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Sample records for recombinant platelet-derived growth

  1. Safety of recombinant human platelet-derived growth factor-BB in Augment® Bone Graft

    Directory of Open Access Journals (Sweden)

    Luis A Solchaga

    2012-12-01

    Full Text Available This article discusses nonclinical and clinical data regarding the safety of recombinant human platelet-derived growth factor-BB as a component of the Augment® Bone Graft (Augment. Augment is a bone graft substitute intended to be used as an alternative to autologous bone graft in the fusion of hindfoot and ankle joints. Nonclinical studies included assessment of the pharmacokinetic profile of intravenously administered recombinant human platelet-derived growth factor-BB in rat and dog, effects of intravenous administration of recombinant human platelet-derived growth factor-BB in a reproductive and development toxicity study in rats, and chronic toxicity and carcinogenicity of Augment in a 12-month implantation model. These studies showed that systemic exposure was brief and clearance was rapid. No signs of toxicity, carcinogenicity, or tumor promotion were observed even with doses far exceeding the maximum clinical dose. Results of clinical trials (605 participants and commercial use of recombinant human platelet-derived growth factor-BB containing products indicate that these products are not associated with increased incidence of adverse events or cancer. The safety data presented provide evidence that recombinant human platelet-derived growth factor-BB is a safe therapeutic when used in combination products as a single administration during surgical procedures for bone repair and fusion. There is no evidence associating use of recombinant human platelet-derived growth factor-BB in Augment with chronic toxicity, carcinogenicity, or tumor promotion.

  2. The Use of Recombinant Human Platelet-Derived Growth Factor for Maxillary Sinus Augmentation.

    Science.gov (United States)

    Kubota, Atsushi; Sarmiento, Hector; Alqahtani, Mohammed Saad; Llobell, Arturo; Fiorellini, Joseph P

    The maxillary sinus augmentation procedure has become a predictable treatment to regenerate bone for implant placement. The purpose of this study was to evaluate the effect of recombinant human platelet-derived growth factor BB (rhPDGF-BB) combined with a deproteinized cancellous bovine bone graft for sinus augmentation. The lateral window approach was used for maxillary sinuses with minimal residual bone. After a healing period of 4 months, dental implants were placed and then restored following a 2-month osseointegration period. The result demonstrated increased bone height and ISQ values and a 100% survival rate. This study indicates that the addition of rhPDGF-BB to deproteinized cancellous bovine bone accelerated the healing period in maxillary sinuses with minimal native bone.

  3. Human histologic evaluation of anorganic bovine bone mineral combined with recombinant human platelet-derived growth factor BB in maxillary sinus augmentation: case series study.

    Science.gov (United States)

    Nevins, Myron; Garber, David; Hanratty, James J; McAllister, Bradley S; Nevins, Marc L; Salama, Maurice; Schupbach, Peter; Wallace, Steven; Bernstein, Simon M; Kim, David M

    2009-12-01

    The objective of this proof-of-principle study was to examine the potential for improved bone regenerative outcomes in maxillary sinus augmentation procedures when recombinant human platelet-derived growth factor BB (0.3 mg/mL) is combined with particulate anorganic bovine bone mineral. The surgical outcomes in all treated sites were uneventful at 6 to 8 months, with sufficient regenerated bone present to allow successful placement of maxillary posterior implants. Large areas of dense, well-formed lamellar bone were seen throughout the intact core specimens in more than half of the grafted sites. Abundant numbers of osteoblasts were noted in concert with significant osteoid in all sites, indicating ongoing osteogenesis. A number of cores demonstrated efficient replacement of the normally slowly resorbing anorganic bovine bone mineral matrix particles with newly formed bone when the matrix was saturated with recombinant human platelet-derived growth factor BB.

  4. Expressions of the low density lipoprotein receptor and 3-hydroxy-3-methylglutaryl coenzyme A reductase genes are stimulated by recombinant platelet-derived growth factor isomers

    International Nuclear Information System (INIS)

    Roth, M.; Emmons, L.R.; Perruchoud, A.; Block, L.H.

    1991-01-01

    The plausible role that platelet-derived growth factor (PDGF) has in the localized pathophysiological changes that occur in the arterial wall during development of atherosclerotic lesions led the authors to investigate the influence of recombinant (r)PDGF isomers -AA, -AB, and -BB on the expression of low density lipoprotein receptor (LDL-R) and 3-hydroxy-3-methylglutaryl coenzyme A (HMG0CoA) reductase [(S)-mevalonate:NAD + oxidoreductase (CoA-acylating), EC 1.1.1.88] genes. In addition, they clarified the role of protein kinase C (PKC) in expression of the two genes in human skin fibroblasts and vascular smooth muscle cells. The various rPDGF isoforms are distinct in their ability to activate transcription of both genes: (i) both rPDGF-AA and -BB stimulate transcription of the LDL-R gene; in contrast, rPDGF-BB but not -AA, activates transcription of the HMG-CoA reductase gene; (ii) all recombinant isoforms of PDGF activate transcription of the c-fos gene; (iii) while rPDGF-dependent transcription of the lDL-R gene occurs independently of PKC, transcription of the HMG-CoA reductase gene appears to involve the action of that enzyme

  5. Topical application of recombinant platelet-derived growth factor increases the rate of healing and the level of proteins that regulate this response.

    Science.gov (United States)

    Gowda, Santosh; Weinstein, David A; Blalock, Timothy D; Gandhi, Kavita; Mast, Bruce A; Chin, Gloria; Schultz, Gregory S

    2015-10-01

    A bipedicle ischaemic rat skin flap model was used to study the effects of daily topical applications of platelet-derived growth factor (PDGF) on the healing of ischaemic wounds. Levels of tumour necrosis factor-alpha (TNFA), interleukin 1-beta (IL1B) and both the latent and active forms of matrix metalloproteinase 2 (MMP2) and 9 (MMP9) were measured. Full-thickness wounds were made on a total of 72 adult male Sprague-Dawley rats. Each group of 18 rats with normal and ischaemic wounds received either vehicle or 0·01% recombinant PDGF-BB. Additional applications were made on the wounds on a daily basis. Wound areas were measured at 0, 1, 3, 5, 7 9 and 13 days after wounding. Ischaemia caused a delay in wound healing as well as an increase in TNFA, IL1B and both the pro and active forms of MMP2 and MMP9. PDGF accelerated the rate of wound healing in both normal and ischaemic wounds and negated the effect of ischaemia. PDGF reduced the TNFA concentration in both normal and ischaemic wounds, and the rate of wound healing closely resembled the pattern of TNFA protein expression. PDGF also reduced both the magnitude and duration of the increases in IL1B and both the pro and active forms of MMP2 and MMP9 induced by ischaemia. © 2013 The Authors. International Wound Journal © 2013 Medicalhelplines.com Inc and John Wiley & Sons Ltd.

  6. Association assessment of platelet derived growth factor B gene ...

    African Journals Online (AJOL)

    Background: Coronary artery disease (CAD) is the most frequent cause of morbidity and mortality in the world and it is known as a multifactorial disorder which is influenced by both genetic and environmental factors. Based on different assays, the platelet derived growth factor B (PDGF-B) gene is shown to be amongst the ...

  7. Purification of human platelet-derived growth factor

    International Nuclear Information System (INIS)

    Raines, E.W.; Ross, R.

    1985-01-01

    The paper describes a method for purification of human platelet-derived growth factor (PDGF) from outdated platelet-rich plasma (PRP) using commonly available laboratory reagents and yielding a mitogen purified 800,000-fold over the starting material. [ 3 H]thymidine incorporation into DNA of cultured cells responsive to PDGF represents the most readily available method to follow its purification and define the biological activity of a purified preparation. Other assays to quantitate PDGF include radioreceptor assay and radioimmunoassay

  8. Vertical ridge augmentation using an equine bone and collagen block infused with recombinant human platelet-derived growth factor-BB: a randomized single-masked histologic study in non-human primates.

    Science.gov (United States)

    Nevins, Myron; Al Hezaimi, Khalid; Schupbach, Peter; Karimbux, Nadeem; Kim, David M

    2012-07-01

    This study tests the effectiveness of hydroxyapatite and collagen bone blocks of equine origin (eHAC), infused with recombinant human platelet-derived growth factor-BB (rhPDGF-BB), to augment localized posterior mandibular defects in non-human primates (Papio hamadryas). Bilateral critical-sized defects simulating severe atrophy were created at the time of the posterior teeth extraction. Test and control blocks (without growth factor) were randomly grafted into the respective sites in each non-human primate. All sites exhibited vertical ridge augmentation, with physiologic hard- and soft-tissue integration of the blocks when clinical and histologic examinations were done at 4 months after the vertical ridge augmentation procedure. There was a clear, although non-significant, tendency to increased regeneration in the test sites. As in the first two preclinical studies in this series using canines, experimental eHAC blocks infused with rhPDGF-BB proved to be a predictable and technically viable method to predictably regenerate bone and soft tissue in critical-sized defects. This investigation supplies additional evidence that eHAC blocks infused with rhPDGF-BB growth factor is a predictable and technically feasible option for vertical augmentation of severely resorbed ridges.

  9. Effect of platelet-derived growth factor-BB on bone formation in calvarial defects: an experimental study in rabbits

    DEFF Research Database (Denmark)

    Vikjaer, D; Blom, S; Hjørting-Hansen, E

    1997-01-01

    The effect of recombinant human platelet-derived growth factor-BB (rhPDGF-BB) on bone healing was examined in calvarial defects in rabbits. Bicortical circular (critical size) defects were prepared in the calvarial bone of 16 rabbits. The defects were closed on the dural side and covered externally...

