiophene) (PEDOT)/carbon nanotube (CNT) composite and covalently bonded YIGSR and RGD. Its biological effect and electrical characteristics were assessed in vivo on microwire arrays (MWA). The coated electrodes exhibited a significantly higher charge storage capacity (CSC) and lower electrochemical impedance at 1 kHz which are desired to improve the stimulating and recording performances, respectively. Acute neural recording experiments revealed that coated MWA possess a higher signal/noise ratio capturing spikes undetected by uncoated electrodes. Moreover, coated MWA possessed more active sites and single units, and the noise floor of coated electrodes was lower than that of uncoated electrodes. There is little information in the literature concerning the chronic performance of bioactively modified neural interfaces in vivo. Therefore in this work, chronic in vivo tests were conducted and the PEDOT/PSS/MWCNT-polypeptide coated arrays exhibited excellent performances with the highest mean maximal amplitude from day 4 to day 12 during which the acute response severely compromised the performance of the electrodes. In brief, we developed a simple method of covalently bonding YIGSR and RGD to a PEDOT/PSS/MWCNT-COOH composite improving both the biocompatibility and electrical performance of the neural interface. Our findings suggest that YIGSR and RGD modified PEDOT/PSS/MWCNT is a promising bioactivated composite coating for neural recording and stimulating.

  8. Discovery and in Vivo Evaluation of Novel RGD-Modified Lipid-Polymer Hybrid Nanoparticles for Targeted Drug Delivery

    Directory of Open Access Journals (Sweden)

    Yinbo Zhao

    2014-09-01

    Full Text Available In the current study, the lipid-shell and polymer-core hybrid nanoparticles (lpNPs modified by Arg–Gly–Asp(RGD peptide, loaded with curcumin (Cur, were developed by emulsification-solvent volatilization method. The RGD-modified hybrid nanoparticles (RGD–lpNPs could overcome the poor water solubility of Cur to meet the requirement of intravenous administration and tumor active targeting. The obtained optimal RGD-lpNPs, composed of PLGA (poly(lactic-co-glycolic acid–mPEG (methoxyl poly(ethylene- glycol, RGD–polyethylene glycol (PEG–cholesterol (Chol copolymers and lipids, had good entrapment efficiency, submicron size and negatively neutral surface charge. The core-shell structure of RGD–lpNPs was verified by TEM. Cytotoxicity analysis demonstrated that the RGD–lpNPs encapsulated Cur retained potent anti-tumor effects. Flow cytometry analysis revealed the cellular uptake of Cur encapsulated in the RGD–lpNPs was increased for human umbilical vein endothelial cells (HUVEC. Furthermore, Cur loaded RGD–lpNPs were more effective in inhibiting tumor growth in a subcutaneous B16 melanoma tumor model. The results of immunofluorescent and immunohistochemical studies by Cur loaded RGD–lpNPs therapies indicated that more apoptotic cells, fewer microvessels, and fewer proliferation-positive cells were observed. In conclusion, RGD–lpNPs encapsulating Cur were developed with enhanced anti-tumor activity in melanoma, and Cur loaded RGD–lpNPs represent an excellent tumor targeted formulation of Cur which might be an attractive candidate for cancer therapy.

  9. Probing secondary structures of end-capped tripeptide, BOC–Phe–Aib–Leu–OMe by combined study of infrared spectroscopy and quantum chemistry calculation

    International Nuclear Information System (INIS)

    Graphical abstract: Our analysis reveals two secondary structures of BOC–Phe–Aib–Leu–OMe in CCl4 solution, which are largely different from that present in solid crystal of the tripeptide. The figure shows the curve fitting of the amide-I spectral profile of the tripeptide in 0.006 M CCl4 solution at room temperature by voigt band shapes (right). Set 1 (green) corresponds to the amide-I transitions of secondary structure-1, and Set 2 (blue) corresponds to similar transitions of secondary structure-2. Highlights: ► Probed secondary structures of BOC–Phe–Aib–Leu–OMe in CCl4 solution and solid state. ► FTIR spectroscopy and quantum chemistry computations are used in the study. ► Study reveals largely different secondary structures in solid and solution phases. ► A β-turn structure in solid crystal, whereas the solution contains γ-turns. - Abstract: We report conformational analysis of an end-protected tripeptide, BOC–Phe–Aib–Leu–OMe in CCl4 solution and solid crystal at room temperature by infrared spectroscopy and quantum chemistry calculation. The measured spectra in those physical conditions appear different, which has been attributed to different secondary structures of the peptide in the two phases. Our analysis reveals that the peptide has only one secondary structure in the crystal, but in the solution there are two different secondary structures. While the spectrum in solution shows excellent correspondence with the superposition of predicted spectra of the two most favored gas-phase secondary structures, the spectrum of crystal shows best correspondence with the predicted spectrum of a relatively higher energy (∼6 kcal/mol) secondary structure. However, the latter in crystal can gain maximum stability by intermolecular hydrogen bonding. The secondary structure of the peptide in crystal that we have deduced agrees nicely with the one suggested by X-ray crystallographic study.

  10. Tissue-targeting lead generation and optimization from random and directed screening of technetium-99m labeled tripeptide complex libraries in vivo

    Institute of Scientific and Technical Information of China (English)

    ZENG Jun; LIU Ci-yi; XIE Wen-hui; HU Si-long; JIN Mu-xiu

    2006-01-01

    Background Screening libraries against a molecular target in vitro are idealized models that cannot reflect the real state in vivo where biomolecules coexist and interact. C-terminal amide tripeptides labelled with Technetium-99m can provide a unique noninvasive approach to trace a large number of compounds in vivo.Methods The C-terminal amide tripeptide libraries were synthesized on Rink Amide-MBHA resin using iterative and pooling protocol. Technetium (Ⅴ) oxo core [TcO3+] was bound to each tripeptide via 4 deprotonated nitrogen atoms to form a library of 8000 99mTc tripeptoid complexes. The radiocombinatorial screening (RCS) in vivo was carried out on SD rats and A549 tumour bearing mice.Results Signals of tissue distribution and metabolism of libraries were recorded by counting or imaging and tissue targeting leads identified by both random and directed RCS. Among them, 99mTc RPA, 99mTc VIG and 99mTc RES had specific tissue targeting in kidney, liver and tumour respectively. The percent injected dose per gram tissue of 99mTc labelled leads in their target tissue was highly structure dependent. Because the nontarget tissue binding and the metabolism of 99mTc tripeptoid sublibraries were simultaneously monitored successfully by RCS, the interference of background activity was limited to the lowest level. Optimization of renal function agent from the labelled libraries was carried out by directed screening. 99mTc DSG was finally identified the most promising agent for renal function studies.Conclusions RCS in vivo is a powerful tool for the discovery of tissue targeting drugs. The potential screening bias is probably the major limitation of labelled libraries.

  11. Genome-wide DNA polymorphism in the indica rice varieties RGD-7S and Taifeng B as revealed by whole genome re-sequencing.

    Science.gov (United States)

    Fu, Chong-Yun; Liu, Wu-Ge; Liu, Di-Lin; Li, Ji-Hua; Zhu, Man-Shan; Liao, Yi-Long; Liu, Zhen-Rong; Zeng, Xue-Qin; Wang, Feng

    2016-03-01

    Next-generation sequencing technologies provide opportunities to further understand genetic variation, even within closely related cultivars. We performed whole genome resequencing of two elite indica rice varieties, RGD-7S and Taifeng B, whose F1 progeny showed hybrid weakness and hybrid vigor when grown in the early- and late-cropping seasons, respectively. Approximately 150 million 100-bp pair-end reads were generated, which covered ∼86% of the rice (Oryza sativa L. japonica 'Nipponbare') reference genome. A total of 2,758,740 polymorphic sites including 2,408,845 SNPs and 349,895 InDels were detected in RGD-7S and Taifeng B, respectively. Applying stringent parameters, we identified 961,791 SNPs and 46,640 InDels between RGD-7S and Taifeng B (RGD-7S/Taifeng B). The density of DNA polymorphisms was 256.8 SNPs and 12.5 InDels per 100 kb for RGD-7S/Taifeng B. Copy number variations (CNVs) were also investigated. In RGD-7S, 1989 of 2727 CNVs were overlapped in 218 genes, and 1231 of 2010 CNVs were annotated in 175 genes in Taifeng B. In addition, we verified a subset of InDels in the interval of hybrid weakness genes, Hw3 and Hw4, and obtained some polymorphic InDel markers, which will provide a sound foundation for cloning hybrid weakness genes. Analysis of genomic variations will also contribute to understanding the genetic basis of hybrid weakness and heterosis.

  12. Integrin engagement by the helical RGD motif of the Helicobacter pylori CagL protein is regulated by pH-induced displacement of a neighboring helix.

    Science.gov (United States)

    Bonsor, Daniel A; Pham, Kieu T; Beadenkopf, Robert; Diederichs, Kay; Haas, Rainer; Beckett, Dorothy; Fischer, Wolfgang; Sundberg, Eric J

    2015-05-15

    Arginine-aspartate-glycine (RGD) motifs are recognized by integrins to bridge cells to one another and the extracellular matrix. RGD motifs typically reside in exposed loop conformations. X-ray crystal structures of the Helicobacter pylori protein CagL revealed that RGD motifs can also exist in helical regions of proteins. Interactions between CagL and host gastric epithelial cell via integrins are required for the translocation of the bacterial oncoprotein CagA. Here, we have investigated the molecular basis of the CagL-host cell interactions using structural, biophysical, and functional analyses. We solved an x-ray crystal structure of CagL that revealed conformational changes induced by low pH not present in previous structures. Using analytical ultracentrifugation, we found that pH-induced conformational changes in CagL occur in solution and not just in the crystalline environment. By designing numerous CagL mutants based on all available crystal structures, we probed the functional roles of CagL conformational changes on cell surface integrin engagement. Together, our data indicate that the helical RGD motif in CagL is buried by a neighboring helix at low pH to inhibit CagL binding to integrin, whereas at neutral pH the neighboring helix is displaced to allow integrin access to the CagL RGD motif. This novel molecular mechanism of regulating integrin-RGD motif interactions by changes in the chemical environment provides new insight to H. pylori-mediated oncogenesis.

  13. In vivo stimulation of connective tissue accumulation by the tripeptide-copper complex glycyl-L-histidyl-L-lysine-Cu2+ in rat experimental wounds.

    OpenAIRE

    Maquart, F X; Bellon, G.; Chaqour, B; Wegrowski, J; Patt, L M; Trachy, R. E.; Monboisse, J C; Chastang, F; Birembaut, P; Gillery, P.

    1993-01-01

    The tripeptide-copper complex glycyl-L-histidyl-L-lysine-Cu2+ (GHK-Cu) was first described as a growth factor for differentiated cells. Recent in vitro data showed that it possesses several properties of a potential activator of wound repair. We investigated the effects of GHK-Cu in vivo, using the wound chamber model described previously (Schilling, J.A., W. Joel, and M.T. Shurley, 1959. Surgery [St. Louis]. 46:702-710). Stainless steel wire mesh cylinders were implanted subcutaneously on th...

  14. Common interruptions in the repeating tripeptide sequence of non-fibrillar collagens: Sequence analysis and structural studies on triple-helix peptide models

    OpenAIRE

    Thiagarajan, Geetha; Li, Yingjie; Mohs, Angela; Strafaci, Christopher; Popiel, Magdalena; Baum, Jean; Brodsky, Barbara

    2007-01-01

    Interruptions in the repeating (Gly-X1-X2)n amino acid sequence pattern are found in the triple-helix domains of all non-fibrillar collagens, and perturbations to the triple-helix at such sites are likely to play a role in collagen higher order structure and function. This report defines the sequence features and structural consequences of the most common interruption, where one residue is missing in the tripeptide pattern, Gly-X1-X2-Gly-AA1-Gly-X1-X2, designated as G1G interruptions. Residue...

  15. Atomic Structures of the Amino Acids, Glycine, Alanine and Serine and Their Tripeptide, with Bond Lengths as Sums of Atomic Covalent Radii

    CERN Document Server

    Heyrovska, Raji

    2008-01-01

    Recently, the bond lengths of the molecular components of nucleic acids and of caffeine and related molecules were shown to be sums of the appropriate covalent radii of the adjacent atoms. Thus, each atom was shown to have its specific contribution to the bond length. This enabled establishing their atomic structures for the first time. In this work, the known bond lengths for amino acids and the peptide bond are similarly shown to be sums of the atomic covalent radii. Based on this result, the atomic structures of glycine, alanine and serine and their tripeptide have been presented.

  16. The HDAC Inhibitors Scriptaid and LBH589 Combined with the Oncolytic Virus Delta24-RGD Exert Enhanced Anti-Tumor Efficacy in Patient-Derived Glioblastoma Cells.

    Directory of Open Access Journals (Sweden)

    Lotte M E Berghauser Pont

    Full Text Available A phase I/II trial for glioblastoma with the oncolytic adenovirus Delta24-RGD was recently completed. Delta24-RGD conditionally replicates in cells with a disrupted retinoblastoma-pathway and enters cells via αvβ3/5 integrins. Glioblastomas are differentially sensitive to Delta24-RGD. HDAC inhibitors (HDACi affect integrins and share common cell death pathways with Delta24-RGD. We studied the combination treatment effects of HDACi and Delta24-RGD in patient-derived glioblastoma stem-like cells (GSC, and we determined the most effective HDACi.SAHA, Valproic Acid, Scriptaid, MS275 and LBH589 were combined with Delta24-RGD in fourteen distinct GSCs. Synergy was determined by Chou Talalay method. Viral infection and replication were assessed using luciferase and GFP encoding vectors and hexon-titration assays. Coxsackie adenovirus receptor and αvβ3 integrin levels were determined by flow cytometry. Oncolysis and mechanisms of cell death were studied by viability, caspase-3/7, LDH and LC3B/p62, phospho-p70S6K. Toxicity was studied on normal human astrocytes. MGMT promotor methylation status, TCGA classification, Rb-pathway and integrin gene expression levels were assessed as markers of responsiveness.Scriptaid and LBH589 acted synergistically with Delta24-RGD in approximately 50% of the GSCs. Both drugs moderately increased αvβ3 integrin levels and viral infection in responding but not in non-responding GSCs. LBH589 moderately increased late viral gene expression, however, virus titration revealed diminished viral progeny production by both HDACi, Scriptaid augmented caspase-3/7 activity, LC3B conversion, p62 and phospho-p70S6K consumption, as well as LDH levels. LBH589 increased LDH and phospho-p70S6K consumption. Responsiveness correlated with expression of various Rb-pathway genes and integrins. Combination treatments induced limited toxicity to human astrocytes.LBH589 and Scriptaid combined with Delta24-RGD revealed synergistic anti

  17. Study on bioactive di-, and tripeptide content in natural plant sources by HPLC methods with respect to functional food application

    International Nuclear Information System (INIS)

    Complete text of publication follows. Small molecular weight peptides represent an important family of compounds that play significant role in physiological and biochemical processes as well as in clinical and food research. The functional properties can include antioxidant and antimicrobial activity, surfactant and nutritional capabilities, moreover these compounds can contribute to the development of characteristic flavors such as sweetness and bitterness in various types of food. Several publications have dealt with the separation, detection and identification of these compounds, however the published methods carry difficulties in terms of the quantitative analysis, the sensitivity and reproducibility have been proven to be poor mainly because several amino acid moieties have low UV-absorbing properties. Our intention was to develop a reliable and sensitive chromatographic method to detect di-, and tripeptides (Aspartame, L-carnosine, L-glutation, Alanyl-glutamine and γ-glutein) in raw and processed food materials. A HPLC method was developed to analyze peptide containing complex food samples and raw materials using trifluoro-acetic acid-acetonitrile eluent system with gradient elution on a C18 reverse phase chromatographic column. The detection of free peptides was carried out using evaporative light scattering (ELS) detection, while UV detection was accomplished by pre-column derivatization with danzyl-chloride. Pea, rice and garlic samples have been selected for the study, the extraction procedure was optimized with different solvents: 0.1 M phosphoric-acid, 0.1 M hydrochloric-acid, acetic-acid, ethanol and water, the peptide content was analyzed with the newly developed technique. Antioxidant activity (FRAP) was observed only for the sulphur containing derivatives (γ-glutein, L-glutation). Garlic extracts have shown the highest antioxidant activity (46 ppm in ascorbic acid equivalents), pea samples have exhibited lower activity (23 ppm) and the lowest

  18. Theranostic cRGD-BioShuttle Constructs Containing Temozolomide- and Cy7 For NIR-Imaging and Therapy

    Science.gov (United States)

    Wiessler, Manfred; Hennrich, Ute; Pipkorn, Rüdiger; Waldeck, Waldemar; Cao, Liji; Peter, Jörg; Ehemann, Volker; Semmler, Wolfhard; Lammers, Twan; Braun, Klaus

    2011-01-01

    Innovative and personalized therapeutic approaches result from the identification and control of individual aberrantly expressed genes at the transcriptional and post-transcriptional level. Therefore, it is of high interest to establish diagnostic, therapeutic and theranostic strategies at these levels. In the present study, we used the Diels-Alder Reaction with inverse electron demand (DARinv) click chemistry to prepare a series of cyclic RGD-BioShuttle constructs. These constructs carry the near-infrared (NIR) imaging agent Cy7 and the chemotherapeutic agent temozolomide (TMZ). We evaluated their uptake by and their efficacy against integrin αvβ3-expressing MCF7 human breast carcinoma cells. In addition, using a mouse phantom, we analyzed the suitability of this targeted theranostic agent for NIR optical imaging. We observed that the cyclic RGD-based carriers containing TMZ and/or Cy7 were effectively taken up by αvβ3-expressing cells, that they were more effective than free TMZ in inducing cell death, and that they could be quantitatively visualized using NIR fluorescence imaging. Therefore, these targeted theranostic agents are considered to be highly suitable systems for improving disease diagnosis and therapy. PMID:22211144

  19. Theranostic cRGD-BioShuttle Constructs Containing Temozolomide- and Cy7 For NIR-Imaging and Therapy

    Directory of Open Access Journals (Sweden)

    Manfred Wiessler, Ute Hennrich, Rüdiger Pipkorn, Waldemar Waldeck, Liji Cao, Jörg Peter, Volker Ehemann, Wolfhard Semmler, Twan Lammers, Klaus Braun

    2011-01-01

    Full Text Available Innovative and personalized therapeutic approaches result from the identification and control of individual aberrantly expressed genes at the transcriptional and post-transcriptional level. Therefore, it is of high interest to establish diagnostic, therapeutic and theranostic strategies at these levels. In the present study, we used the Diels-Alder Reaction with inverse electron demand (DARinv click chemistry to prepare a series of cyclic RGD-BioShuttle constructs. These constructs carry the near-infrared (NIR imaging agent Cy7 and the chemotherapeutic agent temozolomide (TMZ. We evaluated their uptake by and their efficacy against integrin αvβ3-expressing MCF7 human breast carcinoma cells. In addition, using a mouse phantom, we analyzed the suitability of this targeted theranostic agent for NIR optical imaging. We observed that the cyclic RGD-based carriers containing TMZ and/or Cy7 were effectively taken up by αvβ3-expressing cells, that they were more effective than free TMZ in inducing cell death, and that they could be quantitatively visualized using NIR fluorescence imaging. Therefore, these targeted theranostic agents are considered to be highly suitable systems for improving disease diagnosis and therapy.

  20. RGD-conjugated rod-like viral nanoparticles on 2D scaffold improved bone differentiation of mesenchymal stem cells

    Science.gov (United States)

    Wang, Qian; Pongkwan, Sitasuwan; Lee, L.; Li, Kai; Nguyen, Huong

    2014-05-01

    Viral nanoparticles have uniform and well-defined nano-structures and can be produced in large quantities. Several plant viral nanoparticles have been tested in biomedical applications due to the lack of mammalian cell infectivity. We are particularly interested in using Tobacco mosaic virus (TMV), which has been demonstrated to enhance bone tissue regeneration, as a tuneable nanoscale building block for biomaterials development. Unmodified TMV particles have been shown to accelerate osteogenic differentiation of adult stem cells by synergistically upregulating BMP2 and IBSP expression with dexamethasone. However, the lack of affinity to mammalian cell surface resulted in low initial cell adhesion. In this study, to increase cell binding capacity of TMV based material the chemical functionalization of TMV with arginine-glycine-aspartic acid (RGD) peptide was explored. An azide-derivatized RGD peptide was “clicked” to tyrosine residues on TMV outer surface via an efficient copper(I) catalysed azide-alkyne cycloaddition reaction. The ligand spacing is calculated to be 2-4 nm, which could offer a polyvalent ligand clustering effect for enhanced cell receptor signalling, further promoting the proliferation and osteogenic differentiation of bone marrow derived mesenchymal stem cells.

  1. RGD-conjugated two-photon absorbing near-IR emitting fluorescent probes for tumor vascular imaging (Conference Presentation)

    Science.gov (United States)

    Belfield, Kevin D.; Yue, Xiling; Morales, Alma R.; Githaiga, Grace W.; Woodward, Adam W.; Tang, Simon; Sawada, Junko; Komatsu, Masanobu; Liu, Xuan

    2016-03-01

    Observation of the activation and inhibition of angiogenesis processes is important in the progression of cancer. Application of targeting peptides, such as a small peptide that contains adjacent L-arginine (R), glycine (G) and L-aspartic acid (D) residues can afford high selectivity and deep penetration in vessel imaging. To facilitate deep tissue vasculature imaging, probes that can be excited via two-photon absorption (2PA) in the near-infrared (NIR) and subsequently emit in the NIR are essential. In this study, the enhancement of tissue image quality with RGD conjugates was investigated with new NIR-emitting pyranyl fluorophore derivatives in two-photon fluorescence microscopy. Linear and nonlinear photophysical properties of the new probes were comprehensively characterized; significantly the probes exhibited good 2PA over a broad spectral range from 700-1100 nm. Cell and tissue images were then acquired and examined, revealing deep penetration and high contrast with the new pyranyl RGD-conjugates up to 350 μm in tumor tissue.

  2. The tripeptide-copper complex glycyl-L-histidyl-L-lysine-Cu2+ stimulates matrix metalloproteinase-2 expression by fibroblast cultures.

    Science.gov (United States)

    Siméon, A; Emonard, H; Hornebeck, W; Maquart, F X

    2000-09-22

    Glycyl-histidyl-lysine-Cu2+ (GHK-Cu) is a tripeptide-copper complex known to be a potent wound healing agent. We previously showed its ability to stimulate in vitro and in vivo the synthesis of extracellular matrix components. The aim of this study was to determine the effects of GHK-Cu on MMP-2 synthesis by dermal fibroblasts in culture. We showed that GHK-Cu increased MMP-2 levels in conditioned media of cultured fibroblasts. This effect was reproduced by copper ions but not by the tripeptide GHK alone. This stimulation was accompanied by an increase of MMP-2 mRNA level. We also showed that GHK-Cu increased the secretion of the tissue inhibitors of metalloproteinases, TIMP-1 and TIMP-2. Taken together, our results underline that GHK-Cu is not only an activator of connective tissue production but also of the remodeling of the extracellular matrix. It is able to modulate MMP expression by acting directly on wound fibroblasts. PMID:11045606

  3. In vivo stimulation of connective tissue accumulation by the tripeptide-copper complex glycyl-L-histidyl-L-lysine-Cu2+ in rat experimental wounds.

    Science.gov (United States)

    Maquart, F X; Bellon, G; Chaqour, B; Wegrowski, J; Patt, L M; Trachy, R E; Monboisse, J C; Chastang, F; Birembaut, P; Gillery, P

    1993-11-01

    The tripeptide-copper complex glycyl-L-histidyl-L-lysine-Cu2+ (GHK-Cu) was first described as a growth factor for differentiated cells. Recent in vitro data showed that it possesses several properties of a potential activator of wound repair. We investigated the effects of GHK-Cu in vivo, using the wound chamber model described previously (Schilling, J.A., W. Joel, and M.T. Shurley, 1959. Surgery [St. Louis]. 46:702-710). Stainless steel wire mesh cylinders were implanted subcutaneously on the back of rats. The animals were divided into groups that received sequential injections into the wound chamber of either saline (control group) or various concentrations of GHK-Cu. At the end of the experiments, rats were killed, wound chambers were collected, and their content was analyzed for dry weight, total proteins, collagen, DNA, elastin, glycosaminoglycans, and specific mRNAs for collagens and TGF beta. In the GHK-Cu-injected wound chambers, a concentration-dependent increase of dry weight, DNA, total protein, collagen, and glycosaminoglycan contents was found. The stimulation of collagen synthesis was twice that of noncollagen proteins. Type I and type III collagen mRNAs were increased but not TGF beta mRNAs. An increase of the relative amount of dermatan sulfate was also found. A control tripeptide, L-glutamyl-L-histidyl-L-proline, had no significant effect. These results demonstrate that GHK-Cu is able to increase extracellular matrix accumulation in wounds in vivo. PMID:8227353

  4. Identification and Characterization of a Novel Nontranslated Sequence Variant of the Human Intestinal Di-/Tripeptide Transporter, hPEPT1

    Directory of Open Access Journals (Sweden)

    Helle Bach Søndergaard

    2012-01-01

    Full Text Available The human H+-coupled di-/tripeptide transporter (hPEPT1 mediates intestinal absorption of dietary di- and tripeptides, as well as several peptidomimetic drug compounds. The aim of the present study was to investigate the possible role of the hPEPT1 variant hPEPT1-RF in hPEPT1 regulation. However, the proposed hPEPT1-RF mRNA sequence could not be detected in Caco-2 cells or in human intestinal samples. Instead, a new sequence variant, hPEPT1-RFI, was found, which is almost identical to the proposed hPEPT1-RF, except for two nucleotide insertions and one deletion that resulted in a changed open reading frame as compared to hPEPT1-RF. In vitro translation analysis showed that hPEPT1-RFI was not translated. In conclusion, the existence of hPEPT1-RF could not be confirmed; furthermore, the identified sequence variant, hPEPT1-RFI, does not appear to be translated and is therefore unlikely to have a regulatory effect on hPEPT1 transport activity.

  5. Echistatin is a potent inhibitor of bone resorption in culture.

    Science.gov (United States)

    Sato, M; Sardana, M K; Grasser, W A; Garsky, V M; Murray, J M; Gould, R J

    1990-10-01

    The venom protein, s-echistatin, originally derived from the saw-scaled viper Echis carinatus, was found to be a potent inhibitor of bone resorption by isolated osteoclasts. This Arg24-Gly25-Asp26-(RGD)-containing protein inhibited the excavation of bone slices by rat osteoclasts (IC50 = 0.1 nM). It also inhibited the release of [3H]proline from labeled bone particles by chicken osteoclasts (IC50 = 100 nM). By comparison, the tetrapeptide Arg-Gly-Asp-Ser (RGDS) inhibited resorption by rat or chicken osteoclasts with an IC50 of 0.1 mM while ala24-echistatin was inactive. Video microscopy showed that rat osteoclast attachment to substrate was more sensitive to s-echistatin than was the attachment of mononuclear cells or chicken osteoclasts. The difference in sensitivity of rat and chicken osteoclasts to s-echistatin may be due to differences between receptors on rat and chicken osteoclasts for s-echistatin. Antibody localization of echistatin on these cells showed much greater echistatin binding to rat osteoclasts than to chicken osteoclasts. Laser scanning confocal microscopy after immunohistochemical staining showed that s-echistatin binds to osteoclasts, that s-echistatin receptors are most abundant at the osteoclast/glass interface, and that s-echistatin colocalizes with vinculin. Confocal interference reflection microscopy of osteoclasts incubated with s-echistatin, demonstrated colocalization of s-echistatin with the outer edges of clusters of grey contacts at the tips of some lamellipodia. Identification of the echistatin receptor as an integrin was confirmed by colocalization of echistatin fluorescence with staining for an alpha-like subunit. Attachment of bone particles labeled with [3H]proline to chicken osteoclasts confirmed that the mechanism of action of echistatin was to inhibit osteoclast binding to bone presumably by disrupting adhesion structures. These data demonstrate that osteoclasts bind to bone via an RGD-sequence as an obligatory step in bone

  6. Pharmacokinetics of 99m Tc-EDDA/HYNIC-Lys-D-Phe-RGD in athymic mice with induced malignant tumors for integrin imaging

    International Nuclear Information System (INIS)

    Full text: Nuclear medicine imaging techniques are non-invasive and monitor the spatiotemporal distribution of molecular events. Radiolabeled RGD-peptides are currently investigated to target integrin receptors for in vivo tumor imaging. The αvβ3 integrin is a target structure involved in the angio genesis process which mediates the binding to extracellular matrix via different proteins such as vitronectin, fibronectin and von Willebrand factor. The aim of this research was to prepare [99mTc]-Lys-D-Phe-RGD and to evaluate its pharmacokinetics in athymic mice with three different induced malignant tumors. Tumor uptake values of 99mTc-Lys-D-Phe-RGD labeled via HYNIC and EDDA showed good ability to target αvβ3 integrin receptors in the three different kinds of tumors (breast, prostate and neuroendocrine). A high in vivo stability and favorable pharmacokinetic properties such as fast blood clearance, rapid renal excretion, low liver and muscle uptake and low intestinal excretion were observed. This study demonstrated that 99mTc-EDDA/HYNIC-Lys-D-Phe-RGD is a specific and potential radiopharmaceutical to image αvβ3 integrin receptors in a variety of tumors. (Author)

  7. PET imaging of alpha(v)beta(3) integrin expression in tumours with Ga-68-labelled mono-, di- and tetrameric RGD peptides

    NARCIS (Netherlands)

    Dijkgraaf, Ingrid; Yim, Cheng-Bin; Franssen, Gerben M.; Schuit, Robert C.; Luurtsema, Gert; Liu, Shuang; Oyen, Wim J. G.; Boerman, Otto C.

    2011-01-01

    Due to the restricted expression of alpha(v)beta(3) in tumours, alpha(v)beta(3) is considered a suitable receptor for tumour targeting. In this study the alpha(v)beta(3)-binding characteristics of Ga-68-labelled monomeric, dimeric and tetrameric RGD peptides were determined and compared with their I

  8. Effect of RGD-insulin on activities of bone resorption and the possible mechanism in human osteoclast-like cells in vitro.

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    Objective: In this study, we explored the mechanism of anti-bone resorption of RGD-insulin using osteoclastlike cells(OLCs) from giant cell tumor of bone as an in vitro model. Methods: The function of bone resorption was observed and the staining for tartrate-resistant acid

  9. 整合素靶向性RGD-Luc荧光探针体外显像人肝癌细胞的实验研究

    Institute of Scientific and Technical Information of China (English)

    赵伟娜; 李绍东; 徐凯

    2015-01-01

    目的 构建环状RGD肽偶联荧光素酶(luciferase)蛋白的荧光分子探针RGD-Luc,探讨RGD-Luc在体外与人肝癌细胞结合后分子显像对肝癌早期诊断的价值. 方法 (1)用化学方法偶联luciferase蛋白的氨基与环RGD短肽的巯基,并用Western Blotting鉴定;(2)将RGD-Luc、Luciferase分别与HepG2细胞孵育2h,同时设立未作处理的对照组,使用小动物活体成像系统分别观察靶向结合情况. 结果 (1)Western Blotting分析证明荧光素酶luciferase与cRGDfC肽成功偶联;(2)小动物活体成像系统检测示RGD-Luc荧光探针能够特异性的与HepG2细胞结合,而Luciferase组及对照组未观察到荧光分布. 结论 利用化学偶联方法成功制备新型荧光探针RGD-Luc,其能靶向增强肝癌细胞的分子显像,为肝癌的早期显像提供了一定的实验依据.

  10. Improved targeting of the {alpha}{sub v}{beta}{sub 3} integrin by multimerisation of RGD peptides

    Energy Technology Data Exchange (ETDEWEB)

    Dijkgraaf, Ingrid [Radboud University Nijmegen Medical Centre, Department of Nuclear Medicine, P.O. Box 9101, Nijmegen (Netherlands); Utrecht University, Department of Medicinal Chemistry, Utrecht Institute for Pharmaceutical Sciences, P.O. Box 80082, Utrecht (Netherlands); Kruijtzer, John A.W.; Liskamp, Rob M.J. [Utrecht University, Department of Medicinal Chemistry, Utrecht Institute for Pharmaceutical Sciences, P.O. Box 80082, Utrecht (Netherlands); Liu, Shuang [Purdue University, School of Health Sciences, West Lafayette, IN (United States); Soede, Annemieke C.; Oyen, Wim J.G.; Corstens, Frans H.M.; Boerman, Otto C. [Radboud University Nijmegen Medical Centre, Department of Nuclear Medicine, P.O. Box 9101, Nijmegen (Netherlands)

    2007-02-15

    The integrin {alpha}{sub v}{beta}{sub 3} is expressed on sprouting endothelial cells and on various tumour cell types. Due to the restricted expression of {alpha}{sub v}{beta}{sub 3} in tumours, {alpha}{sub v}{beta}{sub 3} is considered a suitable receptor for tumour targeting. In this study the {alpha}{sub v}{beta}{sub 3} binding characteristics of an {sup 111}In-labelled monomeric, dimeric and tetrameric RGD analogue were compared. A monomeric (E-c(RGDfK)), dimeric (E-[c(RGDfK)]{sub 2}), and tetrameric (E{l_brace}E[c(RGDfK)]{sub 2}{r_brace}{sub 2}) RGD peptide were synthesised, conjugated with DOTA and radiolabelled with {sup 111}In. In vitro {alpha}{sub v}{beta}{sub 3} binding characteristics were determined in a competitive binding assay. In vivo {alpha}{sub v}{beta}{sub 3} targeting characteristics of the compounds were assessed in mice with SK-RC-52 xenografts. The IC{sub 50} values for DOTA-E-c(RGDfK), DOTA-E-[c(RGDfK)]{sub 2}, and DOTA-E{l_brace}E[c(RGDfK)]{sub 2}{r_brace}{sub 2}were 120 nM, 69.9 nM and 19.6 nM, respectively. At all time points, the tumour uptake of the dimer was significantly higher as compared to that of the monomer. At 8 h p.i., tumour uptake of the tetramer (7.40 {+-}1.12%ID/g) was significantly higher than that of the monomer (2.30 {+-}0.34%ID/g), p <0.001, and the dimer (5.17 {+-}1.22%ID/g), p <0.05. At 24 h p.i., the tumour uptake was significantly higher for the tetramer (6.82 {+-}1.41%ID/g) than for the dimer (4.22 {+-}0.96%ID/g), p <0.01, and the monomer (1.90 {+-}0.29%ID/g), p <0.001. Multimerisation of c(RGDfK) resulted in enhanced affinity for {alpha}{sub v}{beta}{sub 3} as determined in vitro. Tumour uptake of a tetrameric RGD peptide was significantly higher than that of the monomeric and dimeric analogues, presumably owing to the enhanced statistical likelihood for rebinding to {alpha}{sub v}{beta}{sub 3}. (orig.)

  11. ETPE发射药与RGD7硝胺发射药燃烧性能及热行为的对比研究%Combustion Properties and Thermal Behavior of ETPE gun propellant and RGD7 Nitramine Gun Propellant

    Institute of Scientific and Technical Information of China (English)

    赵瑛; 刘毅; 杨丽侠; 张邹邹

    2012-01-01

    用密闭爆发器实验、差示扫描量热法(DSC)和扫描电子显微镜(SEM)研究了3,3-二叠氮甲基氧丁环/3-叠氮甲基-3-甲基氧丁环( BAMO/AMMO)基含能热塑性弹性体(ETPE)发射药和RGD7硝胺发射药的燃烧性能及热行为.结果表明:与RGD7硝胺发射药相比,ETPE发射药燃烧时间较长,燃速较低,燃速压力指数n大于1,而RGD7硝胺发射药燃速压力指数小于1.对于RGD7硝胺发射药,RDX的熔融吸热峰(204.8℃)不明显,且分解放热峰(240℃)滞后于硝化棉/硝化甘油(NC/NG) (194℃),而ETPE发射药中poly(BAMO/AMMO)分解温度(263℃)高于RDX(240℃).ETPE发射药和RGD7硝胺发射药的不同燃烧性能归因于发射药中主组分的不同热行为.%The combustion properties and thermal behaviors of energetic thermoplastic elastomer (ETPE) gun propellant and RCD7 nitramine gun propellant were studied by the closed bomb test, differential scanning calorimetry ( DSC) and scanning electron microscopy (SEM). The results show that in comparison with RGD7 nitramine gun propellant, the ETPE gun propellant has lower burning rate, longer burning time and the burning rate pressure exponent larger than 1 , whereas the pressure exponent of RGD7 nitramine gun propellant is less than 1. For RGD7 nitramine gun propellant, the endothermic'melting peak of RDX at 204.8 t in the propellant is inconspicuous and the exothermic decomposition peak of RDX at 240 ℃ is lag behind the exothermic decomposition peak at 194 ℃ of the NC/NG, where as the thermal decomposition temperature at 263 ℃ of poly( BAMO/AMMO) in ETPE gun propellant is higher than the thermal decomposition temperature at 240 ℃ of RDX. The different combustion properties of two gun propellants are caused by the different thermal behaviour of main compounds in the propellants.

  12. Formation of multicellular tumor spheroids induced by cyclic RGD-peptides and use for anticancer drug testing in vitro.

    Science.gov (United States)

    Akasov, Roman; Zaytseva-Zotova, Daria; Burov, Sergey; Leko, Maria; Dontenwill, Monique; Chiper, Manuela; Vandamme, Thierry; Markvicheva, Elena

    2016-06-15

    Development of novel anticancer formulations is a priority challenge in biomedicine. However, in vitro models based on monolayer cultures (2D) which are currently used for cytotoxicity tests leave much to be desired. More and more attention is focusing on 3D in vitro systems which can better mimic solid tumors. The aim of the study was to develop a novel one-step highly reproducible technique for multicellular tumor spheroid (MTS) formation using synthetic cyclic RGD-peptides, and to demonstrate availability of the spheroids as 3D in vitro model for antitumor drug testing. Cell self-assembly effect induced by addition of both linear and cyclic RGD-peptides directly to monolayer cultures was studied for 12 cell lines of various origins, including tumor cells (e.i. U-87 MG, MCF-7, M-3, HCT-116) and normal cells, in particular L-929, BNL.CL2, HepG2. Cyclo-RGDfK and its modification with triphenylphosphonium cation (TPP), namely cyclo-RGDfK(TPP) in a range of 10-100μM were found to induce spheroid formation. The obtained spheroids were unimodal with mean sizes in a range of 60-120μm depending on cell line and serum content in culture medium. The spheroids were used as 3D in vitro model, in order to evaluate cytotoxicity effects of antitumor drugs (doxorubicin, curcumin, temozolomide). The developed technique could be proposed as a promising tool for in vitro test of novel antitumor drugs. PMID:27107900

  13. Anti-EGFR-iRGD recombinant protein conjugated silk fibroin nanoparticles for enhanced tumor targeting and antitumor efficiency

    Directory of Open Access Journals (Sweden)

    Bian X

    2016-05-01

    Full Text Available Xinyu Bian,* Puyuan Wu,* Huizi Sha, Hanqing Qian, Qing Wang, Lei Cheng, Yang Yang, Mi Yang, Baorui LiuComprehensive Cancer Center of Drum-Tower Hospital, Medical School of Nanjing University, Clinical Cancer Institute of Nanjing University, Nanjing, People’s Republic of China*These authors contributed equally to this workAbstract: In this study, we report a novel kind of targeting with paclitaxel (PTX-loaded silk fibroin nanoparticles conjugated with iRGD–EGFR nanobody recombinant protein (anti-EGFR-iRGD. The new nanoparticles (called A-PTX-SF-NPs were prepared using the carbodiimide-mediated coupling procedure and their characteristics were evaluated. The cellular cytotoxicity and cellular uptake of A-PTX-SF-NPs were also investigated. The results in vivo suggested that NPs conjugated with the recombinant protein exhibited more targeting and anti-neoplastic property in cells with high EGFR expression. In the in vivo antitumor efficacy assay, the A-PTX-SF-NPs group showed slower tumor growth and smaller tumor volumes than PTX-SF-NPs in a HeLa xenograft mouse model. A real-time near-infrared fluorescence imaging study showed that A-PTX-SF-NPs could target the tumor more effectively. These results suggest that the anticancer activity and tumor targeting of A-PTX-SF-NPs were superior to those of PTX-SF-NPs and may have the potential to be used for targeted delivery for tumor therapies. Keywords: EGFR, nanobody, iRGD, recombinant protein, targeting drug carriers, antitumor efficiency

  14. Preparation of the radiopharmaceutical 99m Tc-HYNIC-cyclo-Lys-D-Phe-RGD for In vivo image of integrines

    International Nuclear Information System (INIS)

    The diagnostic of some pathological processes by means of images constitutes one of the used methods in the determination of the origin, condition and/or evolution of one illness. The use of contrast agents in conjunction with other techniques help to the obtaining and visualization of complex systems, among these we can find to those radiopharmaceuticals used in nuclear medicine to visualize diverse organs and corporal systems. At the moment it is sought to develop a radiopharmaceutical of third generation that can be used for image In vivo of integrines with the purpose of detecting angio genesis processes, that which would allow to diagnose in way it specifies a wide range of primary tumors and their metastasis. Presently work it developed the radiopharmaceutical 99mTc-HYNIC-cycle-Lys-D-Phe-RGD, likewise the good conditions were determined for the formation of this complex. The HYNIC was employee as chelating agent, using as co ligands EDDA and Tricine for to complete the sphere of coordination of the 99mTc. The conjugated HYNIC-RGD was synthesized, purified, characterized and radiolabelled In situ with 99mTc using High pressure liquid chromatography as analysis method in Reverse Phase (RP-HPLC). By this way it was developed the lyophilized formulation for its instantaneous labelled to which were carried out quality control tests. The one conjugated was obtained free of impurities, showing stability at same as their complex formed with 99mTc. The analysis method was validated turning out to be necessary, exact, lineal and specific for the quantification of the analyte of interest. The lyophilized formulation showed a radiochemical purity bigger than 95%, besides being sterile and free of pyrogens. The biodistribution tests in athymic mice with induced tumors showed that the radiopharmaceutical was united mainly to the tumor and that this it was excreted mainly for renal via. (Author)

  15. Pre- and postirradiation hemopoietic effects of liposomal muramyl tripeptide phosphatidylethanolamine (MTP-PE) administered to C57Bl/6 mice before irradiation

    International Nuclear Information System (INIS)

    The effect was investigated of muramyl tripeptide (MTP-PE) encapsulated in liposomes (multilamellar vesicles, MLV) on hemopoiesis and hemopoietic recovery after sublethal single irradiation with a dose of 6 Gy. MTP-PE/MLV during 24 h after administration stimulated the monocyte/macrophage system and accelerated the recovery from radiation-induced damage of hemopoiesis. The reparative processes were accompanied by hyperplasia of granulocytes in the bone marrow on days 6-8 after irradiation, acceleration of erythropoiesis in the spleen and recovery of granulocyte count in the peripheral blood. The repair of thymic and splenic lymphoid tissues as well as recovery of the number of mononuclear cells in peripheral blood was not influenced by MTP-PE/MLV application. (author) 8 figs., 40 refs

  16. Asociación de índices infecciosos del virus de la fiebre aftosa a receptores celulares en el ganado criollo bon

    Directory of Open Access Journals (Sweden)

    Ariza Botero Manuel Fernando

    2006-06-01

    Full Text Available Los miembros de la familia de receptores celulares, integrinas (·V‚1, ·V‚3 ·V‚6, han sido identificados como factores de adhesión de diferentes virus a las células del hospedero. Para el caso del virus de la Fiebre Aftosa, estas integrinas se unen al sitio de reconocimiento celular en la secuencia tripéptidica Arg-Gly-Asp (RGD e igualmente están localizadas en la proteína VP1 del virus. Evidencia genética de esta interacción ha sido obtenida mediante la mutación de la secuencia RGD en clones infecciosos de ADNc, identificando partículas virales no infectivas incapaces de absorberse a células susceptibles. Cierta clase de resistencia genética natural al virus de la fiebre aftosa ha sido descrita en el ganado bovino Blanco Orejinegro, considerándose que cierto grado de esta resistencia podría ser causado por mutaciones en estos receptores celulares responsables por la adhesión del virus. Uno de los objetivos del estudio fue la identificación de genes candidatos (integrinas, específicamente marcadores moleculares asociados a genes  potencialmente responsables por la adhesión del virus aftoso a la célula hospedera. Un total de 106 individuos pertenecientes al núcleo de ganado criollo (banco de germoplasma bovino, Corpoica fueron genotipados con dos marcadores tipo microsatélite (ILS030 y BM2113 y dos SNPs (SLC11A1 y ITGB6 encontrados en el cromosoma 2 bovino. Estos genotipos fueron asociados a índices de infecciosidad, obtenidos a partir de la infección de cultivos BHK y de fibroblastos con los serotipos O1 Campos y A24 Cruzeiro, para cada uno de los individuos. Dos de los cuatro marcadores (ITGB6 e ILST030S se asociaron significativamente con el fenotipo Índice de Resistencia y mostraron un amplio efecto aditivo (p=0,025 y p=0,001 respectivamente. El presente estudio permitirá identificar aquellas líneas familiares que segregan aquellos alelos benéficos dentro de un programa de selección asistida por marcadores

  17. A cyclic-RGD-BioShuttle functionalized with TMZ by DARinv “Click Chemistry” targeted to αvβ3 integrin for therapy

    Directory of Open Access Journals (Sweden)

    Klaus Braun, Manfred Wiessler, Rüdiger Pipkorn, Volker Ehemann, Tobias Bäuerle, Heinz Fleischhacker, Gabriele Müller, Peter Lorenz, Waldemar Waldeck

    2010-01-01

    Full Text Available Clinical experiences often document, that a successful tumor control requires high doses of drug applications. It is widely believed that unavoidable adverse reactions could be minimized by using gene-therapeutic strategies protecting the tumor-surrounding healthy tissue as well as the bone-marrow. One new approach in this direction is the use of “Targeted Therapies” realizing a selective drug targeting to gain effectual amounts at the target site, even with drastically reduced application doses. MCF-7 breast cancer cells expressing the αvβ3 [alpha(vbeta(3] integrin receptor are considered as appropriate candidates for such a targeted therapy. The modularly composed BioShuttle carrier consisting of different units designed to facilitate the passage across the cell membranes and for subcellular addressing of diagnostic and/or therapeutic molecules could be considered as an eligible delivery platform. Here we used the cyclic RGD-BioShuttle as a carrier for temozolomide (TMZ at the αvβ3 integrin receptor realizing local TMZ concentrations sufficient for cell killing. The IC50 values are 12 µMol/L in the case of cRGD-BioShuttle-TMZ and 100 µMol/L for underivatized TMZ, which confirms the advantage of TMZ reformulation to realize local concentrations sufficient for cell killing.Our paper focuses on the design, synthesis and application of the cRGD-BioShuttle conjugate composed of the cyclic RGD, a αvβ3 integrin-ligand, ligated to the cytotoxic drug TMZ. The ligation was carried out by the Diels Alder Reaction with inverse electron demand (DARinv.