  10. Platelet-derived growth factor receptors in the human central nervous system : autoradiographic distribution and receptor densities in multiple sclerosis

    NARCIS (Netherlands)

    De Keyser, J; Wilczak, N

    1997-01-01

    Platelet derived growth factor (PDGF) receptors were studied in postmortem adult human brain and cervical spinal cord using autoradiography with human recombinant I-125-PDGF-BB. PDGF-BB binds to the three different dimers of PDGF receptors (alpha alpha, alpha beta and beta beta) PDGF receptors were

  11. Platelet-derived growth factor inhibits platelet activation in heparinized whole blood.

    Science.gov (United States)

    Selheim, F; Holmsen, H; Vassbotn, F S

    1999-08-15

    We previously have demonstrated that human platelets have functionally active platelet-derived growth factor alpha-receptors. Studies with gel-filtered platelets showed that an autocrine inhibition pathway is transduced through this tyrosine kinase receptor during platelet activation. The physiological significance of this inhibitory effect of platelet-derived growth factor on gel-filtered platelets activation is, however, not known. In the present study, we investigated whether platelet-derived growth factor inhibits platelet activation under more physiological conditions in heparinized whole blood, which represents a more physiological condition than gel-filtered platelets. Using flow cytometric assays, we demonstrate here that platelet-derived growth factor inhibits thrombin-, thrombin receptor agonist peptide SFLLRN-, and collagen-induced platelet aggregation and shedding of platelet-derived microparticles from the platelet plasma membrane during platelet aggregation in stirred heparinized whole blood. The inhibitory effect of platelet-derived growth factor was dose dependent. However, under nonaggregating conditions (no stirring), we could not demonstrate any significant effect of platelet-derived growth factor on thrombin- and thrombin receptor agonist peptide-induced platelet surface expression of P-selectin. Our results demonstrate that platelet-derived growth factor appears to be a true antithrombotic agent only under aggregating conditions in heparinized whole blood.

  12. Signal transduction by the platelet-derived growth factor receptor

    International Nuclear Information System (INIS)

    Williams, L.T.; Escobedo, J.A.; Keating, M.T.; Coughlin, S.R.

    1988-01-01

    The mitogenic effects of platelet-derived growth factor (PDGF) are mediated by the PDGF receptor. The mouse PDGF receptor was recently purified on the basis of its ability to become tyrosine phosphorylated in response to the A-B human platelet form of PDGF, and the receptor amino acid sequence was determined from a full-length cDNA clone. Both the human and mouse receptor cDNA sequences have been expressed in Chinese hamster ovary fibroblast (CHO) cells that normally lack PDGF receptors. This paper summarizes recent results using this system to study signal transduction by the PDGF receptor. Some of the findings show that the KI domain of the PDGF receptor plays an important role in the stimulation of DNA synthesis by PDGF. Surprisingly, the kinase insert region is not essential for PDGF stimulation of PtdIns turnover, pH change, increase in cellular calcium, and receptor autophosphorylation. In addition, PDGF stimulates a conformational change in the receptor

  13. Serum platelet-derived growth factor and fibroblast growth factor in patients with benign and malignant ovarian tumors

    DEFF Research Database (Denmark)

    Madsen, Christine Vestergaard; Steffensen, Karina Dahl; Olsen, Dorte Aalund

    2012-01-01

    New biological markers with predictive or prognostic value are highly warranted in the treatment of ovarian cancer. The platelet-derived growth factor (PDGF) system and fibroblast growth factor (FGF) system are important components in tumor growth and angiogenesis....

  14. A bioactive molecule in a complex wound healing process: platelet-derived growth factor.

    Science.gov (United States)

    Kaltalioglu, Kaan; Coskun-Cevher, Sule

    2015-08-01

    Wound healing is considered to be particularly important after surgical procedures, and the most important wounds related to surgical procedures are incisional, excisional, and punch wounds. Research is ongoing to identify methods to heal non-closed wounds or to accelerate wound healing; however, wound healing is a complex process that includes many biological and physiological events, and it is affected by various local and systemic factors, including diabetes mellitus, infection, ischemia, and aging. Different cell types (such as platelets, macrophages, and neutrophils) release growth factors during the healing process, and platelet-derived growth factor is a particularly important mediator in most stages of wound healing. This review explores the relationship between platelet-derived growth factor and wound healing. © 2014 The International Society of Dermatology.

  15. Inhibition of platelet-derived growth factor signaling prevents muscle fiber growth during skeletal muscle hypertrophy.

    Science.gov (United States)

    Sugg, Kristoffer B; Korn, Michael A; Sarver, Dylan C; Markworth, James F; Mendias, Christopher L

    2017-03-01

    The platelet-derived growth factor receptors alpha and beta (PDGFRα and PDGFRβ) mark fibroadipogenic progenitor cells/fibroblasts and pericytes in skeletal muscle, respectively. While the role that these cells play in muscle growth and development has been evaluated, it was not known whether the PDGF receptors activate signaling pathways that control transcriptional and functional changes during skeletal muscle hypertrophy. To evaluate this, we inhibited PDGFR signaling in mice subjected to a synergist ablation muscle growth procedure, and performed analyses 3 and 10 days after induction of hypertrophy. The results from this study indicate that PDGF signaling is required for fiber hypertrophy, extracellular matrix production, and angiogenesis that occur during muscle growth. © 2017 Federation of European Biochemical Societies.

  16. Expression of platelet-derived growth factor and its receptors in proliferative disorders of fibroblastic origin.

    OpenAIRE

    Smits, A.; Funa, K.; Vassbotn, F. S.; Beausang-Linder, M.; af Ekenstam, F.; Heldin, C. H.; Westermark, B.; Nistér, M.

    1992-01-01

    Platelet-derived growth factor (PDGF) is known to stimulate the proliferation of connective tissue-derived cells in vitro. Less is known about its functions in vivo, and the role of PDGF in the development of human tumors has not been clarified. The authors have investigated the occurrence of PDGF and PDGF receptors in a series of proliferative disorders of fibroblastic origin using immunohistochemical and in situ hybridization techniques. High expression of PDGF beta-receptor mRNA and protei...

  17. Reversion of autocrine transformation by a dominant negative platelet-derived growth factor mutant.

    Science.gov (United States)

    Vassbotn, F S; Andersson, M; Westermark, B; Heldin, C H; Ostman, A

    1993-07-01

    A non-receptor-binding mutant of the platelet-derived growth factor (PDGF) A chain, PDGF-0, was generated by exchanging 7 amino acids in the sequence. The mutant chains formed dimers that were similar to wild-type PDGF-AA with regard to stability and rate of processing to the mature 30-kDa secreted forms. Moreover, the mutant chains formed disulfide-bonded heterodimers with the PDGF B chain in NIH 3T3 cells heterodimer underwent the same processing and secretion as PDGF-AB. Transfection of c-sis-expressing 3T3 cells with PDGF-0 significantly inhibited the transformed phenotype of these cells, as determined by the following criteria. (i) Compared with PDGF-0-negative clones, PDGF-0-producing clones showed a reverted morphology. (ii) Clones producing PDGF-0 grew more slowly than PDGF-0-negative clones, with a fivefold difference in cell number after 14 days in culture. (iii) The expression of PDGF-0 completely inhibited the ability of the c-sis-expressing 3T3 cells to form colonies in soft agar; this inhibition was overcome by the addition of recombinant PDGF-BB to the culture medium, showing that the lack of colony formation of these cells was not due to a general unresponsiveness to PDGF. The specific expression of a PDGF-0/PDGF wild-type heterodimer in COS cells revealed that the affinity of the mutant heterodimer for the PDGF alpha receptor was decreased by approximately 50-fold compared with that of PDGF-AA. Thus, we show that a non-receptor-binding PDGF A-chain mutant neutralizes in a trans-dominant manner the autocrine transforming potential of the c-sis/PDGF B chain by forming low-affinity heterodimers with wild-type PDGF chains. This method of specifically antagonizing the effect of PDGF may be useful in investigations of the role of PDGF in normal and pathological conditions.

  18. Association of coatomer proteins with the beta-receptor for platelet-derived growth factor

    DEFF Research Database (Denmark)

    Hansen, Klaus; Rönnstrand, L; Rorsman, C

    1997-01-01

    The nonreceptor tyrosine kinase Src binds to and is activated by the beta-receptor for platelet-derived growth factor (PDGF). The interaction leads to Src phosphorylation of Tyr934 in the kinase domain of the receptor. In the course of the functional characterization of this phosphorylation, we...... of intracellular vesicle transport. In order to explore the functional significance of the interaction between alpha- and beta'-COP and the PDGF receptor, a receptor mutant was made in which the conserved histidine residue 928 was mutated to an alanine residue. The mutant receptor, which was unable to bind alpha...

  19. Abnormal platelet-derived growth factor signaling accounting for lung hypoplasia in experimental congenital diaphragmatic hernia.

    Science.gov (United States)

    Dingemann, Jens; Doi, Takashi; Ruttenstock, Elke; Puri, Prem

    2010-10-01

    The pathogenesis of pulmonary hypoplasia in congenital diaphragmatic hernia (CDH) is not fully understood. Platelet-derived growth factor A (PDGFA) and platelet-derived growth factor receptor α (PDGFRα) play a crucial role in lung development. It has been reported that PDGF induces H(2)O(2)-production and that oxidative stress may be an important mechanism for the impaired lung development in the nitrofen rat model. We hypothesized that pulmonary expression of PDGFA and PDGFRα is altered in the nitrofen induced CDH model. Pregnant rats received 100 mg nitrofen or vehicle on gestational day 9 (D9) and were sacrificed on D15, D18 or D21. RNA was extracted from fetal left lungs and mRNA levels of PDGFA and PDGFRα were determined using real-time polymerase chain reaction. Immunohistochemistry for protein expression of PDGFA and PDGFRα was performed. Pulmonary H(2)O(2) was measured colorimetrically. mRNA levels of PDGFRα at D15 (4.50 ± 0.87) and PDGFA at D18 (2.90 ± 1.38) were increased in the nitrofen group (P stress during lung development. Copyright © 2010 Elsevier Inc. All rights reserved.

  20. Are genetic variants in the platelet-derived growth factor [beta] gene associated with chronic pancreatitis?

    Science.gov (United States)

    Muddana, Venkata; Park, James; Lamb, Janette; Yadav, Dhiraj; Papachristou, Georgios I; Hawes, Robert H; Brand, Randall; Slivka, Adam; Whitcomb, David C

    2010-11-01

    Platelet-derived growth factor [beta] (PDGF-[beta]) is a major signal in proliferation and matrix synthesis through activated pancreatic stellate cells, leading to fibrosis of the pancreas. Recurrent acute pancreatitis (RAP) seems to predispose to chronic pancreatitis (CP) in some patients but not others. We tested the hypothesis that 2 known PDGF-[beta] polymorphisms are associated with progression from RAP to CP. We also tested the hypothesis that PDGF-[beta] polymorphisms in combination with environmental risk factors such as alcohol and smoking are associated with CP. Three hundred eighty-two patients with CP (n = 176) and RAP (n = 206) and 251 controls were evaluated. Platelet-derived growth factor [beta] polymorphisms +286 A/G (rs#1800818) seen in 5'-UTR and +1135 A/C (rs#1800817) in first intron were genotyped using single-nucleotide polymorphism polymerase chain reaction approach and confirmed by DNA sequencing. The genotypic frequencies for PDGF-[beta] polymorphisms in positions +286 and +1135 were found to be similar in controls and patients with RAP and CP. There was no difference in genotypic frequencies among RAP, CP, and controls in subjects in the alcohol and smoking subgroups. Known variations in the PDGF-[beta] gene do not have a significant effect on promoting or preventing fibrogenesis in pancreatitis. Further evaluation of this important pathway is warranted.

  1. Platelet-derived growth factor-C and -D in the cardiovascular system and diseases.

    Science.gov (United States)

    Lee, Chunsik; Li, Xuri

    2018-08-01

    The cardiovascular system is among the first organs formed during development and is pivotal for the formation and function of the rest of the organs and tissues. Therefore, the function and homeostasis of the cardiovascular system are finely regulated by many important molecules. Extensive studies have shown that platelet-derived growth factors (PDGFs) and their receptors are critical regulators of the cardiovascular system. Even though PDGF-C and PDGF-D are relatively new members of the PDGF family, their critical roles in the cardiovascular system as angiogenic and survival factors have been amply demonstrated. Understanding the functions of PDGF-C and PDGF-D and the signaling pathways involved may provide novel insights into both basic biomedical research and new therapeutic possibilities for the treatment of cardiovascular diseases. Copyright © 2017 Elsevier Ltd. All rights reserved.

  2. Stage specific requirement of platelet-derived growth factor receptor-α in embryonic development.

    Directory of Open Access Journals (Sweden)

    Chen Qian

    Full Text Available Platelet-derived growth factor receptor alpha (PDGFRα is a cell-surface receptor tyrosine kinase for platelet-derived growth factors. Correct timing and level of Pdgfra expression is crucial for embryo development, and deletion of Pdgfra caused developmental defects of multiple endoderm and mesoderm derived structures, resulting in a complex phenotypes including orofacial cleft, spina bifida, rib deformities, and omphalocele in mice. However, it is not clear if deletion of Pdgfra at different embryonic stages differentially affects these structures.To address the temporal requirement of Pdgfra in embryonic development.We have deleted the Pdgfra in Pdgfra-expressing tissues at different embryonic stages in mice, examined and quantified the developmental anomalies.Current study showed that (i conditional deletion of Pdgfra at different embryonic days (between E7.5 and E10.5 resulted in orofacial cleft, spina bifida, rib cage deformities, and omphalocele, and (ii the day of Pdgfra deletion influenced the combinations, incidence and severities of these anomalies. Deletion of Pdgfra caused apoptosis of Pdgfra-expressing tissues, and developmental defects of their derivatives.Orofacial cleft, spina bifida and omphalocele are among the commonest skeletal and abdominal wall defects of newborns, but their genetic etiologies are largely unknown. The remarkable resemblance of our conditional Pdgfra knockout embryos to theses human congenital anomalies, suggesting that dysregulated PDGFRA expression could cause these anomalies in human. Future work should aim at defining (a the regulatory elements for the expression of the human PDGFRA during embryonic development, and (b if mutations / sequence variations of these regulatory elements cause these anomalies.

  3. Platelets promote osteosarcoma cell growth through activation of the platelet-derived growth factor receptor-Akt signaling axis

    OpenAIRE

    Takagi, Satoshi; Takemoto, Ai; Takami, Miho; Oh-hara, Tomoko; Fujita, Naoya

    2014-01-01

    The interactions of tumor cells with platelets contribute to the progression of tumor malignancy, and the expression levels of platelet aggregation-inducing factors positively correlate with the metastatic potential of osteosarcoma cells. However, it is unclear how tumor-platelet interaction contributes to the proliferation of osteosarcomas. We report here that osteosarcoma-platelet interactions induce the release of platelet-derived growth factor (PDGF) from platelets, which promotes the pro...

  4. Molecular cloning and functional characterization of the human platelet-derived growth factor alpha receptor gene promoter

    NARCIS (Netherlands)

    Afink, G. B.; Nistér, M.; Stassen, B. H.; Joosten, P. H.; Rademakers, P. J.; Bongcam-Rudloff, E.; van Zoelen, E. J.; Mosselman, S.

    1995-01-01

    Expression of the platelet-derived growth factor alpha receptor (PDGF alpha R) is strictly regulated during mammalian development and tumorigenesis. The molecular mechanisms involved in the specific regulation of PDGF alpha R expression are unknown, but transcriptional regulation of the PDGF alpha R

  5. Notch signaling regulates platelet-derived growth factor receptor-β expression in vascular smooth muscle cells

    NARCIS (Netherlands)

    Jin, S.; Hansson, E.M.; Tikka, S.; Lanner, F.; Sahlgren, C.; Farnebo, F.; Baumann, M.; Kalimo, H.; Lendahl, U.

    2008-01-01

    Notch signaling is critically important for proper architecture of the vascular system, and mutations in NOTCH3 are associated with CADASIL, a stroke and dementia syndrome with vascular smooth muscle cell (VSMC) dysfunction. In this report, we link Notch signaling to platelet-derived growth factor

  6. Expression of platelet-derived growth factor and its receptors in proliferative disorders of fibroblastic origin.

    Science.gov (United States)

    Smits, A; Funa, K; Vassbotn, F S; Beausang-Linder, M; af Ekenstam, F; Heldin, C H; Westermark, B; Nistér, M

    1992-03-01

    Platelet-derived growth factor (PDGF) is known to stimulate the proliferation of connective tissue-derived cells in vitro. Less is known about its functions in vivo, and the role of PDGF in the development of human tumors has not been clarified. The authors have investigated the occurrence of PDGF and PDGF receptors in a series of proliferative disorders of fibroblastic origin using immunohistochemical and in situ hybridization techniques. High expression of PDGF beta-receptor mRNA and protein was found in the malignant tumors, and also in some benign lesions, such as dermatofibroma. In all these cases, benign as well as malignant, the PDGF B-chain mRNA, and less clearly, the PDGF A-chain mRNA, were coexpressed with the beta-receptor. In contrast, high expression of PDGF alpha-receptor mRNA was only found in fully malignant lesions, i.e., malignant fibrous histiocytoma. These data indicate that an autocrine growth stimulation via the PDGF beta-receptor could occur in an early phase of tumorigenesis, and may be a necessary but insufficient event for the progression into fully malignant human connective tissue lesions.

  7. Inhibition of the mitogenic response to platelet-derived growth factor by terbinafine

    International Nuclear Information System (INIS)

    St Denny, I.H.; Glinka, K.G.; Nemecek, G.M.; Stuetz, A.

    1987-01-01

    Terbinafine (T;(E)-N-(6,6-dimethyl-2-hepten-4-ynyl)-N-methyl-1-naphthalenemethanamine), an antimycotic which inhibits fungal squalene epoxidase activity, was examined for its effects on platelet-derived growth factor (PDGF)-stimulated mitogenesis. The inclusion of 1.5-5μM T in fibroblast incubation media was associated with increased [ 3 H]thymidine incorporation into DNA in the presence and absence of PDGF. However, T at concentrations above 6μM reduced DNA synthesis in control and PDGF-exposed cultures to nearly undetectable levels. Under a phase-contrast microscope, fibroblasts appeared morphologically normal at T concentrations as high as 25 μM. Neither the uptake of [ 3 H]thymidine nor the specific binding of 125 I-PDGF to fibroblast receptors was significantly affected by 10 μM T. Furthermore, concentrations of T which antagonized the mitogenic response to PDGF also interfered with fibroblast growth factor-induced mitogenesis. Together, these data suggest that T has the ability to inhibit the in vitro action of PDGF via a post-receptor mechanism

  8. Inhibition of the mitogenic response to platelet-derived growth factor by terbinafine

    Energy Technology Data Exchange (ETDEWEB)

    St. Denny, I.H.; Glinka, K.G.; Nemecek, G.M. (Sandoz Research Institute, East Hanover, NJ (USA)); Stuetz, A. (Sandoz Forschungsinstitut, Vienna (Austria))

    1987-05-01

    Terbinafine (T;(E)-N-(6,6-dimethyl-2-hepten-4-ynyl)-N-methyl-1-naphthalenemethanamine), an antimycotic which inhibits fungal squalene epoxidase activity, was examined for its effects on platelet-derived growth factor (PDGF)-stimulated mitogenesis. The inclusion of 1.5-5{mu}M T in fibroblast incubation media was associated with increased ({sup 3}H)thymidine incorporation into DNA in the presence and absence of PDGF. However, T at concentrations above 6{mu}M reduced DNA synthesis in control and PDGF-exposed cultures to nearly undetectable levels. Under a phase-contrast microscope, fibroblasts appeared morphologically normal at T concentrations as high as 25 {mu}M. Neither the uptake of ({sup 3}H)thymidine nor the specific binding of {sup 125}I-PDGF to fibroblast receptors was significantly affected by 10 {mu}M T. Furthermore, concentrations of T which antagonized the mitogenic response to PDGF also interfered with fibroblast growth factor-induced mitogenesis. Together, these data suggest that T has the ability to inhibit the in vitro action of PDGF via a post-receptor mechanism.