  18. A cyclic-RGD-BioShuttle functionalized with TMZ by DARinv “Click Chemistry” targeted to αvβ3 integrin for therapy

    Science.gov (United States)

    Braun, Klaus; Wiessler, Manfred; Pipkorn, Rüdiger; Ehemann, Volker; Bäuerle, Tobias; Fleischhacker, Heinz; Müller, Gabriele; Lorenz, Peter; Waldeck, Waldemar

    2010-01-01

    Clinical experiences often document, that a successful tumor control requires high doses of drug applications. It is widely believed that unavoidable adverse reactions could be minimized by using gene-therapeutic strategies protecting the tumor-surrounding healthy tissue as well as the bone-marrow. One new approach in this direction is the use of “Targeted Therapies” realizing a selective drug targeting to gain effectual amounts at the target site, even with drastically reduced application doses. MCF-7 breast cancer cells expressing the αvβ3 [alpha(v)beta(3)] integrin receptor are considered as appropriate candidates for such a targeted therapy. The modularly composed BioShuttle carrier consisting of different units designed to facilitate the passage across the cell membranes and for subcellular addressing of diagnostic and/or therapeutic molecules could be considered as an eligible delivery platform. Here we used the cyclic RGD-BioShuttle as a carrier for temozolomide (TMZ) at the αvβ3 integrin receptor realizing local TMZ concentrations sufficient for cell killing. The IC50 values are 12 µMol/L in the case of cRGD-BioShuttle-TMZ and 100 µMol/L for underivatized TMZ, which confirms the advantage of TMZ reformulation to realize local concentrations sufficient for cell killing. Our paper focuses on the design, synthesis and application of the cRGD-BioShuttle conjugate composed of the cyclic RGD, a αvβ3 integrin-ligand, ligated to the cytotoxic drug TMZ. The ligation was carried out by the Diels Alder Reaction with inverse electron demand (DARinv). PMID:20922134

  19. Differentiated adipose-derived stem cells act synergistically with RGD-modified surfaces to improve neurite outgrowth in a co-culture model.

    Science.gov (United States)

    de Luca, A C; Faroni, A; Downes, S; Terenghi, G

    2016-08-01

    Peripheral nerve damage is a problem encountered after trauma and during surgery and the development of synthetic polymer conduits may offer a promising alternative to autografts. In order to improve the performance of the polymer to be used for nerve conduits, poly-ε-caprolactone (PCL) films were chemically functionalized with RGD moieties, using a chemical reaction previously developed. In vitro cultures of dissociated dorsal root ganglion (DRG) neurons provide a valid model to study different factors affecting axonal growth. In this work, DRG neurons were cultured on RGD-functionalized PCL films. Adult adipose-derived stem cells differentiated to Schwann cells (dASCs) were initially cultured on the functionalized PCL films, resulting in improved attachment and proliferation. dASCs were also co-cultured with DRG neurons on treated and untreated PCL to assess stimulation by dASCs on neurite outgrowth. Neuron response was generally poor on untreated PCL films, but long neurites were observed in the presence of dASCs or RGD moieties. A combination of the two factors enhanced even further neurite outgrowth, acting synergistically. Finally, in order to better understand the extracellular matrix (ECM)-cell interaction, a β1 integrin blocking experiment was carried out. Neurite outgrowth was not affected by the specific antibody blocking, showing that β1 integrin function can be compensated by other molecules present on the cell membrane. Copyright © 2013 John Wiley & Sons, Ltd. PMID:23950058

  20. Preparation and Application of Tripeptide-cellulose Ester%三肽纤维素酯的制备及应用研究

    Institute of Scientific and Technical Information of China (English)

    李杨; 朱进科; 李连杰; 蒋登高

    2016-01-01

    Tripeptide-cellulose ester ( TPC) was synthesized by sequent acyl chlorination, esterification, peptide elongation and amino modification using microcrystalline cellulose as raw material and N-(9-fluorenylmethoxycarbonyl)-L-proline(FMOC-L-Pro-OH), N-(9-fluorenylmethoxycarbonyl)-L-valinamide-L-phenylalanine(FMOC-L-Val-L-Phe-OH), 3,5-dinitrobenzoic acid as derivatization reagents. The experimental results showed that during the synthesis process, esterification, a key step, was performed firstly, and followed by the extension of peptide chain. The effects of various experimental factors on esterification and substitution degree ( DS ) were investigated. The optimal condition was described as that the mass concentration of avicel in LiCl/DMAc 20 g/L, the molar ratio of —C(O)Cl to the hydroxyl content of cellulose 3∶1, reaction temperature 100℃ and reaction time 20 h. By the further peptide elongation and amino modification, the tripeptide-cellulose ester with DS 2. 15 was synthesized. The structure and properties were then characterized by using Fourier transform infrared spectroscopy, elemental analysis, X-ray diffraction and thermogravimetric analysis. The results showed that the modification of peptide by 3,5-dinitrobenzoic acid was successful and the synthesis process was feasible. With the alteration of precipitants, the intramolecular hydrogen bond was different and the obtained derivatives had different characteristics and functionalities. The tripeptide-cellulose ester precipitated in methanol was good enteric-coating material with good shading effect and rapid disintegration, but the physiological toxicity needed further study. The tripeptide-cellulose ester ( using water as the precipitant) could be coated onto silica gel to get a chiral stationary phase, which has a wider application than previous coated-cellulose chiral stationary phase. In addition, the stable chemical resistance of tripeptide-cellulose ester was particularly suitable for the

  1. Imaging integrin alpha-v-beta-3 expression in tumors with an 18F-labeled dimeric RGD peptide

    Science.gov (United States)

    Dijkgraaf, Ingrid; Terry, Samantha; McBride, William J.; Goldenberg, David M.; Laverman, Peter; Franssen, Gerben M.; Oyen, Wim J. G.; Boerman, Otto C.

    2014-01-01

    Integrin αvβ3 receptors are expressed on activated endothelial cells during neovascularization to maintain tumor growth. Many radiolabeled probes utilize the tight and specific association between the arginine-glycine-aspartatic acid (RGD) peptide and integrin αvβ3, but one main obstacle for any clinical application of these probes is the laborious multistep radiosynthesis of 18F. In this study, the dimeric RGD peptide, E-[c(RGDfK)]2, was conjugated with NODAGA and radiolabeled with 18F in a simple one-pot process with a radiolabeling yield of 20%; the whole process lasting only 45 min. NODAGA-E-[c(RGDfK)]2 labeled with 18F at a specific activity of 1.8 MBq/nmol and a radiochemical purity of 100% could be achieved. Log P value of 18F-labeled NODAGA-E-[c(RGDfK)]2 was −4.26 ± 0.02. In biodistribution studies, 18F-NODAGA-E-[c(RGDfK)]2 cleared rapidly from the blood with 0.03 ± 0.01 %ID/g in the blood at 2 h p.i., mainly via the kidneys and showed good in vivo stability. Tumor uptake of 18F-NODAGA-E-[c(RGDfK)]2 (3.44 ± 0.20 %ID/g, 2 h p.i.) was significantly lower than that of reference compounds 68Ga-labeled NODAGA-E-[c(RGDfK)]2 (6.26 ± 0.76 %ID/g; P <0.001) and 111In-labeled NODAGA-E-[c(RGDfK)]2 (4.99 ± 0.64 %ID/g; P < 0.01). Co-injection of an excess of unlabeled NODAGA-E-[c(RGDfK)]2 along with 18F-NODAGA-E-[c(RGDfK)]2 resulted in significantly reduced radioactivity concentrations in the tumor (0.85 ± 0.13 %ID/g). The αvβ3 integrin-expressing SK-RC-52 tumor could be successfully visualized by microPET with 18F-labeled NODAGA-E-[c(RGDfK)]2. In conclusion, NODAGA-E-[c(RGDfK)]2 could be labeled rapidly with 18F using a direct aqueous, one-pot method and it accumulated specifically in αvβ3 integrin-expressing SK-RC-52 tumors, allowing for visualization by microPET. PMID:23606427

  2. PET imaging of {alpha}{sub v}{beta}{sub 3} integrin expression in tumours with {sup 68}Ga-labelled mono-, di- and tetrameric RGD peptides

    Energy Technology Data Exchange (ETDEWEB)

    Dijkgraaf, Ingrid; Franssen, Gerben M.; Oyen, Wim J.G.; Boerman, Otto C. [Radboud University Nijmegen Medical Centre, Department of Nuclear Medicine, P.O. Box 9101, Nijmegen (Netherlands); Yim, Cheng-Bin [Radboud University Nijmegen Medical Centre, Department of Nuclear Medicine, P.O. Box 9101, Nijmegen (Netherlands); Utrecht University, Department of Medicinal Chemistry and Chemical Biology, Utrecht Institute for Pharmaceutical Sciences, Utrecht (Netherlands); Schuit, Robert C. [VU University Medical Centre, Department of Nuclear Medicine and PET Research, P.O. Box 7057, Amsterdam (Netherlands); Luurtsema, Gert [University Medical Center Groningen, Department of Nuclear Medicine and Molecular Imaging, Hanzeplein 1, P.O. Box 30.001, Groningen (Netherlands); Liu, Shuang [Purdue University, School of Health Sciences, West Lafayette, IN (United States)

    2011-01-15

    Due to the restricted expression of {alpha}{sub v}{beta}{sub 3} in tumours, {alpha}{sub v}{beta}{sub 3} is considered a suitable receptor for tumour targeting. In this study the {alpha}{sub v}{beta}{sub 3}-binding characteristics of {sup 68}Ga-labelled monomeric, dimeric and tetrameric RGD peptides were determined and compared with their {sup 111}In-labelled counterparts. A monomeric (E-c(RGDfK)), a dimeric (E-[c(RGDfK)]{sub 2}) and a tetrameric (E{l_brace}E[c(RGDfK)]{sub 2}{r_brace}{sub 2}) RGD peptide were synthesised, conjugated with DOTA and radiolabelled with {sup 68}Ga. In vitro {alpha}{sub v}{beta}{sub 3}-binding characteristics were determined in a competitive binding assay. In vivo {alpha}{sub v}{beta}{sub 3}-targeting characteristics of the compounds were assessed in mice with subcutaneously growing SK-RC-52 xenografts. In addition, microPET images were acquired using a microPET/CT scanner. The IC{sub 50} values for the Ga(III)-labelled DOTA-E-c(RGDfK), DOTA-E-[c(RGDfK)]{sub 2} and DOTA-E{l_brace}E[c(RGDfK)]{sub 2}{r_brace}{sub 2} were 23.9 {+-} 1.22, 8.99 {+-} 1.20 and 1.74 {+-} 1.18 nM, respectively, and were similar to those of the In(III)-labelled mono-, di- and tetrameric RGD peptides (26.6 {+-} 1.15, 3.34 {+-} 1.16 and 1.80 {+-} 1.37 nM, respectively). At 2 h post-injection, tumour uptake of the {sup 68}Ga-labelled mono-, di- and tetrameric RGD peptides (3.30 {+-} 0.30, 5.24 {+-} 0.27 and 7.11 {+-} 0.67%ID/g, respectively) was comparable to that of their {sup 111}In-labelled counterparts (2.70 {+-} 0.29, 5.61 {+-} 0.85 and 7.32 {+-} 2.45%ID/g, respectively). PET scans were in line with the biodistribution data. On all PET scans, the tumour could be clearly visualised. The integrin affinity and the tumour uptake followed the order of DOTA-tetramer > DOTA-dimer > DOTA-monomer. The {sup 68}Ga-labelled tetrameric RGD peptide has excellent characteristics for imaging of {alpha}{sub v} {beta}{sub 3} expression with PET. (orig.)

  3. Biologically and mechanically driven design of an RGD-mimetic macroporous foam for adipose tissue engineering applications.

    Science.gov (United States)

    Rossi, Eleonora; Gerges, Irini; Tocchio, Alessandro; Tamplenizza, Margherita; Aprile, Paola; Recordati, Camilla; Martello, Federico; Martin, Ivan; Milani, Paolo; Lenardi, Cristina

    2016-10-01

    Despite clinical treatments for adipose tissue defects, in particular breast tissue reconstruction, have certain grades of efficacy, many drawbacks are still affecting the long-term survival of new formed fat tissue. To overcome this problem, in the last decades, several scaffolding materials have been investigated in the field of adipose tissue engineering. However, a strategy able to recapitulate a suitable environment for adipose tissue reconstruction and maintenance is still missing. To address this need, we adopted a biologically and mechanically driven design to fabricate an RGD-mimetic poly(amidoamine) oligomer macroporous foam (OPAAF) for adipose tissue reconstruction. The scaffold was designed to fulfil three fundamental criteria: capability to induce cell adhesion and proliferation, support of in vivo vascularization and match of native tissue mechanical properties. Poly(amidoamine) oligomers were formed into soft scaffolds with hierarchical porosity through a combined free radical polymerization and foaming reaction. OPAAF is characterized by a high water uptake capacity, progressive degradation kinetics and ideal mechanical properties for adipose tissue reconstruction. OPAAF's ability to support cell adhesion, proliferation and adipogenesis was assessed in vitro using epithelial, fibroblast and endothelial cells (MDCK, 3T3L1 and HUVEC respectively). In addition, in vivo subcutaneous implantation in murine model highlighted OPAAF potential to support both adipogenesis and vessels infiltration. Overall, the reported results support the use of OPAAF as a scaffold for engineered adipose tissue construct. PMID:27428768

  4. RGD偶联吉西他滨白蛋白纳米粒对胰腺癌细胞增殖抑制作用的研究%Study on inhibitory effects of RGD-conjugated gemcitabine-loaded albumin nanoparticles on the proliferation of BxPC-3 cells in vitro

    Institute of Scientific and Technical Information of China (English)

    吉顺荣; 吴闻哲; 王浩; 张波; 刘辰; 龙江; 虞先濬; 倪泉兴; 徐近

    2011-01-01

    Objective To explore the inhibitory effects of RGD-conjugated gemcitabine (GEM)-loaded albumin nanoparticles (ANP) on the proliferation of BxPC-3 cells in vitro. Methods Human pancreatic carcinoma BxPC-3 cells were cultured and then treated 5 different combinations of drugs (BSANP controls, RGD-BSANP, BSANP-GEM, GEM and RGD-BSANP-GEM) respectively. The inhibition rate of BxPC-3 cells was detected by MTT. Results The gemcitabine-loaded albumin nanoparticles produced higher inhibitory effects than gemcitabine alone. The RGD-conjugated gemcitabine-loaded albumin nanoparticles produced even higher anti-proliferation rate than those without RGD conjugation. Conclusions RGD peptides can increase the anti-proliferation rate of gemcitabine-loaded albumin nanoparticles on BxPC-3 cells in vitro.%目的:通过体外试验,探讨RGD偶联吉西他滨白蛋白纳米粒(RGD-BSANP-GEM)对胰腺癌细胞株BxPC-3的体外增殖的抑制作用.方法:以人胰腺癌细胞株BxPC-3为研究对象,分为5个给药组:BSANP对照组、RGD-BSANP组、BSANP-GEM组、GEM原药组、RGD-BSANP-GEM组.运用MTT法检测给药后24h、48 h和72 h的增殖抑制率.结果:BSANP-GEM组细胞抑制率高于GEM组(P<0.05),RGD-BSANP-GEM组细胞抑制率高于BSANP-GEM组.同时RGD-BSANP-GEM组对胰腺癌细胞株的抑制率与给药浓度和作用时间呈正相关.结论:体外RGD环肽能够增加BSANP-GEM对胰腺癌细胞的增殖抑制作用.

  5. First hyperpolarizability of the natural aromatic amino acids tryptophan, tyrosine, and phenylalanine and the tripeptide lysine-tryptophan-lysine determined by hyper-Rayleigh scattering.

    Science.gov (United States)

    Duboisset, J; Matar, G; Russier-Antoine, I; Benichou, E; Bachelier, G; Jonin, Ch; Ficheux, D; Besson, F; Brevet, P F

    2010-11-01

    We report the first hyperpolarizability of tryptophan (Trp) and tyrosine (Tyr) and an upper limit for that of phenylalanine (Phe), three natural aromatic amino acids. The measurements were performed with hyper-Rayleigh scattering in an aqueous Tris buffer solution at a pH of 8.5 and 150 mM salt concentration with a fundamental wavelength of 780 nm. A value of (4.7 ± 0.7) × 10(-30) esu is found for Trp and (4.1 ± 0.7) × 10(-30) esu for Tyr whereas the upper limit of 1.4 × 10(-30) esu is found for that of Phe due to its limited solubility. The influence of the presence of lysine (Lys) in close vicinity of Trp is investigated with a measurement of the first hyperpolarizabilty of Trp in an excess of Lys and compared to the first hyperpolarizability obtained for the tripeptide Lys-Trp-Lys. The clear decrease of the values measured in these two cases indicates that the first hyperpolarizabilty of Trp is very sensitive to its local environment.

  6. Dipeptidomimetic ketomethylene isosteres as pro-moieties for drug transport via the human intestinal di-/tripeptide transporter hPEPT1

    DEFF Research Database (Denmark)

    Våbenø, Jon; Nielsen, Carsten Uhd; Ingebrigtsen, Truls;

    2004-01-01

    Five dipeptidomimetic-based model prodrugs containing ketomethylene amide bond replacements were synthesized from readily available alpha,beta-unsaturated gamma-ketoesters. The model drug (BnOH) was attached to the C-terminus or to one of the side chain positions of the dipeptidomimetic. The stab......Five dipeptidomimetic-based model prodrugs containing ketomethylene amide bond replacements were synthesized from readily available alpha,beta-unsaturated gamma-ketoesters. The model drug (BnOH) was attached to the C-terminus or to one of the side chain positions of the dipeptidomimetic....... The stability, the affinity for the di-/tripeptide transporter hPEPT1, and the transepithelial transport properties of the model prodrugs were investigated. ValPsi[COCH(2)]Asp(OBn) was the compound with highest chemical stability in buffers at pH 6.0 and 7.4, with half-lives of 190 and 43 h, respectively. All...... compounds, a finding that was corroborated by hPEPT1-mediated intracellular uptake. The results indicate that the stabilized Phe-Asp and Val-Asp derivatives are promising pro-moieties in a prodrug approach targeting hPEPT1....

  7. A tripeptide Diapin effectively lowers blood glucose levels in male type 2 diabetes mice by increasing blood levels of insulin and GLP-1.

    Directory of Open Access Journals (Sweden)

    Jifeng Zhang

    Full Text Available The prevalence of type 2 diabetes (T2D is rapidly increasing worldwide. Effective therapies, such as insulin and Glucagon-like peptide-1 (GLP-1, require injections, which are costly and result in less patient compliance. Here, we report the identification of a tripeptide with significant potential to treat T2D. The peptide, referred to as Diapin, is comprised of three natural L-amino acids, GlyGlyLeu. Glucose tolerance tests showed that oral administration of Diapin effectively lowered blood glucose after oral glucose loading in both normal C57BL/6J mice and T2D mouse models, including KKay, db/db, ob/ob mice, and high fat diet-induced obesity/T2D mice. In addition, Diapin treatment significantly reduced casual blood glucose in KKay diabetic mice in a time-dependent manner without causing hypoglycemia. Furthermore, we found that plasma GLP-1 and insulin levels in diabetic models were significantly increased with Diapin treatment compared to that in the controls. In summary, our findings establish that a peptide with minimum of three amino acids can improve glucose homeostasis and Diapin shows promise as a novel pharmaceutical agent to treat patients with T2D through its dual effects on GLP-1 and insulin secretion.

  8. Radiolabeled cyclic arginine-glycine-aspartic (RGD)-conjugated iron oxide nanoparticles as single-photon emission computed tomography (SPECT) and magnetic resonance imaging (MRI) dual-modality agents for imaging of breast cancer

    Energy Technology Data Exchange (ETDEWEB)

    Deng, Shengming; Zhang, Wei; Zhang, Bin, E-mail: zbnuclmd@126.com [The First Affiliated Hospital of Soochow University, Department of Nuclear Medicine (China); Hong, Ruoyu [Soochow University, College of Chemistry, Chemical Engineering and Materials Science & Key Laboratory of Organic Synthesis of Jiangsu Province (China); Chen, Qing; Dong, Jiajia [The First Affiliated Hospital of Soochow University, Department of Nuclear Medicine (China); Chen, Yinyiin [The First Affiliated Hospital of Soochow University, Department of Radiology (China); Chen, Zhiqiang; Wu, Yiwei, E-mail: wuyiwei3988@gmail.com [The First Affiliated Hospital of Soochow University, Department of Nuclear Medicine (China)

    2015-01-15

    Ultrasmall superparamagnetic iron oxide nanoparticles (USPIOs) modified with a novel cyclic arginine-glycine-aspartate (RGD) peptide were made and radiolabeled as single-photon emission computed tomography (SPECT) and magnetic resonance imaging (MRI) dual-modality agents for imaging of breast cancer. The probe was tested both in vitro and in vivo to determine its receptor targeting efficacy and feasibility for SPECT and MRI. The radiochemical syntheses of {sup 125}I-cRGD-USPIO were accomplished with a radiochemical purity of 96.05 ± 0.33 %. High radiochemical stability was found in fresh human serum and in phosphate-buffered saline. The average hydrodynamic size of {sup 125}I-cRGD-USPIO determined by dynamic light scattering was 51.3 nm. Results of in vitro experiments verified the specificity of the radiolabeled nanoparticles to tumor cells. Preliminary biodistribution studies of {sup 125}I-radiolabeled cRGD-USPIO in Bcap37-bearing nude mice showed that it had long circulation half-life, high tumor uptake, and high initial blood retention with moderate liver uptake. In vivo tumor targeting and uptake of the radiolabeled nanoparticles in mice model were visualized by SPECT and MRI collected at different time points. Our results strongly indicated that the {sup 125}I-cRGD-USPIO could be used as a promising bifunctional radiotracer for early clinical tumor detection with high sensitivity and high spatial resolution by SPECT and MRI.

  9. Radiolabeled cyclic arginine-glycine-aspartic (RGD)-conjugated iron oxide nanoparticles as single-photon emission computed tomography (SPECT) and magnetic resonance imaging (MRI) dual-modality agents for imaging of breast cancer

    International Nuclear Information System (INIS)

    Ultrasmall superparamagnetic iron oxide nanoparticles (USPIOs) modified with a novel cyclic arginine-glycine-aspartate (RGD) peptide were made and radiolabeled as single-photon emission computed tomography (SPECT) and magnetic resonance imaging (MRI) dual-modality agents for imaging of breast cancer. The probe was tested both in vitro and in vivo to determine its receptor targeting efficacy and feasibility for SPECT and MRI. The radiochemical syntheses of 125I-cRGD-USPIO were accomplished with a radiochemical purity of 96.05 ± 0.33 %. High radiochemical stability was found in fresh human serum and in phosphate-buffered saline. The average hydrodynamic size of 125I-cRGD-USPIO determined by dynamic light scattering was 51.3 nm. Results of in vitro experiments verified the specificity of the radiolabeled nanoparticles to tumor cells. Preliminary biodistribution studies of 125I-radiolabeled cRGD-USPIO in Bcap37-bearing nude mice showed that it had long circulation half-life, high tumor uptake, and high initial blood retention with moderate liver uptake. In vivo tumor targeting and uptake of the radiolabeled nanoparticles in mice model were visualized by SPECT and MRI collected at different time points. Our results strongly indicated that the 125I-cRGD-USPIO could be used as a promising bifunctional radiotracer for early clinical tumor detection with high sensitivity and high spatial resolution by SPECT and MRI

  10. Biomineralization of the Surface of PLGA-(ASP-PEG) Modified with the K_(16) and RGD-containing Peptide

    Institute of Scientific and Technical Information of China (English)

    HAO Jie; HU Zhenming; ZHENG Qixin; GUO Xiaodong; SONG Yulin

    2009-01-01

    K_(16) and RGD-containing peptide was used to modify the surface of three-dimensional PLGA-(ASP-PEG)matrix,then the modified PLGA-(ASP-PEG)was incubated in modified simulated body fluid(SBF).The biomineralization of the modified PLGA-(ASP-PEG)was explored,and the peptide was synthesized with solid phase synthesis technology and linked cova-lently to PLGA-(ASP-PEG)through cross-linker(Sulfo-LC-SPDP),which was characterized with XPS.The modified PLGA-(ASP-PEG)(Experiment group,EG)and PLGA-(ASP-PEG)(Control group,CG)were all incubated into SBF for 10 d,and the growth of hydroxyapatite(HA)nanocrys-tals was confirmed with XRD,EDS and SEM.HPLC shows that peptide purity is 94.13%,while MS analysis shows that molecular value of peptide is 2741.26.Binding energy of the sulphur in EG was 164 eV is detected by XPS,and the ratio of carbon and sulphur is 99.746:0.1014.SEM analysis demonstrates the better growth of bonelike HA nanocrystals in EG than that in CG.The component of mineral in EG consisted mainly of hydroxyapatite containing low crystalline nanocrystals,and the Ca/P ratio is about 1.60,which is similar to that of natural bone,while the Ca/P ratio in CG is 1.52. PLGA-(ASP-PEG)modified with peptide provided enough functional groups for biomineralization, and possessed the bonelike structure.

  11. A comparative study of polyethylene glycol hydrogels derivatized with the RGD peptide and the cell-binding domain of fibronectin.

    Science.gov (United States)

    Zhang, Chen; Hekmatfer, Sogol; Karuri, Nancy W

    2014-01-01

    The goal of our study was to compare the biological responses of cells cultured on polyethylene glycol (PEG) hydrogels functionalized with varying concentrations of the widely used adhesion peptide, RGD, and the cell-binding domain of fibronectin, III(9-10). We used Michael addition chemistry to covalently link cysteines in GRGDSPC and glutathione S-transferase (GST) tagged III(9-10) (GST-III(9-10)), to the acrylate groups in PEG diacrylate (PEGDA). Conjugation of GST-III(9-10) to PEGDA occurred through cysteine residues in GST. Ellman's reagent and immunoblotting studies demonstrated an efficiency of 90% or more for PEG conjugation of 1 μM GST-III(9-10) or GRDGSPC in 10% (wt/vol) PEGDA at 37°C for 1 h. Circular dichroism and limited proteolysis demonstrated that conjugating PEGDA to GST-III(9-10) did not significantly perturb its native secondary structure. Sodium dodecyl sulfate polyacrylamide gel electrophoresis characterization of the wash solution of PEG hydrogels after photopolymerization demonstrated that >95% of the 1 μM GST-III(9-10) was incorporated into the PEG hydrogels after cross-linking. PEG hydrogels derivatized with 1 μM GST-III(9-10) had significantly higher cell adhesion and spreading than PEG hydrogels with 1 μM GRGDSPC. A comparable adhesion response between GRGDSPC and GST-III(9-10) was obtained when the former was at millimolar and the latter at micromolar concentration. The amount and type of conjugate in the PEG hydrogel derivative was statistically more significant than hydrogel rigidity in stimulating the biological responses observed. This report presents new evidence of the robustness of III(9-10) in mediating cell adhesion and spreading on PEG hydrogels.

  12. Synthesis of the Biomimetic Polymer: Aliphatic Diamine and RGDS Modified Poly(d,l-lactic acid)

    Institute of Scientific and Technical Information of China (English)

    Xu Feng NIU; Yuan Liang WANG; Yan Feng LUO; Jun PAN; Juan Fang SHANG; Li Xia GUO

    2005-01-01

    A novel poly(d,l-lactic acid) (PDLLA) based biomimetic polymer was synthesized by grafting maleic anhydride, butanediamine and arg-gly-asp-ser (RGDS) peptides onto the backbone of PDLLA, aiming to overcome the acidity and auto-accelerating degradation of PDLLA during degradation and to improve its biospecificity and biocompatibility. The synthetic copolymer was characterized by FTIR, 13C NMR and amino acid analyzer (AAA).

  13. The replication of human immunodeficiency virus type 1 in macrophages is enhanced after phagocytosis of apoptotic cells

    OpenAIRE

    Lima, Rosangela G; Van Weyenbergh, Johan; Saraiva, Elvira M. B.; Barral-Netto, Manoel; Galvão-Castro, Bernardo; Bou-Habib, Dumith Chequer

    2002-01-01

    Clearance of apoptotic cells increases macrophage secretion of antiinflammatory mediators and might modulate viral replication in human immunodeficiency virus (HIV) type 1-infected macrophages. To study this, primary macrophages were infected with HIV-1 and exposed to apoptotic cells. It was found that phagocytosis of apoptotic cells potently enhanced HIV-1 growth. The peptide Arg-Gly-Asp-Ser, which binds to integrin receptors, inhibited the uptake of apoptotic cells and the subsequent enhanc...

  14. Variants of the cell recognition site of fibronectin that retain attachment-promoting activity

    OpenAIRE

    1985-01-01

    A tetrapeptide sequence, Arg-Gly-Asp-Ser, is the minimal structure recognized by cells in the large, adhesive glycoprotein fibronectin. We now have defined the structural requirements for this cell recognition site by testing several synthetic variants of the active tetrapeptide sequence. The conservative substitutions of lysine for arginine, alanine for glycine, or glutamic acid for aspartic acid each resulted in abrogation of the cell attachment-promoting activity characteristic of the natu...

  15. Biodistribution und Analyse von 68Ga-markierten RGD-Multimeren an tumortragenden Nacktmäusen mit Hilfe der Kleintier-Positronen-Emissions-Tomographie

    OpenAIRE

    Wrubel, Scarlett

    2012-01-01

    Hintergrund und Ziele Die Tumorangiogenese spielt für die Früherkennung von Tumoren und deren Metastasen eine wichtige Rolle. Während dieses Vorgangs sind Funktions-proteine, speziell Integrine, hoch exprimiert und stellen daher ein molekulares Target für die Bildgebung von Tumorgefäßen dar. Als hoch-affine Moleküle, die an die Integrine binden, sind seit langem cyclische Pentapeptide bekannt, die die RGD-Sequenz tragen; diese eignen sich als DOTA-Konjugate für die Radio-markierung mit dem Po...

  16. Noninvasive visualization and quantification of tumor {alpha}{sub V{beta}3} integrin expression using a novel positron emission tomography probe, {sup 64}Cu-cyclam-RAFT-c(-RGDfK-){sub 4}

    Energy Technology Data Exchange (ETDEWEB)

    Jin, Zhao-Hui [Molecular Imaging Center, National Institute of Radiological Sciences, Chiba 263-8555 (Japan); Furukawa, Takako, E-mail: tfuru@nirs.go.j [Molecular Imaging Center, National Institute of Radiological Sciences, Chiba 263-8555 (Japan); Biomedical Imaging Research Center, University of Fukui, Fukui 910-1193 (Japan); Galibert, Mathieu; Boturyn, Didier [Departement de Chimie Moleculaire, UMR 5250, CNRS-Universite Joseph Fourier, 38041 Grenoble Cedex 9 (France); Coll, Jean-Luc [INSERM U823, Institut Albert Bonniot and Universite Joseph Fourier, 38706 La Tronche Cedex, Grenoble (France); Fukumura, Toshimitsu; Saga, Tsuneo [Molecular Imaging Center, National Institute of Radiological Sciences, Chiba 263-8555 (Japan); Dumy, Pascal [Departement de Chimie Moleculaire, UMR 5250, CNRS-Universite Joseph Fourier, 38041 Grenoble Cedex 9 (France); Fujibayashi, Yasuhisa [Molecular Imaging Center, National Institute of Radiological Sciences, Chiba 263-8555 (Japan); Biomedical Imaging Research Center, University of Fukui, Fukui 910-1193 (Japan)

    2011-05-15

    Introduction: The {alpha}{sub V{beta}3} integrin is a well-known transmembrane receptor involved in tumor invasion, angiogenesis and metastasis. Our aim was to evaluate a novel positron emission tomography (PET) probe, {sup 64}Cu-cyclam-RAFT-c(-RGDfK-){sub 4}, for noninvasive visualization and quantification of {alpha}{sub V{beta}3} integrin expression. Methods: RAFT-c(-RGDfK-){sub 4}, a tetrameric cyclic Arg-Gly-Asp (RGD)-based peptide, was conjugated with a bifunctional chelator, 1,4,8,11-tetraazacyclotetradecane (cyclam), radiolabeled with the positron emitter {sup 64}Cu and evaluated in vitro by cell binding and competitive inhibition assays and in vivo by biodistribution and receptor blocking studies, and PET imaging. The following cell lines, human embryonic kidney HEK293({beta}{sub 1}) [{alpha}{sub V{beta}3}-negative] and HEK293({beta}{sub 3}) [{alpha}{sub V{beta}3}-overexpressing] and human glioblastoma U87MG [naturally expressing {alpha}{sub V{beta}3}], together with their subcutaneous xenografts in athymic nude mice, were used for the present study. The expression levels of {alpha}{sub V{beta}3} on these cell lines and tumor xenografts were analyzed by flow cytometry and sodium dodecyl sulfate-polyacrylamide gel electrophoresis/autoradiography, respectively. Results: {sup 64}Cu-cyclam-RAFT-c(-RGDfK-){sub 4} demonstrated the in vitro and in vivo specificity for the {alpha}{sub V{beta}3} integrin and displayed rapid blood clearance, predominantly renal excretion and low uptake in nontumor tissues. Tumor uptake of {sup 64}Cu-cyclam-RAFT-c(-RGDfK-){sub 4} (3 h postinjection) in HEK293({beta}{sub 3}) (high levels of {alpha}{sub V{beta}3}), U87MG (moderate levels of {alpha}{sub V{beta}3}) and HEK293({beta}{sub 1}) (undetectable levels of {alpha}{sub V{beta}3}) tumors was 9.35%{+-}1.19%, 3.46%{+-}0.45% and 1.18%{+-}0.30% injected dose per gram, respectively, with a strong and positive correlation with the tumor {alpha}{sub V{beta}3} expression levels

  17. {alpha}{sub v}{beta}{sub 3} imaging can accurately distinguish between mature teratoma and necrosis in {sup 18}F-FDG-negative residual masses after treatment of non-seminomatous testicular cancer: a preclinical study

    Energy Technology Data Exchange (ETDEWEB)

    Aide, Nicolas [Francois Baclesse Cancer Centre and Caen University, Bioticla Team, EA1772, IFR 146 ICORE, GRECAN, Caen (France); Caen University Hospital and Francois Baclesse Cancer Centre, PET Unit, Caen (France); Centre Francois Baclesse, Service de Medecine Nucleaire, Caen (France); Briand, Melanie; Dutoit, Soizic; Deslandes, Edwiges; Poulain, Laurent [Francois Baclesse Cancer Centre and Caen University, Bioticla Team, EA1772, IFR 146 ICORE, GRECAN, Caen (France); Bohn, Pierre; Rouvet, Jean; Modzelewski, Romain; Vera, Pierre [Henri Becquerel Cancer Center and Rouen University Hospital and QuantIF- LITIS (EA4108), Department of Nuclear Medicine, Rouen (France); Lasnon, Charline [Caen University Hospital and Francois Baclesse Cancer Centre, PET Unit, Caen (France); Chasle, Jacques [Francois Baclesse Cancer Centre and Caen University, Pathology Department, Caen (France); Vela, Antony [Francois Baclesse Cancer Centre and Caen University, Radiophysics Unit, Caen (France); Carreiras, Franck [Universite de Cergy Pontoise, UFR Sciences et Techniques, ERRMECe, EA 1391, Institut des materiaux, Cergy-Pontoise (France)

    2011-02-15

    We assessed whether imaging {alpha}{sub v}{beta}{sub 3} integrin could distinguish mature teratoma from necrosis in human non-seminomatous germ cell tumour (NSGCT) post-chemotherapy residual masses. Human embryonal carcinoma xenografts (six/rat) were untreated (controls) or treated to form mature teratomas with low-dose cisplatin and all-trans retinoic acid (ATRA) over a period of 8 weeks. In another group, necrosis was induced in xenografts with high-dose cisplatin plus etoposide (two cycles).{sup 18}F-Fluorodeoxyglucose ({sup 18}F-FDG) small animal positron emission tomography (SA PET) imaging was performed in three rats (one control and two treated for 4 and 8 weeks with cisplatin+ATRA). Imaging of {alpha}{sub v}{beta}{sub 3} expression was performed in six rats bearing mature teratomas and two rats with necrotic lesions on a microSPECT/CT device after injection of the tracer [{sup 99m}Tc]HYNIC-RGD [6-hydrazinonicotinic acid conjugated to cyclo(Arg-Gly-Asp-D-Phe-Lys)]. Correlative immunohistochemistry studies of human and mouse {alpha}{sub v}{beta}{sub 3} expression were performed. Cisplatin+ATRA induced differentiation of the xenografts. After 8 weeks, some glandular structures and mesenchymal cells were visible; in contrast, control tumours showed undifferentiated tissues. SA PET imaging showed that mature teratoma had very low avidity for {sup 18}F-FDG [mean standardised uptake value (SUV{sub mean}) = 0.48 {+-} 0.05] compared to untreated embryonal carcinoma (SUV{sub mean} = 0.92 {+-} 0.13) (p = 0.005). {alpha}{sub v}{beta}{sub 3} imaging accurately distinguished mature teratoma (tumour to muscle ratio = 4.29 {+-} 1.57) from necrosis (tumour to muscle ratio = 1.3 {+-} 0.26) (p = 0.0002). Immunohistochemistry studies showed that {alpha}{sub v}{beta}{sub 3} integrin expression was strong in the glandular structures of mature teratoma lesions and negative in host stroma. Imaging {alpha}{sub v}{beta}{sub 3} integrin accurately distinguished mature teratoma from

  18. Preparation and Characterization of Drug-Loaded,RGD Peptide-Modified and Reconstituted High Density Lipoprotein Nanoparticles and Their Uptake by Tumor Cells%RGD肽修饰重组高密度脂蛋白载药纳米粒的制备和表征以及肿瘤细胞对其摄取作用

    Institute of Scientific and Technical Information of China (English)

    周欣; 王伟; 王玉; 冯美卿; 丁杨; 刘聪燕; 周建平

    2014-01-01

    目的:制备和表征RGD肽修饰的重组高密度脂蛋白(RGD-rHDL)载药纳米粒,考察肿瘤细胞对其摄取作用.方法:合成RGD与载脂蛋白A I (ApoA I)的偶联物(RGD-ApoA I),并以藤黄酸(GA)作为模型药物,采用薄膜分散法制得GA脂质体(LP-GA),再将RGD-ApoA I与LP-GA共孵育,制备载有GA的RGD-rHDL纳米粒(RGD-rHDL-GA);随后,对RGD-rHDL-GA进行表征,且采用荧光标记示踪法,通过RGD-rHDL-香豆素-6来考察人肝癌细胞HepG2对载药RGD-rHDL纳米粒的摄取作用.结果:制备的RGD-rHDL-GA呈现规则圆整的类球形,粒径分布均一[(110.70±3.25)nm],Zeta电位为(-39.21±0.10)mV,包封率为(92.20±0.28)%,载药量为(9.03±0.75)%,且其体外释药缓慢,稳定性良好;RGD-rHDL-香豆素-6的肿瘤细胞摄取率明显高于rHDL-香豆素-6.结论:rHDL经RGD修饰后可有效促进所载药物进入肿瘤细胞,提高其肿瘤靶向性.