  9. Biologic effects of platelet-derived growth factor receptor α blockade in uterine cancer.

    Science.gov (United States)

    Roh, Ju-Won; Huang, Jie; Hu, Wei; Yang, XiaoYun; Jennings, Nicholas B; Sehgal, Vasudha; Sohn, Bo Hwa; Han, Hee Dong; Lee, Sun Joo; Thanapprapasr, Duangmani; Bottsford-Miller, Justin; Zand, Behrouz; Dalton, Heather J; Previs, Rebecca A; Davis, Ashley N; Matsuo, Koji; Lee, Ju-Seog; Ram, Prahlad; Coleman, Robert L; Sood, Anil K

    2014-05-15

    Platelet-derived growth factor receptor α (PDGFRα) expression is frequently observed in many kinds of cancer and is a candidate for therapeutic targeting. This preclinical study evaluated the biologic significance of PDGFRα and PDGFRα blockade (using a fully humanized monoclonal antibody, 3G3) in uterine cancer. Expression of PDGFRα was examined in uterine cancer clinical samples and cell lines, and biologic effects of PDGFRα inhibition were evaluated using in vitro (cell viability, apoptosis, and invasion) and in vivo (orthotopic) models of uterine cancer. PDGFRα was highly expressed and activated in uterine cancer samples and cell lines. Treatment with 3G3 resulted in substantial inhibition of PDGFRα phosphorylation and of downstream signaling molecules AKT and mitogen-activated protein kinase (MAPK). Cell viability and invasive potential of uterine cancer cells were also inhibited by 3G3 treatment. In orthotopic mouse models of uterine cancer, 3G3 monotherapy had significant antitumor effects in the PDGFRα-positive models (Hec-1A, Ishikawa, Spec-2) but not in the PDGFRα-negative model (OVCA432). Greater therapeutic effects were observed for 3G3 in combination with chemotherapy than for either drug alone in the PDGFRα-positive models. The antitumor effects of therapy were related to increased apoptosis and decreased proliferation and angiogenesis. These findings identify PDGFRα as an attractive target for therapeutic development in uterine cancer. ©2014 American Association for Cancer Research.

  10. Synthesis and secretion of platelet-derived growth factor by human breast cancer cell lines

    International Nuclear Information System (INIS)

    Bronzert, D.A.; Pantazis, P.; Antoniades, H.N.; Kasid, A.; Davidson, N.; Dickson, R.B.; Lippman, M.E.

    1987-01-01

    The authors report that human breast cancer cells secrete a growth factor that is biologically and immunologically similar to platelet-derived growth factor (PDGF). Serum-free medium conditioned by estrogen-independent MDA-MB-231 or estrogen-dependent MCF-7 cells contains a mitogenic or competence activity that is capable of inducing incorporation of [ 3 H] thymidine into quiescent Swiss 3T3 cells in the presence of platelet-poor plasma. Like authentic PDGF, the PDGF-like activity produced by breast cancer cells is stable after acid and heat treatment (95 0 C) and inhibited by reducing agents. The mitogenic activity comigrates with a material of ≅30 kDa on NaDodSO 4 /polyacrylamide gels. Immunoprecipitation with PDGF antiserum of proteins from metabolically labeled cell lysates and conditioned medium followed by analysis on nonreducing NaDodSO 4 /polyacrylamide gels identified proteins of 30 and 34 kDa. Upon reduction, the 30- and 34-kDa bands were converted to 15- and 16-kDa bands suggesting that the immunoprecipitated proteins were made up of two disulfide-linked polypeptides similar to PDGF. Hybridization studies with cDNA probes for the A chain PDGF and the B chain of PDGF/SIS identified transcripts for both PDGF chains in the MCF-7 and MDA-MB-231 cells. The data summarized above provide conclusive evidence for the synthesis and hormonally regulated secretion of a PDGF-like mitogen by breast carcinoma cells. Production of a PDGF-like growth factor by breast cancer cell lines may be important in mediating paracrine stimulation of tumor growth

  11. Platelet-derived growth factor induces phosphorylation of a 64-kDa nuclear protein

    International Nuclear Information System (INIS)

    Shawver, L.K.; Pierce, G.F.; Kawahara, R.S.; Deuel, T.F.

    1989-01-01

    The platelet-derived growth factor (PDGF) stimulated the phosphorylation of a nuclear protein of 64 kDa (pp64) in nuclei of nontransformed normal rat kidney (NRK) cells. Low levels of phosphorylation of pp64 were observed in nuclei of serum-starved NRK cells. Fetal calf serum (FCS), PDGF, and homodimeric v-sis and PDGF A-chain protein enhanced the incorporation of 32P into pp64 over 4-fold within 30 min and over 8-fold within 2 h of exposure of NRK cells to the growth factors. In contrast, constitutive phosphorylation of 32P-labeled pp64 in nuclei of NRK cells transformed by the simian sarcoma virus (SSV) was high and only minimally stimulated by PDGF and FCS. 32P-Labeled pp64 was isolated from nuclei of PDGF-stimulated nontransformed NRK cells; the 32P of pp64 was labile in 1 M KOH, and pp64 was not significantly recognized by anti-phosphotyrosine antisera, suggesting that the PDGF-induced phosphorylation of pp64 occurred on serine or on threonine residues. However, pp64 from SSV-transformed NRK cell nuclei was significantly stable to base hydrolysis and was immunoprecipitated with anti-phosphotyrosine antisera, suggesting that pp64 from SSV-transformed cell nuclei is phosphorylated also on tyrosine. FCS, PDGF, and PDGF A- and B-chain homodimers thus stimulate the rapid time-dependent phosphorylation of a 64-kDa nuclear protein shortly after stimulation of responsive cells. The growth factor-stimulated phosphorylation of pp64 and the constitutive high levels of pp64 phosphorylation in cells transformed by SSV suggest important roles for pp64 and perhaps regulated nuclear protein kinases and phosphatases in cell division and proliferation

  12. Platelets promote osteosarcoma cell growth through activation of the platelet-derived growth factor receptor-Akt signaling axis.

    Science.gov (United States)

    Takagi, Satoshi; Takemoto, Ai; Takami, Miho; Oh-Hara, Tomoko; Fujita, Naoya

    2014-08-01

    The interactions of tumor cells with platelets contribute to the progression of tumor malignancy, and the expression levels of platelet aggregation-inducing factors positively correlate with the metastatic potential of osteosarcoma cells. However, it is unclear how tumor-platelet interaction contributes to the proliferation of osteosarcomas. We report here that osteosarcoma-platelet interactions induce the release of platelet-derived growth factor (PDGF) from platelets, which promotes the proliferation of osteosarcomas. Co-culture of platelets with MG63 or HOS osteosarcoma cells, which could induce platelet aggregation, enhanced the proliferation of each cell line in vitro. Analysis of phospho-antibody arrays revealed that co-culture of MG63 cells with platelets induced the phosphorylation of platelet derived growth factor receptor (PDGFR) and Akt. The addition of supernatants of osteosarcoma-platelet reactants also increased the growth of MG63 and HOS cells as well as the level of phosphorylated-PDGFR and -Akt. Sunitinib or LY294002, but not erlotinib, significantly inhibited the platelet-induced proliferation of osteosarcoma cells, indicating that PDGF released from platelets plays an important role in the proliferation of osteosarcomas by activating the PDGFR and then Akt. Our results suggest that inhibitors that specifically target osteosarcoma-platelet interactions may eradicate osteosarcomas. © 2014 The Authors. Cancer Science published by Wiley Publishing Asia Pty Ltd on behalf of Japanese Cancer Association.

  13. Platelet-Derived Growth Factor BB Influences Muscle Regeneration in Duchenne Muscle Dystrophy.

    Science.gov (United States)

    Piñol-Jurado, Patricia; Gallardo, Eduard; de Luna, Noemi; Suárez-Calvet, Xavier; Sánchez-Riera, Carles; Fernández-Simón, Esther; Gomis, Clara; Illa, Isabel; Díaz-Manera, Jordi

    2017-08-01

    Duchenne muscular dystrophy (DMD) is characterized by a progressive loss of muscle fibers, and their substitution by fibrotic and adipose tissue. Many factors contribute to this process, but the molecular pathways related to regeneration and degeneration of muscle are not completely known. Platelet-derived growth factor (PDGF)-BB belongs to a family of growth factors that regulate proliferation, migration, and differentiation of mesenchymal cells. The role of PDGF-BB in muscle regeneration in humans has not been studied. We analyzed the expression of PDGF-BB in muscle biopsy samples from controls and patients with DMD. We performed in vitro experiments to understand the effects of PDGF-BB on myoblasts involved in the pathophysiology of muscular dystrophies and confirmed our results in vivo by treating the mdx murine model of DMD with repeated i.m. injections of PDGF-BB. We observed that regenerating and necrotic muscle fibers in muscle biopsy samples from DMD patients expressed PDGF-BB. In vitro, PDGF-BB attracted myoblasts and activated their proliferation. Analysis of muscles from the animals treated with PDGF-BB showed an increased population of satellite cells and an increase in the number of regenerative fibers, with a reduction in inflammatory infiltrates, compared with those in vehicle-treated mice. Based on our results, PDGF-BB may play a protective role in muscular dystrophies by enhancing muscle regeneration through activation of satellite cell proliferation and migration. Copyright © 2017 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.