  19. Tripeptide amide L-pyroglutamyl-histidyl-L-prolineamide (L-PHP-thyrotropin-releasing hormone, TRH) promotes insulin-producing cell proliferation.

    Science.gov (United States)

    Luo, LuGuang; Luo, John Z Q; Jackson, Ivor

    2013-02-01

    A very small tripeptide amide L-pyroglutamyl-L-histidyl-L-prolineamide (L-PHP, Thyrotropin-Releasing Hormone, TRH), was first identified in the brain hypothalamus area. Further studies found that L-PHP was expressed in pancreas. The biological role of pancreatic L-PHP is still not clear. Growing evidence indicates that L-PHP expression in the pancreas may play a pivotal role for pancreatic development in the early prenatal period. However, the role of L-PHP in adult pancreas still needs to be explored. L-PHP activation of pancreatic β cell Ca2+ flow and stimulation of β-cell insulin synthesis and release suggest that L-PHP involved in glucose metabolism may directly act on the β cell separate from any effects via the central nervous system (CNS). Knockout L-PHP animal models have shown that loss of L-PHP expression causes hyperglycemia, which cannot be reversed by administration of thyroid hormone, suggesting that the absence of L-PHP itself is the cause. L-PHP receptor type-1 has been identified in pancreas which provides a possibility for L-PHP autocrine and paracrine regulation in pancreatic function. During pancreatic damage in adult pancreas, L-PHP may protect beta cell from apoptosis and initiate its regeneration through signal pathways of growth hormone in β cells. L-PHP has recently been discovered to affect a broad array of gene expression in the pancreas including growth factor genes. Signal pathways linked between L-PHP and EGF receptor phosphorylation suggest that L-PHP may be an important factor for adult β-cell regeneration, which could involve adult stem cell differentiation. These effects suggest that L-PHP may benefit pancreatic β cells and diabetic therapy in clinic.

  20. A vacuolar membrane protein affects drastically the biosynthesis of the ACV tripeptide and the beta-lactam pathway of Penicillium chrysogenum.

    Science.gov (United States)

    Fernández-Aguado, Marta; Teijeira, Fernando; Martín, Juan F; Ullán, Ricardo V

    2013-01-01

    The knowledge about enzymes' compartmentalization and transport processes involved in the penicillin biosynthesis in Penicillium chrysogenum is very limited. The genome of this fungus contains multiple genes encoding transporter proteins, but very little is known about them. A bioinformatic search was made to find major facilitator supefamily (MFS) membrane proteins related to CefP transporter protein involved in the entry of isopenicillin N to the peroxisome in Acremonium chrysogenum. No strict homologue of CefP was observed in P. chrysogenum, but the penV gene was found to encode a membrane protein that contained 10 clear transmembrane spanners and two other motifs COG5594 and DUF221, typical of membrane proteins. RNAi-mediated silencing of penV gene provoked a drastic reduction of the production of the δ-(L-α-aminoadipyl-L-cysteinyl-D-valine) (ACV) and isopenicillin N intermediates and the final product of the pathway. RT-PCR and northern blot analyses confirmed a reduction in the expression levels of the pcbC and penDE biosynthetic genes, whereas that of the pcbAB gene increased. Localization studies by fluorescent laser scanning microscopy using Dsred and GFP fluorescent fusion proteins and the FM 4-64 fluorescent dye showed clearly that the protein was located in the vacuolar membrane. These results indicate that PenV participates in the first stage of the beta-lactam biosynthesis (i.e., the formation of the ACV tripeptide), probably taking part in the supply of amino acids from the vacuolar lumen to the vacuole-anchored ACV synthetase. This is in agreement with several reports on the localization of the ACV synthetase and provides increased evidence for a compartmentalized storage of precursor amino acids for non-ribosomal peptides. PenV is the first MFS transporter of P. chrysogenum linked to the beta-lactam biosynthesis that has been located in the vacuolar membrane.

  1. Motifs in the C-terminal region of the Penicillium chrysogenum ACV synthetase are essential for valine epimerization and processivity of tripeptide formation.

    Science.gov (United States)

    Wu, Xiaobin; García-Estrada, Carlos; Vaca, Inmaculada; Martín, Juan-Francisco

    2012-02-01

    The first step in the penicillin biosynthetic pathway is the non-ribosomal condensation of L-α-aminoadipic acid, L-cysteine and L-valine into the tripeptide δ-(L-α-aminoadipyl)-L-cysteinyl-D-valine (ACV). This reaction is catalysed by the multienzyme ACV synthetase (ACVS), which is encoded in the filamentous fungus Penicillium chrysogenum by the pcbAB gene. This enzyme contains at least ten catalytic domains. The precise role of the C-terminal domain of this multidomain NRPS still remains obscure. The C-terminal region of ACVS bears the epimerase and the thioesterase domains and may be involved in the epimerization of LLL-ACV to LLD-ACV and in the hydrolysis of the thioester bond. In this work, the conserved motifs (3371)EGHGRE(3376) (located in the putative epimerase domain) and (3629)GWSFG(3633) (located in the thioesterase domain) were changed by site-directed-mutagenesis to LGFGLL and GWAFG, respectively. In addition, the whole thioesterase domain (230 amino acids) and the different parts of this domain were deleted. The activity of these mutant enzymes was assessed in vivo by two different procedures: i) through the quantification of bisACV produced by the fungus and ii) by quantifying the benzylpenicillin production using tailored strains of P. chrysogenum, which lack the pcbAB gene, as host strains. All indicated mutant enzymes showed lower or null activity than the control strain confirming that E3371, H3373, R3375 and E3376 belong to the epimerase active centre. Different fragments included in the C-terminal region of ACVS control thioester hydrolysis. Overexpression of the sequence encoding the ACVS integrated thioesterase domain as a separate (stand-alone) transcriptional unit complemented mutants lacking the integrated thioesterase domain, although with low ACV releasing activity, suggesting that the stand-alone thioesterease interacts with the other ACVS domains.

  2. Utilization of a novel electrochemical {sup 90}Sr/{sup 90}Y generator for the preparation of {sup 90}Y-labeled RGD peptide dimer in clinically relevant dose

    Energy Technology Data Exchange (ETDEWEB)

    Chakraborty, Sudipta; Chakravarty, Rubel; Pillai, Maroor Raghavan Ambikalmajan; Dash, Ashutosh [Bhabha Atomic Research Centre, Mumbai (India). Radiopharmaceuticals Div.; Sarma, Haladhar Dev [Bhabha Atomic Research Centre, Mumbai (India). Radiation Biology and Health Sciences Div.

    2014-09-01

    The work reported in this paper provides a systematic study towards the development of an optimized strategy for preparation of a clinically relevant dose of {sup 90}Y-labeled dimeric RGD peptide derivative, DOTA-E[c(RGDfK)]{sub 2} [DOTA-(RGD){sub 2}] for in vivo targeted therapy utilizing {sup 90}Y obtained from a novel electrochemical {sup 90}Sr/{sup 90}Y generator. The performance of the generator was evaluated to ensure its suitability for providing {sup 90}Y in adequate quantity and purity required for formulation of clinically relevant dose for PRRT. {sup 90}Y-DOTA-(RGD){sub 2} was synthesized in high yield (86.2 ± 2.5%) and radiochemical purity (98.4 ± 0.5%) using clinically relevant dose (∝ 3.8 GBq) of {sup 90}Y. In vitro stability studies revealed that the radiolabeled conjugate retained its radiochemical purity in normal saline and human serum. Preliminary biodistribution studies carried out in C57/BL6 mice bearing melanoma tumors showed that the preparation exhibited significant tumor uptake (5.30 ± 0.78% of injected activity at 30 min post-injection) with good tumor to background ratio. The optimized radiolabeling protocol seems to be an attractive strategy which is largely viewed as a springboard to realize scope of developing {sup 90}Y labeled cyclic RGD peptides for targeted therapy of tumors over-expressing integrin-α{sub ν}β{sub 3} receptors. (orig.)

  3. MicroPET/CT imaging of {alpha}{sub v}{beta}{sub 3} integrin via a novel {sup 68}Ga-NOTA-RGD peptidomimetic conjugate in rat myocardial infarction

    Energy Technology Data Exchange (ETDEWEB)

    Menichetti, Luca; Kusmic, Claudia; Panetta, Daniele; Petroni, Debora; Salvadori, Piero A. [CNR-Institute of Clinical Physiology (IFC), Pisa (Italy); Arosio, Daniela; Manzoni, Leonardo [CNR-Institute of Molecular Science and Technologies (ISTM), Milan (Italy); Matteucci, Marco [Scuola Superiore Sant' Anna, Pisa (Italy); Casagrande, Cesare [University of Milan, Department of Chemistry, Milan (Italy); L' Abbate, Antonio [CNR-Institute of Clinical Physiology (IFC), Pisa (Italy); Scuola Superiore Sant' Anna, Pisa (Italy)

    2013-08-15

    The {alpha}{sub v}{beta}{sub 3} integrin is expressed in angiogenic vessels and is a potential target for molecular imaging of evolving pathological processes. Its expression is upregulated in cancer lesions and metastases as well as in acute myocardial infarction (MI) as part of the infarct healing process. The purpose of our study was to determine the feasibility of a new imaging approach with a novel {sup 68}Ga-2,2',2''-(1,4,7-triazonane-1,4,7-triyl)triacetic acid (NOTA)-arginine-glycine-aspartic acid (RGD) construct to assess integrin expression in the evolving MI. A straightforward labelling chemistry to attach the radionuclide {sup 68}Ga to a NOTA-based chelating agent conjugated with a cyclic RGD peptidomimetic is described. Affinity for {alpha}{sub v}{beta}{sub 3} integrin was assessed by in vitro receptor binding assay. The proof-of-concept in vivo studies combined the {sup 68}Ga-NOTA-RGD with the flow tracer {sup 13}N-NH{sub 3} imaging in order to obtain positron emission tomography (PET)/CT imaging of both integrin expression and perfusion defect at 4 weeks after infarction. Hearts were then processed for immunostaining of integrin {beta}{sub 3}. NOTA-RGD conjugate displayed a binding affinity for {alpha}{sub v}{beta}{sub 3} integrin of 27.9 {+-} 6.8 nM. {sup 68}Ga-NOTA-RGD showed stability without detectable degradation or formation of by-products in urine up to 2 h following injection in the rat. MI hearts exhibited {sup 68}Ga-NOTA-RGD uptake in correspondence to infarcted and border zone regions. The tracer signal drew a parallel with vascular remodelling due to ischaemia-induced angiogenesis as assessed by immunohistochemistry. As compared to similar imaging approaches using the {sup 18}F-galacto-derivative, we documented for the first time with microPET/CT imaging the {sup 68}Ga-NOTA-RGD derivative that appears eligible for PET imaging in animal models of vascular remodelling during evolving MI. The simple chemistry employed to

  4. Separation of N-derivatized di- and tri-peptide stereoisomers by micro-liquid chromatography using a quinidine-based monolithic column - Analysis of l-carnosine in dietary supplements.

    Science.gov (United States)

    Wang, Qiqin; Sánchez-López, Elena; Han, Hai; Wu, Huihui; Zhu, Peijie; Crommen, Jacques; Marina, Maria Luisa; Jiang, Zhengjin

    2016-01-01

    In the present study, a new analytical methodology was developed enabling the enantiomeric determination of N-derivatized di- and tri-peptides in dietary supplements using chiral micro-LC on a monolithic column consisting of poly(O-9-[2-(methacryloyloxy)-ethylcarbamoyl]-10,11-dihydroquinidine-co-2-hydroxyethyl methacrylate-co-ethylene dimethacrylate) (poly(MQD-co-HEMA-co-EDMA)). After optimization of the mobile phase conditions, a baseline resolution of the stereoisomers of 24 out of 53 N-derivatized di- and tri-peptides was obtained. 3,5-Dinitrobenzoyl- and 3,5-dichlorobenzoyl-peptide stereoisomers were separated with exceptionally high selectivity and resolution. The monolithic column was then applied to the quantitative analysis of l-carnosine and its enantiomeric impurity in three different commercial dietary supplements. Method validation demonstrated satisfactory results in terms of linearity, precision, selectivity, accuracy and limits of detection and quantification. The determined amounts of l-carnosine in commercial formulations were in agreement with the labeled content for all analyzed samples, and the enantiomeric impurity was found to be below the limit of detection (LOD), showing the potential of the poly(MQD-co-HEMA-co-EDMA) monolithic column as a reliable tool for the quality control of l-carnosine in dietary supplements by micro-LC.

  5. Separation of N-derivatized di- and tri-peptide stereoisomers by micro-liquid chromatography using a quinidine-based monolithic column - Analysis of l-carnosine in dietary supplements.

    Science.gov (United States)

    Wang, Qiqin; Sánchez-López, Elena; Han, Hai; Wu, Huihui; Zhu, Peijie; Crommen, Jacques; Marina, Maria Luisa; Jiang, Zhengjin

    2016-01-01

    In the present study, a new analytical methodology was developed enabling the enantiomeric determination of N-derivatized di- and tri-peptides in dietary supplements using chiral micro-LC on a monolithic column consisting of poly(O-9-[2-(methacryloyloxy)-ethylcarbamoyl]-10,11-dihydroquinidine-co-2-hydroxyethyl methacrylate-co-ethylene dimethacrylate) (poly(MQD-co-HEMA-co-EDMA)). After optimization of the mobile phase conditions, a baseline resolution of the stereoisomers of 24 out of 53 N-derivatized di- and tri-peptides was obtained. 3,5-Dinitrobenzoyl- and 3,5-dichlorobenzoyl-peptide stereoisomers were separated with exceptionally high selectivity and resolution. The monolithic column was then applied to the quantitative analysis of l-carnosine and its enantiomeric impurity in three different commercial dietary supplements. Method validation demonstrated satisfactory results in terms of linearity, precision, selectivity, accuracy and limits of detection and quantification. The determined amounts of l-carnosine in commercial formulations were in agreement with the labeled content for all analyzed samples, and the enantiomeric impurity was found to be below the limit of detection (LOD), showing the potential of the poly(MQD-co-HEMA-co-EDMA) monolithic column as a reliable tool for the quality control of l-carnosine in dietary supplements by micro-LC. PMID:26410182

  6. RGD修饰的葫芦素B纳米脂质载体对乳腺癌荷瘤裸鼠的抑瘤作用%Inhibitory Effect of RGD Modified B Nano Lipid Carrier on Breast Cancer Bearing Nude Mice

    Institute of Scientific and Technical Information of China (English)

    唐淑洁; 赵旭伟; 张佳琦; 刘洋; 王梓丞

    2016-01-01

    Objective:To prepare RGD conjugated cucurbitacin B loaded liposome(RGD-CuB-NLC) and observe RGD-CuB-NLC’s anti-cancer effect on the treatment with the cells of breast cancer in vitro and RGD-CuB-NLC’s anti-cancer effect on nude mice of human breast cancer.Method:The liposomes were prepared by thin film hydration method,a single cell suspension liquid of the MCF-7 cells of breast cancer was made according to the cultivation of the adherent cell method,then it was inoculated into 96 hole cell culture plate,the group of RGD-CuB-NLC was designed,the control groups were CuB-NLC,RGD-NLC and the blank group of NLC were designed,the rate of suppression in different condition were calculated.Nude mouse transplantation tumor were established,30 qualified nude mice models of human breast cancer xenograft were randomly divided into 5 groups, each group was 6,the mice of 5 groups were respectively treated by CuB,CuB-NLC,RGD-NLC,RGD-CuB-NLC and 0.9% sodium chloride solution every 24 hours per time,7 times in total.The body weight of nude mice and the weight of the transplanted tumor growth were recorded,and the inhibition rate of tumor growth were calculated. Result:For RGD-CuB-NLC group,the form was the circular,the encapsulation rate>80%,partical size was less than 150 nm,the average particle size was (115.6±2.1)nm,polydispersity index was (0.217±0.033),the electric potential was(-12.31±0.76)mV.The inhibition ratio of different liposome on breast cancer cells in the group of NLC was 0,the group of CuB-NLC was (46.3±2.21)%,the group of RGD-NLC was (55.7±3.22)%, the group of RGD-CuB-NLC was (75.6±5.67)%,the differences were statistically significant(P80%,粒径<150 nm,平均为(115.6±2.1)nm,多分散系数为(0.217±0.033),电位为(-12.31±0.76)mV。不同脂质体对乳腺癌细胞体外抑制率分别为:NLC组为0,CuB-NLC组为(46.3±2.21)%,RGD-NLC组为(55.7±3.22)%, RGD-CuB-NLC组为(75.6±5.67)%,比较

  7. Optimization of the production process of hybrid and multivalent formulation Bombesin/RGD for the opportune detection of breast cancer; Optimizacion del proceso de fabricacion de la formulacion hibrida y multivalente Bombesina/RGD para la deteccion oportuna de cancer de mama

    Energy Technology Data Exchange (ETDEWEB)

    Robles M, M.

    2013-07-01

    The radiopharmaceuticals of third generation are used in nuclear medicine to obtain images of specific molecular targets, and they are unique in their capacity to detect in vivo specific biochemical sites as receptors that are over-expressed in diverse illness. In cancer cells several types of receptors are over-expressed, as the integrin s α(v)β(3) and α(v)β(5) that specifically recognize the sequence RGD (Arginine-Glycin-Ac. Aspartic) and gastrin-releasing peptide that recognizes specifically to the peptide Lys{sup 3}-Bombesin. The integrin s α(v)β(3) and α(v)β(5) are involved in the tumor angio genesis processes and the gastrin-releasing peptide is over-expressed in breast and prostate cancer. The molecular recognition of the specific receptors is the basis to be utilized as targets of the radiopharmaceuticals {sup 99m}Tc-HYNIC-Bombesin and {sup 99m}Tc-HYNIC-RGD. In this work was developed a lyophilized pharmaceutical formulation effective, stable and safe for the simultaneous obtaining of the radiopharmaceuticals {sup 99m}Tc-HYNIC-Bombesin ({sup 99m}Tc-EDDA/HYNIC-Lys{sup 3}-Bombesin) and {sup 99m}Tc-HYNIC-RGD ({sup 99m}Tc EDDA/HYNIC-E-[c(RGDfK)]{sub 2}). Later on the production process of the product HYNIC-Bombesin/RGD-Sn was optimized using a factorial design and the formulation was transferred to the production plant of radiopharmaceuticals of the Instituto Nacional de Investigaciones Nucleares (ININ). The optimized formulation is described in the following chart: HYNIC-[Lys{sup 3}]-Bombesin - 12.5 μg; HYNIC-E-c[RGDfK]{sub 2} - 12.5 μg; Stannous chloride (SnCl{sub 2}) - 20 μg; Ethylenediamine diacetic acid (EDDA) - 10 mg; N-tris(hydroxymethyl)methyl glycin (Tricine) - 20 mg; Mannitol - 50 mg. The production process was validated and were carried out the stability studies under refrigeration conditions. (Author)

  8. Self-assembled Biodegradable Nanoparticles and Polysaccharides as Biomimetic ECM Nanostructures for the Synergistic effect of RGD and BMP-2 on Bone Formation.

    Science.gov (United States)

    Wang, Zhenming; Dong, Li; Han, Lu; Wang, Kefeng; Lu, Xiong; Fang, Liming; Qu, Shuxin; Chan, Chun Wai

    2016-01-01

    Producing biomimetic extracellular matrix (ECM) is an effective approach to improve biocompatibility of medical devices. In this study, biomimetic ECM nanostructures are constructed through layer-by-layer self-assembling positively charged chitosan (Chi), negatively charged oxidized sodium alginate (OAlg), and positively charged bovine serum albumin (BSA)-based nanoparticles. The BSA-based nanoparticles in the self-assembled films not only result in porous nanostructures similar to natural ECM, but also preserve the activity and realize the sustained release of Bone morphogenetic protein-2 (BMP-2). The results of bone marrow stem cells (BMSCs) culture demonstrate that the penta-peptide glycine-arginine-glycine-aspartate-serine (GRGDS) grafted Chi/OAlg films favor cell adhesion and proliferation. GRGDS and BMP-2 in biomimetic ECM nanostructures synergistically promote BMSC functions and new bone formation. The RGD and BMP incorporated biomimetic ECM coatings could be applied on a variety of biomedical devices to improve the bioactivity and biocompatibility. PMID:27121121

  9. 酶催化合成CCK-4三肽片段%Enzymatic synthesis of a CCK-4 tripeptide fragment

    Institute of Scientific and Technical Information of China (English)

    郭丽; 吕子敏; Heiner Eckstein

    2003-01-01

    Objective To synthesize a tripeptide derivative Phac-Met-Asp(OMe)-Phe -NH2, which is a fragment of the gastrin C-terminal tetrapeptide CCK-4, by enzymatic reaction. Methods Three free enzymes, α-chymotrypsin, papain and thermolysin from acyl donor Phac-Met-OCam was involved in three steps. The choice of appropriate enzymes and solvents was selected.Results All enzymatic reactions were obtained in reasonable yields (63%-92%). FAB-MS and FD-MS verified the correct molecular mass of the peptides. Conclusion Studies on the α-chymotrypsin catalyzed coupling reaction between Phac-Met-OCam and H-Asp(OMe)2 have focused on the low water content media. By papain catalyzed saponification of Phac-Met-Asp(OMe)2, α-methyl ester of aspartic acid is selectively hydrolyzed to retain β-methyl ester, and Phac-Met-Asp (OMe)-OH and H-Phc-NH2 can be coupled efficiently by thermolysin.%目的探讨酶法合成胃泌激素CCK-4 C-末端片段三肽衍生物Phac-Met-Asp(OMe)-Phe-NH2.方法以Phac-Met-OCam为起始原料,采用三种蛋白酶,即α-糜蛋白酶、木瓜蛋白酶、嗜热菌蛋白酶(α-chymotrypsin,papain,thermolysin),经过三步酶促反应得到目标三肽化合物;研究了酶催化条件下Met-Asp肽键的形成以及天门冬氨酸双酯中α-酯的酶催化条件下的选择性水解;对酶以及反应介质的选择等酶促合成条件进行了研究.结果三步酶促反应均可得到较合适的收率(63%~92%),产物经FAB-MS、FD-MS质谱确认.结论以α-糜蛋白酶为酶催化剂,在含有1.5%0.05 mol/LT^s-HCl缓冲液的乙酸乙酯介质中,Phac-Met-OCam和H-Asp(OMe)2缩合可得到收率大于63%的缩合产物;木瓜蛋白酶催化下可选择性水解Phac-Met-Asp(OMe)2中的α-酯而保留β-酯;嗜热菌蛋白酶能有效地催化Phac-Met-Asp(OMe)-OH与H-Phe-NH2生成肽键.

  10. Imaging of 99Tcm-labeled new cyclic RGDfK Dimer in nude mice bearing U87MG human glioma xenografts%99Tcm标记新型RGD环肽在神经胶质瘤动物模型的实验研究

    Institute of Scientific and Technical Information of China (English)

    靳晓娜; 史继云; 刘妍; 朱朝晖; 贾兵; 刘昭飞; 石希敏; 王凡; 李方

    2010-01-01

    目的 评价引入2个聚乙二醇(PEG4)对精氨酸-甘氨酸-天冬氨酸(RGD)环肽二聚体(Dimer:E[c(RGDfK)]2)体外受体结合亲和力和体内药代动力学特征的影响,以及99Tcm标记2PEG4-Dimer用于整合素αvβ3阳性肿瘤显像的前景.方法 用免疫组织化学实验测定U87MG人神经胶质瘤细胞以及肿瘤组织中整合素αvβ3的表达.通过U87MG细胞受体竞争结合实验测定RGD环肽单体c(RGDyK)、联肼尼克酰胺(HYNIC)-Dimer和HYNIC-2PEG4-Dimer对125I-c(RGDyK)的半数抑制浓度(IC50).采用无亚锡一步法制备99Tcm-HYNIC-Dimer和99Tcm-HYNIC-2PEG4-Dimer,评价"TcmHYNIC-2PEG4-Dimer在荷U87MG瘤裸鼠的生物分布并进行γ显像.采用非配对t检验法对实验数据进行分析.结果 U87MG细胞和荷瘤裸鼠肿瘤组织中高表达整合素αvβ3.HYNIC-2PEG4-Dimer比c(RGDyK)和HYNIC-Dimer有更高的整合素αvβ3亲和力(IC50分别是0.8,27和2.4 nmol/L).99Tcm-HYNIC-Dimer和99Tcm-HYNIC-2PEG4-Dimer的99Tcm标记率均>95%,经Sep-Pek C18柱纯化后其放化纯>99%.生物分布实验显示,2种标记物均主要经肾排泄,在注射后2h,肿瘤对99Tcm-HYNIC-2PEG4-Dimer的摄取为99Tcm-HYNIC-Dimer的2.7倍[(5.71±0.96)%ID/g和(2.10±0.50)%ID/g],t=4.80,P<0.05,与体外受体竞争结合实验数据相一致.γ显像结果显示,注射99Tcm-HYNIC-2PEG4-Dimer后0.5 h肿瘤即清晰可见,随时间延长,体内放射性本底明显减低,显像对比度增高.结论 99Tcm-HYNIC-2PEG4-Dimer有希望用于整合素αvβ3阳性肿瘤显像.%Objective (1) To evaluate the effect of insertion of two 15-amino-4,7,10,13-tetraoxapentadecanoic (2 PEG4 ) linkers into cyclic Arg-Gly-Asp (RGD) Dimer E [c(RGDfK)]2 on receptor binding in vitro, (2) to assess its biodistribution in vivo and (3) to investigate the value of 99Tcm labeled 2PEG4-Dimer for integrin αvβ3-positive tumors imaging.Methods The expression of U87 human glioma cells and integrin αv β3 was determined by immunofluorescence staining

  11. 整合素受体和细胞穿膜肽共修饰紫杉醇脂质体抑制食管癌Ec9706细胞的研究%The Inhibition Effect of RGD and R8 Co-modified Paclitaxel Loaded Liposome on the Esophagus Carcinoma EC9706 Cells

    Institute of Scientific and Technical Information of China (English)

    吕行; 印滇

    2014-01-01

    目的:制备整合素受体RGD和细胞穿膜肽R8共修饰载紫杉醇( PTX)脂质体( RGD/R8-LP-PTX),对其理化性质进行表征,并观察脂质体与食管癌Ec9706细胞的亲和力和增殖抑制作用。方法:采用薄膜分散法制备RGD/R8-LP-PTX,观察脂质体的粒径,电位以及包封率;通过定量细胞摄取实验观察食管癌Ec9706细胞对RGD/R8-LP的摄取效率以及对脂质体摄取的影响因素。定性共聚焦实验观察肿瘤细胞对脂质体的摄取。 MTT实验观察RGD/R8-LP-PTX对食管癌Ec9706细胞的细胞毒性;构建食管癌Ec9706细胞肿瘤球模型,观察脂质体对肿瘤球的生长抑制能力。结果: RGD/R8-LP-PTX的粒径在124.8±9.4 nm,电位为21.35±3.55 mV。食管癌Ec9706细胞对RGD/R8-LP的摄取以及 RGD/R8-LP-PTX 对食管癌 Ec9706细胞的增殖抑制率具有时间依赖性;食管癌Ec9706细胞对RGD/R8-LP的摄取效率显著高于R8-LP、RGD-LP和LP,差异有统计学意义(P<0.01);在给药48 h后,R8-LP-PTX、RGD-LP-PTX和LP-PTX的细胞存活率分别是RGD/R8-LP-PTX组的1.6倍、1.7倍和2.2倍,差异有统计学意义(P<0.01)。给药7天后,生理盐水组肿瘤球持续生长,体积增大1.48倍,LP-PTX组肿瘤球体积增大到原体积的1.12倍, RGD/R8-LP-PTX组、R8-LP-PTX组和 RGD-LP-PTX组肿瘤球体积减小到原体积的36%、59%和64%,差异具有统计学意义(P<0.01)。结论:整合素受体RGD和细胞穿膜肽R8共修饰载紫杉醇(PTX)脂质体能够有效穿透肿瘤细胞膜进入肿瘤细胞,是一种有效的食管癌化疗靶向给药系统。%Objective: To prepare RGD and R8 co-modified paclitaxel loaded liposome(RGD/R8-LP-PTX)for EC9706 cells’ treatment. Methods: The co-modified liposome was prepared by film-ultrasonic method. The appearance, particle size,Zeta potential were evaluated. The cellular uptake by EC9706 cells in vitro was used to evaluate the targeting efficiency. The anti-proliferation efficiency of

  12. Evaluation of two novel {sup 64}Cu-labeled RGD peptide radiotracers for enhanced PET imaging of tumor integrin α{sub v}β{sub 3}

    Energy Technology Data Exchange (ETDEWEB)

    Hernandez, Reinier; Graves, Stephen A.; Nickles, Robert J. [University of Wisconsin, Department of Medical Physics, Madison, WI (United States); Czerwinski, Andrzej; Valenzuela, Francisco [Peptides International, Inc., Louisville, KY (United States); Chakravarty, Rubel; Yang, Yunan; England, Christopher G. [University of Wisconsin, Department of Radiology, Madison, WI (United States); Cai, Weibo [University of Wisconsin, Department of Medical Physics, Madison, WI (United States); University of Wisconsin, Department of Radiology, Madison, WI (United States); University of Wisconsin Carbone Cancer Center, Madison, WI (United States)

    2015-11-15

    Our goal was to demonstrate that suitably derivatized monomeric RGD peptide-based PET tracers, targeting integrin α{sub v}β{sub 3}, may offer advantages in image contrast, time for imaging, and low uptake in nontarget tissues. Two cyclic RGDfK derivatives, (PEG){sub 2}-c(RGDfK) and PEG{sub 4}-SAA{sub 4}-c(RGDfK), were constructed and conjugated to NOTA for {sup 64}Cu labeling. Their integrin α{sub v}β{sub 3}-binding properties were determined via a competitive cell binding assay. Mice bearing U87MG tumors were intravenously injected with each of the {sup 64}Cu-labeled peptides, and PET scans were acquired during the first 30 min, and 2 and 4 h after injection. Blocking and ex vivo biodistribution studies were carried out to validate the PET data and confirm the specificity of the tracers. The IC{sub 50} values of NOTA-(PEG){sub 2}-c(RGDfK) and NOTA-PEG{sub 4}-SAA{sub 4}-c(RGDfK) were 444 ± 41 nM and 288 ± 66 nM, respectively. Dynamic PET data of {sup 64}Cu-NOTA-(PEG){sub 2}-c(RGDfK) and {sup 64}Cu-NOTA-PEG{sub 4}-SAA{sub 4}-c(RGDfK) showed similar circulation t{sub 1/2} and peak tumor uptake of about 4 %ID/g for both tracers. Due to its marked hydrophilicity, {sup 64}Cu-NOTA-PEG{sub 4}-SAA{sub 4}-c(RGDfK) provided faster clearance from tumor and normal tissues yet maintained excellent tumor-to-background ratios. Static PET scans at later time-points corroborated the enhanced excretion of the tracer, especially from abdominal organs. Ex vivo biodistribution and receptor blocking studies confirmed the accuracy of the PET data and the integrin α{sub v}β{sub 3}-specificity of the peptides. Our two novel RGD-based radiotracers with optimized pharmacokinetic properties allowed fast, high-contrast PET imaging of tumor-associated integrin α{sub v}β{sub 3}. These tracers may facilitate the imaging of abdominal malignancies, normally precluded by high background uptake. (orig.)

  13. Development of NOTA/DOTA cyclo-RGD dimers labelled with Ga-68 for cancer diagnosis and therapy follow-up

    International Nuclear Information System (INIS)

    Full text of publication follows. The ανβ3 integrin receptor, expressed on tumor cell membranes can be preferentially targeted by peptides containing the RGD sequence, resulting in a versatile cell recognition system. DOTA-E- [c(RGDfK)2] and NOTA-SCN-Bn-E-[c(RGDyK)2] were labelled with Ga-68 and tested for radiolabelling yield, purity, stability, in vitro binding and ex vivo biodistribution. The radiolabelling was performed using an automated system with in-line quality control and Ga-68 eluate from a tin oxide based Ge-68/Ga-68 generator, purified and concentrated to 400 MBq in 0.1 ml water on an anionic exchanger resin. DOTA/NOTA-derivatized peptides were labeled by heating at 95 C. degrees. Elution, concentration, labeling and purification procedures took 20 min. The ex-vivo biodistribution of 68Ga-DOTA-E-[c(RGDfK)2] and 68Ga-NOTA-SCN-Bn-E-[c(RGDyK)2] was tested in tumor bearing animal models. Real-time quantification of bio-molecular interactions was determined: on- and off-rates, affinity of DOTA/NOTA cyclo-RGD dimers to tumor cell-surface receptors. Nanomoles of peptides were labeled and purified, RCP>98%. The biodistribution pattern of 68Ga- NOTA-SCN-Bn-E-[c(RGDyK)2] in melanoma rats shows high and stable tumor uptake up to 11.6% ID/g. The blood clearance is fast, the renal elimination is more rapid than in the case of 68Ga-DOTA-E-[c(RGDfK)2]. Higher tumor to background ratios was observed. Binding to receptors is achieved in the first 3 min of incubation and remains stable for 30 min. The radiolabelling with Ga-68 of very promising candidates for imaging targets of interest in cancer diagnosis and therapy follow-up such as ανβ3 receptors, were successfully adapted on the automated module, reducing reaction time and operator exposure. The biological evaluations of 68Ga-DOTA-E-[c(RGDfK)2] and 68Ga-NOTA-SCN-Bn-E-[c(RGDyK)2] show a high and stable tumor uptake of both compounds. (authors)

  14. Preparation of the radiopharmaceutical {sup 99m} Tc-HYNIC-cyclo-Lys-D-Phe-RGD for In vivo image of integrines; Preparacion del radiofarmaco {sup 99m} Tc-HYNIC-ciclo-Lys-D-Phe-RGD para imagen In vivo de integrinas

    Energy Technology Data Exchange (ETDEWEB)

    Hernandez H, E. [ININ, 52750 La Marquesa, Estado de Mexico (Mexico)

    2007-07-01

    The diagnostic of some pathological processes by means of images constitutes one of the used methods in the determination of the origin, condition and/or evolution of one illness. The use of contrast agents in conjunction with other techniques help to the obtaining and visualization of complex systems, among these we can find to those radiopharmaceuticals used in nuclear medicine to visualize diverse organs and corporal systems. At the moment it is sought to develop a radiopharmaceutical of third generation that can be used for image In vivo of integrines with the purpose of detecting angio genesis processes, that which would allow to diagnose in way it specifies a wide range of primary tumors and their metastasis. Presently work it developed the radiopharmaceutical {sup 99m}Tc-HYNIC-cycle-Lys-D-Phe-RGD, likewise the good conditions were determined for the formation of this complex. The HYNIC was employee as chelating agent, using as co ligands EDDA and Tricine for to complete the sphere of coordination of the {sup 99m}Tc. The conjugated HYNIC-RGD was synthesized, purified, characterized and radiolabelled In situ with {sup 99m}Tc using High pressure liquid chromatography as analysis method in Reverse Phase (RP-HPLC). By this way it was developed the lyophilized formulation for its instantaneous labelled to which were carried out quality control tests. The one conjugated was obtained free of impurities, showing stability at same as their complex formed with {sup 99m}Tc. The analysis method was validated turning out to be necessary, exact, lineal and specific for the quantification of the analyte of interest. The lyophilized formulation showed a radiochemical purity bigger than 95%, besides being sterile and free of pyrogens. The biodistribution tests in athymic mice with induced tumors showed that the radiopharmaceutical was united mainly to the tumor and that this it was excreted mainly for renal via. (Author)

  15. Correlation of breast cancer subtypes, based on estrogen receptor, progesterone receptor, and HER2, with functional imaging parameters from {sup 68}Ga-RGD PET/CT and {sup 18}F-FDG PET/CT

    Energy Technology Data Exchange (ETDEWEB)

    Yoon, Hai-Jeon [Seoul National University College of Medicine, Department of Nuclear Medicine, Seoul (Korea, Republic of); Seoul National University College of Medicine, The Institute of Radiation Medicine, Seoul (Korea, Republic of); Ewha Womans University School of Medicine, Department of Nuclear Medicine, Seoul (Korea, Republic of); Kang, Keon Wook; Jeong, Jae Min; Chung, June-Key [Seoul National University College of Medicine, Department of Nuclear Medicine, Seoul (Korea, Republic of); Seoul National University College of Medicine, Department of Biomedical Sciences, Seoul (Korea, Republic of); Seoul National University College of Medicine, The Institute of Radiation Medicine, Seoul (Korea, Republic of); Seoul National University, Cancer Research Institute, Seoul (Korea, Republic of); Chun, In Kook [Seoul National University College of Medicine, Department of Nuclear Medicine, Seoul (Korea, Republic of); Kangwon National University Hospital, Department of Nuclear Medicine, Chuncheon, Kangwon-Do (Korea, Republic of); Cho, Nariya [Seoul National University College of Medicine, Department of Radiology, Jongno-gu, Seoul (Korea, Republic of); Im, Seock-Ah [Seoul National University College of Medicine, Department of Internal Medicine, Seoul (Korea, Republic of); Jeong, Sunjoo [Dankook University, Department of Molecular Biology, Yongin (Korea, Republic of); Lee, Song [Seoul National University College of Medicine, Department of Nuclear Medicine, Seoul (Korea, Republic of); Seoul National University College of Medicine, The Institute of Radiation Medicine, Seoul (Korea, Republic of); Jung, Kyeong Cheon [Seoul National University College of Medicine, Department of Pathology, Seoul (Korea, Republic of); Lee, Yun-Sang [Seoul National University College of Medicine, Department of Nuclear Medicine, Seoul (Korea, Republic of); Seoul National University College of Medicine, Department of Molecular Medicine and Biopharmaceutical Sciences, Graduate School of Convergence Science and Technology, Seoul (Korea, Republic of); Lee, Dong Soo [Seoul National University College of Medicine, Department of Nuclear Medicine, Seoul (Korea, Republic of); Seoul National University College of Medicine, The Institute of Radiation Medicine, Seoul (Korea, Republic of); Seoul National University, Department of Molecular Medicine and Biopharmaceutical Sciences, Seoul (Korea, Republic of); Moon, Woo Kyung [Seoul National University College of Medicine, Department of Radiology, Jongno-gu, Seoul (Korea, Republic of); Seoul National University College of Medicine, Department of Biomedical Sciences, Seoul (Korea, Republic of); Seoul National University College of Medicine, The Institute of Radiation Medicine, Seoul (Korea, Republic of)

    2014-08-15

    Imaging biomarkers from functional imaging modalities were assessed as potential surrogate markers of disease status. Specifically, in this prospective study, we investigated the relationships between functional imaging parameters and histological prognostic factors and breast cancer subtypes. In total, 43 patients with large or locally advanced invasive ductal carcinoma (IDC) were analyzed (47.6 ± 7.5 years old). {sup 68}Ga-Labeled arginine-glycine-aspartic acid (RGD) and {sup 18}F-fluorodeoxyglucose (FDG) positron emission tomography/computed tomography (PET/CT) were performed. The maximum and average standardized uptake values (SUV{sub max} and SUV{sub avg}) from RGD PET/CT and SUV{sub max} and SUV{sub avg} from FDG PET/CT were the imaging parameters used. For histological prognostic factors, estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2) expression was identified using immunohistochemistry (IHC) or fluorescent in situ hybridization (FISH). Four breast cancer subtypes, based on ER/PR and HER2 expression (ER/PR+,Her2-, ER/PR+,Her2+, ER/PR-,Her2+, and ER/PR-,Her2-), were considered. Quantitative FDG PET parameters were significantly higher in the ER-negative group (15.88 ± 8.73 vs 10.48 ± 6.01, p = 0.02 for SUV{sub max}; 9.40 ± 5.19 vs 5.92 ± 4.09, p = 0.02 for SUV{sub avg}) and the PR-negative group (8.37 ± 4.94 vs 4.79 ± 3.93, p = 0.03 for SUV{sub avg}). Quantitative RGD PET parameters were significantly higher in the HER2-positive group (2.42 ± 0.59 vs 2.90 ± 0.75, p = 0.04 for SUV{sub max}; 1.60 ± 0.38 vs 1.95 ± 0.53, p = 0.04 for SUV{sub avg}) and showed a significant positive correlation with the HER2/CEP17 ratio (r = 0.38, p = 0.03 for SUV{sub max} and r = 0.46, p < 0.01 for SUV{sub avg}). FDG PET parameters showed significantly higher values in the ER/PR-,Her2- subgroup versus the ER/PR+,Her2- or ER/PR+,Her2+ subgroups, while RGD PET parameters showed significantly lower values in the ER

  16. In vivo pharmacokinetics, biodistribution and the anti-tumor effect of cyclic RGD-modified doxorubicin-loaded polymers in tumor-bearing mice.

    Science.gov (United States)

    Wang, Chen; Li, Yuan; Chen, Binbin; Zou, Meijuan

    2016-10-01

    In our previous study, we successfully produced and characterized a multifunctional drug delivery system with doxorubicin (RC/GO/DOX), which was based on graphene oxide (GO) and cyclic RGD-modified chitosan (RC). Its characteristics include: pH-responsiveness, active targeting of hepatocarcinoma cells, and efficient loading with controlled drug release. Here, we report the pharmacokinetics, biodistribution, and anti-tumor efficacy of RC/GO/DOX polymers in tumor-bearing nude mice. The objective of this study is to assess its targeting potential for tumors. Pharmacokinetic and biodistribution profiles demonstrated that tumor accumulation of RC/GO/DOX polymers was almost three times higher than the others, highlighting the efficacy of the active targeting strategy. Furthermore, the tumor inhibition rate of RC/GO/DOX polymers was 56.64%, 2.09 and 2.93 times higher than that of CS/GO/DOX polymers (without modification) and the DOX solution, respectively. Anti-tumor efficacy results indicated that the tumor growth was better controlled by RC/GO/DOX polymers than the others. Hematoxylin and eosin (H&E) staining showed remarkable changes in tumor histology. Compared with the saline group, the tumor section from the RC/GO/DOX group revealed a marked increase in the quantity of apoptotic and necrotic cells, and a reduction in the quantity of the blood vessels. Together, these studies show that this new system could be regarded as a suitable form of DOX-based treatment of the hepatocellular carcinoma. PMID:27244048

  17. Surface Modification of Biomimetic PLGA-(ASP-PEG) Matrix with RGD-Containing Peptide: a New Non-Viral Vector for Gene Transfer and Tissue Engineering

    Institute of Scientific and Technical Information of China (English)

    GUO Xiaodong; SONG Yulin; ZHENG Qixin; YANG Shuhua; SHAO Zengwu; WU Yongchao; HAO Jie; QUAN Daping

    2006-01-01

    RGD-containing peptide (K16-GRGDSPC), characterized as non-viral gene vectors, was fabricated to modify the surface of PLGA-[ASP-PEG] matrix, which offered the foundation for gene transfer with porous matrix of gene activated later. Peptide was synthesized and matrix was executed into chips A,B and chip C. Chip C was regarded as control. Chips A and B were reacted with cross-linker. Then chip A was reacted with peptide. MS and HPLC were used to detect the MW and purity of peptide. Sulphur, existing on the surface of biomaterials, was detected by XPS. The purity of un-reacted peptide in residual solution was detected by a spectrophotometer. HPLC shows that the peptide purity was 94%-95%, and MS shows that the MW was 2 741.3307. XPS reveals that the binding energy of sulphur was 164 eV and the ratio of carbon to sulphur (C/S) was 99.746∶0.1014 in reacted chip A. The binding energy of sulphur in reacted chip B was 164 eV and 162 eV, C/S was 99.574∶0.4255, and there was no sulphur in chip C. Peptide was manufactured and linked to the surface of biomimetic and 3-D matrix, which offered the possibilities for gene transfer and tissue engineering with this new kind of non-viral gene vector.