  14. Platelet-derived growth factor predicts prolonged relapse-free period in multiple sclerosis.

    Science.gov (United States)

    Stampanoni Bassi, Mario; Iezzi, Ennio; Marfia, Girolama A; Simonelli, Ilaria; Musella, Alessandra; Mandolesi, Georgia; Fresegna, Diego; Pasqualetti, Patrizio; Furlan, Roberto; Finardi, Annamaria; Mataluni, Giorgia; Landi, Doriana; Gilio, Luana; Centonze, Diego; Buttari, Fabio

    2018-04-14

    In the early phases of relapsing-remitting multiple sclerosis (RR-MS), a clear correlation between brain lesion load and clinical disability is often lacking, originating the so-called clinico-radiological paradox. Different factors may contribute to such discrepancy. In particular, synaptic plasticity may reduce the clinical expression of brain damage producing enduring enhancement of synaptic strength largely dependent on neurotrophin-induced protein synthesis. Cytokines released by the immune cells during acute inflammation can alter synaptic transmission and plasticity possibly influencing the clinical course of MS. In addition, immune cells may promote brain repair during the post-acute phases, by secreting different growth factors involved in neuronal and oligodendroglial cell survival. Platelet-derived growth factor (PDGF) is a neurotrophic factor that could be particularly involved in clinical recovery. Indeed, PDGF promotes long-term potentiation of synaptic activity in vitro and in MS and could therefore represent a key factor improving the clinical compensation of new brain lesions. The aim of the present study is to explore whether cerebrospinal fluid (CSF) PDGF concentrations at the time of diagnosis may influence the clinical course of RR-MS. At the time of diagnosis, we measured in 100 consecutive early MS patients the CSF concentrations of PDGF, of the main pro- and anti-inflammatory cytokines, and of reliable markers of neuronal damage. Clinical and radiological parameters of disease activity were prospectively collected during follow-up. CSF PDGF levels were positively correlated with prolonged relapse-free survival. Radiological markers of disease activity, biochemical markers of neuronal damage, and clinical parameters of disease progression were instead not influenced by PDGF concentrations. Higher CSF PDGF levels were associated with an anti-inflammatory milieu within the central nervous system. Our results suggest that PDGF could promote a

  15. Observations on human smooth muscle cell cultures from hyperplastic lesions of prosthetic bypass grafts: Production of a platelet-derived growth factor-like mitogen and expression of a gene for a platelet-derived growth factor receptor--a preliminary study

    International Nuclear Information System (INIS)

    Birinyi, L.K.; Warner, S.J.; Salomon, R.N.; Callow, A.D.; Libby, P.

    1989-01-01

    Prosthetic bypass grafts placed to the distal lower extremity often fail because of an occlusive tissue response in the perianastomotic region. The origin of the cells that comprise this occlusive lesion and the causes of the cellular proliferation are not known. To increase our understanding of this process we cultured cells from hyperplastic lesions obtained from patients at the time of reexploration for lower extremity graft failure, and we studied their identity and growth factor production in tissue culture. These cultures contain cells that express muscle-specific actin isoforms, shown by immunohistochemical staining, consistent with vascular smooth muscle origin. These cultures also released material that stimulated smooth muscle cell growth. A portion of this activity was similar to platelet-derived growth factor, since preincubation with antibody-to-human platelet-derived growth factor partially blocked the mitogenic effect of medium conditioned by human anastomotic hyperplastic cells. These conditioned media also contained material that competed with platelet-derived growth factor for its receptor, as measured in a radioreceptor assay. Northern blot analysis showed that these cells contain messenger RNA that encodes the A chain but not the B chain of platelet-derived growth factor. In addition, these cells contain messenger RNA that encodes a platelet-derived growth factor receptor. We conclude that cultured smooth muscle cells from human anastomotic hyperplastic lesions express genes for platelet-derived growth factor A chain and a platelet-derived growth factor receptor and secrete biologically active molecules similar to platelet-derived growth factor

  16. Expression of platelet-derived growth factor B is upregulated in patients with thoracic aortic dissection.

    Science.gov (United States)

    Meng, Weixin; Liu, Shangdian; Li, Dandan; Liu, Zonghong; Yang, Hui; Sun, Bo; Liu, Hongyu

    2018-04-20

    Thoracic aortic dissection (TAD) is a serious condition requiring urgent treatment to avoid catastrophic consequences. The inflammatory response is involved in the occurrence and development of TAD, possibly potentiated by platelet-derived growth factors (PDGFs). This study aimed to determine whether expression of PDGF-B (a subunit of PDGF-BB) was increased in TAD patients and to explore the factors responsible for its upregulation and subsequent effects on TAD. Full-thickness ascending aorta wall specimens from TAD patients (n = 15) and control patients (n = 10) were examined for expression of PDGF-B and its receptor (PDGFRB) and in terms of morphology, inflammation, and fibrosis. Blood samples from TAD and control patients were collected to detect plasma levels of PDGF-BB and soluble elastins. Expression levels of PDGF-B, PDGFRB, and collagen I were significantly enhanced in ascending aorta wall specimens from TAD patients compared with controls. Furthermore, soluble elastic fragments and PDGF-BB were significantly increased in plasma from TAD patients compared with controls, and numerous irregular elastic fibers and macrophages were seen in the ascending aorta wall in TAD patients. An increase in elastic fragments in the aorta wall might be responsible for inducing the activation and migration of macrophages to injured sites, leading to elevated expression of PDGF-B, which in turn induces deposition of collagen, disrupts extracellular matrix homeostasis, and increases the stiffness of the aorta wall, resulting in compromised aorta compliance. Copyright © 2018 Society for Vascular Surgery. Published by Elsevier Inc. All rights reserved.

  17. Platelet-derived growth factor regulates K-Cl cotransport in vascular smooth muscle cells.

    Science.gov (United States)

    Zhang, Jing; Lauf, Peter K; Adragna, Norma C

    2003-03-01

    Platelet-derived growth factor (PDGF), a potent serum mitogen for vascular smooth muscle cells (VSMCs), plays an important role in membrane transport regulation and in atherosclerosis. K-Cl cotransport (K-Cl COT/KCC), the coupled-movement of K and Cl, is involved in ion homeostasis. VSMCs possess K-Cl COT activity and the KCC1 and KCC3 isoforms. Here, we report on the effect of PDGF on K-Cl COT activity and mRNA expression in primary cultures of rat VSMCs. K-Cl COT was determined as the Cl-dependent Rb influx and mRNA expression by semiquantitative RT-PCR. Twenty four-hour serum deprivation inhibited basal K-Cl COT activity. Addition of PDGF increased total protein content and K-Cl COT activity in a time-dependent manner. PDGF activated K-Cl COT in a dose-dependent manner, both acutely (10 min) and chronically (12 h). AG-1296, a selective inhibitor of the PDGF receptor tyrosine kinase, abolished these effects. Actinomycin D and cycloheximide had no effect on the acute PDGF activation of K-Cl COT, suggesting posttranslational regulation by the drug. Furthermore, PDGF increased KCC1 and decreased KCC3 mRNA expression in a time-dependent manner. These results indicate that chronic activation of K-Cl COT activity by PDGF may involve regulation of the two KCC mRNA isoforms, with KCC1 playing a dominant role in the mechanism of PDGF-mediated activation.

  18. Platelet-derived growth factor (PDGF)-C inhibits neuroretinal apoptosis in a murine model of focal retinal degeneration.

    Science.gov (United States)

    Wang, Yujuan; Abu-Asab, Mones S; Yu, Cheng-Rong; Tang, Zhongshu; Shen, Defen; Tuo, Jingsheng; Li, Xuri; Chan, Chi-Chao

    2014-06-01

    Platelet-derived growth factor (PDGF)-C is a member of the PDGF family and is critical for neuronal survival in the central nervous system. We studied the possible survival and antiapoptotic effects of PDGF-C on focal retinal lesions in Ccl2(-/-)/Cx3cr1(-/-) on C57BL/6N [Crb1(rd8)] (DKO rd8) background mice, a model for progressive and focal retinal degeneration. We found no difference in transcript and protein expression of PDGF-C in the retina between DKO rd8 mice and wild type (WT, C57BL/6N). Recombinant PDGF-CC protein (500 ng/eye) was injected intravitreally into the right eye of DKO rd8 mice with phosphate-buffered saline as controls into the left eye. The retinal effects of PDGF-C were assessed by fundoscopy, ocular histopathology, A2E levels, apoptotic molecule analysis, and direct flat mount retinal vascular labeling. We found that the PDGF-CC-treated eyes showed slower progression or attenuation of the focal retinal lesions, lesser photoreceptor and retinal pigment epithelial degeneration resulting in better-preserved photoreceptor structure. Lower expression of apoptotic molecules was detected in the PDGF-CC-treated eyes than in controls. In addition, no retinal neovascularization was observed after PDGF-CC treatment. Our results demonstrate that PDGF-C potently ameliorates photoreceptor degeneration via the suppression of apoptotic pathways without inducing retinal angiogenesis. The protective effects of PDGF-C suggest a novel alternative approach for potential age-related retinal degeneration treatment.