  18. Hydrothermal Synthesis and Characterization of Pyrochlore Titanate R2 Ti2O7(R=Gd3+, Tb3+, Dy3+)

    Institute of Scientific and Technical Information of China (English)

    PENG Wen; HU Bin; CHEN Yan; HU Wei-wei; GUO Li; YUAN Hong-ming; FENG Shou-hua

    2011-01-01

    Pyrochlore titanate oxides, R2Ti2O7(R=Gd3+, Tb3+, Dy3+), were synthesized under mild hydrothermal conditions. The crystal growth of pyrochlore titanate oxides and taking place of chemical reaction in the hydrothermal processing were sensitive to the alkalinity, temperature, reaction time, the nature of the rare earth ion and the composition of initial reaction mixture. The as-prepared samples were characterized by powder X-ray diffraction, scanning electron microscopy, Raman spectrum and variable temperature dc magnetic susceptibility(Superconductivity quantum interference device, SQUIDS). The magnetic studies gave 7.29× 10-23 A·m2/Gd3+ and -8.28 K, 8.75 × 10-23 A·m2/ Tb3+ and -19.7 K, and 8.85×10-23 A·m2/Dy3+ and 0.84 K effective moments and Weiss constants for Gd2Ti2O7,Tb2Ti2O7 and Dy2Ti2O7, respectively.

  19. Complex based on Isthmin and RGD motif against glioma U251 cells%基于Isthmin和RGD基序的复合物对胶质瘤U251的治疗作用

    Institute of Scientific and Technical Information of China (English)

    陈宏颉; 曹磊; 王守森; 郑兆聪; 王如密; 汪君

    2011-01-01

    Objective To study the effect of complex based on Isthmin and RGD motif against glioma U251 cells.Methods The complex based on Isthmin and RGD motif was prepared,and the glioma U251 cell xenografts were established to observe the antitumor and antiangiogenesis effects of the complex in vivo and in vivo.Results The complex could induce U251 cell apoptosis and attack tumor endothelium cells,which inhibited the growth of glioma U251 cells and improved the lifespan of tumor bearing mice.Conclusion The complex based on Isthmin and RGD motif could dually target tumor and endothelium cells,which provided a promising strategy for glioma gene therapy.%目的 探讨基于Isthmin和BGD基序的复合物对胶质瘤U251的治疗效应.方法 构建Isthmin和RGD基序的复合物,并建立胶质瘤U251移植瘤模型,体内外观察其对U251细胞和血管内皮细胞的影响.结果 该复合物能有效诱导U251细胞的凋亡,抑制肿瘤血管的生成,从而抑制肿瘤的生长和延长荷瘤小鼠的平均生存率.结论 该复合物能同时靶向肿瘤和血管内皮细胞,为胶质瘤的基因治疗提供了较理想的策略.

  20. Milk-derived tripeptides IPP (Ile-Pro-Pro and VPP (Val-Pro-Pro promote adipocyte differentiation and inhibit inflammation in 3T3-F442A cells.

    Directory of Open Access Journals (Sweden)

    Subhadeep Chakrabarti

    Full Text Available Milk derived tripeptides IPP (Ile-Pro-Pro and VPP (Val-Pro-Pro have shown promise as anti-hypertensive agents due to their inhibitory effects on angiotensin converting enzyme (ACE. Due to the key inter-related roles of hypertension, chronic inflammation and insulin resistance in the pathogenesis of metabolic syndrome, there is growing interest in investigating established anti-hypertensive agents for their effects on insulin sensitivity and inflammation. In this study, we examined the effects of IPP and VPP on 3T3-F442A murine pre-adipocytes, a widely used model for studying metabolic diseases. We found that both IPP and VPP induced beneficial adipogenic differentiation as manifested by intracellular lipid accumulation, upregulation of peroxisome proliferator-activated receptor gamma (PPARγ and secretion of the protective lipid hormone adiponectin by these cells. The observed effects were similar to those induced by insulin, suggesting potential benefits in the presence of insulin resistance. IPP and VPP also inhibited cytokine induced pro-inflammatory changes such as reduction in adipokine levels and activation of the nuclear factor kappa B (NF-κB pathway. Taken together, our findings suggest that IPP and VPP exert insulin-mimetic adipogenic effects and prevent inflammatory changes in adipocytes, which may offer protection against metabolic disease.

  1. Involvement of integrins in the adhesion of osteoblastic cells to a type-I collagen matrix

    Directory of Open Access Journals (Sweden)

    Antonio Desmond McCarthy

    2006-01-01

    Full Text Available Se han desarrollado varios biomateriales con potencial aplicaci ón en la reconstrucción de tejidos. En este sentido, existe un creciente interés en el diseño de materiales de implante óseo con máxima biocompatibilidad y adecuada adhesividad celular. El objetivo de este trabajo fue estudiar cuáles receptores integrinas participan en la adhesión de osteoblastos a una matriz de colágeno tipo-I. Se analizaron dos líneas celulares osteoblásticas: UMR106, derivada de osteosarcoma de rata; y MC3T3E1, derivada de calvaria de rata. Las células se cultivaron durante una hora sobre plástico, o sobre un gel de colágeno tipo-I, solas o co-incubadas con diferentes péptidos: (a péptido-b, un oligopéptido de 13 aminoácidos que corresponde a la secuencia conservada 113-125 de la subunidad b de los receptores integrinas; (b RGD (Arg-Gly-Asp, que corresponde a la secuencia de reconocimiento de la subunidad a de las integrinas a1,5b1; o (c DGEA (Asp-Gly-Glu-Ala, la secuencia de reconocimiento de la subunidad a de las integrinas a2b1. La adhesión y la inducción de extensiones celulares se evaluaron microscópicamente luego de lavar, fijar y colorear las células con Giemsa. Los resultados demostraron que las células osteoblásticas se adhieren más fácilmente a un sustrato de colágeno tipo-I que al plástico (86 ± 5 células/campo vs. 69 ± 4 células/campo para colágeno tipo-I vs. plástico, respectivamente, p < 0,02. El péptido-b inhibió la adhesión de ambas líneas celulares a una matriz de colágeno y el efecto inhibidor mostró dependencia dosis-respuesta. Por otro lado, este péptido indujo en las células tipo osteoblastos UMR106 un aumento del agrupamiento intercelular, y una reducción en la inducción de extensiones celulares. Estos cambios morfol ógicos podrían estar indicando un incremento en las interacciones célulac élula y una disminución en las interacciones célula-matriz, posiblemente inducidos por el péptido-b. De

  2. Anti-tumor activity of miRNA through dual-targeting carrier of RGD and cell permeable peptides conjugated liposomes%RGD与细胞穿膜肽共修饰miRNA脂质体的构建及抗肿瘤研究

    Institute of Scientific and Technical Information of China (English)

    蔺伟; 李庭; 杜金瑞

    2015-01-01

    目的 制备RGD和细胞穿膜肽TAT共修饰载microRNA-34a脂质体(RGD/TAT-miLPs-34a),对其理化性质进行表征,研究脂质体与人骨肉瘤MG63细胞的亲和力和增殖抑制作用.方法 采用薄膜分散法制备RGD/TAT-miLPs-34a;定量细胞摄取实验研究MG63细胞对RGD/TAT-miLPs-34a的摄取效率以及对脂质体摄取的影响因素.定性共聚焦实验观察肿瘤细胞对脂质体的摄取.MTT实验研究RGD/TAT-miLPs-34a对MG63细胞的细胞毒性;构建骨肉瘤异位瘤模型,研究不同脂质体对肿瘤的生长抑制率.结果 MG63细胞在与脂质体共同孵育4h后,RGD/TAT-miLPs-34a组摄取效率为(185.3±17.5)%,TAT-miLPs-34a组为(67.5±8.6)%,RGD-miLPs-34a组为(82.7±9.5)%,miLPs-34a组为(40.9±7.8)%,差异有统计学意义,H=27.93,P<0.001.RGD/TAT-miLPs-34a 4 h摄取效率是2h摄取效率(101.3±12.4)%的1.83倍,差异有统计学意义,z=5.58,P=0.001.骨肉瘤MG63细胞给药48 h后,RGD/TAT-miLPs-34a组MG63细胞存活率为(18.5±5.3)%,TAT-miLPs-34a组为(36.1±5.2)%、RGD-miIPs-34a组为(44.8±6.7)%,miLPs-34a组为(67.7±8.9)%,生理盐水组为(98.7±3.6)%,差异有统计学意义,H=19.271,P<0.001.荷瘤裸鼠治疗实验miLPs-34a组对肿瘤生长的抑制率为(19.4±3.2)%,RGD-miLPs-34a组为(39.6±4.8)%,TAT-miLPs-34a组为(42.6±6.5)%,RGD/TAT-miLPs-34a组为(73.7±7.1)%,生理盐水组为0,差异有统计学意义,F=145.237,P<0.001.结论 细胞穿膜肽与RGD共修饰载miRNA脂质体能够有效识别并穿透肿瘤细胞膜进入肿瘤细胞,是一种潜在高效的骨肉瘤靶向给药系统.%OBJECTIVE To prepare RGD and cell permeable peptides TAT conjugated miRNA loaded liposomes and evaluates the anti-cancer effect on the treatment of osteosarcoma.METHODS The RGD/TAT-miLPs-34a was prepared by thin film hydration method.The anti-proliferation efficiency of RGD/TAT-miLPs-34a was evaluated by MTT assay.The efficiency of cellular uptake on MG63 cells in vitro was evaluated

  3. Radiochemical and biological evaluation of Ter-Cys-RGD derivative labeled with the precursor [{sup 99m}Tc(H{sub 2}O){sub 3}(CO){sub 3}]{sup +}

    Energy Technology Data Exchange (ETDEWEB)

    Teodoro, Rodrigo; Faintuch, Bluma L.; Queiroz, Rodrigo G.; Muramoto, Emiko; Morganti, Ligia; Nascimento, Nanci do [Instituto de Pesquisas Energeticas e Nucleares (IPEN/CNEN-SP), Sao Paulo, SP (Brazil)]. E-mail: teodoro_rodrigo@yahoo.com.br; blfaintuch@hotmail.com

    2007-07-01

    Introduction: Endothelial {alpha}{sub v}{beta}{sub 3} integrin is considered a marker of tumor-induced angiogenesis. This integrin can bind to the arginine-glycine-aspartic acid (RGD) aminoacid sequence present in extracellular matrix proteins. Cyclic peptides containing RGD sequences have high affinity and selectivity for {alpha}{sub v}{beta}{sub 3} integrin. Aim: The aim of this study was the labelling of Ter-Cys-RGD derivative using the {sup 99m}Tc-tricarbonyl technique and the exploration of its potential as a radiopharmaceutical in vivo. Methods: The preparation of the {sup 99m}Tc-precursor consisted in flushing the mixture of 4.4 mg of sodium carbonate, 5.5 mg of sodium borohydride and 20 mg of sodium-potassium tartrate tetrahydrate with CO gas during 30 min. Pertechnetate was added and the vial was heated for 35 min at 75 deg C. The reaction was stopped in ice bath, and pH was adjusted to 7. Then 100 {mu}L of the precursor was added to 1.1 mM (50 {mu}g) of the ligand and heated at 75 deg C for 60 min. The reaction was stopped in ice bath. Radiochemical purity of the precursor and the final product was assessed by paper and thin layer chromatography, as well as by HPLC. Biodistribution studies were performed in healthy Swiss mice at 1 and 4 hours post-injection and also in nude mice bearing A549 lung-cancer cells, 4 hours post-injection. Planar gamma-camera imaging was also done in nude athymic mice bearing tumor. Results: The precursor [{sup 99m}Tc(H{sub 2}O){sub 3}(CO){sub 3}]{sup +} was quantitatively formed by carbonylation with high yield (94.8 {+-} 2.6%). Radiochemical purity of {sup 99m}Tc(CO){sub 3}-Ter-Cys-RGD was 86.9% {+-} 4.3 %. A specific activity of 6.7 MBq/nmol was achieved. Biodistribution findings 4 hours post-injection revealed the highest uptake by respectively kidneys (12.1 {+-} 0.7 %ID/g), liver (11.8 {+-} 0.8 %ID/g) and intestines (10.0 {+-} 1.5 %ID/g). Biodistribution in mice bearing lung tumor also showed highest uptake by kidneys and

  4. Implications of a quasispecies genome structure: effect of frequent, naturally occurring amino acid substitutions on the antigenicity of foot-and-mouth disease virus.

    OpenAIRE

    Mateu, M G; Martínez, M A; Rocha, E.; Andreu, D; Parejo, J; Giralt, E.; Sobrino, F; Domingo, E

    1989-01-01

    We provide evidence that the quasispecies nature (extreme genetic heterogeneity) of foot-and-mouth disease virus is relevant to the virus evading an immune response. A monoclonal antibody neutralizing the viral infectivity (clone SD6) recognizes an epitope located around a highly conserved sequence (amino acid sequence Arg-Gly-Asp-Leu-Ala at positions 141-145) in the capsid protein VP1 of foot-and-mouth disease virus of serotype C1. The amino acid substitutions Ala-138----Thr and Leu-147----I...

  5. A monoclonal antibody (8H3) that binds to rat T lineage cells and augments in vitro proliferative responses

    OpenAIRE

    1990-01-01

    A murine monoclonal antibody, designated 8H3, recognizes a cell surface antigen expressed exclusively on rat T lineage cells. 8H3 antibody immunoprecipitated 180-, 120-, and 90-kD components from rat thymocytes as well as splenic T cells under nonreducing conditions. 8H3 antibody specifically inhibited the binding of thymocytes to fibronectin. Furthermore, binding of rat thymocytes to immobilized synthetic peptide Gly-Arg-Gly-Asp-Ser-Pro-Cys-BSA was inhibited by 8H3 antibody as was Gly-Arg-Gl...

  6. Synthesis and evaluation of {sup 99m}Tc-labeled folate-tripeptide conjugate as a folate receptor-targeeted imaging agent in a tumor-bearing mouse model

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Myoung Hyoun; Kim, Chang Guhn; Kim, Dae Weung [Dept. of Nuclear Medicine, Wonkwang University School of Medicine, Iksan (Korea, Republic of); Kim, Woo Hyoung [Dept. of Nuclear Medicine, Seoul National University Hospital, Seoul (Korea, Republic of)

    2015-09-15

    The folate receptor (FR) is an attractive molecular target since it is overexpressed in a variety of human tumors. The purpose of the present study was to synthesize and evaluate the feasibility of a novel {sup 99m}Tc-ECG-EDA (Glu-Cys-Gly-ethylenediamine)-folate as an FR-positive tumor imaging agent in a mouse tumor model. ECG-EDA-folate was synthesized using solid phase peptide synthesis (SPPS) and radiolabeled with {sup 99m}Tc using tripeptide ECG as a chelator. FR-positive KB cells were inoculated in athymic nude mice. Following injection of {sup 99m}Tc-ECG-EDA-folate, serial scintigraphy and micro-SPECT/CT imaging were performed at various time points with and without pre-administration of excess free folate. Mean count densities (MCD) for regions of interest drawn on KB tumors and major normal organs at each time point were measured, and uptake ratios of tumor to normal organs were calculated. ECG-EDA-folate was labeled with {sup 99m}Tc with high radiolabeling efficiency and stability (>96 %). FR-positive tumors were clearly visualized on both scintigraphy and micro-SPECT/CT images and the tumor uptake of {sup 99m}Tc-ECG-EDA-folate was markedly suppressed with faint visualization of tumors by pre-administration of excess free folate on serial planar scintigraphy, indicating FR-specific binding of the agent. Furthermore, semiquantitative analysis of MCD data showed again that both tumor MCD and tumor-to-normal organ ratios decreased considerably by pre-administration of excess free folate, supporting FR-specific tumor uptake. Tumor-to-normal organ ratios approximately increased with time after injection until 4 h. The present study demonstrated that 9{sup 99m}Tc-ECG-EDA-folate can bind specifically to FR with clear visualization of FR-positive tumors in a mouse tumor model.

  7. RGD多肽耦联活化小牛松质骨骨组织工程材料的研究%RGD polypeptide coupling activation dallas cancellous bone tissue engineering materials.

    Institute of Scientific and Technical Information of China (English)

    仝健; 高平; 张澜君; 李怀芬; 牛惠生

    2011-01-01

    目的:利用肽羧合剂EDC[1-乙基-3-(3-二甲基氨丙基)-碳二亚胺盐酸盐]将粘附蛋白类八肽(谷氨酸-脯氨酸-精氨酸-甘氨酸-天冬氨酸-天冬酰胺-酪氨酸-精氨酸)耦联到经紫外线辐照活化的小牛松质骨片上,制备出用于骨缺损性疾病的骨修复与重建的组织工程新方法,为口腔医学中牙齿和颌骨缺损性疾病的治疗提供骨移植的新型材料.方法:采用I-RGD放射性示踪技术、碳二亚胺羧合、紫外线辐照活化的小牛松质骨的羧基及蛋白质染色技术.结果:牢固地将RGD耦联到小牛松质骨上.结论:随着骨组织工程研究的发展和这一新材料的深入研究,将改变骨缺损临床采用羟基磷灰石或骨水泥等材料将缺损部位封闭,从而使其丧失功能的治疗方法,引领骨移植临床应用的到来.%Objective: to use peptide carboxy mixture EDC (1-ethyl-3-(3-2 methyl ammonia isopropyl)-carbon two imine)will adhesion protein class eight peptide (Glu-Pro-Arg-Gly-Asn-Asn-Tyr-Arg RGD) by ultraviolet irradiation coupling to activate the mavericks cancellous bone chip, which is prepared for bone defect sex disease of rehabilitation and reconstruction of bone tissue engineering,especially a new method of oral medicine teeth and jaws defect sexual diseases treatment provides bone grafts new material.Method: using 125I-RGD radioactive tracer technology, carbon two imine carboxy close, the mavericks ultraviolet irradiation activation cancellous bone carboxyl and protein dyeing of technology.Result:firmly coupling to Dallas will RGD cancellous bone.Conclusion: along with the development of tissue engineering research and the new materials of thorough research,will change bone defect clinical use a hydroxyapatite or bone cement materials will defect site sealed dead, so the loss of function treatment.The clinical application of bone graft coming.

  8. Co-delivery of adipose-derived stem cells and growth factor-loaded microspheres in RGD-grafted N-methacrylate glycol chitosan gels for focal chondral repair.

    Science.gov (United States)

    Sukarto, Abby; Yu, Claire; Flynn, Lauren E; Amsden, Brian G

    2012-08-13

    The coencapsulation of growth factor-loaded microspheres with adipose-derived stem cells (ASCs) within a hydrogel matrix was studied as a potential means to enhance ASC chondrogenesis in the development of a cell-based therapeutic strategy for the regeneration of partial thickness chondral defects. A photopolymerizable N-methacrylate glycol chitosan (MGC) was employed to form an in situ gel used to encapsulate microspheres loaded with bone morphogenetic protein 6 (BMP-6) and transforming growth factor-β3 (TGF-β3) with human ASCs. ASC viability and retention were enhanced when the Young's modulus of the MGC ranged between 225 and 380 kPa. Grafting an RGD-containing peptide onto the MGC backbone (RGD-MGC) improved ASC viability within the gels, remaining at greater than 90% over 14 days in culture. The effects of BMP-6 and TGF-β3 released from the polymer microspheres on ASC chondrogenesis were assessed, and the level of differentiation was compared to ASCs in control gels containing nongrowth factor-loaded microspheres cultured with and without the growth factors supplied in the medium. There was enhanced expression of chondrogenic markers at earlier time points when the ASCs were induced with the sustained and local release of BMP-6 and TGF-β3 from the microspheres. More specifically, the normalized glycosaminoglycan and collagen type II protein expression levels were significantly higher than in the controls. In addition, the ratio of collagen type II to type I was significantly higher in the microsphere delivery group and increased over time. End-point RT-PCR analysis supported that there was a more rapid induction and enhancement of ASC chondrogenesis in the controlled release group. Interestingly, in all of the assays, there was evidence of chondrogenic differentiation when the ASCs were cultured in the gels in the absence of growth factor stimulation. Overall, the co-delivery of growth-factor-loaded microspheres and ASCs in RGD-modified MGC gels

  9. Induced thermal ablation with a radiofrequency field in breast cancer cells using gold nanoparticles conjugated to the peptide cycle[RGDfK(C)

    International Nuclear Information System (INIS)

    The conjugation of peptides to gold nanoparticles (AuNP) produces biocompatible and stable multimeric systems with target-specific molecular recognition. Peptides based on the cyclic Arg-Gly-Asp (RGD) sequence have been reported as high-affinity agents for the α(v)β(3) and α(v)β(v) integrin s over expressed in breast cancer cells. AuNP have also been proposed as localized heat sources for cancer treatment using laser irradiation or radiofrequency (RF). The objective of this research was to evaluate the thermo ablative effect of the AuNP-c [RGDfK(C)] system on MCF7 breast cancer cell viability after exposure to a radiofrequency field and to compare it with that produced by the laser irradiation. The effect of the 13.56 MHz RF (using a power from 0 to 200 W at intervals of 50 W) over the temperature increase in AuNP-colloidal system of 5 and 20 nm at two different concentrations was evaluated. The absorption cross sections (Cabs) of the AuNP-c [RGDfK(C)] nano system when it interacts with low frequency electromagnetic waves (13.56 MHz, λ = 22 m) and optical frequency waves (laser at λ = 532 nm) was analyzed based on the Mi e theory. The effect on the MCF7 cell viability was assessed using two thermal conversion sources (Laser and RF) on AuNP-c [RGDfK(C)] located inside the cytoplasm of the cells. MCF7 cells were treated with AuNP-c [RGDfK(C)] or water after exposure to the RF field (200 W, 100 V/cm) or laser irradiation (Irradiance 0.65 W/cm2). In both cases (RF and laser) the presence of nanoparticles internalized inside the cells caused a significant increase in the temperature of the medium (RF: ΔT = 29.9 ± 1.7 grades C for AuNP compared toΔT = 13.0 ± 1.4 grades C for water; laser: ΔT = 13.5 ± 0.7 grades C for AuNP compared to 3.3 ± 0.5 grades C for water). Although RF induced a higher increase in the temperature of the medium with nanoparticles, the largest effect on the cell viability was produced by laser when nanoparticles were located inside the

  10. Induced thermal ablation with a radiofrequency field in breast cancer cells using gold nanoparticles conjugated to the peptide cycle[RGDfK(C)]; Termoablacion inducida con un campo de radiofrecuencia en celulas de cancer de mama utilizando nanoparticulas de oro conjugadas al peptido ciclo[RGDfK(C)

    Energy Technology Data Exchange (ETDEWEB)

    Sanchez H, L.

    2014-07-01

    The conjugation of peptides to gold nanoparticles (AuNP) produces biocompatible and stable multimeric systems with target-specific molecular recognition. Peptides based on the cyclic Arg-Gly-Asp (RGD) sequence have been reported as high-affinity agents for the α(v)β(3) and α(v)β(v) integrin s over expressed in breast cancer cells. AuNP have also been proposed as localized heat sources for cancer treatment using laser irradiation or radiofrequency (RF). The objective of this research was to evaluate the thermo ablative effect of the AuNP-c [RGDfK(C)] system on MCF7 breast cancer cell viability after exposure to a radiofrequency field and to compare it with that produced by the laser irradiation. The effect of the 13.56 MHz RF (using a power from 0 to 200 W at intervals of 50 W) over the temperature increase in AuNP-colloidal system of 5 and 20 nm at two different concentrations was evaluated. The absorption cross sections (C{sub abs}) of the AuNP-c [RGDfK(C)] nano system when it interacts with low frequency electromagnetic waves (13.56 MHz, λ = 22 m) and optical frequency waves (laser at λ = 532 nm) was analyzed based on the Mi e theory. The effect on the MCF7 cell viability was assessed using two thermal conversion sources (Laser and RF) on AuNP-c [RGDfK(C)] located inside the cytoplasm of the cells. MCF7 cells were treated with AuNP-c [RGDfK(C)] or water after exposure to the RF field (200 W, 100 V/cm) or laser irradiation (Irradiance 0.65 W/cm{sup 2}). In both cases (RF and laser) the presence of nanoparticles internalized inside the cells caused a significant increase in the temperature of the medium (RF: ΔT = 29.9 ± 1.7 grades C for AuNP compared toΔT = 13.0 ± 1.4 grades C for water; laser: ΔT = 13.5 ± 0.7 grades C for AuNP compared to 3.3 ± 0.5 grades C for water). Although RF induced a higher increase in the temperature of the medium with nanoparticles, the largest effect on the cell viability was produced by laser when nanoparticles were located

  11. Comparison of Predicted pKa Values for Some Amino-Acids, Dipeptides and Tripeptides, Using COSMO-RS, ChemAxon and ACD/Labs Methods Comparaison des valeurs de pKa de quelques acides aminés, dipeptides et tripeptides, prédites en utilisant les méthodes COSMO-RS, ChemAxon et ACD/Labs

    Directory of Open Access Journals (Sweden)

    Toure O.

    2013-05-01

    Full Text Available Liquid-phase pKa values play a key role in food science. Chemical properties of molecules depend largely on whether they are ionized or not. Most organic molecules are capable of gaining and/or losing a proton in aqueous solutions. Proton transfer most. frequently occurs between water and any ionizable atom of the organic molecule. The molecule’s response to profanation or deprotonation depends significantly on the site that was disturbed by proton transfer. Partial charge distribution in the molecule also varies with protonation of the acidlbase active sites. Then it can he used to determine the pKa of a molecule. First, we use the COSMO-RS method, a combination of the quantum chemical dielectric continuum solvation model COSMO with a statistical thermodynamics treatment fin- more Realistic Solvation (RS simulations, for the direct prediction of pKa constants of about 50 molecules (amino-acids, dipeptides and tripeptides. Then, we compare our results with experimental data and the pKa values predicted using two other methods. We used respectively the ChemAxon method using a program based on the calculation of partial charge of atoms in the molecule and the ACD/Labs method that enables to calculate single pKa values. for all possible dissociation centers when the rest of the molecule is considered neutral, using an internal database containing chemical structures and their experimental pKa values. The averaged Root Mean Square Error (RMSE of the predicted pKa values for each method compared to experimental results were respectively 0.596 for COSMO-RS, 0.445 for ChemAxon and 0.490 for ACD/Labs. While ACDILabs and ChemAxon are parameterized using a large set ofexperimental data (including several of the studied molecules, the COSMO- RS method was used in a fully predictive way. Regarding these results, COSMO-RS appears as a promising method to predict the pKa values of molecules of interest in food science with scarce available pKa values such

  12. NGR和RGD配体修饰脂质体对人脐静脉内皮细胞靶向效果的比较%A comparison study of the targeting properties of NGR-liposomes and RGD-liposomes towards human umbilical vein endothelial cells

    Institute of Scientific and Technical Information of China (English)

    陈晓梅; 王殉; 黄跃; 张烜; 张强

    2009-01-01

    Endothelial cells in the angiogenic vessels of solid tumors over-express several proteins, which could be recognized by some peptide ligands. In this study, the targeting properties of two peptides, RGD (argininc-glycine-aspartic acid) and NGR (asparagine-glycine-arginine), towards human umbilical vein endothelial cells (HUVEC) were compared in vitro using doxorubicin entrapped liposomes as vehicles. The doxorubicin-loaded sterically stabilized liposomes (SSL-DOX) and RGD or NGR modified liposomes (RGD-SSL-DOX or NGR-SSL-DOX) were prepared and characterized. The studied properties included particle size, zeta potential, encapsulation efficiency and in vitro release rate. Flow eytometry, confocal microscopy and SRB assay were used on HUVEC to assess the targeting effect of the two peptides towards endothelial cells of tumor vasculature. All of the liposomes prepared in this study were obtained with encapsulation efficiencies of above 98%, particle sizes of about 65-75 nm and slight negative surface charges. The in vitro release results demonslrated that the modification of RGD or NGR did not alter the release behaviors of liposomes. It was observed in flow cytometry that the uptake of doxorubicin by HUVEC from SSL-DOX, NGR-SSL-DOX, RGD-SSL-DOX and doxorubicin solution followed the order of doxorubicin solution>RGD-SSL-DOX >NGR-SSL-DOX>SSL-DOX,and the intemalized doxorubicin distributed in both nuclei and cytoplasm for ligand modified SSL and only in nuclei for non-targeted SSL. The order of cytotoxicity in SRB assay was the same as that of the uptake study. The characterization study indicated that modifications did not significantly change the properties of the sterically stabilized liposomes. HUVEC treated with both modified liposomes showed higher uptake of doxorubicin as compared to those with SSL-DOX as a result of the receptor-mediated enducytosis. Moreover, RGD-SSL-DOX exhibited better targeting effect than NGR-SSL-DOX.%肿瘤新生血管内皮细胞特异

  13. Retargeting FX-binding-ablated HAdV-5 to vascular cells by inclusion of the RGD-4C peptide in hexon hypervariable region 7 and the HI loop.

    Science.gov (United States)

    Robertson, Stacy; Parker, Alan L; Clarke, Carolyn; Duffy, Margaret R; Alba, Raul; Nicklin, Stuart A; Baker, Andrew H

    2016-08-01

    Recent studies have generated interest in the function of human adenovirus serotype 5 (HAdV-5) hexon:  factor X (FX) binding and subsequent hepatocyte transduction and interaction with the immune system. Here, we retargeted adenovirus serotype 5 vectors, ablated for FX interaction, by replacing amino acids in hexon HVR7 with RGD-4C or inserting the peptide into the fibre HI loop. These genetic modifications in the capsid were compatible with virus assembly, and could efficiently retarget transduction of the vector via the αvβ3/5 integrin-mediated pathway, but did not alter immune recognition by pre-existing human neutralizing anti-HAdV-5 antibodies or by natural antibodies in mouse serum. Thus, FX-binding-ablated HAdV-5 can be retargeted but remain sensitive to immune-mediated attack. These findings further refine HAdV-5-based vectors for human gene therapy and inform future vector development. PMID:27189759

  14. 量子点-RGD探针光动力疗法联合吉西他滨治疗胰腺癌荷瘤裸鼠初探%Efficacy of Quantum Dots-RGD Based Photodynamic Therapy Combined with Gemcitabine for Treatment of Pancreatic Cancer Xenograft in Nude Mice

    Institute of Scientific and Technical Information of China (English)

    高双; 倪倩雯; 周敏; 徐雷鸣

    2014-01-01

    Background:Pancreatic cancer is obscure in onset and progresses rapidly with very poor prognosis. Photodynamic therapy( PDT)has been developed as a novel anti-tumor treatment modality since 1980s. At present,there are only limited researches on pancreatic cancer treated with PDT in vivo. Aims:To investigate the efficacy of quantum dots-RGD ( QDs-RGD)based PDT combined with gemcitabine for treatment of pancreatic cancer xenograft in nude mice. Methods:QDs-RGD probe was synthesized and nude mice bearing pancreatic cancer xenograft was established. Nude mice were imaged at 1,5,10 and 24 hours after injection of QDs-RGD and QDs by in vivo imaging system. Forty model nude mice were randomly divided into five groups:control group( without any treatment),simple illumination group( laser 630 nm, 120 J/cm2,20 min),PDT group(QDs-RGD 0. 5 nmol+laser irradiation),gemcitabine group(gemcitabine 50 mg/kg)and combination group(QDs-RGD 0. 5 nmol+laser irradiation+gemcitabine 50 mg/kg). All the nude mice were sacrificed 18 days later. Tumor weight and volume were measured and tumor inhibition rate was calculated. Results:Fluorescence of tumor was shown 1 hour after injection and became clearest at the 5th hour,then showing a decrescendo trend. Density of QDs surrounding tumor was significantly less than that of QDs-RGD and faded away at the 10th hour. Tumor weight and volume in PDT group,gemcitabine group and combination group were all significantly lower than those in control group and simple illumination group(P0. 05),as well as between PDT group and gemcitabine group(P >0. 05). Tumor inhibition rate in combination group,gemcitabine group and PDT group was 70. 5%,43. 5% and 37. 1%, respectively. Conclusions:QDs-RGD based PDT combined with gemcitabine can inhibit the growth of pancreatic cancer xenograft in nude mice,which introduces a new idea to the treatment of pancreatic cancer.%背景:胰腺癌发病隐匿,进展迅速,预后极差。光动力疗法( PDT)是20

  15. Dimer of the peptide cycle (Ar-Gly-Asp-D-Phe-Lys) radiolabeled with 99mTc for the integrin s over-expression image: formulation, biokinetics and dosimetry

    International Nuclear Information System (INIS)

    In breast cancer, α(v)β(3) and/or α(v)β(5) integrin s are over-expressed in both endothelial and tumour cells. Radiolabeled peptides based on the RGD (Arg-Gly-Asp) sequence are radiopharmaceuticals with high affinity and selectivity for those integrin s. The RGD-dimer peptide (E-[c(RGDfK)]2) radiolabeled with 99mTc has been reported as a radiopharmaceutical with 10-fold higher affinity for the α(v)β(3) integrin as compared to the RGD-monomer. EDDA (Ethylenediamine-N,N-diacetic acid) is a hydrophilic molecule that may favours renal excretion when used as coligand in the 99mTc labelling of HYNIC-peptides and can easily be formulated in a lyophilized kit. Aim: Establish a biokinetic model for 99mTc-EDDA/HYNIC-E-[c(RGDfK)]2 prepared from lyophilized kits and evaluate the dosimetry as breast cancer imaging agent. Methods: 99mTc labelling was performed by addition of sodium pertechnetate solution and 0.2 M phosphate buffer ph 7.0 to a lyophilized formulation containing E-[c(RGDfK)]2, EDDA, tricine, mannitol and stannous chloride. Radiochemical purity was evaluated by reversed phase HPLC and ITLC-SG analyses. Stability studies in human serum were carried out by size-exclusion HPLC. In-vitro cell uptake was tested using breast cancer cells (MCF7, T47D and MDA-MB-231) with blocked and non-blocked receptors. Biodistribution and tumour uptake were determined in MCF7 tumour-bearing nude mice with blocked and non-blocked receptors, and images were obtained using a micro-SPECT/CT. Whole-body images from seven healthy women were acquired at 1, 3, 6 and 24 h after 99mTc-EDDA/HYNIC-E-[c(RGDfK)]2 administration obtained with radiochemical purities of >94 %. Regions of interest (ROIs) were drawn around source organs on each time frame. Each ROI was converted to activity using the conjugate view counting method. The image sequence was used to extrapolate 99mTc-EDDA/HYNIC-E-[c(RGDfK)]2 time-activity curves in each organ in order to adjust the biokinetic model and to calculate the

  16. Regulation of proliferation of embryonic heart mesenchyme: Role of transforming growth factor-beta 1 and the interstitial matrix

    International Nuclear Information System (INIS)

    Proliferation of atrioventricular cushion mesenchyme of the embryonic avian heart maintained in three-dimensional aggregate culture is stimulated by interaction with the interstitial matrix. Chicken serum or transforming growth factor-beta 1, which stimulates proliferation, induces matrix deposition in regions of the aggregate showing high labeling indices with tritiated thymidine. Dispersed heart mesenchyme interstitial matrix introduced into serum-free culture is incorporated into the aggregate and stimulates cellular proliferation similar to serum or transforming growth factor-beta 1. Proliferation is reversibly inhibited by the peptide Gly-Arg-Gly-Asp-Ser-Pro. It is suggested that transforming growth factor-beta 1 stimulates the production of interstitial matrix and that a sufficient stimulus for proliferation in this system is the presence of the matrix, which acts as the adhesive support for cellular anchorage

  17. Chromopeptides from phycoerythrocyanin. Structure and linkage of the three bilin groups

    International Nuclear Information System (INIS)

    Phycoerythrocyanin carries two covalently attached phycocyanobilin (PCB) groups on the β subunit and a phycobiliviolinoid (PXB) group on the α subunit. Three distinct bilipeptides were obtained by proteolytic digestion of this protein: Asn-Gln-Ala-Ala-Cys(PCB)-Ile-Arg, Gly-Asp-Cys(PCB)-Ser-Gln, and Cys(PXB)-Val-Arg. Correlation 500-MHz 1H NMR analyses showed that the heptapeptide and pentapeptide were attached by cysteinyl thioether linkage to the A ring of the PCB moiety. 1H NMR and mass spectrometry determinations led to structural assignment for the hitherto uncharacterized PXB moiety, with peptide-thioether bonding possible to either ring A or D. Amino acid sequence homologies strongly favor A-ring linkage

  18. A composite role of vitronectin and urokinase in the modulation of cell morphology upon expression of the urokinase receptor

    DEFF Research Database (Denmark)

    Engelholm, L.H.; Ingvarsen, S.; Madsen, D.H.;

    2008-01-01

    a different single-site mutant, uPAR(Y57A), in the presence of a synthetic uPAR-binding peptide, as well as with wild-type uPAR, which underwent cytoskeletal rearrangements even when cultivated in uPA-deficient serum. Blocking of integrins with an Arg-Gly-Asp-containing peptide counteracted the matrix...... adhesion and cytoskeletal rearrangements, whereas a mutant uPAR, uPAR(W32A) with defective vitronectin binding, failed to induce both phenomena. However, upon saturation of uPAR(W32A) with the protease ligand, pro-uPA, or its receptor-binding domain, the ability to induce cytoskeletal rearrangements......, and this event is followed by downstream effects including changes in the cytoskeletal organization. However, it remains unclear whether the adhesion through uPAR-vitronectin is the only event capable of initiating these morphological rearrangements or whether lateral interactions between uPAR and integrins can...

  19. Chromopeptides from phycoerythrocyanin. Structure and linkage of the three bilin groups. [Mastigocladus laminosus; Anabaena variabilis

    Energy Technology Data Exchange (ETDEWEB)

    Bishop, J.E.; Rapoport, H.; Klotz, A.V.; Chan, C.F.; Glazer, A.N.; Fueglistaller, P.; Zuber, H.

    1987-01-01

    Phycoerythrocyanin carries two covalently attached phycocyanobilin (PCB) groups on the ..beta.. subunit and a phycobiliviolinoid (PXB) group on the ..cap alpha.. subunit. Three distinct bilipeptides were obtained by proteolytic digestion of this protein: Asn-Gln-Ala-Ala-Cys(PCB)-Ile-Arg, Gly-Asp-Cys(PCB)-Ser-Gln, and Cys(PXB)-Val-Arg. Correlation 500-MHz /sup 1/H NMR analyses showed that the heptapeptide and pentapeptide were attached by cysteinyl thioether linkage to the A ring of the PCB moiety. /sup 1/H NMR and mass spectrometry determinations led to structural assignment for the hitherto uncharacterized PXB moiety, with peptide-thioether bonding possible to either ring A or D. Amino acid sequence homologies strongly favor A-ring linkage.

  20. Identification by Site-directed Mutagenesis of Amino Acid Residues Flanking RGD Motifs of Snake Venom Disintegrins for Their Structure and Function%用定点诱变技术鉴定蛇毒解联蛋白RGD侧翼氨基酸残基的结构和功能

    Institute of Scientific and Technical Information of China (English)

    徐存拴; RAHMAN Salman

    2001-01-01

    In order to demonstrate that the amino acid residues flanking the RGD sequence were important for inhibiting the ADP-induced platelet aggregation, we analyzed the role of the amino acid residues in the domain preceding the RGD loop on the activity of disintegrins. Our approach was to develop hybrids between the disintegrins kistrin and elegantin targeting residues in this domain and within the RGD loop. The basic sequence within elegantin KKKR44T45I46/A50RGDN54P55 was changed by mutagensis to KAG44T45I46/P50 RGDM54P55 and to SKAG44I46/P50RGDM54P55, thereby resembling the corresponding S39 RAGT43/P50 RGDM52 P53 sequence in kistrin. This changed KKKR44 T45 I46/A50RGDN54P55→SKAG44T45 I46/P50RGDM54 P55 dramatically reduced the activity of elegantin as an inhibitor of platelet aggregation. In contrast, deletion of T45 (KKKR44T45 I46/A50RGDN54P55→SKAG44 T45 I46/P50RGDM54P55) increased activity of elegantin as an inhibitor platelet aggregation. It was further shown that their electrophoresis properties were very different. These data highlight thc importance of the domain encompassing residues 39--45 and the amimo acid residues flanking the RGD sequence on disintegrin structure-function.%为了证实蛇毒蛋白RGD侧翼氨基酸残基对ADP诱导的血小板凝聚抑制作用的重要性,分析了RGD环上游氨基酸残基对解联蛋白活性的影响。将位于两种解联蛋白蝮蛇毒素(kistrin)和华丽蛇毒素(elegantin)之间的氨基酸残基和RGD环内的氨基酸残基进行杂交。用定点诱变技术将elegantin一级结构内的KKKR44T45I46/A50RGDN54P55分别突变为SKAG44T45I46/P50RGDM54P55和SKAG44I46/P50RGDM54P55,这些序列和kistrin中相应的序列S39RAGT43/P50RGDM52P53有相似之处,序列由KKKR44T45I46/A50RGDN54P55突变为SKAG44T45I46/P50RGDM54P55,显著地降低了elegantin对血小板凝聚的抑制因子的活性,而T45的缺失(KKKR44T45I46/A50RGDN54P55突变为SKAG44I46/P50RGDM54P55)却增强了elegantin对血小板

  1. The human tri-peptide GHK and tissue remodeling.

    Science.gov (United States)

    Pickart, Loren

    2008-01-01

    Tissue remodeling follows the initial phase of wound healing and stops inflammatory and scar-forming processes, then restores the normal tissue morphology. The human peptide Gly-(L-His)-(L-Lys) or GHK, has a copper 2+ (Cu(2+)) affinity similar to the copper transport site on albumin and forms GHK-Cu, a complex with Cu(2+). These two molecules activate a plethora of remodeling related processes: (1) chemoattraction of repair cells such as macrophages, mast cells, capillary cells; (2) anti-inflammatory actions (suppression of free radicals, thromboxane formation, release of oxidizing iron, transforming growth factor beta-1, tumor necrosis factor alpha and protein glycation while increasing superoxide dismutase, vessel vasodilation, blocking ultraviolet damage to skin keratinocytes and improving fibroblast recovery after X-ray treatments); (3) increases protein synthesis of collagen, elastin, metalloproteinases, anti-proteases, vascular endothelial growth factor, fibroblast growth factor 2, nerve growth factor, neutrotropins 3 and 4, and erythropoietin; (4) increases the proliferation of fibroblasts and keratinocytes; nerve outgrowth, angiogenesis, and hair follicle size. GHK-Cu stimulates wound healing in numerous models and in humans. Controlled studies on aged skin demonstrated that it tightens skin, improves elasticity and firmness, reduces fine lines, wrinkles, photodamage and hyperpigmentation. GHK-Cu also improves hair transplant success, protects hepatic tissue from tetrachloromethane poisoning, blocks stomach ulcer development, and heals intestinal ulcers and bone tissue. These results are beginning to define the complex biochemical processes that regulate tissue remodeling. PMID:18644225

  2. Aggregation behavior of a gemini surfactant with a tripeptide spacer.

    Science.gov (United States)

    Wang, Meina; Han, Yuchun; Qiao, Fulin; Wang, Yilin

    2015-02-28

    A peptide gemini surfactant, 12-G(NH2)LG(NH2)-12, has been constructed with two dodecyl chains separately attached to the two terminals of a glutamic acid-lysine-glutamic acid peptide and the aggregation behavior of the surfactant was studied in aqueous solution. The 12-G(NH2)LG(NH2)-12 molecules form fiber-like precipitates around pH 7.0, and the precipitation range is widened on increasing the concentration. At pHs 3.0 and 11.0, 12-G(NH2)LG(NH2)-12 forms soluble aggregates because each molecule carries two positively charged amino groups at the two ends of the peptide spacer at pH 3.0, while each molecule carries one negatively charged carboxyl group in the middle of the peptide spacer at pH 11.0. 12-G(NH2)LG(NH2)-12 displays a similar concentration-dependent process at these two pHs: forming small micelles above the critical micelle concentration and transferring to fibers at pH 3.0 or twisted ribbons at pH 11.0 above the second critical concentration. The fibers formed at pH 3.0 tend to aggregate into bundles with twisted structure. Both the twisted fibers at pH 3.0 and the twisted ribbons at pH 11.0 contain β-sheet structure formed by the peptide spacer. PMID:25588349

  3. 99Tcm标记的PEG4/2PEG4修饰的环状RGD二聚体体内外性质的对比%Comparison of in Vitro and in Vivo Characteristics of 99Tcm-Labeled Cyclic RGD Dimers With PEG4/2PEG4 Linkers

    Institute of Scientific and Technical Information of China (English)

    靳存敬; 史继云; 刘妍; 靳晓娜; 黄金铭; 赵慧云; 李方; 王凡

    2010-01-01

    采用2种方法修饰RGD环肽二聚体,在2个RGD模序之间引入2个PEG4分子,以及在RGD二聚体与双功能螯合剂HYNIC之间引入1个PEG4分子,比较在不同部位引入PEG4分子对99Tcm标记的RGD环肽二聚体体内外性质的影响.研究结果表明,在2个RGD模序之间引入2个PEG4分子能显著提高RGD环肽二聚体与整合素αvβ3的亲和力;PEG4的引入能够增强标记物的水溶性.99Tcm-HYNIC-2PEG4-RGD dimer具有更好的体内外性质,作为一种潜在的肿瘤显像剂,具有进一步研究开发的价值.

  4. Development of a specific radiopharmaceutical based on gold nanoparticles functionalized with HYNIC-peptide/mannose for the sentinel lymph node detection in breast cancer; Desarrollo de un radiofarmaco especifico basado en nanoparticulas de oro funcionalizadas con HYNIC-peptido/manosa para la deteccion de ganglio centinela en cancer de mama

    Energy Technology Data Exchange (ETDEWEB)

    Ocampo G, B. E.

    2012-07-01

    ) which was retained during 24 h with minimal kidney accumulation (0.98 {+-} 0.10% Id) and negligible uptake in all other tissues. In order to design a pharmaceutical formulation for the instant preparation of stable m ultimeric systems with target-specific molecular recognition based on gold nanoparticles, a freeze-dried kit formulation of {sup 99m}Tc-ethylenediamine-N, N-diacetic acid (EDDA)/hydrazino nicotinyl (HYNIC)-Tyr{sup 3}-octreotide ({sup 99m}Tc-EDDA/HYNIC-TOC, previously approved by the Mexican Ministry of Health) (vial 1) and a second vial containing 1.5 ml of Au-Np solution plus 10 {mu}L of thiol-mannose, Lys{sup 3}-bombesin, or cyclo[Arg-Gly-Asp-D-Phe-Lys-(Cys)] (c[RGDfK(C)] (approximately 285 molecules per Au-Np) (vial 2) were prepared. M ultimeric radiopharmaceuticals prepared from kit showed a radiochemical purity of 96 {+-} 2%. The far-infrared spectra showed a characteristic band at 279 {+-} 1 cm{sup -1}, which was assigned to the Au-S bond. UV-Vis and XP S also indicated that the Au-Np were functionalized with peptides or mannose. Radiopharmaceuticals showed specific recognition for receptors expressed in cancer cells or rat liver cells. Micro-SPECT/CT images showed clear tumour uptake and lymph node accumulation. The kit demonstrated excellent stability during storage at 4 C for 6 months. This study demonstrated that {sup 99m}Tc-Au-Np-mannose remains within the first lymph node during 24 h and therefore might be useful as a target-specific radiopharmaceutical for SLND using 1-day or 2-day conventional protocols. Likewise, m ultimeric systems of {sup 99m}Tc-Au-Np-mannose, {sup 99m}Tc-Au-Np-RGD and {sup 99m}Tc-Au-Np-Lys{sup 3}-bombesin prepared from kits exhibited properties suitable as target-specific agents for molecular imaging of tumours and sentinel lymph node. (Author)

  5. Development of a specific radiopharmaceutical based on gold nanoparticles functionalized with HYNIC-peptide/mannose for the sentinel lymph node detection in breast cancer

    International Nuclear Information System (INIS)

    kidney accumulation (0.98 ± 0.10% Id) and negligible uptake in all other tissues. In order to design a pharmaceutical formulation for the instant preparation of stable m ultimeric systems with target-specific molecular recognition based on gold nanoparticles, a freeze-dried kit formulation of 99mTc-ethylenediamine-N, N-diacetic acid (EDDA)/hydrazino nicotinyl (HYNIC)-Tyr3-octreotide (99mTc-EDDA/HYNIC-TOC, previously approved by the Mexican Ministry of Health) (vial 1) and a second vial containing 1.5 ml of Au-Np solution plus 10 μL of thiol-mannose, Lys3-bombesin, or cyclo[Arg-Gly-Asp-D-Phe-Lys-(Cys)] (c[RGDfK(C)] (approximately 285 molecules per Au-Np) (vial 2) were prepared. M ultimeric radiopharmaceuticals prepared from kit showed a radiochemical purity of 96 ± 2%. The far-infrared spectra showed a characteristic band at 279 ± 1 cm-1, which was assigned to the Au-S bond. UV-Vis and XP S also indicated that the Au-Np were functionalized with peptides or mannose. Radiopharmaceuticals showed specific recognition for receptors expressed in cancer cells or rat liver cells. Micro-SPECT/CT images showed clear tumour uptake and lymph node accumulation. The kit demonstrated excellent stability during storage at 4 C for 6 months. This study demonstrated that 99mTc-Au-Np-mannose remains within the first lymph node during 24 h and therefore might be useful as a target-specific radiopharmaceutical for SLND using 1-day or 2-day conventional protocols. Likewise, m ultimeric systems of 99mTc-Au-Np-mannose, 99mTc-Au-Np-RGD and 99mTc-Au-Np-Lys3-bombesin prepared from kits exhibited properties suitable as target-specific agents for molecular imaging of tumours and sentinel lymph node. (Author)

  6. Bioimprinted Polymer Scaffolds for Selective Recognition of RGD Peptides

    Science.gov (United States)

    Bergmann, Nicole; Peppas, Nicholas A.

    2003-03-01

    Fibronectin and a number of other plasma and extracellular matrix (ECM) adhesion proteins contain the tetrapeptide arginine-glycine-aspartic acid-serine (RGDS), and this sequence can be summarily recognized and bound by integrins present on cell membranes. Upon integrin binding, cells adhere to the substrate, and this adherence encourages ECM deposition and other cellular remodeling events. By targeting specific chemical functional groups on the peptide using non-covalent molecular imprinting, biomimetic polymeric scaffolds can be designed to mimic protein-ECM binding both on the surface and in the bulk during polymer degradation. Methacrylic acid-ethylene glycol dimethacrylate (MAA-g-EGDMA) copolymer films were prepared by free-radical ultraviolet polymerization in the presence of RGDS to create novel imprinted matrices for possible tissue engineering scaffolds. SEM analysis revealed a highly macroporous structure in peptide-imprinted polymers compared to controls. Optimal crosslinking ratios for peptide imprinting were determined using a small molecular weight fluorescent tag, 4-chloro-7-nitrobenzofurazan, and analyzed using fluorescent microscopy. Higher crosslinking ratios yielded better template recognition and gels exhibited specific recognition in aqueous media to RGDS molecules when in the presence of similar tetrapeptides.

  7. Macrocyclic Chelator Assembled RGD Multimers for Tumor Targeting

    OpenAIRE

    Zhang, Xiaofen; Liu, Hongguang; Miao, Zheng; Kimura, Richard; Fan, Feiyue; Cheng, Zhen

    2011-01-01

    Macrocyclic chelators have been extensively used for complexation of metal ions. A widely used chelator, DOTA, has been explored as a molecular platform to assemble multiple bioactive peptides in this paper. The multivalent DOTA-peptide bioconjugates demonstrate promising tumor targeting ability.

  8. Biodistribution in healthy KM mice and micro PET/CT imaging in U87MG tumor-bearing nude mice of a new 18F-labeled cyclic RGD dimer%新型18F-RGD二聚体的正常生物分布及U87MG荷瘤裸鼠小动物PET/CT显像研究

    Institute of Scientific and Technical Information of China (English)

    2013-01-01

    Background and purpose:Integrinαvβ3 receptor plays an important role in promoting, sustaining and regulating the angiogenesis. It is overexpressed on neovascular endothelial cells and tumor cells. RGD peptide specifically binds to integrinαvβ3, which could evaluate growth status and invasiveness of tumor. This study aimed to investigate the biodistribution in healthy KM mice and micro PET/CT imaging in U87MG tumor-bearing mice of 18F-E[c(RGDfK)2]. Methods: 18F-E[c(RGDfK)2] was produced using an automated synthesis module via a simple one-step 18F-labeling strategy of the precursor 4-NO2-3-TFMBz-E[c(RGDfK)2]. The percentage activity of injection dose per gram of tissue (%ID/g) was calculated at 0.5, 1, 2, 4 h post injection of the probe. Micro PET/CT images of U87MG tumor-bearing nude mice with or without 18F-E[c(RGDfK)2] blocking were acquired at each time point. Results: The labeling efficiency and radiochemical purity of 18F-E[c(RGDfK)2] were 10% and 98%, respectively. 18F-E[c(RGDfK)2] was excreted via renal route, with a high blood clearance. The other organs had background-level activity accumulation. At 1 h, the%ID/g of kidney, liver, intestine, muscle and blood was (1.02±0.16)%ID/g,(0.24±0.06)%ID/g, (0.35±0.03)%ID/g, (0.13±0.03)%ID/g and (0.11±0.03)%ID/g 18F-E[c(RGDfK)2] had initial high tumor uptake [(5.2±0.56)%ID/g] and good tumor-to-background contrast (5.36) at 1 h post injection. Tumor uptake for blocking group was lower than those without blocking, and T/M reduced to 1.57. Conclusion: 18F-E[c(RGDfK)2] appears a promising PET molecular imaging probe targeting integrin αvβ3, with high tumor uptake. It could be suitable for prognosis evaluation of integrin-positive tumor, selection of vascular targeting therapy and therapy effect monitoring.%  背景与目的:整合素αvβ3受体在促进、维持以及调节血管生成的过程中有着至关重要的作用,高表达于多种肿瘤细胞及新生血管内皮细胞。RGD多肽

  9. Targeting ligand-functionalized and redox-sensitive heparin-Pluronic nanogels for intracellular protein delivery

    Energy Technology Data Exchange (ETDEWEB)

    Nguyen, Dai Hai; Joung, Yoon Ki; Choi, Jong Hoon; Park, Ki Dong [Department of Molecular Science and Technology, Ajou University, 5 Wonchon, Yeoungtong, Suwon 443-749 (Korea, Republic of); Moon, Hyun Tae, E-mail: kdp@ajou.ac.kr [Research Institute of Pharmaceutical Science, College of Pharmacy, Seoul National University, Seoul 151-742 (Korea, Republic of)

    2011-10-15

    The heparin-Pluronic (HP) conjugate was coupled via redox-sensitive disulfide bond and contains a vinyl sulfone (VS) group with high reactivity to some functional groups such as thiol group. Heparin was conjugated with cystamine and the terminal hydroxyl groups of Pluronic were activated with the VS group, followed by coupling of VS groups of Pluronic with cystamine of heparin. The chemical structure, heparin content and VS group content of the resulting product were determined by {sup 1}H NMR, FT-IR, toluidine blue assay and Ellman's method. The HP conjugate formed a type of nanogel in an aqueous medium, showing a critical micelle concentration of approximately 129.35 mg L{sup -1}, a spherical shape and the mean diameter of 115.7 nm, which were measured by AFM and DLS. The release test demonstrated that HP nanogel was rapidly degraded when treated with glutathione. Cytotoxicity results showed a higher viability of drug-free HP nanogel than that of drug-loaded one. Cyclo(Arg-Gly-Asp-D-Phe-Cys) (cRGDfC) peptide was efficiently conjugated to VS groups of HP nanogel and exhibited higher cellular uptake than unmodified nanogels. All results suggest a novel multi-functional nanocarrier delivery and effective release of proteins to the intracellular region in a redox-sensitive manner.

  10. Combinational Effect of Cell Adhesion Biomolecules and Their Immobilized Polymer Property to Enhance Cell-Selective Adhesion

    Directory of Open Access Journals (Sweden)

    Rio Kurimoto

    2016-01-01

    Full Text Available Although surface immobilization of medical devices with bioactive molecules is one of the most widely used strategies to improve biocompatibility, the physicochemical properties of the biomaterials significantly impact the activity of the immobilized molecules. Herein we investigate the combinational effects of cell-selective biomolecules and the hydrophobicity/hydrophilicity of the polymeric substrate on selective adhesion of endothelial cells (ECs, fibroblasts (FBs, and smooth muscle cells (SMCs. To control the polymeric substrate, biomolecules are immobilized on thermoresponsive poly(N-isopropylacrylamide-co-2-carboxyisopropylacrylamide (poly(NIPAAm-co-CIPAAm-grafted glass surfaces. By switching the molecular conformation of the biomolecule-immobilized polymers, the cell-selective adhesion performances are evaluated. In case of RGDS (Arg-Gly-Asp-Ser peptide-immobilized surfaces, all cell types adhere well regardless of the surface hydrophobicity. On the other hand, a tri-Arg-immobilized surface exhibits FB-selectivity when the surface is hydrophilic. Additionally, a tri-Ile-immobilized surface exhibits EC-selective cell adhesion when the surface is hydrophobic. We believe that the proposed concept, which is used to investigate the biomolecule-immobilized surface combination, is important to produce new biomaterials, which are highly demanded for medical implants and tissue engineering.

  11. Molecular dynamics and docking simulation of a natural variant of Activated Protein C with impaired protease activity: implications for integrin-mediated antiseptic function.

    Science.gov (United States)

    D'Ursi, Pasqualina; Orro, Alessandro; Morra, Giulia; Moscatelli, Marco; Trombetti, Gabriele; Milanesi, Luciano; Rovida, Ermanna

    2015-01-01

    Activated Protein C (APC) is a multifunctional serine protease, primarily known for its anticoagulant function in the coagulation system. Several studies have already elucidated its role in counteracting apoptosis and inflammation in cells, while significant effort is still ongoing for defining its involvement in sepsis. Earlier literature has shown that the antiseptic function of APC is mediated by its binding to leukocyte integrins, which is due to the presence of the integrin binding motif Arg-Gly-Asp at the N-terminus of the APC catalytic chain. Many natural mutants have been identified in patients with Protein C deficiency diagnosis including a variant of specificity pocket (Gly216Asp). In this work, we present a molecular model of the complex of APC with αVβ3 integrin obtained by protein-protein docking approach. A computational analysis of this variant is hereby presented, based on molecular dynamics and docking simulations, aiming at investigating the effects of the Gly216Asp mutation on the protein conformation and inferring its functional implications. Our study shows that such mutation is likely to impair the protease activity while preserving the overall protein fold. Moreover, superposition of the integrin binding motifs in wild-type and mutant forms suggests that the interaction with integrin can still occur and thus the mutant is likely to retain its antiseptic function related to the neutrophyl integrin binding. Therapeutic applications could result in this APC mutant which retains antiseptic function without anticoagulant side effects.

  12. Carbon-Dot-Decorated Carbon Nitride Nanoparticles for Enhanced Photodynamic Therapy against Hypoxic Tumor via Water Splitting.

    Science.gov (United States)

    Zheng, Di-Wei; Li, Bin; Li, Chu-Xin; Fan, Jin-Xuan; Lei, Qi; Li, Cao; Xu, Zushun; Zhang, Xian-Zheng

    2016-09-27

    Hypoxia, a typical feature of solid tumors, remarkably restricts the efficiency of photodynamic therapy (PDT). Here, a carbon nitride (C3N4)-based multifunctional nanocomposite (PCCN) for light-driven water splitting was used to solve this problem. Carbon dots were first doped with C3N4 to enhance its red region absorption because red light could be used to trigger the in vivo water splitting process. Then, a polymer containing a protoporphyrin photosensitizer, a polyethylene glycol segment, and a targeting Arg-Gly-Asp motif was synthesized and introduced to carbon-dot-doped C3N4 nanoparticles. In vitro study showed that PCCN, thus obtained, could increase the intracellular O2 concentration and improve the reactive oxygen species generation in both hypoxic and normoxic environments upon light irradiation. Cell viability assay demonstrated that PCCN fully reversed the hypoxia-triggered PDT resistance, presenting a satisfactory growth inhibition of cancer cells in an O2 concentration of 1%. In vivo experiments also indicated that PCCN had superior ability to overcome tumor hypoxia. The use of water splitting materials exhibited great potential to improve the intratumoral oxygen level and ultimately reverse the hypoxia-triggered PDT resistance and tumor metastasis.

  13. Fabrication of Thermo-Responsive Molecular Layers from Self-Assembling Elastin-Like Oligopeptides Containing Cell-Binding Domain for Tissue Engineering

    Directory of Open Access Journals (Sweden)

    Tomoyuki Koga

    2015-01-01

    Full Text Available Novel thermo-responsive elastin-like oligopeptides containing cell-binding epitope (Arg-Gly-Asp-Ser sequence; arginine-glycine-aspartic acid-serine (RGDS-elastin-like peptides (ELP and RGDS-deg-ELP; were newly prepared as building blocks of self-assembled molecular layer for artificial extra cellular matrix. A detailed analysis of the conformation of the oligo(ELPs in water and their self-assembling behavior onto hydrophobic surfaces were performed by using circular dichroism, Fourier transform infrared spectroscopy (FTIR, atomic force microscopy and water contact angle measurements. The experimental results revealed that both oligo(ELPs self-assembled onto hydrophobic surfaces and formed molecular layers based on their thermo-responsive conformational change from hydrous random coil to dehydrated β-turn structure. Effective cell adhesion and spreading behaviors were observed on these self-assembled oligo(ELP layers. In addition, attached cells were found to be recovered successfully as a cell-sheet by temperature-induced disassembly of oligo(ELP layer. This achievement provides an important insight to construct novel oligopeptide-based nano-surfaces for the design of smart artificial extra-cellular matrix.

  14. Gracilaria lemaneiformis polysaccharide as integrin-targeting surface decorator of selenium nanoparticles to achieve enhanced anticancer efficacy.

    Science.gov (United States)

    Jiang, Wenting; Fu, Yuanting; Yang, Fang; Yang, Yufeng; Liu, Ting; Zheng, Wenjie; Zeng, Lilan; Chen, Tianfeng

    2014-08-27

    The poor permeability of glioma parenchyma represents a major limit for antiglioblastoma drug delivery. Gracilaria lemaneiformis polysaccharide (GLP), which has a high binding affinity to αvβ3 integrin overexpressed in glioma cells, was employed in the present study to functionalize selenium nanoparticles (SeNPs) to achieve antiglioblastoma efficacy. GLP-SeNPs showed satisfactory size distribution, high stability, and selectivity between cancer and normal cells. In U87 glioma cell membrane, which has a high integrin expression level, GLP-SeNPs exhibited significantly higher cellular uptake than unmodified SeNPs. As expected, U87 cells exhibited a greater uptake of GLP-SeNPs than C6 cells with low integrin expression level. Furthermore, the internalization of GLP-SeNPs was inhibited by cyclo-(Arg-Gly-Asp-Phe-Lys) peptides, suggesting that cellular uptake into U87 cells and C6 cells occurred via αvβ3 integrin-mediated endocytosis. For U87 cells, the cytotoxicity of SeNPs decorated by GLP was enhanced significantly because of the induction of various apoptosis signaling pathways. Internalized GLP-SeNPs triggered intracellular reactive oxygen species downregulation. Therefore, p53, MAPKs, and AKT pathways were activated to advance cell apoptosis. These findings suggest that surface decoration of nanomaterials with GLP could be an efficient strategy for design and preparation of glioblastoma targeting nanodrugs.

  15. Acid-responsive PEGylated doxorubicin prodrug nanoparticles for neuropilin-1 receptor-mediated targeted drug delivery.

    Science.gov (United States)

    Song, Huijuan; Zhang, Ju; Wang, Weiwei; Huang, Pingsheng; Zhang, Yumin; Liu, Jianfeng; Li, Chen; Kong, Deling

    2015-12-01

    Self-assembled prodrug nanoparticles have demonstrated great promise in cancer chemotherapy. In the present study, we developed a new kind of prodrug nanoparticles for targeted drug delivery. PEGylated doxorubicin conjugate with an acid-cleavable cis-aconityl spacer was prepared. Then it was functionalized with a tumor-penetrating peptide, Cys-Arg-Gly-Asp-Lys (CRGDK), providing the prodrug nanoparticles with the specific binding ability to neurophilin-1 receptor. In acid mediums, doxorubicin could be released from the prodrug nanoparticles with an accumulative release around 60% through the acid-triggered hydrolysis of cis-aconityl bond and nanoparticle disassembly. Whereas, drug release was slow under a neutral pH and the accumulative drug release was less than 16%. In the cell culture tests, our prodrug nanoparticles showed enhanced endocytosis and cytotoxicity in cancer cells including HepG2, MCF-7 and MDA-MB-231 cells, but lower cytotoxicity in human cardiomyocyte H2C9. In the animal experiments, the prodrug nanoparticles were intravenously injected into Balb/c nude mice bearing MDA-MB-231 tumors. Enhanced drug penetration and accumulation in tumors, accompanying with a rapid early tumor-binding behavior, was observed after intravenous injection of the peptide modified prodrug nanoparticles. These data suggests that the acid-sensitive and tumor-targeting PEGylated doxorubicin prodrug nanoparticle may be an efficient drug delivery system for cancer chemotherapy.

  16. Experimental Study on Transarterial Administration of GRGDSP Combined with Transarterial Chemoembolization in Rats with Hepatic Carcinoma

    International Nuclear Information System (INIS)

    Purpose. To evaluate the effects of transarterial administration of an integrin antagonist, GRGDSP (Gly-Arg-Gly-Asp-Ser-Pro), combined with transarterial chemoembolization (TACE) to treat hepatic carcinoma in rats. Methods. Walker-256 tumor was implanted beneath the liver capsule in 26 Wistar rats. Animal subjects were assigned to groups based on which treatment was injected into the hepatic artery: group A, GRGDSP + TACE; group B, TACE alone; and group C, normal saline. Magnetic resonance imaging (MRI), tumor pathology, and immunohistochemistry were performed to assess each treatment. Results. The ratios of the post-treatment to pretreatment tumor volumes (V2/V1) in groups A, B, and C were 4.42 ± 0.48, 6.98 ± 1.09, and 13.00 ± 1.68, respectively. The metastatic potential of the tumors was assessed by tumor cell nest counts, which were 5.00 ± 1.25, 6.63 ± 1.60, and 7.22 ± 1.92 in groups A, B, and C, respectively. Microvessel density (MVD) was quantified by measuring von Willebrand factor density values, which were 0.18 ± 0.02, 0.22 ± 0.02, and 0.23 ± 0.02 in groups A, B, and C, respectively. Conclusions. Transarterial infusion of GRGDSP combined with TACE noticeably inhibited the growth of hepatic carcinoma and intrahepatic metastases in rats

  17. Tailored polymer-lipid hybrid nanoparticles for the delivery of drug conjugate: dual strategy for brain targeting.

    Science.gov (United States)

    Agrawal, Udita; Chashoo, Gousia; Sharma, Parduman Raj; Kumar, Ashok; Saxena, Ajit Kumar; Vyas, S P

    2015-02-01

    The object of the present study was to investigate the glioma targeting propensity of folic acid (F) decorated polymer-lipid hybrid nanoparticles (PLNs) encapsulating cyclo-[Arg-Gly-Asp-D-Phe-Lys] (cRGDfK) modified paclitaxel (PtxR-FPLNs). The prepared PLNs were supposed to bypass the blood-brain barrier (BBB) efficiently and subsequently target integrin rich glioma cells. The developed formulations were characterized for size, shape, drug entrapment efficiency, and in vitro release profile. PtxR-FPLNs demonstrated highest in vitro inhibitory effect, cell apoptosis and cell uptake. Pharmacokinetics and biodistribution studies showed efficacy of PtxR-FPLNs in vivo. In vivo anti-tumor studies clearly revealed that the median survival time for Balb/C mice treated with PtxR-FPLNs (42 days) was extended significantly as compared to PtxR-PLNs (35 days), free PtxR (18 days), Ptx-FPLNs (38 days), Ptx-PLNs (30 days), free Ptx (14 days) and control group (12 days). From the results it can be concluded that the developed dual targeted nanoformulation was able to efficiently cross the BBB and significantly deliver higher amounts of drug to brain tumor for better therapeutic outcome.

  18. Fibrillin-1 mutations causing Weill-Marchesani syndrome and acromicric and geleophysic dysplasias disrupt heparan sulfate interactions.

    Directory of Open Access Journals (Sweden)

    Stuart A Cain

    Full Text Available The extracellular glycoprotein fibrillin-1 forms microfibrils that act as the template for elastic fibers. Most mutations in fibrillin-1 cause Marfan syndrome with severe cardiovascular and ocular symptoms, and tall stature. This is in contrast to mutations within a heparin-binding TB domain (TB5, which is downstream of the arg-gly-asp cell adhesion domain, which can cause Weill-Marchesani syndrome (WMS or Acromicric (AD and Geleophysic Dysplasias (GD. WMS is characterized by short limbs, joint stiffness and ocular defects, whilst fibrillin-1 AD and GD have severe short stature, joint defects and thickened skin. We previously showed that TB5 binds heparin. Here, we show that the corresponding region of fibrillin-2 binds heparin very poorly, highlighting a novel functional difference between the two isoforms. This finding enabled us to map heparin/heparan sulfate binding to two sites on fibrillin-1 TB5 using a mutagenesis approach. Once these sites were mapped, we were able to investigate whether disease-causing mutations in this domain disrupt binding to HS. We show that a WMS deletion mutant, and five AD and GD point mutants all have disrupted heparin binding to TB5. These data provide insights into the biology of fibrillins and the pathologies of WMS, AD and GD.

  19. Enhanced Intracellular Uptake of CdTe Quantum Dots by Conjugation of Oligopeptides

    Directory of Open Access Journals (Sweden)

    Moon-Jeong Choi

    2013-01-01

    Full Text Available Arg-Gly-Asp-Ser (RGDS, a typical membrane-permeable carrier peptide, was conjugated with mercaptoisobutyric acid-immobilized CdTe quantum dot (CTNPs to enhance the intracellular uptake of quantum dots. Mean size of mercaptoisobutyric acid-immobilized quantum dots (37 nm as determined by dynamic light scattering was increased up to 54 nm after RGDS immobilization. It was found, from in vitro cell culture experiment, that fibroblast (NIH 3T3 cells were well proliferated in the presence of RGDS-conjugated quantum dots (RCTNPs, and the intracellular uptake of CTNPs and RCTNPs was studied by means of ICP and fluorescence microscopy. As a result, the RCTNPs specifically bound to the membrane of NIH 3T3 cells and almost saturated after 6 hours incubation. The amount of RCTNPs uptaken by the cells was higher than that of CTNPs, demonstrating the enhancing effect of RGDS peptide conjugation on the intracellular uptake of quantum dots (QDs.

  20. Surface modifications of photocrosslinked biodegradable elastomers and their influence on smooth muscle cell adhesion and proliferation.

    Science.gov (United States)

    Ilagan, Bernadette G; Amsden, Brian G

    2009-09-01

    Photocrosslinked, biodegradable elastomers based on aliphatic polyesters have many desirable features as scaffolds for smooth muscle tissue engineering. However, they lack cell adhesion motifs. To address this shortcoming, two different modification procedures were studied utilizing a high and a low crosslink density elastomer: base etching and the incorporation of acryloyl-poly(ethylene glycol) (PEG)-Gly-Arg-Gly-Asp-Ser (GRGDS) into the elastomer network during photocrosslinking. Base etching improved surface hydrophilicity without altering surface topography, but did not improve bovine aortic smooth muscle cell adhesion. Incorporation of PEG-GRGDS into the elastomer network significantly improved cell adhesion for both high and low crosslink density elastomers, with a greater effect with the higher crosslink density elastomer. Incorporation of GRGDS into the high crosslink density elastomer also enhanced smooth muscle cell proliferation, while proliferation on the low crosslink density unmodified, base etched, and PEG-GRGDS incorporated elastomers was significantly greater than on the high crosslink density unmodified and base etched elastomer. PMID:19375999

  1. Fibrin, a scaffold material for islet transplantation and pancreatic endocrine tissue engineering.

    Science.gov (United States)

    Riopel, Matthew; Trinder, Mark; Wang, Rennian

    2015-02-01

    Fibrin is derived from fibrinogen during injury to produce a blood clot and thus promote wound repair. Composed of different domains, including Arg-Gly-Asp amino acid motifs, fibrin is used extensively as a hydrogel and sealant in the clinic. By binding to cell surface receptors like integrins and acting as a supportive 3D scaffold, fibrin has been useful in promoting cell differentiation, proliferation, function, and survival. In particular, fibrin has been able to maintain islet cell architecture, promote beta cell insulin secretion, and islet angiogenesis, as well as inducing a protective effect against cell death. During islet transplantation, fibrin improved neovascularization and islet function. These improvements resulted in reduced number of transplanted islets necessary to reverse diabetes. Therefore, fibrin, as a biocompatible and biodegradable scaffold, should be considered during subcutaneous islet transplantation and beta cell expansion in vitro to ensure maintenance of islet cell function, proliferation, and survival to develop effective cell-based therapies for the treatment of diabetes. PMID:24947304

  2. Peptide modified polymer poly (glycerol- dodecanedioate co-fumarate) for efficient control of motor neuron differentiation.

    Science.gov (United States)

    Dai, Xizi; Huang, Yen-Chih; Leichner, Jared; Nair, Madhvan; Lin, Wei-Chiang; Li, Chen-Zhong

    2015-12-01

    Neural tissue engineering is one of the most promising approaches for healing nerve damage, which bypasses the limits of contemporary conventional treatments. In a previous study, we developed a fibrous scaffold via electrospinning poly (glycerol dodecanedioate) (PGD) and gelatin that mimics the structure of a native extracellular matrix (ECM) for soft tissue engineering application. In this study, fumaric acid (FA) was incorporated into the PGD synthesis process, which produced a PGD derivative referred to as poly (glycerol dodecanedioate co-fumarate) (PGDF). This introduced a new functional group, a double bond, into the polymer thus providing new modification possibilities. Arg-Gly-Asp-Cys (RGDC) and laminin peptides were chosen as biomolecules to modify the fiber and facilitate cell attachment and differentiation efficiency. The release of FA into the medium was quantified to investigate the bioreactivity of the derived scaffolds. In combination with UV crosslinking, the developed PGDF fiber mats were able to withstand degradation processes for up to 2 months, which ensures that neural tissue engineering applications are viable. Cell viability and motor neuron differentiation efficiency were demonstrated to be significantly improved with the addition of FA, RGDC and laminin peptides. PMID:26584592

  3. Controlled surface modification of tissue culture polystyrene for selective cell binding using resilin-inspired polypeptides

    International Nuclear Information System (INIS)

    Modified tissue culture polystyrene (TCP) surfaces have been fabricated by attachment of recombinant polypeptides based on Drosophila melanogaster resilin and the Anopheles gambiae resilin-like protein. The D. melanogaster polypeptide (Rec-1) was from the first exon of resilin and consisted of 17 very similar repeats of a 15 residue sequence. The A. gambiae polypeptide consisted of 16 repeats of an 11 residue consensus sequence (An16). Polypeptides were attached to the TCP surface through tyrosine-based photo-crosslinking using blue light in combination with (RuII(bpy)3)Cl2 and sodium persulfate. TCP that has been manufactured by mild oxidation has surface phenolic groups that are believed to participate in this crosslinking process. X-ray photoelectron spectroscopy and contact angle analyses were used to demonstrate polypeptide binding. At higher coating concentrations of Rec-1 and An16, the surface was passivated and fibroblasts no longer attached and spread. At coating concentrations of 1 mg ml−1 for Rec-1 and 0.1 mg ml−1 for An16, where the surface was fully passivated against fibroblast attachment, addition of a cell attachment peptide, cyclo(Arg-Gly-Asp-D-Tyr-Lys) during coating and photo-crosslinking at >0.1 mg ml−1, led to the restoration of fibroblast binding that was dependent on the integrin αV chain. (paper)

  4. Gracilaria lemaneiformis polysaccharide as integrin-targeting surface decorator of selenium nanoparticles to achieve enhanced anticancer efficacy.

    Science.gov (United States)

    Jiang, Wenting; Fu, Yuanting; Yang, Fang; Yang, Yufeng; Liu, Ting; Zheng, Wenjie; Zeng, Lilan; Chen, Tianfeng

    2014-08-27

    The poor permeability of glioma parenchyma represents a major limit for antiglioblastoma drug delivery. Gracilaria lemaneiformis polysaccharide (GLP), which has a high binding affinity to αvβ3 integrin overexpressed in glioma cells, was employed in the present study to functionalize selenium nanoparticles (SeNPs) to achieve antiglioblastoma efficacy. GLP-SeNPs showed satisfactory size distribution, high stability, and selectivity between cancer and normal cells. In U87 glioma cell membrane, which has a high integrin expression level, GLP-SeNPs exhibited significantly higher cellular uptake than unmodified SeNPs. As expected, U87 cells exhibited a greater uptake of GLP-SeNPs than C6 cells with low integrin expression level. Furthermore, the internalization of GLP-SeNPs was inhibited by cyclo-(Arg-Gly-Asp-Phe-Lys) peptides, suggesting that cellular uptake into U87 cells and C6 cells occurred via αvβ3 integrin-mediated endocytosis. For U87 cells, the cytotoxicity of SeNPs decorated by GLP was enhanced significantly because of the induction of various apoptosis signaling pathways. Internalized GLP-SeNPs triggered intracellular reactive oxygen species downregulation. Therefore, p53, MAPKs, and AKT pathways were activated to advance cell apoptosis. These findings suggest that surface decoration of nanomaterials with GLP could be an efficient strategy for design and preparation of glioblastoma targeting nanodrugs. PMID:25073123

  5. Disintegrins from Hematophagous Sources

    Directory of Open Access Journals (Sweden)

    Ivo M. B. Francischetti

    2012-04-01

    Full Text Available Bloodsucking arthropods are a rich source of salivary molecules (sialogenins which inhibit platelet aggregation, neutrophil function and angiogenesis. Here we review the literature on salivary disintegrins and their targets. Disintegrins were first discovered in snake venoms, and were instrumental in our understanding of integrin function and also for the development of anti-thrombotic drugs. In hematophagous animals, most disintegrins described so far have been discovered in the salivary gland of ticks and leeches. A limited number have also been found in hookworms and horseflies, and none identified in mosquitoes or sand flies. The vast majority of salivary disintegrins reported display a RGD motif and were described as platelet aggregation inhibitors, and few others as negative modulator of neutrophil or endothelial cell functions. This notably low number of reported disintegrins is certainly an underestimation of the actual complexity of this family of proteins in hematophagous secretions. Therefore an algorithm was created in order to identify the tripeptide motifs RGD, KGD, VGD, MLD, KTS, RTS, WGD, or RED (flanked by cysteines in sialogenins deposited in GenBank database. The search included sequences from various blood-sucking animals such as ticks (e.g., Ixodes sp., Argas sp., Rhipicephalus sp., Amblyomma sp., tabanids (e.g., Tabanus sp., bugs (e.g., Triatoma sp., Rhodnius prolixus, mosquitoes (e.g., Anopheles sp., Aedes sp., Culex sp., sand flies (e.g., Lutzomyia sp., Phlebotomus sp., leeches (e.g., Macrobdella sp., Placobdella sp. and worms (e.g., Ancylostoma sp.. This approach allowed the identification of a remarkably high number of novel putative sialogenins with tripeptide motifs typical of disintegrins (>450 sequences whose biological activity remains to be verified. This database is accessible

  6. Chronic use of the immunomodulating tripeptide feG-COOH in experimental feline asthma.

    Science.gov (United States)

    Eberhardt, Jason M; DeClue, Amy E; Reinero, Carol R

    2009-12-15

    We have previously documented that a single dose of feG-COOH prior to allergen challenge significantly decreased eosinophilic airway inflammation in cats with experimental asthma, but did not result in complete resolution of airway inflammation. This study was undertaken to determine if a chronic (2 weeks) course of feG-COOH in experimentally asthmatic cats would induce complete remission of airway inflammation and clinical signs of asthma. Experimental asthma was induced using Bermuda grass allergen (BGA) and cats were randomly selected to receive either feG-COOH (1mg/kg, PO) or saline for 2 weeks, followed by a 2-week washout period. Cats then received the alternate treatment. Aerosol challenge with BGA was performed weekly throughout the study and bronchoalveolar lavage fluid (BALF) and blood were collected prior to and after each of the 2-week treatment periods. Regular use of feG-COOH had no significant effect on airway inflammation, BALF and plasma TNF bioactivity or a clinical sign compared to placebo. Regular use of feG-COOH can thus not be recommended as the sole therapy for feline allergic asthma.

  7. Safety evaluation of an IPP tripeptide-containing milk protein hydrolysate

    NARCIS (Netherlands)

    Ponstein-Simarro Doorten, A.Y.; Wiel, J.A.G. van de; Jonker, D.

    2009-01-01

    Tensguard™ is a milk protein hydrolysate containing the lactotripeptide IPP. It is derived from cow's milk, which is present in the human diet and has a safe history of consumption. The final Tensguard™ product, a supplement or a functional food ingredient, is intended for use by people who want to

  8. Structural flexibility of Aib-containing peptides: the N-terminal tripeptide of trichotoxin.

    Science.gov (United States)

    Gessmann, R; Brueckner, H; Kokkinidis, M

    1991-01-31

    The sequence Aib-Gly-Aib which corresponds to the N-terminus of the microheterogeneous peptide antibiotic trichotoxin has been studied crystallographically in the context of different protecting groups. Peptides Ac-Aib-Gly-Aib-OH (A) and Z-Aib-Gly-Aib-OH (B) form beta-turns. Both peptides show a remarkable conformational flexibility forming a large variety of beta-turns of different types.

  9. Tyroserleutide tripeptide affects calcium homeostasis of human hepatocarcinoma BEL-7402 cells

    Institute of Scientific and Technical Information of China (English)

    FU Zheng; LU Rong; LI Guoli; ZHAO Lan; GAO Weizhen; CHE Xuchun; JIAN Xu; ZHOU Chunlei; YAO Zhi

    2005-01-01

    This study aimed to observe the effects of tyroserleutide (tyrosyl-seryl-leucine, YSL) on the growth of human hepatocarcinoma BEL-7402 that was transplanted into nude mice, and explore its anti-tumor mechanism preliminarily. YSL, at doses of 80 μg·kg-1·d-1, 160μg·kg-1·d-1 and 320μg·kg-1·d-1 significantly inhibited the growth of the human hepatocarcinoma BEL-7402 tumor in nude mice, producing inhibition of 21.66%, 41.34%, and 34.78%, respectively. Ultra structure of BEL-7402 tumor in nude mice showed that YSL could induce tumor cells apoptosis and necrosis, cell organelle mitochondria and endoplasmic reticulum damage, and calcium overload. By confocal laser scanning microscopy and flow cytometry, we found that 10 μg/mL YSL rapidly induced an increase of the concentration of cytoplasmic free calcium in BEL-7402 cells in vitro, and maintained high concentrations of cytoplasmic free calcium for 1 h. Then the calcium concentration began to decrease after 2 h, and was lower than that of the control group at 4 h and 24 h (p<0.05). YSL also decreased the mitochondrial transmembrane potential of BEL-7402 cells in vitro, but had no effect on the calcium homeostasis or mitochondrial transmembrane potential of Chang liver hepatocytes. So affecting calcium homeostasis, then inducing apoptosis and necrosis may be a mechanism by which YSL inhibits the tumor growth in animal model.

  10. IRMPD spectroscopy b(2) ions from protonated tripeptides with 4-aminomethyl benzoic acid residues

    NARCIS (Netherlands)

    M. J. Kullman; S. Molesworth; G. Berden; J. Oomens; M. van Stipdonk

    2012-01-01

    Collision-induced dissociation (CID) of the peptide alanine-4-aminomethylbenzoic acid-glycine, A(AMBz)G generates a prominent b2 ion despite a previous report [E.R. Talaty, T.J. Cooper, S.M. Osburn, M.J. Van Stipdonk, Collision-induced dissociation of protonated tetrapeptides containing β-alanine, γ

  11. IRMPD spectroscopy b(2) ions from protonated tripeptides with 4-aminomethyl benzoic acid residues

    NARCIS (Netherlands)

    Kullman, M. J.; Molesworth, S.; G. Berden,; Oomens, J.; Van Stipdonk, M.

    2012-01-01

    Collision-induced dissociation (CID) of the peptide alanine-4-aminomethylbenzoic acid-glycine, A(AMBz)G generates a prominent b(2) ion despite a previous report [ER. Talaty, T.J. Cooper, S.M. Osburn, M.J. Van Stipdonk, Collision-induced dissociation of protonated tetrapeptides containing beta-alanin

  12. Vitamin E-TPGS stabilized self-assembled tripeptide nanostructures for drug delivery.

    Science.gov (United States)

    Yadav, Santosh; Rai, Vartika; Mahato, Manohar; Singh, Mahak; Deka, Smriti Rekha; Sharma, Ashwani Kumar

    2015-01-01

    Self-assembled peptides and specifically small peptide based nanostructures have been the focus of research in past decade due to their potential biological applications. In this study, we prepared a protected peptide, Boc-Pro-Phe-Gly-OMe, which self-assembled in aqueous solutions leading to the formation of nanostructures and ability to act as a drug carrier. Dynamic light scattering (DLS) measurements showed nanostructures with average size of 119.6 nm containing hydrophobic core, wherein hydrophobic drugs, viz, eosin, aspirin and curcumin, were successfully encapsulated. These encapsulated nanostructures, were further stabilized with Vitamin E-TPGS. In-vitro drug release studies revealed the release of drugs in controlled fashion from the nanostructures. The results advocate the potential of the proposed peptide nanostructures as controlled drug delivery systems and could be used in other biomedical applications.