  19. Immunohistochemical examination of effects of kefir, koumiss and commercial probiotic capsules on platelet derived growth factor-c and platelet derived growth factor receptor-alpha expression in mouse liver and kidney.

    Science.gov (United States)

    Bakir, B; Sari, E K; Aydin, B D; Yildiz, S E

    2015-04-01

    We investigated using immunohistochemistry the effects of kefir, koumiss and commercial probiotic capsules on the expression of platelet derived growth factor-c (PDGF-C) and platelet derived growth factor receptor-alpha (PDGFR-α) in mouse liver and kidney. Mice were assigned to four groups: group 1 was given commercial probiotic capsules, group 2 was given kefir, group 3 was given koumiss and group 4 was untreated. After oral administration for 15 days, body weights were recorded and liver and kidney tissue samples were obtained. Hematoxylin and eosin staining was used to examine histology. PDGF-C and PDGFR-α in liver and kidney were localized using the streptavidin-biotin peroxidase complex method (ABC). We found that the weights of the mice in the kefir, koumiss and commercial probiotic capsules groups increased compared to the control group. No differences in liver and kidney histology were observed in any of the experimental groups. Kefir, koumiss and the commercial probiotic preparation increased PDGF-C and PDGFR-α expression.

  20. Injectable Biodegradable Polyurethane Scaffolds with Release of Platelet-derived Growth Factor for Tissue Repair and Regeneration

    Science.gov (United States)

    Hafeman, Andrea E.; Li, Bing; Yoshii, Toshitaka; Zienkiewicz, Katarzyna; Davidson, Jeffrey M.; Guelcher, Scott A.

    2013-01-01

    Purpose The purpose of this work was to investigate the effects of triisocyanate composition on the biological and mechanical properties of biodegradable, injectable polyurethane scaffolds for bone and soft tissue engineering. Methods Scaffolds were synthesized using reactive liquid molding techniques, and were characterized in vivo in a rat subcutaneous model. Porosity, dynamic mechanical properties, degradation rate, and release of growth factors were also measured. Results Polyurethane scaffolds were elastomers with tunable damping properties and degradation rates, and they supported cellular infiltration and generation of new tissue. The scaffolds showed a two-stage release profile of platelet-derived growth factor, characterized by a 75% burst release within the first 24 h and slower release thereafter. Conclusions Biodegradable polyurethanes synthesized from triisocyanates exhibited tunable and superior mechanical properties compared to materials synthesized from lysine diisocyanates. Due to their injectability, biocompatibility, tunable degradation, and potential for release of growth factors, these materials are potentially promising therapies for tissue engineering. PMID:18516665

  1. Platelet-Derived Growth Factor-Receptor α Strongly Inhibits Melanoma Growth In Vitro and In Vivo

    Directory of Open Access Journals (Sweden)

    Debora Faraone

    2009-08-01

    Full Text Available Cutaneous melanoma is the most aggressive skin cancer; it is highly metastatic and responds poorly to current therapies. The expression of platelet-derived growth factor receptors (PDGF-Rs is reported to be reduced in metastatic melanoma compared with benign nevi or normal skin; we then hypothesized that PDGF-Rα may control growth of melanoma cells. We show here that melanoma cells overexpressing PDGF-Rα respond to serum with a significantly lower proliferation compared with that of controls. Apoptosis, cell cycle arrest, pRb dephosphorylation, and DNA synthesis inhibition were also observed in cells overexpressing PDGF-Rα. Proliferation was rescued by PDGF-Rα inhibitors, allowing to exclude nonspecific toxic effects and indicating that PDGF-Rα mediates autocrine antiproliferation signals in melanoma cells. Accordingly, PDGF-Rα was found to mediate staurosporine cytotoxicity. A protein array-based analysis of the mitogen-activated protein kinase pathway revealed that melanoma cells overexpressing PDGF-Rα show a strong reduction of c-Jun phosphorylated in serine 63 and of protein phosphatase 2A/Bα and a marked increase of p38γ, mitogen-activated protein kinase kinase 3, and signal regulatory protein α1 protein expression. In a mouse model of primary melanoma growth, infection with the Ad-vector overexpressing PDGF-Rα reached a significant 70% inhibition of primary melanoma growth (P < .001 and a similar inhibition of tumor angiogenesis. All together, these data demonstrate that PDGF-Rα strongly impairs melanoma growth likely through autocrine mechanisms and indicate a novel endogenous mechanism involved in melanoma control.

  2. Bacteria-induced release of white cell--and platelet-derived vascular endothelial growth factor in vitro

    DEFF Research Database (Denmark)

    Nielsen, Hans Jørgen; Werther, K; Mynster, T

    2001-01-01

    BACKGROUND AND OBJECTIVES: Poor prognosis after resection of primary colorectal cancer may be related to the combination of perioperative blood transfusion and subsequent development of infectious complications. White blood cell--and platelet-derived cancer growth substances, including vascular...... endothelial growth factor (VEGF), may be involved in this process. Therefore, we studied the in vitro release of VEGF from white blood cells and platelets stimulated by bacterial antigens and supernatants from stored red cell components. MATERIALS AND METHODS: Eight units of whole blood (WB) and eight units...... of buffy-coat-depleted red cell (SAGM) blood were donated by healthy blood donors. Subsequently, half of every unit was leucocyte depleted by filtration, and all 32 half-units were stored under standard conditions for 35 days. Just after storage, and on days 7, 21 and 35 during storage, aliquots...

  3. Neer Award 2018: Platelet-derived growth factor receptor α co-expression typifies a subset of platelet-derived growth factor receptor β-positive progenitor cells that contribute to fatty degeneration and fibrosis of the murine rotator cuff.

    Science.gov (United States)

    Jensen, Andrew R; Kelley, Benjamin V; Mosich, Gina M; Ariniello, Allison; Eliasberg, Claire D; Vu, Brandon; Shah, Paras; Devana, Sai K; Murray, Iain R; Péault, Bruno; Dar, Ayelet; Petrigliano, Frank A

    2018-04-10

    After massive tears, rotator cuff muscle often undergoes atrophy, fibrosis, and fatty degeneration. These changes can lead to high surgical failure rates and poor patient outcomes. The identity of the progenitor cells involved in these processes has not been fully elucidated. Platelet-derived growth factor receptor β (PDGFRβ) and platelet-derived growth factor receptor α (PDGFRα) have previously been recognized as markers of cells involved in muscle fibroadipogenesis. We hypothesized that PDGFRα expression identifies a fibroadipogenic subset of PDGFRβ + progenitor cells that contribute to fibroadipogenesis of the rotator cuff. We created massive rotator cuff tears in a transgenic strain of mice that allows PDGFRβ + cells to be tracked via green fluorescent protein (GFP) fluorescence. We then harvested rotator cuff muscle tissues at multiple time points postoperatively and analyzed them for the presence and localization of GFP + PDGFRβ + PDGFRα + cells. We cultured, induced, and treated these cells with the molecular inhibitor CWHM-12 to assess fibrosis inhibition. GFP + PDGFRβ + PDGFRα + cells were present in rotator cuff muscle tissue and, after massive tears, localized to fibrotic and adipogenic tissues. The frequency of PDGFRβ + PDGFRα + cells increased at 5 days after massive cuff tears and decreased to basal levels within 2 weeks. PDGFRβ + PDGFRα + cells were highly adipogenic and significantly more fibrogenic than PDGFRβ + PDGFRα - cells in vitro and localized to adipogenic and fibrotic tissues in vivo. Treatment with CWHM-12 significantly decreased fibrogenesis from PDGFRβ + PDGFRα + cells. PDGFRβ + PDGFRα + cells directly contribute to fibrosis and fatty degeneration after massive rotator cuff tears in the mouse model. In addition, CWHM-12 treatment inhibits fibrogenesis from PDGFRβ + PDGFRα + cells in vitro. Clinically, perioperative PDGFRβ + PDGFRα + cell inhibition may limit rotator cuff tissue degeneration and, ultimately

  4. Growth/differentiation factor-5 significantly enhances periodontal wound healing/regeneration compared with platelet-derived growth factor-BB in dogs.

    Science.gov (United States)

    Kwon, Hyuk-Rak; Wikesjö, Ulf M E; Park, Jung-Chul; Kim, Young-Taek; Bastone, Patrizia; Pippig, Susanne D; Kim, Chong-Kwan

    2010-08-01

    Recombinant human growth/differentiation factor-5 (rhGDF-5) in a particulate beta-tricalcium phosphate (beta-TCP) carrier is being evaluated to support periodontal regeneration. The objective of this study was to evaluate periodontal wound healing/regeneration following an established clinical (benchmark) protocol including surgical implantation of rhGDF-5/beta-TCP in comparison with that following implantation of recombinant human platelet-derived growth factor-BB (rhPDGF) combined with a particulate beta-TCP biomaterial using an established canine defect model. Bilateral, 4 x 5 mm (width x depth), one-wall, critical-size, intrabony periodontal defects were surgically created at the mandibular second and fourth pre-molar teeth in five adult Beagle dogs. Defect sites were randomized to receive rhGDF-5/beta-TCP or the rhPDGF construct followed by wound closure for primary intention healing. The animals were sacrificed following an 8-week healing interval for histological and histometric examination. Clinical healing was generally uneventful. Sites receiving rhGDF-5/beta-TCP exhibited a significantly enhanced cementum formation compared with sites receiving the rhPDGF construct, averaging (+/-SD) 4.49+/-0.48 versus 2.72+/-0.91 mm (palveolar bone. Both protocols displayed beta-TCP residues apparently undergoing resorption. Application of both materials appears safe, as they were associated with limited, if any, adverse events. rhGDF-5/beta-TCP shows a significant potential to support/accelerate periodontal wound healing/regeneration. Application of rhGDF-5/beta-TCP appears safe and should be further evaluated in human clinical trials.