  13. Preparation and anti-tumor activity of a novel liposome-loaded drug%RGD脂肪醇与17-AAG脂质体的制备及抗肿瘤活性研究

    Institute of Scientific and Technical Information of China (English)

    李雪梅; 王玉记; 吴建辉; 崔纯莹

    2015-01-01

    Objective To prepare an Arg-Gly-Asp-Phe-fatty alcohol ( RGDFOC12 ) liposomes-loaded 17-allylamino-17-demethoxygeldanamycin(17-AAG). Methods RGDFOC12 liposomes-loaded 17-AAG(RLAs) was prepared by film dispersion method and evaluated by particle size analysis, Zeta potential, encapsulation efficiency, the release in vitro, plasma stability, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide( MTT) assay and the anti-tumor activity in vivo. Results The RLAs was stable colloidal dispersion system in spherical shape of (130.6±0.6)nm in diameter and the Zeta potential was (-28.37±1.67)mV. The release of RLAs in vitro showed that the released percentage of RLAs in pH 5. 4 is more than that in pH 7. 4. The MTT assay proved that RLAs inhibited the proliferation of cancer cells. The anti-tumor assay showed that RLAs inhibited tumor growth and reduced the toxicity. Conclusion The RLAs were prepared by film dispersion method. RLAs showed anti-tumor activity in vivo and good potential in cancer therapy.%目的:制备一种新的精氨酸-甘氨酸-天冬氨酸-苯丙氨酸-脂肪醇( Arg-Gly-Asp-Phe-fatty alcohol,RGDFOC12)与17-丙烯氨基-17-去甲氧基格尔德霉素(17-allylamino-17-demethoxygeldanamycin,17-AAG)的脂质体(RGDFOC12 liposomes-loaded 17-AAG, RLAs)。方法采用薄膜分散-探头超声法制备;采用激光纳米粒度仪、透射电镜和扫描电镜测定粒径,Zeta电位和外观形态;采用动态透析法测定药物释放;采用四甲基偶氮唑盐[3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide,MTT]考察其对5种人肿瘤细胞株增生的抑制作用;通过瘤质量、存活数、体质量、脏器指数比评价其在小鼠体内抗肿瘤效果。结果制备得到的RLAs的粒径为(130.6±0.6)nm,Zeta电位为(-28.37±1.67)mV,外观形态为球形,包封率为80%以上。 RLAs在pH 5.4环境的累积释放百分数大于在pH 7.4环境的累积释放百分数。 RLAs在血浆中可稳定存在,12 h

  14. The evolution of tenascins and fibronectin.

    Science.gov (United States)

    Adams, Josephine C; Chiquet-Ehrismann, Ruth; Tucker, Richard P

    2015-01-01

    Tenascins are extracellular matrix glycoproteins that act both as integrin ligands and as modifiers of fibronectin-integrin interactions to regulate cell adhesion, migration, proliferation and differentiation. In tetrapods, both tenascins and fibronectin bind to integrins via RGD and LDV-type tripeptide motifs found in exposed loops in their fibronectin-type III domains. We previously showed that tenascins appeared early in the chordate lineage and are represented by single genes in extant cephalochordates and tunicates. Here we have examined the genomes of the coelacanth Latimeria chalumnae, the elephant shark Callorhinchus milii as well as the lampreys Petromyzon marinus and Lethenteron japonicum to learn more about the evolution of the tenascin gene family as well as the timing of the appearance of fibronectin during chordate evolution. The coelacanth has 4 tenascins that are more similar to tetrapod tenascins than are tenascins from ray-finned fishes. In contrast, only 2 tenascins were identified in the elephant shark and the Japanese lamprey L. japonicum. An RGD motif exposed to integrin binding is observed in tenascins from many, but not all, classes of chordates. Tetrapods that lack this RGD motif in tenascin-C have a similar motif in the paralog tenascin-W, suggesting the potential for some overlapping function. A predicted fibronectin with the same domain organization as the fibronectin from tetrapods is found in the sea lamprey P. marinus but not in tunicates, leading us to infer that fibronectin first appeared in vertebrates. The motifs that recognize LDV-type integrin receptors are conserved in fibronectins from a broad spectrum of vertebrates, but the RGD integrin-binding motif may have evolved in gnathostomes.

  15. Up-regulation and profibrotic role of osteopontin in human idiopathic pulmonary fibrosis.

    Directory of Open Access Journals (Sweden)

    2005-09-01

    Full Text Available BACKGROUND: Idiopathic pulmonary fibrosis (IPF is a progressive and lethal disorder characterized by fibroproliferation and excessive accumulation of extracellular matrix in the lung. METHODS AND FINDINGS: Using oligonucleotide arrays, we identified osteopontin as one of the genes that significantly distinguishes IPF from normal lungs. Osteopontin was localized to alveolar epithelial cells in IPF lungs and was also significantly elevated in bronchoalveolar lavage from IPF patients. To study the fibrosis-relevant effects of osteopontin we stimulated primary human lung fibroblasts and alveolar epithelial cells (A549 with recombinant osteopontin. Osteopontin induced a significant increase of migration and proliferation in both fibroblasts and epithelial cells. Epithelial growth was inhibited by the pentapeptide Gly-Arg-Gly-Asp-Ser (GRGDS and antibody to CD44, while fibroproliferation was inhibited by GRGDS and antibody to alphavbeta3 integrin. Fibroblast and epithelial cell migration were inhibited by GRGDS, anti-CD44, and anti-alphavbeta3. In fibroblasts, osteopontin up-regulated tissue inhibitor of metalloprotease-1 and type I collagen, and down-regulated matrix metalloprotease-1 (MMP-1 expression, while in A549 cells it caused up-regulation of MMP-7. In human IPF lungs, osteopontin colocalized with MMP-7 in alveolar epithelial cells, and application of weakest link statistical models to microarray data suggested a significant interaction between osteopontin and MMP-7. CONCLUSIONS: Our results provide a potential mechanism by which osteopontin secreted from the alveolar epithelium may exert a profibrotic effect in IPF lungs and highlight osteopontin as a potential target for therapeutic intervention in this incurable disease.

  16. Platelets may inhibit leucotriene biosynthesis by human neutrophils at the integrin level.

    Science.gov (United States)

    Chabannes, Bernard; Moliere, Patrick; Merhi-Soussi, Faten; Poubelle, Patrice E; Lagarde, Michel

    2003-04-01

    Polymorphonuclear leucocytes and blood platelets co-operate in several pathophysiological processes, and arachidonic acid (AA) metabolites produced in response to the activation of these cells are potent mediators of their functions. We studied the role of platelets in the formation of 5-lipoxygenase products from AA by autologous neutrophils, especially the chemotactic agent leucotriene (LT) B4. The formation of all products, namely 5-hydroxy-eicosatetraenoic acid (5-HETE), LTB4 and the other LTA4-derived metabolites, in response to the calcium ionophore A23187 was evaluated by high-performance liquid chromatography. All the 5-lipoxygenase products were significantly diminished by physiological concentrations of platelets. This inhibitory effect was lost when platelets were previously degranulated by thrombin in non-aggregating conditions. Peptides containing the Arg-Gly-Asp-Ser or His-His-Leu-Gly-Gly-Ala-Lys-Gln-Ala-Gly-Asp-Val sequence, which prevent the adhesion of platelets to neutrophils via the fibrinogen released from platelet granules and the integrin glycoprotein IIb/IIIa, markedly decreased the inhibitory effect of non-degranulated platelets. The production of transcellular metabolites of AA such as LTC4, the dual 5- and 12-lipoxygenase product 5,12-diHETE and lipoxins could not account for the decreased formation of 5-HETE and LTA4-derived metabolites. It is concluded that platelets may inhibit the neutrophil 5-lipoxygenase activity at the integrin level and in turn may play a role in slowing down the production of LTB4 in the course of inflammation.

  17. Enhanced compatibility of chemically modified titanium surface with periodontal ligament cells

    Science.gov (United States)

    Kado, T.; Hidaka, T.; Aita, H.; Endo, K.; Furuichi, Y.

    2012-12-01

    A simple chemical modification method was developed to immobilize cell-adhesive molecules on a titanium surface to improve its compatibility with human periodontal ligament cells (HPDLCs).The polished titanium disk was immersed in 1% (v/v) p-vinylbenzoic acid solution for 2 h to introduce carboxyl groups onto the surface. After rinsing with distilled deionized water, the titanium disk was dipped into 1.47% 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide solution containing 0.1 mg/ml Gly-Arg-Gly-Asp-Ser (GRGDS), human plasma fibronectin (pFN), or type I collagen from calf skin (Col) to covalently immobilize the cell-adhesive molecules on the titanium surface via formation of peptide bonds. X-ray photoelectron spectroscopy analyses revealed that cell-adhesive molecules were successfully immobilized on the titanium surfaces. The Col-immobilized titanium surface revealed higher values regarding nano rough characteristics than the as-polished titanium surface under scanning probe microscopy. The number of HPDLCs attached to both the pFN- and Col-immobilized titanium surfaces was twice that attached to the as-polished titanium surfaces. The cells were larger with the cellular processes that stretched to a greater extent on the pFN- and Col-immobilized titanium surfaces than on the as-polished titanium surface (p < 0.05). HPDLCs on the Col-immobilized titanium surfaces showed more extensive expression of vinculin at the tips of cell projections and more contiguously along the cell outline than on the as-polished, GRGDS-immobilized and pFN-immobilized titanium surfaces. It was concluded that cell-adhesive molecules successfully immobilized on the titanium surface and improved the compatibility of the surface with HPDLCs. The Col-immobilized titanium surface could be used for forming ligament-like tissues around titanium dental implants.

  18. Chemical synthesis of echistatin, a potent inhibitor of platelet aggregation from Echis carinatus: synthesis and biological activity of selected analogs.

    Science.gov (United States)

    Garsky, V M; Lumma, P K; Freidinger, R M; Pitzenberger, S M; Randall, W C; Veber, D F; Gould, R J; Friedman, P A

    1989-06-01

    Echistatin, a polypeptide from the venom of the saw-scaled viper, Echis carinatus, containing 49 amino acids and 4 cystine bridges was synthesized by solid-phase methodology in 4% yield. In the final step, air oxidation of the octahydroderivative was found to be optimal at pH 8. The synthetic product was shown to be physically and biologically indistinguishable from native material. It inhibits fibrinogen-dependent platelet aggregation stimulated by ADP with IC50 = 3.3 x 10(-8) M and also prevents aggregation initiated by thrombin, epinephrine, collagen, or platelet-activating factor. Reduction of purified synthetic echistatin to octahydroechistatin with dithiothreitol followed by air oxidation regenerated homogeneous echistatin in quantitative yield. This highly specific refolding strongly suggests that the linear sequence of octahydroechistatin contains all of the information that is required for the proper folding of the peptide. The sequence Arg24-Gly-Asp of echistatin occurs also in adhesive glycoproteins that bind to the platelet fibrinogen receptor--a heterodimeric complex composed of glycoproteins IIb and IIIa. In an effort to evaluate the role of this putative binding site we have synthesized analogs of echistatin with substitution of Arg-24. Replacement with ornithine-24 (Orn-24) resulted in an analog having a platelet aggregation inhibitory activity with IC50 = 1.05 x 10(-7) M. Substitution with Ala-24 gave IC50 = 6.1 x 10(-7) M. The inhibitory activity of the corresponding short sequence analogs Arg-Gly-Asp-Phe (IC50 = 6 x 10(-6) M), Orn-Gly-Asp-Phe (IC50 = 1.3 x 10(-4) M), and Ala-Gly-Asp-Phe (IC50 = 5.0 x 10(-4) M) was also determined. These results suggest that arginine plays a more important role in the binding of the tetrapeptide than in that of echistatin. PMID:2726764

  19. Crystal structure of schistatin, a disintegrin homodimer from saw-scaled viper (Echis carinatus) at 2.5 A resolution.

    Science.gov (United States)

    Bilgrami, Sameeta; Tomar, Shailly; Yadav, Savita; Kaur, Punit; Kumar, Janesh; Jabeen, Talat; Sharma, Sujata; Singh, Tej P

    2004-08-13

    This is the first structure of a biological homodimer of disintegrin. Disintegrins are a class of small (4-14 kDa) proteins that bind to transmembrane integrins selectively. The present molecule is the first homodimer that has been isolated from the venom of Echis carinatus. The monomeric chain contains 64 amino acid residues. The three-dimensional structure of schistatin has been determined by the multiple isomorphous replacement method. It has been refined to an R-factor of 0.190 using all the data to 2.5 A resolution. The two subunits of the disintegrin homodimer are related by a 2-fold crystallographic symmetry. Thus, the crystallographic asymmetric unit contains a monomer of disintegrin. The monomer folds into an up-down topology with three sets of antiparallel beta-strands. The structure is well ordered with four intramolecular disulfide bonds. the two monomers are firmly linked to each other through two intermolecular disulfide bridges at their N termini together with several other interactions. This structure has corrected the error in the disulfide bond pattern of the two intermolecular disulfide bridges that was reported earlier using chemical methods. Unique sequence and structural features of the schistatin monomers suggest that they have the ability to bind well with both alphaIIb beta3 and alphav beta3 integrins. The N termini anchored two chains of the dimer diverge away at their C termini exposing the Arg-Gly-Asp motif into opposite directions thus enhancing their binding efficiency to integrins. This is one of the unique features of the present disintegrin homodimer and seems to be responsible for the clustering of integrin molecules. The homodimer binds to integrins apparently with a higher affinity than the monomers and also plays a role in the signaling pathway. PMID:15317139

  20. c-Abl is an upstream regulator of acid sphingomyelinase in apoptosis induced by inhibition of integrins αvβ3 and αvβ5.

    Directory of Open Access Journals (Sweden)

    Xiuhai Ren

    Full Text Available Inhibition of integrins αvβ3/αvβ5 by the cyclic function-blocking peptide, RGDfV (Arg-Gly-Asp-Phe-Val can induce apoptosis in both normal cells and tumor cells. We show that RGDfV induced apoptosis in ECV-304 carcinoma cells, increased activity and mRNA expression of acid sphingomyelinase (ASM, and increased ceramides C(16, C(18:0, C(24:0 and C(24:1 while decreasing the corresponding sphingomyelins. siRNA to ASM decreased RGDfV-induced apoptosis as measured by TUNEL, PARP cleavage, mitochondrial depolarization, and caspase-3 and caspase-8 activities, as well as by annexinV in a 3D collagen model. These findings indicate a causal role for ASM in RGDfV-induced apoptosis in ECV-304. We have shown that c-Abl, a non-receptor tyrosine kinase, also mediates RGDfV-induced apoptosis. However, c-Abl, has not been previously linked to ASM in any system. Here we show that STI-571 (imatinib, inhibitor of c-Abl inhibited RGDfV-induced ASM activity. Furthermore, STI-571 and c-Abl-siRNA both inhibited RGDfV-induced increase in ASM mRNA, but ASM-siRNA did not affect c-Abl phosphorylation or expression, supporting that c-Abl regulates the RGDfV-induced increase in ASM expression. These studies implicate ASM as a mediator of apoptosis induced by inhibition of integrins αvβ3/αvβ5, and for the first time place c-Abl as an upstream regulator of ASM expression and activity.

  1. Evaluation of treatment response of cilengitide in an experimental model of breast cancer bone metastasis using dynamic PET with 18F-FDG.

    Science.gov (United States)

    Cheng, Caixa; Komljenovic, Dorde; Pan, Leyun; Dimitrakopoulou-Strauss, Antonia; Strauss, Ludwig; Bäuerle, Tobias

    2011-01-01

    The purpose of this study was the assessment of the feasibility of dynamic positron emission tomography (PET) studies with fluorine-18 fluorodeoxyglucose ((18)F-FDG) to quantify effects of the cyclic Arg-Gly-Asp peptide cilengitide, which targets the ανβ 3 and ανβ 5 integrin receptors in rats with breast cancer bone metastases. Rats were inoculated with MDA-MB-231 breast cancer cells, followed by the development of lytic lesions in the hind leg. Rats with lytic lesions were treated with cilengitide five times weekly on a continuous basis from days 30 to 55 after tumor cell inoculation. Dynamic PET studies with (18)F-FDG were performed in untreated (n=9), controlled (n=4) and treated rats (n=6). The data were assessed using learning-machine two-tissue compartmental analysis. The (18)F-FDG kinetic parameters obtained by two-tissue compartmental model learning-machine showed significant differences when individual parameters were compared between the control group and treated animals. Quantitative assessment of the tracer kinetics and the application of classification analysis to the data provided us with evidence to identify those tumors that demonstrated effect of cilengitide treatment. The transport rate K1 and the phosphorylation rate k3 were significantly different (P=0.033 and 0.038, respectively). Classification analysis based on support vector machines ranking feature elimination of the combination of PET parameters revealed an overall accuracy of 80.0% between treated animals and the control group. We were able to identify 83.3% treated animals compared with the control group based on k2 and VB. In conclusion, the results revealed that cilengitide treatment of experimental breast cancer bone metastases had a significant therapeutic impact on (18)F-FDG kinetics. PMID:21512659

  2. Síntese de derivados cíclicos de RGD

    OpenAIRE

    Vilaça, Maria Helena Pereira

    2010-01-01

    Dissertação de mestrado em Química Medicinal Neste trabalho foram sintetizados, com diversos bis-aminoácidos através da reacção de ciclo-adição 1,3-dipolar de Huisgen entre azido- e alquinilaminoácidos e entre alquinilaminoácidos e bis-azidas. Os azido-aminoácidos foram obtidos pelo método de Wong, usando uma transferência diazo entre a azida de trifluorometanossulfonilo e a função amina do respectivo aminoácido. As bis-azidas foram sintetizadas a partir dos respectivos ácid...

  3. AFM Imaging of RGD Presenting Synthetic Extracellular Matrix using Gold Nanoparticles

    Science.gov (United States)

    Cell-interactive polymers have been widely used as synthetic extracellular matrices (sECM) to regulate cell function and promote tissue regeneration. Although it is known that adhesion ligand density and distribution influence the proliferation and differentiation of various cell types, currently a...

  4. Improved targeting of the alpha(v)beta (3) integrin by multimerisation of RGD peptides.

    NARCIS (Netherlands)

    Dijkgraaf, I.; Kruijtzer, J.A.; Liu, S.; Soede, A.C.; Oyen, W.J.G.; Corstens, F.H.M.; Liskamp, R.M.; Boerman, O.C.

    2007-01-01

    PURPOSE: The integrin alpha(v)beta(3) is expressed on sprouting endothelial cells and on various tumour cell types. Due to the restricted expression of alpha(v)beta(3) in tumours, alpha(v)beta(3) is considered a suitable receptor for tumour targeting. In this study the alpha(v)beta(3) binding charac

  5. Investigating Surface Topology and Cyclic-RGD Peptide functionalization on Vascular Endothelialization

    OpenAIRE

    McNichols, Colton; Wilkins, Justin; Kubota, Atsu; Shiu, Yan T.; Aouadi, Samir M.; Kohli, Punit

    2013-01-01

    The advantages of endothelialization of a stent surface in comparison with the bare metal and drug eluting stents used today include reduced late-stent restenosis and in-stent thrombosis. In this paper, we study the effect of surface topology and functionalization of tantalum (Ta) with cyclic-(arginine-glycine-aspartic acid-D-phenylalanine-lysine (cRGDfK)) on the attachment, spreading, and growth of vascular endothelial cells. Self-assembled nano-dimpling on Ta surfaces was performed using a ...

  6. Investigating surface topology and cyclic-RGD peptide functionalization on vascular endothelialization.

    Science.gov (United States)

    McNichols, Colton; Wilkins, Justin; Kubota, Atsutoshi; Shiu, Yan T; Aouadi, Samir M; Kohli, Punit

    2014-02-01

    The advantages of endothelialization of a stent surface in comparison with the bare metal and drug-eluting stents used today include reduced late-stent restenosis and in-stent thrombosis. In this article, we study the effect of surface topology and functionalization of tantalum (Ta) with cyclic-(arginine-glycine-aspartic acid-d-phenylalanine-lysine) (cRGDfK) on the attachment, spreading, and growth of vascular endothelial cells. Self-assembled nanodimpling on Ta surfaces was performed using a one-step electropolishing technique. Next, cRGDfK was covalently bonded onto the surface using silane chemistry. Our results suggest that nanotexturing alone was sufficient to enhance cell spreading, but the combination of a nanodimpled surfaces along with the cRGDfK peptide may produce a better endothelialization coating on the surface in terms of higher cell density, better cell spreading, and more cell-cell interactions, when compared to using cRGDfK peptide functionalization alone or nanotexturing alone. We believe that future research should look into how to implement both modifications (topographic and chemical modifications) to optimize the stent surface for endothelialization. PMID:23505215

  7. Noninvasive and Quantitative Assessment of In Vivo Fetomaternal Interface Angiogenesis Using RGD-Based Fluorescence

    Directory of Open Access Journals (Sweden)

    M. Keramidas

    2014-01-01

    Full Text Available Angiogenesis is a key process for proper placental development and for the success of pregnancy. Although numerous in vitro methods have been developed for the assessment of this process, relatively few reliable in vivo methods are available to evaluate this activity throughout gestation. Here we report an in vivo technique that specifically measures placental neovascularization. The technique is based on the measurement of a fluorescent alpha v beta 3 (αvβ3 integrin-targeting molecule called Angiolone-Alexa-Fluor 700. The αvβ3 integrin is highly expressed by endothelial cells during the neovascularization and by trophoblast cells during their invasion of the maternal decidua. Angiolone was injected to gravid mice at 6.5 and 11.5 days post coitus (dpc. The fluorescence was analyzed one day later at 7.5 and 12.5 dpc, respectively. We demonstrated that (i Angiolone targets αvβ3 protein in the placenta with a strong specificity, (ii this technique is quantitative as the measurement was correlated to the increase of the placental size observed with increasing gestational age, and (iii information on the outcome is possible, as abnormal placentation could be detected early on during gestation. In conclusion, we report the validation of a new noninvasive and quantitative method to assess the placental angiogenic activity, in vivo.

  8. Studies of R2Ti2O7 (R=Gd and Yb); new results

    International Nuclear Information System (INIS)

    Specific heat and muon spin rotation and relaxation data are presented for two geometrically frustrated systems: Gd2Ti2O7 which antiferromagnetically orders and Yb2Ti2O7 which presents dynamical short range correlations at low temperature. The muon data help to characterize the spin dynamics of these two compounds

  9. RGD-modified lipid disks as drug carriers for tumor targeted drug delivery

    Science.gov (United States)

    Gao, Jie; Xie, Cao; Zhang, Mingfei; Wei, Xiaoli; Yan, Zhiqiang; Ren, Yachao; Ying, Man; Lu, Weiyue

    2016-03-01

    Melittin, the major component of the European bee venom, is a potential anticancer candidate due to its lytic properties. However, in vivo applications of melittin are limited due to its main side effect, hemolysis, especially when applied through intravenous administration. The polyethylene glycol-stabilized lipid disk is a novel type of nanocarrier, and the rim of lipid disks has a high affinity to amphiphilic peptides. In our study, a c(RGDyK) modified lipid disk was developed as a tumor targeted drug delivery system for melittin. Cryo-TEM was used to confirm the shape and size of lipid disks with or without c(RGDyK) modification. In vitro and in vivo hemolysis analyses revealed that the hemolysis effect significantly decreased after melittin associated with lipid disks. Importantly, the results of our in vivo biodistribution and tumor growth inhibitory experiments showed that c(RGDyK) modification increased the distribution of lipid disks in the tumor and the anticancer efficacy of melittin loaded lipid disks. Thus, we successfully achieved a targeted drug delivery system for melittin and other amphiphilic peptides with a good therapeutic effect and low side effects.

  10. Studies of R2Ti2O7 ( R=Gd and Yb); new results

    Science.gov (United States)

    Dalmas de Réotier, P.; Glazkov, V.; Marin, C.; Yaouanc, A.; Gubbens, P. C. M.; Sakarya, S.; Bonville, P.; Amato, A.; Baines, C.; King, P. J. C.

    2006-03-01

    Specific heat and muon spin rotation and relaxation data are presented for two geometrically frustrated systems: Gd2Ti2O7 which antiferromagnetically orders and Yb2Ti2O7 which presents dynamical short range correlations at low temperature. The muon data help to characterize the spin dynamics of these two compounds.

  11. Alpha v beta 3 integrin-targeting of intraperitoneally growing tumors with a radiolabeled RGD peptide.

    NARCIS (Netherlands)

    Dijkgraaf, I.; Kruijtzer, J.A.; Frielink, C.; Corstens, F.H.M.; Oyen, W.J.G.; Liskamp, R.M.; Boerman, O.C.

    2007-01-01

    Ovarian cancer is the fourth most common cause of cancer deaths among females in the western world after cancer of the breast, colon and lung. The inability to control the disease within the peritoneal cavity is the major cause of treatment failure in patients with ovarian cancer. The majority of ov

  12. Covalent RGD modification of the inner pore surface of polycaprolactone scaffolds

    NARCIS (Netherlands)

    M. Gabriel; K. Nazmi; M. Dahm; A. Zentner; C.F. Vahl; D. Strand

    2012-01-01

    Scaffold production for tissue engineering was demonstrated by means of a hot compression molding technique and subsequent particulate leaching. The utilization of spherical salt particles as the pore-forming agent ensured complete interconnectivity of the porous structure. This method obviated the

  13. Effect of tripeptide-copper complexes on the process of skin wound healing and on cultured fibroblasts.

    Science.gov (United States)

    Buffoni, F; Pino, R; Dal Pozzo, A

    1995-01-01

    The effects of Gly-His-Lys-Cu and of three synthetic analogues (I, II and III) on wound healing of the guinea-pig dorsal skin, as well as on cultured fibroblasts, were examined. Gly-His-Lys-Cu and peptide I-Cu were tested in vivo. Hydroxyproline, proteins, DNA and semicarbazide-sensitive amine oxidase, with a high affinity for benzylamine, were measured, and the histology of the wounds was observed after staining with hematoxylin/eosin. Another set of wounds was treated in parallel with equivalent amounts of copper acetate. Gly-His-Lys-Cu and the analogues caused a decrease of the activity of semicarbazide-sensitive amine oxidase, with a high affinity for benzylamine, 4-8 days after surgery, followed by an increase on day 11 that was higher than in the control group. No significant difference was found between the two peptides. A slower reorganization of the skin and a delayed activation of fibroblasts are the main effects observed with these peptides-Cu complexes. Preliminary studies on cultured fibroblasts were monitored to see whether these peptides had a direct effect on fibroblasts. The products studied at a concentration of 10(-7) M, decreased cell reproduction and increased collagen expression. PMID:8836453

  14. The Human Tripeptide GHK-Cu in Prevention of Oxidative Stress and Degenerative Conditions of Aging: Implications for Cognitive Health

    OpenAIRE

    Loren Pickart; Jessica Michelle Vasquez-Soltero; Anna Margolina

    2012-01-01

    Oxidative stress, disrupted copper homeostasis, and neuroinflammation due to overproduction of proinflammatory cytokines are considered leading causative factors in development of age-associated neurodegenerative conditions. Recently, a new mechanism of aging—detrimental epigenetic modifications—has emerged. Thus, compounds that possess antioxidant, anti-inflammatory activity as well as compounds capable of restoring copper balance and proper gene functioning may be able to prevent age-associ...

  15. Enhanced β-turn conformational stability of tripeptides containing ΔPhe in cis over trans configuration.

    Science.gov (United States)

    Jaremko, Mariusz; Jaremko, Lukasz; Mazur, Adam; Makowski, Maciej; Lisowski, Marek

    2013-10-01

    Conformations of three pairs of dehydropeptides with the opposite configuration of the ΔPhe residue, Boc-Gly-Δ(Z/E)Phe-Phe-p-NA (Z- p -NA and E- p -NA), Boc-Gly-Δ(Z/E)Phe-Phe-OMe (Z-OMe and E-OMe), and Boc-Gly-Δ(Z/E)Phe-Phe-OH (Z-OH and E-OH) were compared on the basis of CD and NMR studies in MeOH, TFE, and DMSO. The CD results were used as the additional input data for the NMR-based calculations of the detailed solution conformations of the peptides. It was found that Z- p -NA, E- p -NA, Z-OMe, and Z-OH adopt the β-turn conformations and E-OMe and E-OH are unordered. There are two overlapping type III β-turns in Z- p -NA, type II' β-turn in E- p -NA, and type II β-turn in Z-OMe and Z-OH. The results obtained indicate that in the case of methyl esters and peptides with a free carboxyl group, Δ(Z)Phe is a much stronger inducer of ordered conformations than Δ(E)Phe. It was also found that temperature coefficients of the amide protons are not reliable indicators of intramolecular hydrogen bonds donors in small peptides. PMID:23812673

  16. Asia1型口蹄疫病毒受体结合位点多样性

    Institute of Scientific and Technical Information of China (English)

    李平花; 白兴文; 孙普; 卢曾军; 曹伟军; 吴润; 殷宏; 刘在新

    2011-01-01

    【目的】口蹄疫病毒(Foot-and-Mouth Disease Virus,FMDV)通过结构蛋白VP1 G-H环上高度保守的精氨酸-甘氨酸-天门冬氨酸(Arg-Gly-Asp,RGD)基序与整联蛋白结合起始病毒的感染,但FMDV是RNA病毒,在环境条件变化时,FMDV能够以非RGD的途径起始病毒的感染。为了研究FMDV Asia1/JS/China/05田间舌皮毒经两种不同的途径短期传代后细胞受体结合基序RGD的变异。【方法】采用RT-PCR方法扩增FMDV Asia1/JS/China/05田间毒、田间毒的乳鼠适应毒第四代(MF4)和接种田间毒的牛同居感染的猪水泡病料适应细胞的第八代毒(PBF8)结构蛋白VP1基因,并对不同病毒VP1基因的PCR产物测序和cDNA文库测序。【结果】以含RGD受体结合基序为优势的田间毒在乳鼠上短期传代后出现了含精氨酸-丝氨酸-天门冬氨酸(Arg-Ser-Asp,RSD)和RGD受体结合基序的混合种群,而同居感染后的细胞传代病毒种群则以含精氨酸-天门冬氨酸-天门冬氨酸(Arg-Asp-Asp,RDD)受体结合基序为优势种群。【结论】发现了含RGD受体结合位点为优势的FMDV种群,经不同的宿主短期传代后产了含RSD或RDD受体结合基序的优势种群,该发现不仅增加了保守基序RGD发生替换的FMDV变异株的数量,而且为FMDV的细胞识别和宿主嗜性的改变等进一步研究奠定了物质基础。

  17. Enhanced compatibility of chemically modified titanium surface with periodontal ligament cells

    Energy Technology Data Exchange (ETDEWEB)

    Kado, T.; Hidaka, T. [Division of Periodontology and Endodontology, Department of Oral Rehabilitation, School of Dentistry, Health Sciences University of Hokkaido, 1757 Kanazawa, Ishikari-Tobetsu, Hokkaido 061-0293 (Japan); Aita, H. [Division of Occlusion and Removable Prosthodontics, Department of Oral Rehabilitation, School of Dentistry, Health Sciences University of Hokkaido, 1757 Kanazawa, Ishikari-Tobetsu, Hokkaido 061-0293 (Japan); Endo, K. [Division of Biomaterials and Bioengineering, Department of Oral Rehabilitation, School of Dentistry, Health Sciences University of Hokkaido, 1757 Kanazawa, Ishikari-Tobetsu, Hokkaido 061-0293 (Japan); Furuichi, Y., E-mail: furuichi@hoku-iryo-u.ac.jp [Division of Periodontology and Endodontology, Department of Oral Rehabilitation, School of Dentistry, Health Sciences University of Hokkaido, 1757 Kanazawa, Ishikari-Tobetsu, Hokkaido 061-0293 (Japan)

    2012-12-01

    Highlights: Black-Right-Pointing-Pointer Cell-adhesive molecules were covalently immobilized on a Ti surface. Black-Right-Pointing-Pointer Immobilized cell-adhesive molecules maintained native function on the Ti surface. Black-Right-Pointing-Pointer Immobilized collagen enhanced adhesion of periodontal ligament cells to the Ti. - Abstract: A simple chemical modification method was developed to immobilize cell-adhesive molecules on a titanium surface to improve its compatibility with human periodontal ligament cells (HPDLCs).The polished titanium disk was immersed in 1% (v/v) p-vinylbenzoic acid solution for 2 h to introduce carboxyl groups onto the surface. After rinsing with distilled deionized water, the titanium disk was dipped into 1.47% 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide solution containing 0.1 mg/ml Gly-Arg-Gly-Asp-Ser (GRGDS), human plasma fibronectin (pFN), or type I collagen from calf skin (Col) to covalently immobilize the cell-adhesive molecules on the titanium surface via formation of peptide bonds. X-ray photoelectron spectroscopy analyses revealed that cell-adhesive molecules were successfully immobilized on the titanium surfaces. The Col-immobilized titanium surface revealed higher values regarding nano rough characteristics than the as-polished titanium surface under scanning probe microscopy. The number of HPDLCs attached to both the pFN- and Col-immobilized titanium surfaces was twice that attached to the as-polished titanium surfaces. The cells were larger with the cellular processes that stretched to a greater extent on the pFN- and Col-immobilized titanium surfaces than on the as-polished titanium surface (p < 0.05). HPDLCs on the Col-immobilized titanium surfaces showed more extensive expression of vinculin at the tips of cell projections and more contiguously along the cell outline than on the as-polished, GRGDS-immobilized and pFN-immobilized titanium surfaces. It was concluded that cell-adhesive molecules successfully

  18. Transarterial administration of integrin inhibitor loaded nanoparticles combined with transarterial chemoembolization for treating hepatocellular carcinoma in a rat model

    Science.gov (United States)

    Qian, Jun; Oppermann, Elsie; Tran, Andreas; Imlau, Ulli; Qian, Kun; Vogl, Thomas Josef

    2016-01-01

    AIM: To compare the effect of transarterial chemoembolization (TACE) plus GRGDSP (Gly-Arg-Gly-Asp-Ser-Pro, integrin-inhibitor) loaded nanoparticles with TACE alone or TACE + GRGDSP in a rat model of liver tumor. METHODS: Morris hepatoma 3924A tumors were implanted in the livers of 30 ACI rats. The ACI rats were divided randomly into three groups (10 animals each). Tumor volume before treatment (V1) was examined by magnetic resonance imaging (MRI), and then, after laparotomy and placement of a PE-10 catheter into the hepatic artery, the following interventional protocols were performed: TACE (mitomycin C + lipiodol + degradable starch microspheres) + GRGDSP loaded nanoparticles for group A; TACE + GRGDSP for group B (control group 1); TACE alone for group C (control group 2). Tumor volume (V2) was assessed by MRI and the mean ratio of the post-treatment to pretreatment tumor volumes (V2/V1) was calculated. Immunohistochemical analysis was performed to assess the quantification of matrix metalloprotein 9 (MMP-9) and vascular endothelial growth factor (VEGF) positive tumor cells in each treatment group. RESULTS: The mean tumor growth ratios (V2/V1) were 1.3649 ± 0.1194 in group A, 2.0770 ± 0.1595 in group B, and 3.2148 ± 0.1075 in group C. Compared with groups B and C, group A showed a significant reduction in tumor volume. Lower expression of MMP-9 and VEGF in hepatocellular carcinoma was observed in group A than in groups B and C. The angiogenesis of tumor was evaluated using anti-VEGF antibodies, and the metastasis of tumor was assessed using anti-MMP-9 antibody. MMP-9 and VEGF were expressed in all specimens. The immunoexpression of these proteins was confirmed by the presence of red cytoplasmic staining in tumor cells. Lower expression of MMP-9 and VEGF in hepatocellular carcinoma was observed in group A than in groups B and C. CONCLUSION: Transarterial administration of integrin inhibitor loaded nanoparticles combined with TACE evidently retards tumor growth

  19. SPECT/CT of lung nodules using 111In-DOTA-c(RGDfK) in a mouse lung carcinogenesis model

    International Nuclear Information System (INIS)

    Lung cancer is one of the leading causes of cancer-related deaths worldwide, including Japan. Although computed tomography (CT) can detect small lung lesions such as those appearing as ground glass opacity, it cannot differentiate between malignant and non-malignant lesions. Previously, we have shown that single photon emission computed tomography (SPECT) imaging using 111In-1,4,7,10-tetraazacyclododecane-N,N',N'',N'''-tetraacetic acid-cyclo-(Arg-Gly-Asp-D-Phe-Lys) (DOTA-c(RGDfK)), an imaging probe of αvβ3 integrin, is useful for the early detection of pancreatic cancer in a hamster pancreatic carcinogenesis model. In this study, we aimed to assess the usefulness of SPECT/CT with 111In-DOTA-c(RGDfK) for the evaluation of the malignancy of lung cancer. Lung tumors were induced by a single intraperitoneal injection (250 mg/kg) of urethane in male A/J mice. Twenty-six weeks after the urethane treatment, SPECT was performed an hour after injection of 111In-DOTA-c(RGDfK). Following this, the radioactivity ratios of tumor to normal lung tissue were measured by autoradiography (ARG) in the excised lung samples. We also examined the expression of αvβ3 integrin in mouse and human lung samples. Urethane treatment induced 5 hyperplasias, 41 adenomas and 12 adenocarcinomas in the lungs of 8 A/J mice. SPECT with 111In-DOTA-c(RGDfK) could clearly visualize lung nodules, though we failed to detect small lung nodules like adenoma and hyperplasias (adenocarcinoma: 66.7%, adenoma: 33.6%, hyperplasia: 0.0%). ARG analysis revealed significant uptake of 111In-DOTA-c(RGDfK) in all the lesions. Moreover, tumor to normal lung tissue ratios increased along with the progression of carcinogenesis. Histopathological examination using human lung tissue samples revealed clear up-regulation of αvβ3 integrin in well-differentiated adenocarcinoma (Noguchi type B and C) rather than atypical adenomatous hyperplasia. Although there are some limitations in evaluating the malignancy of small lung

  20. Enhanced transduction of fibroblasts in transplanted kidney with an adenovirus having an RGD motif in the HI loop

    NARCIS (Netherlands)

    Sandovici, M; Deelman, LE; Oosten, AS; van Goor, H; Rots, MG; de Zeeuw, D; Henning, RH

    2006-01-01

    Application of gene therapy to the renal graft has a powerful potential to improve the outcome of kidney transplantation and eliminate detrimental side effects associated with systemic therapy, through local expression of immunoregulatory or other protective molecules. However, the search for the op

  1. A pRb-responsive, RGD-modified, and Hyaluronidase-armed Canine Oncolytic Adenovirus for Application in Veterinary Oncology

    OpenAIRE

    Laborda, Eduardo; Puig-Saus, Cristina; Rodriguez-García, Alba; Moreno, Rafael; Cascalló, Manel; Pastor, Josep; Alemany, Ramon

    2014-01-01

    Human and canine cancer share similarities such as genetic and molecular aspects, biological complexity, tumor epidemiology, and targeted therapeutic treatment. Lack of good animal models for human adenovirotherapy has spurred the use of canine adenovirus 2-based oncolytic viruses. We have constructed a canine oncolytic virus that mimics the characteristics of our previously published human adenovirus ICOVIR17: expression of E1a controlled by E2F sites, deletion of the pRb-binding site of E1a...

  2. Evaluation the effects of an angiogenesis inhibitor of endostar with a novel RGD dimer probe labeled by 99Tcm

    International Nuclear Information System (INIS)

    Objective: To study the feasibility of a novel probe 99Tcm-HYNIC-2(poly-(ethylene glycol), PEG)4-Dimer (Dimer:E-[c (RGDfK)2]) as a potential imaging agent for integrin αvβ3 positive tumors, and also to observe the influence of an angiogenesis inhibitor, endostar, on the biodistribution and tumor uptake of the tracer in tumor bearing nude mice. Methods: The expression of integrin αvβ3 in human glioma cells U87MG was determined with immunofluorescence staining before and after treatment with endostar.99Tcm-HYNIC-2PEG4-Dimer was prepared and administered in U87MG tumor bearing mice in 6 h after either administration of endostar (200 μl) or saline (control group) and then biodistribution study was performed. Other 16 mice were divided into endostar treated group (20 mg/kg) and control group (saline) and then gamma imaging was performed in the two groups. Statistical significance of differences between the two groups was assessed using two-sample t test. Results: Radiochemical purity of 99Tcm-HYNIC-2PEG4-Dimer was exceeded 95%. The expression of integrin αvβ3 in U87MG cell was high and gradually decreased after treatment with endostar. There was a negative dose-effect relationship between the dose of endostar and the expression of integrin αvβ3 with the peak effect at the dose of 400 μg/ml. The distribution study in vivo showed that the tracer uptake of U87MG tumors was high, but it decreased after injection of endostar. At 90 min, the %ID/g of endostar and control groups were 1.50±0.08 and 6.27±0.33, respectively (t=40.23, P<0.05). The average T/NT ratios of 99Tcm-HYNIC-2PEG4-Dimer uptake in the endostar and control groups were 1.02±0.11 and 2.58±0.36, respectively (t=10.25, P<0.05). The integrin αvβ3 positive expression ratios of tumor in endostar and control groups were (33.1 ±2.7)% and (81.5±3.2)%,respectively (t=32.60, P<0.05). Conclusions: The novel probe 99Tcm-HYNIC-2PEG4-Dimer may be a promising radiotracer for integrin αvβ3-positive tumor imaging. It may be used for monitoring the therapeutic effect of endostar and may be potentially used for screening the candidates of anti-angiogenesis therapy. (authors)

  3. Material properties and bone marrow stromal cells response to in situ crosslinkable RGD-functionlized lactide-co-glycolide scaffolds.

    Science.gov (United States)

    Jabbari, Esmaiel; He, Xuezhong; Valarmathi, Mani T; Sarvestani, Alireza S; Xu, Weijie

    2009-04-01

    In situ crosslinkable biomaterials with degradation profiles that can be tailored to a particular application are indispensable for treating irregularly shaped defects and for fabrication of shape-selective scaffolds. The objective of this work was to synthesize ultra low molecular weight functionalized PLA and PLGA macromers that can be grafted with bioactive peptides and crosslinked in situ to fabricate biodegradable functional scaffolds. In situ crosslinkable lactide-co-glycolide macromer (cMLGA; "c" for crosslinkable, "M" for macromer, and "LGA" for lactide-co-glycolide) was synthesized by anionic polymerization of lactide and glycolide monomers followed by condensation polymerization with fumaryl chloride. The cMLA (100% L-lactide) and cMLGA macromers formed porous crosslinked scaffolds with NVP as the crosslinker. The mass loss of the crosslinked cMLA and cMLGA was linear with incubation time in vitro (zero-order degradation) and the degradation rate depended on the ratio of lactide to glycolide. cMLGA scaffold with 1:1 lactide to glycolide ratio completely degraded after 4 weeks while the cMLA lost less than 40% of its initial mass after 35 weeks. When cMLA scaffold was functionalized with acrylated integrin-binding Ac-GRGD amino acid sequence, bone marrow stromal (BMS) cells attached and spread on the cMLA scaffold and exhibited focal-point cell adhesion. The mRNA expression levels of collagen-1alpha, osteonectin, and osteopontin for BMS cells seeded in the scaffolds with 1 and 5% Ac-GRGD were upregulated compared with those without Ac-GRGD. cMLGA is attractive as in situ crosslinkable macromer for fabrication of functional scaffolds with degradation characteristics that can be tailored to a particular application. PMID:18431754

  4. Physicochemical characterization of native glycyl-l-histidyl-l-lysine tripeptide for wound healing and anti-aging: a preformulation study for dermal delivery.

    Science.gov (United States)

    Badenhorst, Travis; Svirskis, Darren; Wu, Zimei

    2016-03-01

    This study investigates the physicochemical properties of glycyl-histidyl-lysine-copper (GHK-Cu) to support the development of a formulation for effective topical delivery. The solubility and distribution coefficients (log D) were investigated using conventional methods and GHK concentrations were quantified with a validated stability-indicating reversed phase high performance liquid chromatography (RP-HPLC) method. In addition, the stability of GHK-Cu under stressed conditions and the compatibility with some potential formulation components were assessed. The peptide was susceptible to hydrolytic cleavage under basic and oxidative stressors and to a lesser extent acidic stress with first-order degradation profiles. Surprisingly, the peptide was stable in water and in pH (4.5-7.4) buffers for at least two weeks at 60  °C. The HPLC in conjunction with mass spectrometry identified three key degradation products, one of which was the constituent amino acid histidine. The distribution coefficients in octanol-phosphate buffered saline indicated the highly hydrophilic nature of GHK-Cu with log D values between -2.38 and -2.49 at pH range of 4.5-7.4. Furthermore, GHK-Cu was compatible with Span 60 based niosomes but less stable in the presence of the negatively charged lipid dicetyl phosphate. In summary, the preformulation studies provided information useful to deliver the GHK-Cu complex by carrier. PMID:25384620

  5. The co-crystal structure of ubiquitin carboxy-terminal hydrolase L1 (UCHL1) with a tripeptide fluoromethyl ketone (Z-VAE(OMe)-FMK)

    Energy Technology Data Exchange (ETDEWEB)

    Davies, Christopher W.; Chaney, Joseph; Korbel, Gregory; Ringe, Dagmar; Petsko, Gregory A.; Ploegh, Hidde; Das, Chittaranjan (Whitehead); (Purdue); (Rosenstiel)

    2012-07-25

    UCHL1 is a 223 amino acid member of the UCH family of deubiquitinating enzymes (DUBs), found abundantly and exclusively expressed in neurons and the testis in normal tissues. Two naturally occurring variants of UCHL1 are directly involved in Parkinson's disease (PD). Not only has UCHL1 been linked to PD, but it has oncogenic properties, having been found abnormally expressed in lung, pancreatic, and colorectal cancers. Although inhibitors of UCHL1 have been described previously the co-crystal structure of the enzyme bound to any inhibitor has not been reported. Herein, we report the X-ray structure of UCHL1 co-crystallized with a peptide-based fluoromethylketone inhibitor, Z-VAE(OMe)-FMK (VAEFMK) at 2.35 {angstrom} resolution. The co-crystal structure reveals that the inhibitor binds in the active-site cleft, irreversibly modifying the active-site cysteine; however, the catalytic histidine is still misaligned as seen in the native structure, suggesting that the inhibitor binds to an inactive form of the enzyme. Our structure also reveals that the inhibitor approaches the active-site cleft from the opposite side of the crossover loop as compared to the direction of approach of ubiquitin's C-terminal tail, thereby occupying the P1{prime} (leaving group) site, a binding site perhaps used by the unknown C-terminal extension of ubiquitin in the actual in vivo substrate(s) of UCHL1. This structure provides a view of molecular contacts at the active-site cleft between the inhibitor and the enzyme as well as furnishing structural information needed to facilitate further design of inhibitors targeted to UCHL1 with high selectivity and potency.

  6. Atomic Structures of all the Twenty Essential Amino Acids and a Tripeptide, with Bond Lengths as Sums of Atomic Covalent Radii

    OpenAIRE

    Heyrovska, Raji

    2008-01-01

    Recently, the bond lengths of the molecular components of nucleic acids and of caffeine and related molecules were shown to be sums of the appropriate covalent radii of the adjacent atoms. Thus, each atom was shown to have its specific contribution to the bond length. This enabled establishing their atomic structures for the first time. In this work, the known bond lengths for amino acids and the peptide bond are similarly shown to be sums of the atomic covalent radii. Based on this result, t...

  7. Design, RNA cleavage and antiviral activity of new artificial ribonucleases derived from mono-, di- and tripeptides connected by linkers of different hydrophobicity.

    Science.gov (United States)

    Tamkovich, Nikolay; Koroleva, Lyudmila; Kovpak, Mikhail; Goncharova, Elena; Silnikov, Vladimir; Vlassov, Valentin; Zenkova, Marina

    2016-03-15

    A novel series of metal-free artificial ribonucleases (aRNases) was designed, synthesized and assessed in terms of ribonuclease activity and ability to inactivate influenza virus WSN/A33/H1N1 in vitro. The compounds were built of two short peptide fragments, which include Lys, Ser, Arg, Glu and imidazole residues in various combinations, connected by linkers of different hydrophobicity (1,12-diaminododecane or 4,9-dioxa-1,12-diaminododecane). These compounds efficiently cleaved different RNA substrates under physiological conditions at rates three to five times higher than that of artificial ribonucleases described earlier and displayed RNase A-like cleavage specificity. aRNases with the hydrophobic 1,12-diaminododecane linker displayed ribonuclease activity 3-40 times higher than aRNases with the 4,9-dioxa-1,12-diaminododecane linker. The assumed mechanism of RNA cleavage was typical for natural ribonucleases, that is, general acid-base catalysis via the formation of acid/base pairs by functional groups of amino acids present in the aRNases; the pH profile of cleavage confirmed this mechanism. The most active aRNases under study exhibited high antiviral activity and entirely inactivated influenza virus A/WSN/33/(H1N1) after a short incubation period of viral suspension under physiological conditions. PMID:26899594

  8. Pregnancy-specific glycoproteins bind integrin αIIbβ3 and inhibit the platelet-fibrinogen interaction.

    Directory of Open Access Journals (Sweden)

    Daniel K Shanley

    Full Text Available Pregnancy-specific glycoproteins (PSGs are immunoglobulin superfamily members encoded by multigene families in rodents and primates. In human pregnancy, PSGs are secreted by the syncytiotrophoblast, a fetal tissue, and reach a concentration of up to 400 ug/ml in the maternal bloodstream at term. Human and mouse PSGs induce release of anti-inflammatory cytokines such as IL-10 and TGFβ1 from monocytes, macrophages, and other cell types, suggesting an immunoregulatory function. RGD tri-peptide motifs in the majority of human PSGs suggest that they may function like snake venom disintegrins, which bind integrins and inhibit interactions with ligands. We noted that human PSG1 has a KGD, rather than an RGD motif. The presence of a KGD in barbourin, a platelet integrin αIIbβ3 antagonist found in snake venom, suggested that PSG1 may be a selective αIIbβ3 ligand. Here we show that human PSG1 binds αIIbβ3 and inhibits the platelet - fibrinogen interaction. Unexpectedly, however, the KGD is not critical as multiple PSG1 domains independently bind and inhibit αIIbβ3 function. Human PSG9 and mouse Psg23 are also inhibitory suggesting conservation of this function across primate and rodent PSG families. Our results suggest that in species with haemochorial placentation, in which maternal blood is in direct contact with fetal trophoblast, the high expression level of PSGs reflects a requirement to antagonise abundant (3 mg/ml fibrinogen in the maternal circulation, which may be necessary to prevent platelet aggregation and thrombosis in the prothrombotic maternal environment of pregnancy.

  9. Molecular evolution of dentin phosphoprotein among toothed and toothless animals

    Directory of Open Access Journals (Sweden)

    Fisher Larry W

    2009-12-01

    Full Text Available Abstract Background Dentin sialophosphoprotein (DSPP is the largest member of the SIBLING family and is the most abundant noncollagenous protein in dentin. DSPP is also expressed in non-mineralized tissues including metabolically active ductal epithelia and some cancers. Its function, however, is poorly defined. The carboxy-terminal fragment, dentin phosphoprotein (DPP is encoded predominantly by a large repetitive domain that requires separate cloning/sequencing reactions and is, therefore, often incomplete in genomic databases. Comparison of DPP sequences from at least one member of each major branch in the mammalian evolutionary tree (including some "toothless" mammals as well as one reptile and bird may help delineate its possible functions in both dentin and ductal epithelia. Results The BMP1-cleavage and translation-termination domains were sufficiently conserved to permit amplification/cloning/sequencing of most species' DPP. While the integrin-binding domain, RGD, was present in about half of species, only vestigial remnants of this tripeptide were identified in the others. The number of tandem repeats of the nominal SerSerAsp phosphorylation motif in toothed mammals (including baleen whale and platypus which lack teeth as adults, ranged from ~75 (elephant to >230 (human. These repeats were not perfect, however, and patterns of intervening sequences highlight the rapidity of changes among even closely related species. Two toothless anteater species have evolved different sets of nonsense mutations shortly after their BMP1 motifs suggesting that while cleavage may be important for DSPP processing in other tissues, the DPP domain itself may be required only in dentin. The lizard DSPP had an intact BMP1 site, a remnant RGD motif, as well as a distinctly different Ser/Asp-rich domain compared to mammals. Conclusions The DPP domain of DSPP was found to change dramatically within mammals and was lost in two truly toothless animals. The

  10. A new three dimensional biomimetic hydrogel to deliver factors secreted by human mesenchymal stem cells in spinal cord injury.

    Science.gov (United States)

    Caron, Ilaria; Rossi, Filippo; Papa, Simonetta; Aloe, Rossella; Sculco, Marika; Mauri, Emanuele; Sacchetti, Alessandro; Erba, Eugenio; Panini, Nicolò; Parazzi, Valentina; Barilani, Mario; Forloni, Gianluigi; Perale, Giuseppe; Lazzari, Lorenza; Veglianese, Pietro

    2016-01-01

    Stem cell therapy with human mesenchymal stem cells (hMSCs) represents a promising strategy in spinal cord injury (SCI). However, both systemic and parenchymal hMSCs administrations show significant drawbacks as a limited number and viability of stem cells in situ. Biomaterials able to encapsulate and sustain hMSCs represent a viable approach to overcome these limitations potentially improving the stem cell therapy. In this study, we evaluate a new agarose/carbomer based hydrogel which combines different strategies to optimize hMSCs viability, density and delivery of paracrine factors. Specifically, we evaluate a new loading procedure on a lyophilized scaffold (soaked up effect) that reduces mechanical stress in encapsulating hMSCs into the hydrogel. In addition, we combine arginine-glycine-aspartic acid (RGD) tripeptide and 3D extracellular matrix deposition to increase the capacity to attach and maintain healthy hMSCs within the hydrogel over time. Furthermore, the fluidic diffusion from the hydrogel toward the injury site is improved by using a cling film that oriented efficaciously the delivery of paracrine factors in vivo. Finally, we demonstrate that an improved combination as here proposed of hMSCs and biomimetic hydrogel is able to immunomodulate significantly the pro-inflammatory environment in a SCI mouse model, increasing M2 macrophagic population and promoting a pro-regenerative environment in situ.

  11. Selective inhibition of MG-63 osteosarcoma cell proliferation induced by curcumin-loaded self-assembled arginine-rich-RGD nanospheres

    Directory of Open Access Journals (Sweden)

    Chang R

    2015-05-01

    Full Text Available Run Chang,1 Linlin Sun,1 Thomas J Webster1,2 1Department of Chemical Engineering, Northeastern University, Boston, MA, USA; 2Center of Excellence for Advanced Materials Research, King Abdulaziz University, Jeddah, Saudi Arabia Abstract: Osteosarcoma is the most frequent primary malignant form of bone cancer, comprising 30% of all bone cancer cases. The objective of this in vitro study was to develop a treatment against osteosarcoma with higher selectivity toward osteosarcoma cells and lower cytotoxicity toward normal healthy osteoblast cells. Curcumin (or diferuloylmethane has been found to have antioxidant and anticancer effects by multiple cellular pathways. However, it has lower water solubility and a higher degradation rate in alkaline conditions. In this study, the amphiphilic peptide C18GR7RGDS was used as a curcumin carrier in aqueous solution. This peptide contains a hydrophobic aliphatic tail group leading to their self-assembly by hydrophobic interactions, as well as a hydrophilic head group composed of an arginine-rich and an arginine-glycine-aspartic acid structure. Through characterization by transmission electron microscopy, self-assembled structures of spherical amphiphilic nanoparticles (APNPs with diameters of 10–20 nm in water and phosphate-buffered saline were observed, but this structure dissociated when the pH value was reduced to 4. Using a method of codissolution with acetic acid and dialysis tubing, the solubility of curcumin was enhanced and a homogeneous solution was formed in the presence of APNPs. Successful encapsulation of curcumin in APNPs was then confirmed by Fourier transform infrared and X-ray diffraction analyses. The cytotoxicity and cellular uptake of the APNP/curcumin complexes on both osteosarcoma and normal osteoblast cell lines were also evaluated by methyl-thiazolyl-tetrazolium assays and confocal fluorescence microscopy. The results showed that the curcumin-loaded APNPs had significant selective cytotoxicity against MG-63 osteosarcoma cells when compared with normal osteoblasts. We have demonstrated for the first time that APNPs can encapsulate hydrophobic curcumin in their hydrophobic cores, and curcumin-loaded APNPs could be an innovative treatment for the selective inhibition of osteosarcoma cells. Keywords: osteosarcoma, selective inhibition, curcumin, arginine-rich, arginine-glycine-aspartic acid, self-assembly

  12. A cyclic-RGD-BioShuttle functionalized with TMZ by DARinv “Click Chemistry” targeted to αvβ3 integrin for therapy

    OpenAIRE

    Braun, Klaus; Wiessler, Manfred; Pipkorn, Rüdiger; Ehemann, Volker; Bäuerle, Tobias; Fleischhacker, Heinz; Müller, Gabriele; Lorenz, Peter; Waldeck, Waldemar

    2010-01-01

    Clinical experiences often document, that a successful tumor control requires high doses of drug applications. It is widely believed that unavoidable adverse reactions could be minimized by using gene-therapeutic strategies protecting the tumor-surrounding healthy tissue as well as the bone-marrow. One new approach in this direction is the use of “Targeted Therapies” realizing a selective drug targeting to gain effectual amounts at the target site, even with drastically reduced application do...

  13. A cyclic-RGD-BioShuttle functionalized with TMZ by DARinv “Click Chemistry” targeted to αvβ3 integrin for therapy

    OpenAIRE

    Klaus Braun, Manfred Wiessler, Rüdiger Pipkorn, Volker Ehemann, Tobias Bäuerle, Heinz Fleischhacker, Gabriele Müller, Peter Lorenz, Waldemar Waldeck

    2010-01-01

    Clinical experiences often document, that a successful tumor control requires high doses of drug applications. It is widely believed that unavoidable adverse reactions could be minimized by using gene-therapeutic strategies protecting the tumor-surrounding healthy tissue as well as the bone-marrow. One new approach in this direction is the use of “Targeted Therapies” realizing a selective drug targeting to gain effectual amounts at the target site, even with drastically reduced ap...

  14. Suppression of ITGB4 Gene Expression in PC-3 Cells with Short Interfering RNA Induces Changes in the Expression of β-Integrins Associated with RGD-Receptors.

    Science.gov (United States)

    Knyazev, E N; Nyushko, K M; Alekseev, B Ya; Samatov, T R; Shkurnikov, M Yu

    2015-08-01

    We studied the effect of transfection of PC-3 prostate cancer cells with a plasmid encoding shRNA complimentary to a fragment of integrin β4 (ITGB4). The results attest to considerable changes in the transcriptome of transfected cells. For instance, compensatory changes in the expression of integrin family genes were found. PMID:26395630

  15. Synthesis of [F-18]RGD-K5 by catalyzed [3+2] cycloaddition for imaging integrin alpha(v)beta(3) expression in vivo

    NARCIS (Netherlands)

    Mirfeizi, Leila; Walsh, Joe; Kolb, Hartmuth; Campbell-Verduyn, Lachlan; Dierckx, Rudi A.; Feringa, Ben L.; Elsinga, Philip H.; de Groot, Tjibbe; Sannen, Ivan; Bormans, Guy; Celen, Sofie

    2013-01-01

    In the last few years click chemistry reactions, and in particular copper-catalyzed cycloadditions have been used extensively for the preparation of new bioconjugated molecules such as F-18-radiolabeled radiopharmaceuticals for positron emission tomography (PET). This study is focused on the synthes

  16. Lattice specific heat for the RMIn5 (R=Gd, La, Y; M=Co, Rh) compounds: Non-magnetic contribution subtraction

    Science.gov (United States)

    Facio, Jorge I.; Betancourth, D.; Cejas Bolecek, N. R.; Jorge, G. A.; Pedrazzini, Pablo; Correa, V. F.; Cornaglia, Pablo S.; Vildosola, V.; García, D. J.

    2016-06-01

    We analyze theoretically a common experimental process used to obtain the magnetic contribution to the specific heat of a given magnetic material. In the procedure, the specific heat of a non-magnetic analog is measured and used to subtract the non-magnetic contributions, which are generally dominated by the lattice degrees of freedom in a wide range of temperatures. We calculate the lattice contribution to the specific heat for the magnetic compounds GdMIn5 (M=Co, Rh) and for the non-magnetic YMIn5 and LaMIn5 (M=Co, Rh), using density functional theory based methods. We find that the best non-magnetic analog for the subtraction depends on the magnetic material and on the range of temperatures. While the phonon specific heat contribution of YRhIn5 is an excellent approximation to the one of GdCoIn5 in the full temperature range, for GdRhIn5 we find a better agreement with LaCoIn5, in both cases, as a result of an optimum compensation effect between masses and volumes. We present measurements of the specific heat of the compounds GdMIn5 (M=Co, Rh) up to room temperature where it surpasses the value expected from the Dulong-Petit law. We obtain a good agreement between theory and experiment when we include anharmonic effects in the calculations.

  17. ANTIMETASTATIC EFFECT OF INTEGRIN IIb/IIIa INHIBITORS ON SALIVARY ADENOID CYSTIC CARCINOMA

    Institute of Scientific and Technical Information of China (English)

    LI; Feng-he

    2001-01-01

    [1]Gasic GJ, Gasic TB, Stewart CC, et al. Antimetastatic effects associated with platelet reduction [J]. Proc Natl Acad Sci 1968; 61:46.[2]Gu YZ, Qiu WL, He RG, et al. An experimental study of the effects of aspirin on the adhesion of salivary adenoid cystic carcinoma cells [J]. Shanghai Stomatol 1999; 8:65.[3]Im SY, Ko HM, Ko JW, et al. Augmentation of tumor metastasis by platelet-activating factor [J]. Cancer Res 1996; 56:2262.[4]Tang DG, Onoda JM, Steinert BW. Phenotypic properties of cultured tumor cells: Integrin IIb/b 3 expression, tumor-cell-induced platelet aggrega-tion, and tumor cell adhesion to endothelium as an important parameters of experimental metastasis [J]. Int J Cancer 1993; 54:338.[5]Oleksowicz L, Mrowiec Z, Schwartz E, et al. Characterization of tumor-induced platelet aggregafion: The role of immunorelated GP IIb/IIIa expression by MCF-7 breast cancer cells [J]. Thromb Rest 1995; 79:261.[6]Nierodzik ML, Klepfish A, Karptkin S, et al. Role of platelet thrombin, integrin IIb-IIIa, fibronectin and von Willebrand factors on tumor adhesion in vitro and metastasis in vivo [J]. Thromb Haemost 1995; 74:282.[7]Guan Xiao-feng, Qiu Wei-liu, He Rong-gen, et al. The selection of a highly pulmonary metastatic cell line of salivary adenoid cystic carcinoma [J]. Chi J Stomatol 1996; 31:74.[8]Bhatti RA, Gadarowski J, Ray P, et al. Potential role of platelet and coagulation factors in the metastasis of prostatic cancer [J]. Invasion Metastasis 1996; 16:49.[9]Li Sheng-lin, Liu Xiu-Ping, Zhang Kui-hua. Establishment of a human salivary adenoid cystic carcinoma cell line and its characteristics [J]. Chi J Stomatol 1990; 25:29.[10]Chang HS, Yang RS, Huang TF. The Arg-Gly-Asp-containing peptide, rhodostomin, inhibits in vitro cell adhesion to extracellular matrices and platelet aggregation caused by Sao-2 human osteosarcoma cells [J]. Br J Cancer 1995; 71:265.[11]Karptkin S, Pearlstein E, Ambrogio C, et al. Role of adhesive

  18. Protein hydrogels with engineered biomolecular recognition

    Science.gov (United States)

    Mi, Lixin

    water soluble and swelling. The RGD cell adhesion tripeptide has been inserted into the polyelectrolyte region by site-directed mutagenesis. Two dimensional human foreskin fibroblast cultures have shown that the RGD-containing protein surface is bioactive in promoting cell attachment, cell signaling, and cytoskeleton organization. The protein and the cell recognize and interact at molecular level. Collectively, these findings indicate that this bioactive protein hydrogel system is a promising biomaterial for mammalian cell culture. This research may provide insights for the rational development of bioactive ECM for specific cell and tissue engineering applications.

  19. Rapid and sensitive analysis of 27 underivatized free amino acids, dipeptides, and tripeptides in fruits of Siraitia grosvenorii Swingle using HILIC-UHPLC-QTRAP(®)/MS (2) combined with chemometrics methods.

    Science.gov (United States)

    Zhou, Guisheng; Wang, Mengyue; Li, Yang; Peng, Ying; Li, Xiaobo

    2015-08-01

    In the present study, a new strategy based on chemical analysis and chemometrics methods was proposed for the comprehensive analysis and profiling of underivatized free amino acids (FAAs) and small peptides among various Luo-Han-Guo (LHG) samples. Firstly, the ultrasound-assisted extraction (UAE) parameters were optimized using Plackett-Burman (PB) screening and Box-Behnken designs (BBD), and the following optimal UAE conditions were obtained: ultrasound power of 280 W, extraction time of 43 min, and the solid-liquid ratio of 302 mL/g. Secondly, a rapid and sensitive analytical method was developed for simultaneous quantification of 24 FAAs and 3 active small peptides in LHG at trace levels using hydrophilic interaction ultra-performance liquid chromatography coupled with triple-quadrupole linear ion-trap tandem mass spectrometry (HILIC-UHPLC-QTRAP(®)/MS(2)). The analytical method was validated by matrix effects, linearity, LODs, LOQs, precision, repeatability, stability, and recovery. Thirdly, the proposed optimal UAE conditions and analytical methods were applied to measurement of LHG samples. It was shown that LHG was rich in essential amino acids, which were beneficial nutrient substances for human health. Finally, based on the contents of the 27 analytes, the chemometrics methods of unsupervised principal component analysis (PCA) and supervised counter propagation artificial neural network (CP-ANN) were applied to differentiate and classify the 40 batches of LHG samples from different cultivated forms, regions, and varieties. As a result, these samples were mainly clustered into three clusters, which illustrated the cultivating disparity among the samples. In summary, the presented strategy had potential for the investigation of edible plants and agricultural products containing FAAs and small peptides. PMID:25976704

  20. Rapid and sensitive analysis of 27 underivatized free amino acids, dipeptides, and tripeptides in fruits of Siraitia grosvenorii Swingle using HILIC-UHPLC-QTRAP(®)/MS (2) combined with chemometrics methods.

    Science.gov (United States)

    Zhou, Guisheng; Wang, Mengyue; Li, Yang; Peng, Ying; Li, Xiaobo

    2015-08-01

    In the present study, a new strategy based on chemical analysis and chemometrics methods was proposed for the comprehensive analysis and profiling of underivatized free amino acids (FAAs) and small peptides among various Luo-Han-Guo (LHG) samples. Firstly, the ultrasound-assisted extraction (UAE) parameters were optimized using Plackett-Burman (PB) screening and Box-Behnken designs (BBD), and the following optimal UAE conditions were obtained: ultrasound power of 280 W, extraction time of 43 min, and the solid-liquid ratio of 302 mL/g. Secondly, a rapid and sensitive analytical method was developed for simultaneous quantification of 24 FAAs and 3 active small peptides in LHG at trace levels using hydrophilic interaction ultra-performance liquid chromatography coupled with triple-quadrupole linear ion-trap tandem mass spectrometry (HILIC-UHPLC-QTRAP(®)/MS(2)). The analytical method was validated by matrix effects, linearity, LODs, LOQs, precision, repeatability, stability, and recovery. Thirdly, the proposed optimal UAE conditions and analytical methods were applied to measurement of LHG samples. It was shown that LHG was rich in essential amino acids, which were beneficial nutrient substances for human health. Finally, based on the contents of the 27 analytes, the chemometrics methods of unsupervised principal component analysis (PCA) and supervised counter propagation artificial neural network (CP-ANN) were applied to differentiate and classify the 40 batches of LHG samples from different cultivated forms, regions, and varieties. As a result, these samples were mainly clustered into three clusters, which illustrated the cultivating disparity among the samples. In summary, the presented strategy had potential for the investigation of edible plants and agricultural products containing FAAs and small peptides.

  1. iRGD靶向载药脂质体-微泡复合物的制备及其靶向性研究%Preparation and targeting effect of iRGD modified liposome-microbubble complex

    Institute of Scientific and Technical Information of China (English)

    张婧; 严飞; 李莉

    2016-01-01

    目的:制备iRGD靶向载药脂质体-微泡复合物,研究其靶向性.方法:采用薄膜-超声分散法制备生物素化的iRGD靶向载药脂质体和生物素化的超声微泡.利用生物素-亲和素系统(Biotin-avidin-system,BAS)连接脂质体与微泡,构建并表征iRGD靶向载药脂质体-微泡复合物.细胞黏附实验验证复合物的体外靶向结合性能;构建小鼠乳腺癌移植瘤模型,通过靶组织的药物荧光强度验证复合物的体内靶向性.结果:iRGD靶向载药脂质体的粒径为(165.07±4.01)nm,电位为(-12.92±0.26)mv,复合物的载药量为每108个复合物载紫杉醇(46.22±1.95)μg;黏附实验表明靶向组复合物与血管内皮细胞结合数量明显多于非靶向组复合物(7.8±1.1,0.2±0.45,P<0.01);荷瘤小鼠活体成像实验显示靶向组复合物的肿瘤组织荧光明显强于非靶向组复合物.结论:iRGD靶向载药脂质体-微泡复合物,作为一种靶向给药系统,可以实现超声分子成像与超声给药的有机结合,显著提高药物靶向递送的效率.

  2. Delivery of the p38 MAPkinase inhibitor SB202190 to angiogenic endothelial cells : Development of novel RGD-equipped and PEGylated drug - Albumin conjugates using platinum(II)-based drug linker technology

    NARCIS (Netherlands)

    Temming, Kai; Lacombe, Marie; van der Hoeven, Paul; Prakash, Jai; Gonzalo, Teresa; Dijkers, Eli C. F.; Orfi, Laszlo; Keri, Gyorgy; Poelstra, Klaas; Molema, Grietje; Kok, Robbert J.

    2006-01-01

    Endothelial cells play an important role in inflammatory disorders, as they control the recruitment of leukocytes into inflamed tissue and the formation of new blood vessels. Activation of p38MAP kinase results in the production of proinflammatory cytokines and the expression of adhesion molecules.

  3. Interferon-gamma up-regulates a unique set of proteins in human keratinocytes. Molecular cloning and expression of the cDNA encoding the RGD-sequence-containing protein IGUP I-5111

    DEFF Research Database (Denmark)

    Honoré, B; Leffers, H; Madsen, Peder;

    1993-01-01

    transformation and because peptide sequences were available in the microsequence database. The cDNA was cloned from a fibroblast cDNA library using degenerate oligodeoxyribonucleotides and expressed in AMA cells using the vaccinia virus expression system. Database searches indicated that the predicted protein......Treatment of proliferating and quiescent primary human keratinocytes with interferon-gamma (IFN-gamma) (100 U/ml, 23.5 h) followed by two-dimensional gel analysis revealed three proteins, IGUP I-3421 (M(r) = 48,200, pI = 6.06); IGUP I-3524 (M(r) = 56,900, pI = 5.92), a protein homologous to peptide......-chain-release factor and tryptophanyl-tRNA synthetase; and IGUP I-5111 (M(r) = 30,400, pI = 5.76) recorded in the keratinocyte protein database (Celis et al., 1991, 1992) that are highly and specifically up-regulated by IFN-gamma among several agents tested including 14 other cytokines, second messengers [dibutyryl c...

  4. Effect of rare earth ion in the thermopower of R{sub 5}(Si{sub x}Ge{sub 1-x}){sub 4} compounds with x{approx}0.5 and R=Gd and Tb

    Energy Technology Data Exchange (ETDEWEB)

    Pereira, A.M. [IFIMUP, Rua do Campo Alegre, 678, 4169-007 Porto (Portugal)]. E-mail: ampereira@fc.up.pt; Magen, C. [Instituto de Ciencia de Materiales de Aragon Universidad de Zaragoza and Consejo Superior de Investigaciones Cient icas, 50009 Zaragoza (Spain); Araujo, J.P. [IFIMUP, Rua do Campo Alegre, 678, 4169-007 Porto (Portugal); Algarabel, P.A. [Instituto de Ciencia de Materiales de Aragon Universidad de Zaragoza and Consejo Superior de Investigaciones Cient icas, 50009 Zaragoza (Spain); Morellon, L. [Instituto de Ciencia de Materiales de Aragon Universidad de Zaragoza and Consejo Superior de Investigaciones Cient icas, 50009 Zaragoza (Spain); Instituto de Nanociencia de Aragon, 50009 Zaragoza (Spain); Braga, M.E. [IFIMUP, Rua do Campo Alegre, 678, 4169-007 Porto (Portugal); Pinto, R.P. [IFIMUP, Rua do Campo Alegre, 678, 4169-007 Porto (Portugal); Ibarra, M.R. [Instituto de Ciencia de Materiales de Aragon Universidad de Zaragoza and Consejo Superior de Investigaciones Cient icas, 50009 Zaragoza (Spain); Instituto de Nanociencia de Aragon, 50009 Zaragoza (Spain); Sousa, J.B. [IFIMUP, Rua do Campo Alegre, 678, 4169-007 Porto (Portugal)

    2007-03-15

    The magnetocaloric ferromagnetic compounds with R{sub 5}(Si{sub x}Ge{sub 1-x}){sub 4} composition are very sensitive to the Si:Ge ratio (x) and the particular Rare earth (R) ion. Here we study the thermopower S(T) behavior of samples with a similar x{approx}0.5 ratio but different rare earth ions: Gd{sub 5}(Si{sub 0.45}Ge{sub 0.55}){sub 4} and Tb{sub 5}(Si{sub 0.5}Ge{sub 0.5}){sub 4}, between 10 and 290K. Above the Curie temperature (T{sub C}) both S and its T-dependence do not change with the R-ion but, upon cooling, a first-order magneto-structural transition occurs at T{sub S} (close to T{sub C}) causing an abruptness in thermopower ({delta}S/S{approx}32%). At low temperatures very distinct S(T) behavior is observed, and a spin reorientation transition occurs in the Tb compound.

  5. A novel dual-targeted ultrasound contrast agent provides improvement of gene delivery efficiency in vitro.

    Science.gov (United States)

    Xu, Jinfeng; Zeng, Xinxin; Liu, Yingying; Luo, Hui; Wei, Zhanghong; Liu, Huiyu; Zhou, Yuli; Zheng, Hairong; Zhou, Jie; Tan, Guanghong; Yan, Fei

    2016-07-01

    Ultrasound-targeted microbubble destruction (UTMD) has become a novel gene/drug delivery method in cancer therapeutic application. However, the gene transfection efficiency mediated by UTMD is still unsatisfactory. Here, we introduced iRGD/CCR2 dual-targeted cationic microbubbles (MBiRGD/CCR2) which was modified with PEI-600 and coated with iRGD peptides and anti-CCR-2 antibodies. It showed that MBiRGD/CCR2 had a 25.83 ± 1.57 mV surface zeta potential and good stability. The experiments in vitro showed MBiRGD/CCR2 had higher binding efficiency with both bEnd.3 cells and MCF-7 cells than that of iRGD or CCR2 single-targeted cationic microbubbles (MBiRGD or MBCCR2) (P plasmid DNA. Compared with the plain MBs (MBcontrol) or single-targeted cationic MBs including MBiRGD and MBCCR2 (P < 0.05 for all), the dual-targeted cationic MBiRGD/CCR2 groups had higher gene transfection efficiency under US exposure. It showed that the dual-targeted cationic MBiRGD/CCR2 has a potential value to be used as an ultrasound imaging probe for ultrasound image-guided tumor gene therapy. PMID:26733178

  6. 新型活性修饰对聚乳酸组织工程骨支架上种子细胞生物学行为的影响%Effects of bioactive modification of poly-D,L-lactide acid scaffolds on the biological behaviors of the seed cells

    Institute of Scientific and Technical Information of China (English)

    许子星; 陈建庭; 李涛; 查丁胜; 张鑫鑫; 姜晓锐; 肖文德; 朱青安

    2011-01-01

    Objective To study the changes in the biological behavior of bone marrow mesenchymal stem cells (BMSCs) transfected with red fluorescent protein by lentivirus (RFP-BMSCs) seeded on in poly-D, L-lactide acid (PDLLA) scaffolds with bioactive modification by ammonia plasma and Gly-Arg-Gly-Asp-Ser (GRGDS) in vitro. Methods Circular sheets of PDLLA scaffolds (8 mm in diameter and 1 mm in thickness) were prepared and aminated with PDLLA (group A) or modified with the peptide conjugate A/PDLLA (group PA), with untreated PDLLA as the control (group P). The RFP-BMSCs were seeded on the scaffold materials and their proliferation and metabolic activity were detected using CyQuant NF and Alamar blue staining. The mineralization on the scaffolds was observed using calcein fluorescent dye under a fluorescent microscope. The adhesion and proliferation of RFP-BMSCs were observed by fluorescent microscope, and scanning electron microscope (SEM) was used to confirm the observed adhesion of the seed cells. Results The RFP-BMSCs seeded on the 3 scaffolds all showed proliferative activity at different time points after cell seeding, and the cell numbers decreased significantly in the order of PA>A>P (P0.001). The cell number was significantly greater in group PA than in group A at all the time points except for days 10 (P=0.077) and 12 (P=0.491), and gradually became similar with the passage of time. The metabolic changes of the cells follow a similar pattern of cell proliferation. RFP-BMSCs showed more active proliferation in group A and group PA than in group P. On days 14 and 21, the intensity of green fluorescence decreased in the order of group PA, A and P. The RFP-BMSCs showed better adhesion in group PA than in group A, and the cells in group P appeared more scattered under scanning electron microscope. Conclusion Bioactive modification of PDLLA by ammonia treatment and conjugation withGRGDS peptides may promotes the adhesion, proliferation, metabolism and mineralization of RFP

  7. Sensitive gas chromatographic detection of acetaldehyde and acetone using a reduction gas detector

    Science.gov (United States)

    O'Hara, Dean; Singh, Hanwant B.

    1988-01-01

    The response of a newly available mercuric oxide Reduction Gas Detector (RGD-2) to subpicomole and larger quantities of acetaldehyde and acetone is tested. The RGD-2 is found to be capable of subpicomole detection for these carbonyls and is more sensitive than an FID (Flame Ionization Detector) by an order of magnitude. Operating parameters can be further optimized to make the RGD-2 some 20-40 times more sensitive than an FID. The detector is linear over a wide range and is easily adapted to a conventional gas chromatograph (GC). Such a GC-RGD-2 system should be suitable for atmospheric carbonyl measurements in clean as well as polluted environments.

  8. 气相中环糊精与甘氨酰-苯丙氨酰-苯丙氨酸和甘氨酸三肽非共价复合物的质谱研究%Investigation on Non-Covalent Complexes of Cyclodextrins with GGG and GFF Tripeptides in Gas Phase by Mass Spectrometry

    Institute of Scientific and Technical Information of China (English)

    何小丹; 许崇晟; 储艳秋; 丁传凡

    2013-01-01

    为了探索环糊精和寡肽的非共价相互作用,一定化学计量比的a-,β-,γ-环糊精(CD)分别和甘氨酸三肽(GGG)、甘氨酰-苯丙氨酰-苯丙氨酸三肽(GFF)在室温下反应达到平衡并用正离子模式质谱检测.实验结果显示GGG,GFF均可以和α-,β-,γ-CD生成1∶1配合比的非共价复合物.碰撞诱导解离实验进一步验证了α-,β-,γ-CD与GGG,GFF非共价复合物的形成.质谱滴定法测得的结合常数结果表明环糊精和两种三肽形成非共价复合物的结合强度均按照γ-,β-,α-CD的次序逐渐增大.GGG和α-,β-,γ-CD复合物的结合常数分别为2799.96,2528.73,1697.11L·mol-1,GFF和α-,β-,γ-CD复合物的结合常数分别为2773.94,2134.03,1330.68 L·mol-1对于α-,β-或γ-CD,含有苯基的GFF+ CD复合物的结合强度要小于相应的脂肪族的GGG+ CD复合物,表明虽然在气相GFF+CD复合物的构象与溶液中的构象有所变化,但是苯基仍然参与和环糊精疏水腔体的键合作用.%To investigate the non-covalent interaction between α-, β-, y-cyclodextrins and peptides, a stoichiometry of α-, β-, y-cyclodextrins (CD) with GGG (Gly-Gly-Gly) or GFF (Gly-Phe-Phe) was mixed respectively, and then incubated at room temperature for 12 h to reach the equilibrium. In positive mode, the electrospray ionization mass spectrometry (ESI-MS) results indicated that α-, β-, γ-CD with GGG or GFF could form non-covalent complexes, respectively. The binding of cyclodextrins with GGG or GFF was further confirmed by collision induced dissociation (CID) in a tandem mass spectrometer. The formation constants of six complexes (GGG+CD and GFF+CD) were determined by mass spectrometric titration. The results showed the formation constants for both GGG's and GFF's complexes increased according to the order γ-CD, β-CD, α-CD. The formation constants Kst values for GGG complexes with α-CD, β-CD or γ-CD are 2799.96, 2528.73, 1697.11 L·mol-1, respectively. While the formation constants Kst values for GFF complexes with α-CD, β-CD or γ-CD are 2773.94, 2134.03, 1330.68 L·mol-1 respectively. For α-CD, β-CD or γ-CD, the Kst values of GFF complexes containing aromatic group are smaller than those of GGG complexes only containing aliphatic group. The main reason is that in gas phase, with the weakening of hydrophobic force, Van der Waals force plays an important role in the conjugation process of GFF with CD, the coordinating group of GFF is still phenyl group. While in GGG's complexes, the hydrogen bond dominates in the conjugation process. Our convincing results from the formation constants provides a new evidence, indicating that although the conformations for GFF+CD complexes change slightly when the analysts transfer from solution to gas phase, the phenyl group still takes part in coordinating.

  9. The study of irradiation combined with targeted suicide gene therapy for prostate cancer xenografts

    International Nuclear Information System (INIS)

    Objective: To study whether RGD-4C AAVP HSV-TK/GCV, one of suicide gene therapy targeting to Integrin αv, can enhance radiotherapeutic effect for DU145 prostate cancer xenografts or not. Methods: When the diameter of tumor in 48 nude mice bearing DU145 prostate cancer in the right leg attained 6.0 mm (5.8-6.3 mm), the mice were entered into the experiment. There were 6 experimental groups (8 mice per group), including the control, radiotherapy only (RT), RGD-4C AAVP HSV-TK/GCV only (Targeted, RGD-4C), AAVP HSV-TK/GCV (Non-targeted, non RGD-4C ), radiotherapy plus RGD- 4C AAVP HSV-TK/GCV(XRT + RGD-4C) and radiotherapy plus AAVP HSV-TK/GCV group (XRT + Non RGD-4C). The effect of treatment was assessed by tumor growth delay ( the time required when tumor grew from 6.0 mm to 12.0 mm) and tumor cure. Results: Five mice died during the treatment course. There were 6 mice without tumor after treatment, including 1 in RT group, 1 in RGD-4C group, 1 in non RGD-4C group and 3 in XRT + RGD-4C group, respectively. For tumor growth delay analysis in 37 mice, the absolute growth delay (AGD) for RGD-4C, non RGD-4C and RT group was 24.4 ± 9.0, 22.6±11.3 and 28.3 ±5.5 days, respectively. When RGD-4C AAVP HSV-TK/GCV or AAVP HSV-TK/GCV combined with radiotherapy, their AGD was 64.7±23.8 and 35.4±9.6 days, and nominal growth delay (NGD) was 40.3 ± 23.8 and 12.8 ± 9.6 days, respectively. The enhancement factor of RGD-4C AAVP HSV-TK/GCV and AAVP HSV-TK/GCV for radiotherapy were 1.42 and 0.45. Conclusion: RGD-4C AAVP HSV-TK/GCV can enhance radiotherapeutic effect for DU145 prostate cancer xenografts. Further study is needed. (authors)

  10. Tumor-penetrating peptide fused EGFR single-domain antibody enhances cancer drug penetration into 3D multicellular spheroids and facilitates effective gastric cancer therapy

    Science.gov (United States)

    Sha, Huizi; Zou, Zhengyun; Xin, Kai; Bian, Xinyu; Cai, Xueting; Lu, Wuguang; Chen, Jiao; Chen, Gang; Huang, Leaf; Blair, Andrew M.; Cao, Peng; Liu, Baorui

    2016-01-01

    Human tumors, including gastric cancer, frequently express high levels of epidermal growth factor receptors (EGFRs), which are associated with a poor prognosis. Targeted delivery of anticancer drugs to cancerous tissues shows potential in sparing unaffected tissues. However, it has been a major challenge for drug penetration in solid tumor tissues due to the complicated tumor microenvironment. We have constructed a recombinant protein named anti-EGFR-iRGD consisting of an anti-EGFR VHH (the variable domain from the heavy chain of the antibody) fused to iRGD, a tumor-specific binding peptide with high permeability. Anti-EGFR-iRGD, which targets EGFR and αvβ3, spreads extensively throughout both the multicellular spheroids and the tumor mass. The recombinant protein anti-EGFR-iRGD also exhibited antitumor activity in tumor cell lines, multicellular spheroids, and mice. Moreover, anti-EGFR-iRGD could improve anticancer drugs, such as doxorubicin (DOX), bevacizumab, nanoparticle permeability and efficacy in multicellular spheroids. This study draws attention to the importance of iRGD peptide in the therapeutic approach of anti-EGFR-iRGD. As a consequence, anti-EGFR-iRGD could be a drug candidate for cancer treatment and a useful adjunct of other anticancer drugs. PMID:25553823

  11. Peptide-Conjugated Quantum Dots Act as the Target Marker for Human Pancreatic Carcinoma Cells

    Directory of Open Access Journals (Sweden)

    Shuang-ling Li

    2016-03-01

    Full Text Available Background/Aims: In the present study, we describe a novel and straightforward approach to produce a cyclic- arginine-glycine-aspartic (RGD-peptide-conjugated quantum dot (QD probe as an ideal target tumor biomarker. Due to its specific structure, the probe can be used for targeted imaging of pancreatic carcinoma cells. Methods: Pancreatic carcinoma cells were routinely cultured and marked with QD-RGD probe. The QD-RGD probe on the fluorescence-labeled cancer cell was observed by fluorescence microscopy and laser confocal microscopy. Cancer cell viability was detected by MTT assay after culturing with QD-RGD probe. Results: Fluorescence microscopy and laser confocal microscopy displayed that 10nmol/L QD-RGD probe was able to effectively mark pancreatic carcinoma cells. In comparison with organic dyes and fluorescent proteins, the quantum dot-RGD probe had unique optical and electronic properties. Conclusion: QD-RGD probe has a low cytotoxicity with an excellent optical property and biocompatibility. These findings support further evaluation of QD-RGD probes for the early detection of pancreatic cancer.