  5. Use of multitarget tyrosine kinase inhibitors to attenuate platelet-derived growth factor signalling in lung disease

    Directory of Open Access Journals (Sweden)

    Rana Kanaan

    2017-10-01

    Full Text Available Platelet-derived growth factors (PDGFs and their receptors (PDGFRs play a fundamental role in the embryonic development of the lung. Aberrant PDGF signalling has been documented convincingly in a large variety of pulmonary diseases, including idiopathic pulmonary arterial hypertension, lung cancer and lung fibrosis. Targeting PDGF signalling has been proven to be effective in these diseases. In clinical practice, the most effective way to block PDGF signalling is to inhibit the activity of the intracellular PDGFR kinases. Although the mechanism of action of such drugs is not specific for PDGF signalling, the medications have a broad therapeutic index that allows clinical use. The safety profile and therapeutic opportunities of these and future medications that target PDGFs and PDGFRs are reviewed.

  6. Cloning and expression of a cDNA coding for the human platelet-derived growth factor receptor: Evidence for more than one receptor class

    International Nuclear Information System (INIS)

    Gronwald, R.G.K.; Grant, F.J.; Haldeman, B.A.; Hart, C.E.; O'Hara, P.J.; Hagen, F.S.; Ross, R.; Bowen-Pope, D.F.; Murray, M.J.

    1988-01-01

    The complete nucleotide sequence of a cDNA encoding the human platelet-derived growth factor (PDGF) receptor is presented. The cDNA contains an open reading frame that codes for a protein of 1106 amino acids. Comparison to the mouse PDGF receptor reveals an overall amino acid sequence identity of 86%. This sequence identity rises to 98% in the cytoplasmic split tyrosine kinase domain. RNA blot hybridization analysis of poly(A) + RNA from human dermal fibroblasts detects a major and a minor transcript using the cDNA as a probe. Baby hamster kidney cells, transfected with an expression vector containing the receptor cDNA, express an ∼ 190-kDa cell surface protein that is recognized by an anti-human PDGF receptor antibody. The recombinant PDGF receptor is functional in the transfected baby hamster kidney cells as demonstrated by ligand-induced phosphorylation of the receptor. Binding properties of the recombinant PDGF receptor were also assessed with pure preparations of BB and AB isoforms of PDGF. Unlike human dermal fibroblasts, which bind both isoforms with high affinity, the transfected baby hamster kidney cells bind only the BB isoform of PDGF with high affinity. This observation is consistent with the existence of more than one PDGF receptor class

  7. Forced expression of platelet-derived growth factor B in the mouse cerebellar primordium changes cell migration during midline fusion and causes cerebellar ectopia

    NARCIS (Netherlands)

    Andrae, Johanna; Afink, Gijs; Zhang, Xiao-Qun; Wurst, Wolfgang; Nistér, Monica

    2004-01-01

    The platelet-derived growth factor (PDGF) and receptors are expressed in the developing central nervous system and in brain tumors. To investigate the role of PDGF during normal cerebellar development, we created transgenic mice where PDGF-B was introduced into the endogenous Engrailed1 locus (En1).

  8. Two signaling molecules share a phosphotyrosine-containing binding site in the platelet-derived growth factor receptor.

    Science.gov (United States)

    Nishimura, R; Li, W; Kashishian, A; Mondino, A; Zhou, M; Cooper, J; Schlessinger, J

    1993-11-01

    Autophosphorylation sites of growth factor receptors with tyrosine kinase activity function as specific binding sites for Src homology 2 (SH2) domains of signaling molecules. This interaction appears to be a crucial step in a mechanism by which receptor tyrosine kinases relay signals to downstream signaling pathways. Nck is a widely expressed protein consisting exclusively of SH2 and SH3 domains, the overexpression of which causes cell transformation. It has been shown that various growth factors stimulate the phosphorylation of Nck and its association with autophosphorylated growth factor receptors. A panel of platelet-derived growth factor (PDGF) receptor mutations at tyrosine residues has been used to identify the Nck binding site. Here we show that mutation at Tyr-751 of the PDGF beta-receptor eliminates Nck binding both in vitro and in living cells. Moreover, the Y751F PDGF receptor mutant failed to mediate PDGF-stimulated phosphorylation of Nck in intact cells. A phosphorylated Tyr-751 is also required for binding of phosphatidylinositol-3 kinase to the PDGF receptor. Hence, the SH2 domains of p85 and Nck share a binding site in the PDGF receptor. Competition experiments with different phosphopeptides derived from the PDGF receptor suggest that binding of Nck and p85 is influenced by different residues around Tyr-751. Thus, a single tyrosine autophosphorylation site is able to link the PDGF receptor to two distinct SH2 domain-containing signaling molecules.

  9. The detection of platelet derived growth factor using decoupling of quencher-oligonucleotide from aptamer/quantum dot bioconjugates

    International Nuclear Information System (INIS)

    Kim, Gang-Il; Sung, Yun-Mo; Kim, Kyung-Woo; Oh, Min-Kyu

    2009-01-01

    High-sensitivity, high-specificity detection of platelet derived growth factor (PDGF)-BB was realized using the change in fluorescence resonance energy transfer (FRET) occurring between quantum dot (QD) donors and black hole quencher (BHQ) acceptors. CdSe/ZnS QD/mercaptoacetic acid (MAA)/PDGF aptamer bioconjugates were successfully synthesized using ligand exchange. Black hole quencher (BHQ)-bearing oligonucleotide molecules showing partial sequence matching to PDGF aptamer were attached to PDGF aptamers and photoluminescence (PL) quenching was obtained through FRET. By adding target PDGF-BB to the bioconjugates containing BHQs, PL recovery was detected due to detachment of BHQ-bearing oligonucleotide from the PDGF aptamer as a result of the difference in affinity to the PDGF aptamer. The detection limit of the sensor was ∼0.4 nM and the linearity was maintained up to 1.6 nM in the PL intensity versus concentration curve. Measurement of PL recovery was suggested as a strong tool for high-sensitivity detection of PDGF-BB. Epidermal growth factor (EGF), the negative control molecule, did not contribute to PL recovery due to lack of binding affinity to the PDGF aptamers, which demonstrates the selectivity of the biosensor.

  10. Platelet-Derived Growth Factor (PDGF/PDGF Receptors (PDGFR Axis as Target for Antitumor and Antiangiogenic Therapy

    Directory of Open Access Journals (Sweden)

    Anca Maria Cimpean

    2010-03-01

    Full Text Available Angiogenesis in normal and pathological conditions is a multi-step process governed by positive and negative endogenous regulators. Many growth factors are involved in different steps of angiogenesis, like vascular endothelial growth factors (VEGF, fibroblast growth factor (FGF-2 or platelet-derived growth factors (PDGF. From these, VEGF and FGF-2 were extensively investigated and it was shown that they significantly contribute to the induction and progression of angiogenesis. A lot of evidence has been accumulated in last 10 years that supports the contribution of PDGF/PDGFR axis in developing angiogenesis in both normal and tumoral conditions. The crucial role of PDGF-B and PDGFR-β in angiogenesis has been demonstrated by gene targeting experiments, and their expression correlates with increased vascularity and maturation of the vascular wall. PDGF and their receptors were identified in a large variety of human tumor cells. In experimental models it was shown that inhibition of PDGF reduces interstitial fluid pressure in tumors and enhances the effect of chemotherapy. PDGFR have been involved in the cardiovascular development and their loss leads to a disruption in yolk sac blood vessels development. PDGFRβ expression by pericytes is necessary for their recruitment and integration in the wall of tumor vessels. Endothelial cells of tumor-associated blood vessels can express PDGFR. Based on these data, it was suggested the potential benefit of targeting PDGFR in the treatment of solid tumors. The molecular mechanisms of PDGF/PDGFR-mediated angiogenesis are not fully understood, but it was shown that tyrosine kinase inhibitors reduce tumor growth and angiogenesis in experimental xenograft models, and recent data demonstrated their efficacy in chemoresistant tumors. The in vivo effects of PDGFR inhibitors are more complex, based on the cross-talk with other angiogenic factors. In this review, we summarize data regarding the mechanisms and

  11. The Oligo Fucoidan Inhibits Platelet-Derived Growth Factor-Stimulated Proliferation of Airway Smooth Muscle Cells

    Directory of Open Access Journals (Sweden)

    Chao-Huei Yang

    2016-01-01

    Full Text Available In the pathogenesis of asthma, the proliferation of airway smooth muscle cells (ASMCs is a key factor in airway remodeling and causes airway narrowing. In addition, ASMCs are also the effector cells of airway inflammation. Fucoidan extracted from marine brown algae polysaccharides has antiviral, antioxidant, antimicrobial, anticlotting, and anticancer properties; however, its effectiveness for asthma has not been elucidated thus far. Platelet-derived growth factor (PDGF-treated primary ASMCs were cultured with or without oligo-fucoidan (100, 500, or 1000 µg/mL to evaluate its effects on cell proliferation, cell cycle, apoptosis, and Akt, ERK1/2 signaling pathway. We found that PDGF (40 ng/mL increased the proliferation of ASMCs by 2.5-fold after 48 h (p < 0.05. Oligo-fucoidan reduced the proliferation of PDGF-stimulated ASMCs by 75%–99% after 48 h (p < 0.05 and induced G1/G0 cell cycle arrest, but did not induce apoptosis. Further, oligo-fucoidan supplementation reduced PDGF-stimulated extracellular signal-regulated kinase (ERK1/2, Akt, and nuclear factor (NF-κB phosphorylation. Taken together, oligo-fucoidan supplementation might reduce proliferation of PDGF-treated ASMCs through the suppression of ERK1/2 and Akt phosphorylation and NF-κB activation. The results provide basis for future animal experiments and human trials.