  12. Potential therapeutic radiotracers: preparation, biodistribution and metabolic characteristics of 177Lu-labeled cyclic RGDfK dimer.

    Science.gov (United States)

    Shi, Jiyun; Liu, Zhaofei; Jia, Bing; Yu, Zilin; Zhao, Huiyun; Wang, Fan

    2010-06-01

    In this study, we reported the preparation and evaluation of (177)Lu-DOTA-RGD2, (177)Lu-DOTA-Bz-RGD2 and (177)Lu-DTPA-Bz-RGD2 (RGD2 = E[c(RGDfK)](2)) as a potential therapeutic radiotracers for the treatment of integrin alpha(v)beta(3)-positive tumors. The BALB/c nude mice bearing the U87MG human glioma xenografts were used to evaluate the biodistribution characteristics and excretion kinetics of (177)Lu-DOTA-RGD2, (177)Lu-DOTA-Bz-RGD2 and (177)Lu-DTPA-Bz-RGD2. It was found that there were no major differences in their lipophilicity and biodistribution characteristics, particularly at latter time points. A major advantage of using DTPA-Bz as the bifunctional chelator (BFC) was its high radiolabeling efficiency (fast and high yield radiolabeling) at room temperature. Using DOTA and DOTA-Bz as BFCs, the radiolabeling kinetics was slow, and heating at 100 degrees C and higher DOTA-conjugate concentration were needed for successful (177)Lu-labeling. Therefore, DTPA-Bz is an optimal BFC for routine preparation of (177)Lu-labeled cyclic RGDfK peptides, and (177)Lu-DTPA-Bz-RGD2 is worthy of further investigation for targeted radiotherapy of integrin alpha(v)beta(3)-positive tumors.

  13. Anaerobic and leucine-dependent expression of a peptide transport gene in Salmonella typhimurium.

    OpenAIRE

    Jamieson, D J; Higgins, C F

    1984-01-01

    Using Mu d1-mediated lac operon fusions, we studied the transcriptional regulation of the genes encoding two peptide transport systems, the oligopeptide permease and the tripeptide permease. The four opp genes were found to be constitutively expressed, whereas the genes encoding the tripeptide permease are under a complex set of regulatory controls. Two loci, tppA and tppB, are required for tripeptide permease function. Locus tppA is shown to be a positive regulator of tppB expression. In add...

  14. Altered Cell Cycle Arrest by Multifunctional Drug-Loaded Enzymatically-Triggered Nanoparticles.

    Science.gov (United States)

    Huang, Can; Sun, Ying; Shen, Ming; Zhang, Xiangyu; Gao, Pei; Duan, Yourong

    2016-01-20

    cRGD-targeting matrix metalloproteinase (MMP)-sensitive nanoparticles [PLGA-PEG1K-cRGD/PLGA-peptide-PEG5K (NPs-cRGD)] were successfully developed. Au-Pt(IV) nanoparticles, PTX, and ADR were encapsulated into NPs-RGD separately. The effects of the drug-loaded nanoparticles on the cell cycle were investigated. Here, we showed that higher cytotoxicity of drug-loaded nanoparticles was related to the cell cycle arrest, compared to that of free drugs. The NPs-cRGD studied here did not disrupt cell cycle progression. The cell cycle of Au-Pt(IV)@NPs-cRGD showed a main S phase arrest in all phases of the cell cycle phase, especially in G0/G1 phase. PTX@NPs-cRGD and ADR@NPs-cRGD showed a higher ratio of G2/M and S phase arrest than the free drugs, respectively. Cells in G0/G1 and S phases of the cell cycle had a higher uptake ratio of NPs-cRGD. A nutrient deprivation or an increase in the requirement of nutrients in tumor cells could promote the uptake of nanoparticles from the microenvironments. In vivo, NPs-cRGD could efficiently accumulate at tumor sites. The inhibition of tumor growth coupled with cell cycle arrest is in line with that in vitro. On the basis of our results, we propose that future studies on nanoparticle action mechanism should consider the cell cycle, which could be different from free drugs. Understanding the actions of cell cycle arrest could affect the application of nanomedicine in the clinic.

  15. Direct asymmetric aldol reaction using MBHA resin-supported peptide containing L-proline unit

    Institute of Scientific and Technical Information of China (English)

    Liang Zhang; Wen Bo Ding; Yong Ping Yu; Hong Bin Zou

    2009-01-01

    MBHA resin-supported tripeptide catalyst system containing L-proline unit has been developed for use in the direct asymmetric aldol reaction of acetone and aldehydes,which afford the corresponding products with satisfactory isolated yields and enantiomeric excesses.

  16. Expression, purification, and characterization of scar tissue neovasculature endothelial cell-targeted rhIL10 in Escherichia coli.

    Science.gov (United States)

    Shi, Jihong; Wan, Yi; Shi, Shan; Zi, Jing; Guan, Hao; Zhang, Yuejuan; Zheng, Zhao; Jia, Yanhui; Bai, Xiaozhi; Cai, Weixia; Su, Linlin; Zhu, Xiongxiang; Hu, Dahai

    2015-01-01

    Interleukin 10 (IL10) plays a pivotal role in the anti-inflammatory response and immunosuppressive reactions. It has also been identified as a new promising therapy for scar formation. Treatment of scars with IL10 has significant effects, but there are some shortcomings, including poor tissue-binding specificity and low effectiveness. RGD peptide has been demonstrated to bind specifically to αvβ3 integrin on neovasculature endothelial cells, and the excess production of neovasculature is crucial to scar formation. To increase efficacy against scar formation and to decrease the side effects on normal tissues, a novel hybrid protein combining human IL10 with RGD was designed. The DNA sequence encoding the recombinant fusion protein IL10-RGD (rhIL10-RGD) was subcloned into a pET22b (+) vector for protein expression in E. coli strain BL21 (DE3). SDS-PAGE analysis displayed an induced expression product band at a molecular weight of 19.3 kDa, which constituted 30 % of the total bacterial protein. We developed a procedure to purify rhIL10-RGD from inclusion bodies and then renatured the protein using dialysis against urea with a step-down concentration procedure. Hypertrophic scar fibroblasts (HSFs) were treated with rhIL10-RGD, and the fibrosis-related protein levels were assessed by Western blotting. The results indicated that rhIL10-RGD can downregulate the expression levels of Col1 and α-SMA in HSFs and suppress tube formation of HUVECs. These results indicate that rhIL10-RGD has anti-fibrosis effects and can potentially be used to treat the neovasculature in scar formation and improve the abnormal deposition of the extracellular matrix (ECM). Thus, rhIL10-RGD may be a more effective candidate for scar-improvement and anti-fibrosis therapy.

  17. Toxicity assessment of repeated intravenous injections of arginine–glycine–aspartic acid peptide conjugated CdSeTe/ZnS quantum dots in mice

    Directory of Open Access Journals (Sweden)

    Wang YW

    2014-10-01

    Full Text Available You-Wei Wang, Kai Yang, Hong Tang, Dan Chen, Yun-Long Bai Department of Oral and Maxillofacial Surgery, The First Affiliated Hospital of Chongqing Medical University, Chongqing, People’s Republic of China Background: Nanotechnology-based near-infrared quantum dots (NIR QDs have many excellent optical properties, such as high fluorescence intensity, good fluorescence stability, and strong tissue-penetrating ability. Integrin αvß3 is highly and specifically expressed in tumor angiogenic vessel endothelial cells of almost all carcinomas. Recent studies have shown that NIR QDs linked to peptides containing the arginine–glycine–aspartic acid (RGD sequence (NIR QDs-RGD can specifically target integrin αvß3 expressed in endothelial cells of tumor angiogenic vessels in vivo, and they offer great potential for early cancer diagnosis, in vivo tumor imaging, and tumor individualized therapy. However, the toxicity profile of NIR QDs-RGD has not been reported. This study was conducted to investigate the toxicity of NIR QDs-RGD when intravenously administered to mice singly and repeatedly at the dose required for successful tumor imaging in vivo.Materials and methods: A NIR QDs-RGD probe was prepared by linking NIR QDs with the maximum emission wavelength of 800 nm (QD800 to the RGD peptide (QD800-RGD. QD800-RGD was intravenously injected to BALB/C mice once or twice (200 pmol equivalent of QD800 for each injection. phosphate-buffered saline solution was used as control. Fourteen days postinjection, toxicity tests were performed, including complete blood count (white blood cell, red blood cell, hemoglobin, platelets, lymphocytes, and neutrophils and serum biochemical analysis (total protein, albumin, albumin/globulin, aspartate aminotransferase, alanine aminotransferase, and blood urea nitrogen. The coefficients of liver, spleen, kidney, and lung weight to body weight were measured, as well as their oxidation and antioxidation indicators, including

  18. Novel Method for Radiolabeling and Dimerizing Thiolated Peptides Using (18)F-Hexafluorobenzene.

    Science.gov (United States)

    Jacobson, Orit; Yan, Xuefeng; Ma, Ying; Niu, Gang; Kiesewetter, Dale O; Chen, Xiaoyuan

    2015-10-21

    Hexafluorobenzene (HFB) reacts with free thiols to produce a unique and selective perfluoroaromatic linkage between two sulfurs. We modified this chemical reaction to produce dimeric (18)F-RGD-tetrafluorobenzene (TFB)-RGD, an integrin αvβ3 receptor ligand. (18)F-HFB was prepared by a fluorine exchange reaction using K(18)F/K2.2.2 at room temperature. The automated radiofluorination was optimized to minimize the amount of HFB precursor and, thus, maximize the specific activity. (18)F-HFB was isolated by distillation and subsequently reacted with thiolated c(RGDfk) peptide under basic and reducing conditions. The resulting (18)F-RGD-TFB-RGD demonstrated integrin receptor specific binding, cellular uptake, and in vivo tumor accumulation.(18)F-HFB can be efficiently incorporated into thiol-containing peptides at room temperature to provide novel imaging agents. PMID:26086295

  19. Arginine-glycine-aspartic acid-polyethylene glycol-polyamidoamine dendrimer conjugate improves liver-cell aggregation and function in 3-D spheroid culture.

    Science.gov (United States)

    Chen, Zhanfei; Lian, Fen; Wang, Xiaoqian; Chen, Yanling; Tang, Nanhong

    2016-01-01

    The polyamidoamine (PAMAM) dendrimer, a type of macromolecule material, has been used in spheroidal cell culture and drug delivery in recent years. However, PAMAM is not involved in the study of hepatic cell-spheroid culture or its biological activity, particularly in detoxification function. Here, we constructed a PAMAM-dendrimer conjugate decorated by an integrin ligand: arginine-glycine-aspartic acid (RGD) peptide. Our studies demonstrate that RGD-polyethylene glycol (PEG)-PAMAM conjugates can promote singly floating hepatic cells to aggregate together in a sphere-like growth with a weak reactive oxygen species. Moreover, RGD-PEG-PAMAM conjugates can activate the AKT-MAPK pathway in hepatic cells to promote cell proliferation and improve basic function and ammonia metabolism. Together, our data support the hepatocyte sphere treated by RGD-PEG-PAMAM conjugates as a potential source of hepatic cells for a biological artificial liver system. PMID:27621619

  20. Dual targeting luminescent gold nanoclusters for tumor imaging and deep tissue therapy.

    Science.gov (United States)

    Chen, Dan; Li, Bowen; Cai, Songhua; Wang, Peng; Peng, Shuwen; Sheng, Yuanzhi; He, Yuanyuan; Gu, Yueqing; Chen, Haiyan

    2016-09-01

    Dual targeting towards both extracellular and intracellular receptors specific to tumor is a significant approach for cancer diagnosis and therapy. In the present study, a novel nano-platform (AuNC-cRGD-Apt) with dual targeting function was initially established by conjugating gold nanocluster (AuNC) with cyclic RGD (cRGD) that is specific to αvβ3integrins over-expressed on the surface of tumor tissues and aptamer AS1411 (Apt) that is of high affinity to nucleolin over-expressed in the cytoplasm and nucleus of tumor cells. Then, AuNC-cRGD-Apt was further functionalized with near infrared (NIR) fluorescence dye (MPA), giving a NIR fluorescent dual-targeting probe AuNC-MPA-cRGD-Apt. AuNC-MPA-cRGD-Apt displays low cytotoxicity and favorable tumor-targeting capability at both in vitro and in vivo level, suggesting its clinical potential for tumor imaging. Additionally, Doxorubicin (DOX), a widely used clinical chemotherapeutic drug that kill cancer cells by intercalating DNA in cellular nucleus, was immobilized onto AuNC-cRGD-Apt forming a pro-drug, AuNC-DOX-cRGD-Apt. The enhanced tumor affinity, deep tumor penetration and improved anti-tumor activity of this pro-drug were demonstrated in different tumor cell lines, tumor spheroid and tumor-bearing mouse models. Results in this study suggest not only the prospect of non-toxic AuNC modified with two targeting ligands for tumor targeted imaging, but also confirm the promising future of dual targeting AuNC as a core for the design of prodrug in the field of cancer therapy. PMID:27236844

  1. Cell adhesion to agrin presented as a nanopatterned substrate is consistent with an interaction with the extracellular matrix and not transmembrane adhesion molecules

    OpenAIRE

    Wolfram Tobias; Spatz Joachim P; Burgess Robert W

    2008-01-01

    Abstract Background Molecular spacing is important for cell adhesion in a number of ways, ranging from the ordered arrangement of matrix polymers extracellularly, to steric hindrance of adhesion/signaling complexes intracellularly. This has been demonstrated using nanopatterned RGD peptides, a canonical extracellular matrix ligand for integrin interactions. Cell adhesion was greatly reduced when the RGD-coated nanoparticles were separated by more than 60 nm, indicating a sharp spacing-depende...

  2. Exploring human disease using the Rat Genome Database

    Science.gov (United States)

    Laulederkind, Stanley J. F.; De Pons, Jeff; Nigam, Rajni; Smith, Jennifer R.; Tutaj, Marek; Petri, Victoria; Hayman, G. Thomas; Wang, Shur-Jen; Ghiasvand, Omid; Thota, Jyothi; Dwinell, Melinda R.

    2016-01-01

    ABSTRACT Rattus norvegicus, the laboratory rat, has been a crucial model for studies of the environmental and genetic factors associated with human diseases for over 150 years. It is the primary model organism for toxicology and pharmacology studies, and has features that make it the model of choice in many complex-disease studies. Since 1999, the Rat Genome Database (RGD; http://rgd.mcw.edu) has been the premier resource for genomic, genetic, phenotype and strain data for the laboratory rat. The primary role of RGD is to curate rat data and validate orthologous relationships with human and mouse genes, and make these data available for incorporation into other major databases such as NCBI, Ensembl and UniProt. RGD also provides official nomenclature for rat genes, quantitative trait loci, strains and genetic markers, as well as unique identifiers. The RGD team adds enormous value to these basic data elements through functional and disease annotations, the analysis and visual presentation of pathways, and the integration of phenotype measurement data for strains used as disease models. Because much of the rat research community focuses on understanding human diseases, RGD provides a number of datasets and software tools that allow users to easily explore and make disease-related connections among these datasets. RGD also provides comprehensive human and mouse data for comparative purposes, illustrating the value of the rat in translational research. This article introduces RGD and its suite of tools and datasets to researchers – within and beyond the rat community – who are particularly interested in leveraging rat-based insights to understand human diseases. PMID:27736745

  3. Biomolecular modification of zirconia surfaces for enhanced biocompatibility

    Energy Technology Data Exchange (ETDEWEB)

    Hsu, Shih-Kuang; Hsu, Hsueh-Chuan [Department of Dental Technology and Materials Science, Central Taiwan University of Science and Technology, Taichung 40601, Taiwan, ROC (China); Ho, Wen-Fu [Department of Chemical and Materials Engineering, National University of Kaohsiung, Taiwan, ROC (China); Yao, Chun-Hsu [Department of Biomedical Imaging and Radiological Science, China Medical University, Taichung 40402, Taiwan, ROC (China); Chang, Pai-Ling [Taoyuan General Hospital, Taoyuan 33004, Taiwan, ROC (China); Wu, Shih-Ching, E-mail: scwu@ctust.edu.tw [Department of Dental Technology and Materials Science, Central Taiwan University of Science and Technology, Taichung 40601, Taiwan, ROC (China)

    2014-12-01

    Yttria-tetragonal zirconia polycrystal (Y-TZP) is a preferred biomaterial due to its good mechanical properties. In order to improve the biocompatibility of zirconia, RGD-peptide derived from extracellular matrix proteins was employed to modify the surface of Y-TZP to promote cell adhesion in this study. The surface of Y-TZP specimens was first modified using a hydrothermal method for different lengths of time. The topographies of modified Y-TZP specimens were analyzed by contact angle, XRD, FTIR, AFM, and FE-SEM. The mechanical properties were evaluated using Vickers hardness and three point bending strength. Then, the RGD-peptide was immobilized on the surface of the Y-TZP by chemical treatment. These RGD-peptide immobilized Y-TZP specimens were characterized by FTIR and AFM, and then were cocultured with MG-63 osteoblast cells for biocompatibility assay. The cell morphology and proliferation were evaluated by SEM, WST-1, and ALP activity assay. The XRD results indicated that the phase transition, from tetragonal phase to monoclinic phase, was increased with a longer incubation time of hydrothermal treatment. However, there were no significant differences in mechanical strengths after RGD-peptide was successfully grafted onto the Y-TZP surface. The SEM images showed that the MG-63 cells appeared polygonal, spindle-shaped, and attached on the RGD-peptide immobilized Y-TZP. The proliferation and cellular activities of MG-63 cells on the RGD-peptide immobilized Y-TZP were better than that on the unmodified Y-TZP. From the above results, the RGD-peptide can be successfully grafted onto the hydrothermal modified Y-TZP surface. The RGD-peptide immobilized Y-TZP can increase cell adhesion, and thus, improve the biocompatibility of Y-TZP. - Highlights: • Covalent bonding between peptide and Y-TZP was proposed. • Stable biomimetic structures produced on the surface of zirconia. • The biocompatibility was improved.

  4. Dynamic Presentation of Immobilized Ligands Regulated through Biomolecular Recognition

    OpenAIRE

    Liu, Bo; Liu, Yang; Riesberg, Jeremiah J.; Shen, Wei

    2010-01-01

    To mimic the dynamic regulation of signaling ligands immobilized on extracellular matrices or on the surfaces of neighboring cells for guidance of cell behavior and fate selection, we have harnessed biomolecular recognition in combination with polymer engineering to create dynamic surfaces on which the accessibility of immobilized ligands to cell surface receptors can be reversibly interconverted under physiological conditions. The cell-adhesive RGD peptide is chosen as a model ligand. RGD is...

  5. c(RGDyK)-decorated Pluronic micelles for enhanced doxorubicin and paclitaxel delivery to brain glioma

    Science.gov (United States)

    Huang, YuKun; Liu, Wenchao; Gao, Feng; Fang, Xiaoling; Chen, Yanzuo

    2016-01-01

    Brain glioma therapy is an important challenge in oncology. Here, doxorubicin (DOX) and paclitaxel (PTX)-loaded cyclic arginine-glycine-aspartic acid peptide (c(RGDyK))-decorated Pluronic micelles (cyclic arginine-glycine-aspartic acid peptide-decorated Pluronic micelles loaded with doxorubicin and paclitaxel [RGD-PF-DP]) were designed as a potential targeted delivery system to enhance blood–brain barrier penetration and improve drug accumulation via integrin-mediated transcytosis/endocytosis and based on integrin overexpression in blood–brain barrier and glioma cells. The physicochemical characterization of RGD-PF-DP revealed a satisfactory size of 28.5±0.12 nm with uniform distribution and core-shell structure. The transport rates across the in vitro blood–brain barrier model, cellular uptake, cytotoxicity, and apoptosis of U87 malignant glioblastoma cells of RGD-PF-DP were significantly greater than those of non-c(RGDyK)-decorated Pluronic micelles. In vivo fluorescence imaging demonstrated the specificity and efficacy of intracranial tumor accumulation of RGD-PF-DP. RGD-PF-DP displayed an extended median survival time of 39 days, with no serious body weight loss during the regimen. No acute toxicity to major organs was observed in mice receiving treatment doses via intravenous administration. In conclusion, RGD-PF-DP could be a promising vehicle for enhanced doxorubicin and paclitaxel delivery in patients with brain glioma. PMID:27143884

  6. Biophysical characterization of the proton-coupled oligopeptide transporter YjdL

    DEFF Research Database (Denmark)

    Jensen, Johanne Mørch; Simonsen, Fie C.; Mastali, Amir;

    2012-01-01

    Proton-coupled oligopeptide transporters (POTs) utilize the electrochemical proton gradient to facilitate uptake of di- or tripeptide molecules. YjdL is one of four POTs found in Escherichia coli. It has shown an extraordinary preference for di- rather than tripeptides, and is therefore significa...... (IC(50) values of 0.6 compared to 0.3mM). Finally, Ala-Lys binding to Glu388Gln was also undetectable which may support a previously suggested role in interaction with the ligand peptide N-terminus....... calorimetrical studies exhibit a K(d) of 14 µM for binding of Ala-Lys to WT-YjdL. Expectedly, no binding could be detected for the tripeptide Ala-Ala-Lys. Surprisingly however, binding could not be detected for Ala-Gln, although earlier studies indicated inhibitory potencies of Ala-Gln to be comparable to Ala-Lys...

  7. In vitro evaluation of N-methyl amide tripeptidomimetics as substrates for the human intestinal di-/tri-peptide transporter hPEPT1

    DEFF Research Database (Denmark)

    Andersen, Rikke; Nielsen, Carsten Uhd; Begtrup, Mikael;

    2006-01-01

    of structural properties of 6859 N-methyl amide tripeptidomimetics. In vitro extracellular degradation of the selected tripeptidomimetics as well as affinity for and transepithelial transport via hPEPT1 were investigated in Caco-2 cells. Decreased apparent degradation was observed for all tripeptidomimetics......Oral absorption of tripeptides is generally mediated by the human intestinal di-/tri-peptide transporter, hPEPT1. However, the bioavailability of tripeptides is often limited due to degradation in the GI-tract by various peptidases. The aim of the present study was to evaluate the general...... application of N-methyl amide bioisosteres as peptide bond replacements in tripeptides in order to decrease degradation by peptidases and yet retain affinity for and transport via hPEPT1. Seven structurally diverse N-methyl amide tripeptidomimetics were selected based on a principal component analysis...

  8. A new high-through-put in vivo screening method for the discovery of tissue targeting drug

    International Nuclear Information System (INIS)

    Objective: To establish a model of totally random high-through-put in vivo screening for the discovery of tissue targeting drug. Methods: The C-terminal amide tripeptide libraries were synthesized on Rink Amide-4-methylbenzhydrylamine (MBHA) resin in the OXX→O1 OX→O1O2O[O represented one of the 20 natural L-amino acids in a defined position, O1-O2 represented a defined L-amino acid (after screening) in a defined position, and X represented a mixture of all the L-amino acids with the exception of Cys] positional scanning formal and iterative protocol. A technetium (V) oxo core [(TcO)3+] was bound to the N4-triligands of tripeptide libraries via four deprotonated amide nitrogen atoms to form a structure of 99Tcm-tripeptide libraries. The radio combinatorial screening in vivo was then carried out after SD rats and A549 tumor bearing mice injected with 99Tcm-tripeptide libraries. Results: Using 99Tcm-tripeptide libraries as an example, a new method of totally random high-through-put in vivo screening was demonstrated successfully in generation, optimization, and design of tissue targeting drugs. The uptake (%ID/g) of 99Tcm labeled drugs in their target tissues was highly structure dependent. Because the non-target tissue binding and the metabolism of 99Tcm-tripeptide libraries were simultaneously monitored successfully, the background activity was limited to the lowest level when a 99Tcm labeled drug finally generated. Conclusion: The random high-through-put in vivo screening method is sensitive and reliable for the discovery of tissue targeting drug. (authors)

  9. Trackable and Targeted Phage as Positron Emission Tomography (PET Agent for Cancer Imaging

    Directory of Open Access Journals (Sweden)

    Zibo Li, Qiaoling Jin, Chiunwei Huang, Siva Dasa, Liaohai Chen, Li-peng Yap, Shuanglong Liu, Hancheng Cai, Ryan Park, Peter S Conti

    2011-01-01

    Full Text Available The recent advancement of nanotechnology has provided unprecedented opportunities for the development of nanoparticle enabled technologies for detecting and treating cancer. Here, we reported the construction of a PET trackable organic nanoplatform based on phage particle for targeted tumor imaging. Method: The integrin αvβ3 targeted phage nanoparticle was constructed by expressing RGD peptides on its surface. The target binding affinity of this engineered phage particle was evaluated in vitro. A bifunctional chelator (BFC 1,4,7,10-tetraazadodecane-N,N',N",N"'-tetraacetic acid (DOTA or 4-((8-amino-3,6,10,13,16,19-hexaazabicyclo [6.6.6] icosane-1-ylamino methyl benzoic acid (AmBaSar was then conjugated to the phage surface for 64Cu2+ chelation. After 64Cu radiolabeling, microPET imaging was performed in U87MG tumor model and the receptor specificity was confirmed by blocking experiments. Results: The phage-RGD demonstrated target specificity based on ELISA experiment. According to the TEM images, the morphology of the phage was unchanged after the modification with BFCs. The labeling yield was 25 ± 4% for 64Cu-DOTA-phage-RGD and 46 ± 5% for 64Cu-AmBaSar-phage-RGD, respectively. At 1 h time point, 64Cu-DOTA-phage-RGD and 64Cu-AmBaSar-phage-RGD have comparable tumor uptake (~ 8%ID/g. However, 64Cu-AmBaSar-phage-RGD showed significantly higher tumor uptake (13.2 ± 1.5 %ID/g, P<0.05 at late time points compared with 64Cu-DOTA-phage-RGD (10 ± 1.2 %ID/g. 64Cu-AmBaSar-phage-RGD also demonstrated significantly lower liver uptake, which could be attributed to the stability difference between these chelators. There is no significant difference between two tracers regarding the uptake in kidney and muscle at all time points tested. In order to confirm the receptor specificity, blocking experiment was performed. In the RGD blocking experiment, the cold RGD peptide was injected 2 min before the administration of 64Cu-AmBaSar-phage-RGD. Tumor uptake was

  10. Integrin-targeted pH-responsive micelles for enhanced efficiency of anticancer treatment in vitro and in vivo

    Science.gov (United States)

    Liu, Jinjian; Deng, Hongzhang; Liu, Qiang; Chu, Liping; Zhang, Yumin; Yang, Cuihong; Zhao, Xuefei; Huang, Pingsheng; Deng, Liandong; Dong, Anjie; Liu, Jianfeng

    2015-02-01

    The key to developing more nanocarriers for the delivery of drugs in clinical applications is to consider the route of the carrier from the administration site to the target tissue and to look for a simple design to complete this whole journey. We synthesized the amphiphilic copolymer cRGDfK-poly(ethylene glycol)-b-poly(2,4,6-trimethoxybenzylidene-1,1,1-tris(hydroxymethyl) ethane methacrylate) (cRGD-PETM) to construct multifunctional micelles. These micelles combined enhanced drug-loading efficiency with tumor-targeting properties, visual detection and controllable intracellular drug release, resulting in an improved chemotherapeutic effect in vivo. Doxorubicin (DOX) was encapsulated within the cRGD-PETM micelles as a model drug (termed as cRGD-PETM/DOX Ms). The size and morphology of the micelles were characterized systematically. As a result of the hydrophobic interaction and the π-π conjugation between the DOX molecules and the PTTMA copolymers, the cRGD-PETM/DOX Ms showed an excellent drug-loading capacity. The results of in vitro drug-release studies indicated that the cumulative release of DOX from cRGD-PETM/DOX Ms at pH 5.0 was twice that at pH 7.4. The results of fluorescent microscopic analysis showed that the cRGD-PETM/DOX Ms could be internalized by 4T1 and HepG2 cells via receptor-mediated endocytosis with rapid intracellular drug release, which resulted in increased cytotoxicity compared with free DOX. Ex vivo imaging studies showed that the cRGD-PETM/DOX Ms improved the accumulation and retention of the drug in tumor tissues. Studies of the in vivo anticancer effects showed that the cRGD-PETM/DOX Ms had a significantly higher therapeutic efficacy with lower side-effects than free DOX and PETM/DOX Ms. These results show that the multifunctional cRGD-PETM/DOX Ms have great potential as vehicles for the delivery of hydrophobic anticancer drugs.

  11. Comprehensive coverage of cardiovascular disease data in the disease portals at the Rat Genome Database.

    Science.gov (United States)

    Wang, Shur-Jen; Laulederkind, Stanley J F; Hayman, G Thomas; Petri, Victoria; Smith, Jennifer R; Tutaj, Marek; Nigam, Rajni; Dwinell, Melinda R; Shimoyama, Mary

    2016-08-01

    Cardiovascular diseases are complex diseases caused by a combination of genetic and environmental factors. To facilitate progress in complex disease research, the Rat Genome Database (RGD) provides the community with a disease portal where genome objects and biological data related to cardiovascular diseases are systematically organized. The purpose of this study is to present biocuration at RGD, including disease, genetic, and pathway data. The RGD curation team uses controlled vocabularies/ontologies to organize data curated from the published literature or imported from disease and pathway databases. These organized annotations are associated with genes, strains, and quantitative trait loci (QTLs), thus linking functional annotations to genome objects. Screen shots from the web pages are used to demonstrate the organization of annotations at RGD. The human cardiovascular disease genes identified by annotations were grouped according to data sources and their annotation profiles were compared by in-house tools and other enrichment tools available to the public. The analysis results show that the imported cardiovascular disease genes from ClinVar and OMIM are functionally different from the RGD manually curated genes in terms of pathway and Gene Ontology annotations. The inclusion of disease genes from other databases enriches the collection of disease genes not only in quantity but also in quality. PMID:27287925

  12. High efficiency diffusion molecular retention tumor targeting.

    Directory of Open Access Journals (Sweden)

    Yanyan Guo

    Full Text Available Here we introduce diffusion molecular retention (DMR tumor targeting, a technique that employs PEG-fluorochrome shielded probes that, after a peritumoral (PT injection, undergo slow vascular uptake and extensive interstitial diffusion, with tumor retention only through integrin molecular recognition. To demonstrate DMR, RGD (integrin binding and RAD (control probes were synthesized bearing DOTA (for (111 In(3+, a NIR fluorochrome, and 5 kDa PEG that endows probes with a protein-like volume of 25 kDa and decreases non-specific interactions. With a GFP-BT-20 breast carcinoma model, tumor targeting by the DMR or i.v. methods was assessed by surface fluorescence, biodistribution of [(111In] RGD and [(111In] RAD probes, and whole animal SPECT. After a PT injection, both probes rapidly diffused through the normal and tumor interstitium, with retention of the RGD probe due to integrin interactions. With PT injection and the [(111In] RGD probe, SPECT indicated a highly tumor specific uptake at 24 h post injection, with 352%ID/g tumor obtained by DMR (vs 4.14%ID/g by i.v.. The high efficiency molecular targeting of DMR employed low probe doses (e.g. 25 ng as RGD peptide, which minimizes toxicity risks and facilitates clinical translation. DMR applications include the delivery of fluorochromes for intraoperative tumor margin delineation, the delivery of radioisotopes (e.g. toxic, short range alpha emitters for radiotherapy, or the delivery of photosensitizers to tumors accessible to light.

  13. Self-assembled peptide-based hydrogels as scaffolds for anchorage-dependent cells.

    Science.gov (United States)

    Zhou, Mi; Smith, Andrew M; Das, Apurba K; Hodson, Nigel W; Collins, Richard F; Ulijn, Rein V; Gough, Julie E

    2009-05-01

    We report here the design of a biomimetic nanofibrous hydrogel as a 3D-scaffold for anchorage-dependent cells. The peptide-based bioactive hydrogel is formed through molecular self-assembly and the building blocks are a mixture of two aromatic short peptide derivatives: Fmoc-FF (Fluorenylmethoxycarbonyl-diphenylalanine) and Fmoc-RGD (arginine-glycine-aspartate) as the simplest self-assembling moieties reported so far for the construction of small-molecule-based bioactive hydrogels. This hydrogel provides a highly hydrated, stiff and nanofibrous hydrogel network that uniquely presents bioactive ligands at the fibre surface; therefore it mimics certain essential features of the extracellular matrix. The RGD sequence as part of the Fmoc-RGD building block plays a dual role of a structural component and a biological ligand. Spectroscopic and imaging analysis using CD, FTIR, fluorescence, TEM and AFM confirmed that FF and RGD peptide sequences self-assemble into beta-sheets interlocked by pi-pi stacking of the Fmoc groups. This generates the cylindrical nanofibres interwoven within the hydrogel with the presence of RGDs in tunable densities on the fibre surfaces. This rapid gelling material was observed to promote adhesion of encapsulated dermal fibroblasts through specific RGD-integrin binding, with subsequent cell spreading and proliferation; therefore it may offer an economical model scaffold to 3D-culture other anchorage-dependent cells for in-vitro tissue regeneration. PMID:19201459

  14. 精氨酸-甘氨酸-天冬氨酸与核糖核酸酶修饰碲化镉量子点探针对黑素瘤A375细胞的靶向研究%Arginine-glycine-aspartic acid-and RNase A-conjugated CdTe quantum dot-based nanoprobes for active targeting of human A375 malignant melanoma cells in vitro

    Institute of Scientific and Technical Information of China (English)

    陈晓罡; 张振; 费烨; 陈向东

    2013-01-01

    Objective To prepare arginine-glycine-aspartic acid (RGD)-and ribonuclease A (RNase A)-conjugated CdTe quantum dot (QD) nanoprobes,and to observe their capability to target human A375 malignant melanoma cells.Methods RNase A-modified CdTe quantum dots (CdTe RQDs) were obtained by using a microwave-based heating method,and then chemically conjugated to the RGD peptide to prepare RGD-CdTe RQD nanoprobes,which were then physically and chemically characterized by transmission electron microscopy,powder crystal diffraction,fluorescence spectrophotometry,and ultraviolet absorption spectrophotometry.A375 cells were cultured in vitro and incubated with various concentrations (20,40,80 nmol/L) of RGD-CdTe RQD nanoprobes for different durations (12,24,36,72 hours).Then,methyl thiazolyl tetrazolium (MTT) assay was conducted to estimate the proliferative activity of A375 cells.To observe the targeting capability of RGD-CdTe RQD nanoprobes,A375 cells were treated with RGD-CdTe RQD nanoprobes at the concentration determined by MTT assay for one hour followed by laser confocal microscopy.Results CdTe RQDs with good dispersion and biocompatibility were obtained by using a microwave-based heating method,and then successfully conjugated to the RGD peptide to form RGD-CdTe RQD nanoprobes.The treatment with RGD-CdTe RQDs of 20 nmol/L for 12 hours exhibited the weakest effect on the proliferative activity of A375 cells,and hence,20 nmol/L was selected for the fluorescence imaging assay.Laser confocal microscopy revealed that RGD-CdTe RQD nanoprobes were able to actively target A375 cells.Conclusion RGD-CdTe RQD nanoprobes with a favorable capability to actively target A375 cells are successfully prepared in this study.%目的 制备精氨酸-甘氨酸-天冬氨酸(RGD)与核糖核酸酶(RNase A)修饰的碲化镉(CdTe)量子点(quantum dot,QDs)的纳米探针,观察其对恶性黑素瘤A375细胞的靶向效果.方法 利用微波加热方法得到核糖核酸酶修

  15. Improving Blast Resistance of a Thermo-Sensitive Genic Male Sterile Rice Line GD-8S by Molecular Marker-Assisted Selection

    Institute of Scientific and Technical Information of China (English)

    LIU Wu-ge; LIU Yi-bai; JIN Su-juan; ZHU Xiao-yuan; WANG Feng; LI Jin-hua; LIU Zhen-rong; LIAO Yi-long; ZHU Man-shan; HUANG Hui-jun

    2008-01-01

    The broad-spectrum blast resistance gene Pi-1, from donor line BL122, was introduced into a thermo-sensitive genic male sterile rice line GD-8S, which possessed good grain quality but high susceptibility to rice blast, by using backcross breeding and molecular marker-assisted selection. Five elite improved male sterile lines, RGD8S-1, RGD8S-2, RGD8S-3, RGD8S-4 and RGD8S-5, were selected based on the results of molecular marker analysis, spikelet sterility, recovery rate of genetic background and agronomic traits. Thirty-three representative blast isolates collected from Guangdong Province,China were used to inoculate the improved lines and the original line GD-8S artificially. The resistance frequencies of the improved lines ranged from 76.47% to 100%, much higher than that of the original line GD-8S (9.09%). On the agronomic characters, there were no significant differences between the improved lines and GD-8S except for flag leaf length and panicle number per plant. The improved lines could be used for breeding hybrid rice with high blast resistance.

  16. Aerosol Stable Peptide-Coated Liposome Nanoparticles: A Proof-of-Concept Study with Opioid Fentanyl in Enhancing Analgesic Effects and Reducing Plasma Drug Exposure

    Science.gov (United States)

    HOEKMAN, JOHN D; SRIVASTAVA, PRAMOD; HO, RODNEY J Y

    2014-01-01

    Previous we reported a novel pressurized olfactory drug (POD) delivery device that deposit aerosolized drug preferentially to upper nasal cavity. This POD device provided sustained CNS levels of soluble morphine analgesic effects. However, analgesic onset of less soluble fentanyl was more rapid but brief, likely due to hydrophobic fentanyl redistribution readily back to blood. To determine whether fentanyl incorporated into an aerosol stable liposome that binds to nasal epithelial cells will enhance CNS drug exposure and analgesic effects and reduce plasma exposure, we constructed RGD liposomes anchored with acylated integrin binding peptides (palmitoyl-GRGDS). The RGD liposomes, which assume gel-phase membrane structure at 25°C were stable under the stress of aerosolization as only 2.2 ± 0.5 % calcein leakage detected. The RGD mediated integrin binding of liposome is also verified to be unaffected by aerosolization. Rats treated with fentanyl in RGD-liposome and POD device exhibited greater analgesic effect, compared to the free drug counterpart (AUCeffect = 1387.l vs. 760.1 %MPE*min); while ~20% reduced plasma drug exposure was noted (AUC0-120 = 208.2 vs 284.8 ng*min/ml). Collectively, fentanyl incorporated in RGD-liposomes are physically and biologically stable under aerosolization, enhanced the overall analgesic effects and reduced plasma drug exposure for the first 2 hours. PMID:24909764

  17. Core-shell nanocarriers with high paclitaxel loading for passive and active targeting

    Science.gov (United States)

    Jin, Zhu; Lv, Yaqi; Cao, Hui; Yao, Jing; Zhou, Jianping; He, Wei; Yin, Lifang

    2016-06-01

    Rapid blood clearance and premature burst release are inherent drawbacks of conventional nanoparticles, resulting in poor tumor selectivity. iRGD peptide is widely recognized as an efficient cell membrane penetration peptide homing to αVβ3 integrins. Herein, core-shell nanocapsules (NCs) and iRGD-modified NCs (iRGD-NCs) with high drug payload for paclitaxel (PTX) were prepared to enhance the antitumor activities of chemotherapy agents with poor water solubility. Improved in vitro and in vivo tumor targeting and penetration were observed with NCs and iRGD-NCs; the latter exhibited better antitumor activity because iRGD enhanced the accumulation and penetration of NCs in tumors. The NCs were cytocompatible, histocompatible, and non-toxic to other healthy tissues. The endocytosis of NCs was mediated by lipid rafts in an energy-dependent manner, leading to better cytotoxicity of PTX against cancer cells. In contrast with commercial product, PTX-loaded NCs (PTX-NCs) increased area under concentration-time curve (AUC) by about 4-fold, prolonged mean resident time (MRT) by more than 8-fold and reduced the elimination rate constant by greater than 68-fold. In conclusion, the present nanocarriers with high drug-loading capacity represent an efficient tumor-targeting drug delivery system with promising potential for cancer therapy.

  18. 整合素受体靶向显像诊断乳腺癌溶骨性转移%Imaging of osteolytic bone metastasis from breast cancer with new integrin αvβ3 receptor targeted radiotracer

    Institute of Scientific and Technical Information of China (English)

    邵国强; 赵有财; 崔璨; 梁凯; 孙晋; 杨瑞; 王峰

    2015-01-01

    Objective To investigate the value of integrin αvβ3 targeted micro positron emission tomography (microPET)/CT imaging with 68 Ca-1,4,7,10-tetraazacyclododecane-N,N',N",N"'-tetraacetic acid-arginine-glycine-aspartate peptide dimer (68 Ga-DOTA-RGD2) as radiotracer for the detection of breast cancer osteolytic bone metastases.Methods 68 Ga-DOTA-RGD2 was prepared via one-step method at 100 ℃ for 15 min.Animal model with parathyroid hormone (PTH)-induced osteolysis in the calvarium was established and served as PTH group (BP).Biodistribution study of 68 Ga-DOTA-RGD2 was carried out in BP.Integrin receptor block study was done with pre-injection of high dose of DOTA-RGD2.68Ga-DOTA-RGD2 and 18F-NaF micmPET/CT imaging were perfromed respectively.Breast cancer osteolyic bone metastases was established via intrcardial injection of breast cancer cells.68 Ga-DOTA-RGD2 microPET/CT imaging were perfromed for the detection of breast cancer osetolytic bone metastases.Animals were sarcrificed and bone lesions were harvested for pathological examination.Results 68 Ga-DOTA-RGD2 was stable in vitro and its radiopurity was as high as (96.4 ± 2.1) % 3 h after its preparation.Its blood elimination was fast while its uptake by the liver and kidneys were relativelylow.It was discharged soon after its intravenous injection.In the BP