  12. Platelet-derived-growth-factor-induced signalling in human platelets: phosphoinositide-3-kinase-dependent inhibition of platelet activation.

    Science.gov (United States)

    Selheim, F; Fukami, M H; Holmsen, H; Vassbotn, F S

    2000-09-01

    Human platelets release platelet-derived growth factor (PDGF) from alpha-granules during platelet activation. We have previously shown that platelets have PDGF alpha-receptors, a transmembrane tyrosine kinase that takes part in negative feedback regulation during platelet activation. Here we have described a study of PDGF-induced tyrosine phosphorylation of platelet substrates and phosphoinositide 3-kinase (PI-3K) activity in collagen-stimulated platelets. By immunoblotting with phosphotyrosine antibodies of collagen-activated platelets we found that PDGF increased the phosphorylation of several platelet substrates, e.g. pp140, pp120 and pp85. PDGF inhibited collagen-induced platelet activation in the presence of inhibitors of autocrine stimulation, thus blocking the pure collagen-induced signal transduction. PDGF enhanced the collagen-induced formation of PtdIns(3,4)P(2) and PtdIns(3,4,5)P(3) as measured by HPLC. Wortmannin and LY294002, two unrelated inhibitors of PI-3K, were used to investigate the role of PI-3K in PDGF-induced platelet signalling. Incubation of platelets with wortmannin and LY294002 blocked the formation of three phosphorylated inositides as well as the inhibitory effect of PDGF on collagen-induced platelet activation. We conclude that the inhibitory effect of PDGF on platelet activation is PI-3K dependent. This is the first demonstration of a negative regulatory function of 3-phosphorylated inositides in platelets.

  13. Terbinafine: effects on platelet-derived growth factor-stimulated smooth muscle cells in vitro and myointimal proliferation in vivo

    International Nuclear Information System (INIS)

    McCarthy, L.; Van Halen, R.G.; St Denny, I.H.; Glinka, K.G.; Handley, D.A.; Stuetz, A.; Nemecek, G.M.

    1987-01-01

    Terbinafine (T; (E)-N-(6,6-dimethyl-2-hepten-4-ynyl)-N-methyl-1-naphthalenemethanamine), an antimycotic agent with antimitogenic activity in fibroblasts, was examined for its effects on platelet-derived growth factor (PDGF)-stimulated aortic smooth muscle cell DNA synthesis in vitro and myointimal proliferation in vivo. Exposure of smooth muscle cells to 1-25 μM T resulted in a concentration-dependent inhibition of PDGF-induced mitogenesis as determined by [ 3 H]thymidine incorporation or cell number. The IC 50 for T was approximately 5 μM. The inhibitory effect of terbinafine persisted in the presence of 0.4-8.0 μg/ml cholesterol or 130 μg/ml mevalonate. Administration of T to rats for 2 d before and 14 d after balloon catheter carotid injury resulted in a 40% decrease in lesion area. These observations indicate that T is both a potent in vitro antagonist of the smooth muscle cell mitogenic response to PDGF and an effective, well-tolerated, orally active inhibitor of myointimal proliferation in vivo

  14. Morphine induces expression of platelet-derived growth factor in human brain microvascular endothelial cells: implication for vascular permeability.

    Directory of Open Access Journals (Sweden)

    Hongxiu Wen

    Full Text Available Despite the advent of antiretroviral therapy, complications of HIV-1 infection with concurrent drug abuse are an emerging problem. Morphine, often abused by HIV-infected patients, is known to accelerate neuroinflammation associated with HIV-1 infection. Detailed molecular mechanisms of morphine action however, remain poorly understood. Platelet-derived growth factor (PDGF has been implicated in a number of pathological conditions, primarily due to its potent mitogenic and permeability effects. Whether morphine exposure results in enhanced vascular permeability in brain endothelial cells, likely via induction of PDGF, remains to be established. In the present study, we demonstrated morphine-mediated induction of PDGF-BB in human brain microvascular endothelial cells, an effect that was abrogated by the opioid receptor antagonist-naltrexone. Pharmacological blockade (cell signaling and loss-of-function (Egr-1 approaches demonstrated the role of mitogen-activated protein kinases (MAPKs, PI3K/Akt and the downstream transcription factor Egr-1 respectively, in morphine-mediated induction of PDGF-BB. Functional significance of increased PDGF-BB manifested as increased breach of the endothelial barrier as evidenced by decreased expression of the tight junction protein ZO-1 in an in vitro model system. Understanding the regulation of PDGF expression may provide insights into the development of potential therapeutic targets for intervention of morphine-mediated neuroinflammation.

  15. Terbinafine: effects on platelet-derived growth factor-stimulated smooth muscle cells in vitro and myointimal proliferation in vivo

    Energy Technology Data Exchange (ETDEWEB)

    McCarthy, L.; Van Halen, R.G.; St. Denny, I.H.; Glinka, K.G.; Handley, D.A.; Stuetz, A.; Nemecek, G.M.

    1987-05-01

    Terbinafine (T; (E)-N-(6,6-dimethyl-2-hepten-4-ynyl)-N-methyl-1-naphthalenemethanamine), an antimycotic agent with antimitogenic activity in fibroblasts, was examined for its effects on platelet-derived growth factor (PDGF)-stimulated aortic smooth muscle cell DNA synthesis in vitro and myointimal proliferation in vivo. Exposure of smooth muscle cells to 1-25 ..mu..M T resulted in a concentration-dependent inhibition of PDGF-induced mitogenesis as determined by (/sup 3/H)thymidine incorporation or cell number. The IC/sub 50/ for T was approximately 5 ..mu..M. The inhibitory effect of terbinafine persisted in the presence of 0.4-8.0 ..mu..g/ml cholesterol or 130 ..mu..g/ml mevalonate. Administration of T to rats for 2 d before and 14 d after balloon catheter carotid injury resulted in a 40% decrease in lesion area. These observations indicate that T is both a potent in vitro antagonist of the smooth muscle cell mitogenic response to PDGF and an effective, well-tolerated, orally active inhibitor of myointimal proliferation in vivo.

  16. Platelet-derived growth factor-DD targeting arrests pathological angiogenesis by modulating glycogen synthase kinase-3beta phosphorylation.

    Science.gov (United States)

    Kumar, Anil; Hou, Xu; Lee, Chunsik; Li, Yang; Maminishkis, Arvydas; Tang, Zhongshu; Zhang, Fan; Langer, Harald F; Arjunan, Pachiappan; Dong, Lijin; Wu, Zhijian; Zhu, Linda Y; Wang, Lianchun; Min, Wang; Colosi, Peter; Chavakis, Triantafyllos; Li, Xuri

    2010-05-14

    Platelet-derived growth factor-DD (PDGF-DD) is a recently discovered member of the PDGF family. The role of PDGF-DD in pathological angiogenesis and the underlying cellular and molecular mechanisms remain largely unexplored. In this study, using different animal models, we showed that PDGF-DD expression was up-regulated during pathological angiogenesis, and inhibition of PDGF-DD suppressed both choroidal and retinal neovascularization. We also demonstrated a novel mechanism mediating the function of PDGF-DD. PDGF-DD induced glycogen synthase kinase-3beta (GSK3beta) Ser(9) phosphorylation and Tyr(216) dephosphorylation in vitro and in vivo, leading to increased cell survival. Consistently, GSK3beta activity was required for the antiangiogenic effect of PDGF-DD targeting. Moreover, PDGF-DD regulated the expression of GSK3beta and many other genes important for angiogenesis and apoptosis. Thus, we identified PDGF-DD as an important target gene for antiangiogenic therapy due to its pleiotropic effects on vascular and non-vascular cells. PDGF-DD inhibition may offer new therapeutic options to treat neovascular diseases.

  17. A Tumor Suppressor Gene Product, Platelet-Derived Growth Factor Receptor-Like Protein Controls Chondrocyte Proliferation and Differentiation.

    Science.gov (United States)

    Kawata, Kazumi; Kubota, Satoshi; Eguchi, Takanori; Aoyama, Eriko; Moritani, Norifumi H; Oka, Morihiko; Kawaki, Harumi; Takigawa, Masaharu

    2017-11-01

    The platelet-derived growth factor receptor-like (PDGFRL) gene is regarded as a tumor suppressor gene. However, nothing is known about the molecular function of PDGFRL. In this study, we initially clarified its function in chondrocytes. Among all cell lines examined, the PDGFRL mRNA level was the highest in chondrocytic HCS-2/8 cells. Interestingly, the proliferation of chondrocytic HCS-2/8 cells was promoted by PDGFRL overexpression, whereas that of the breast cancer-derived MDA-MB-231 cells was inhibited. Of note, in PDGFRL-overexpressing HCS-2/8 cells, the expression of chondrocyte differentiation marker genes, SOX9, ACAN, COL2A1, COL10A1, and ALP, was decreased. Moreover, we confirmed the expression of PDGFRL mRNA in normal cartilage tissue and chondrocytes. Eventually, the expression of PDGFRL mRNA in condrocytes except in the case of hypertrophic chondrocytes was demonstrated in vivo and in vitro. These findings suggest that PDGFRL plays the different roles, depending upon cell types. Particularly, in chondrocytes, PDGFRL may play a new and important role which is distinct from the function previously reported. J. Cell. Biochem. 118: 4033-4044, 2017. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  18. Evidence that platelet-derived growth factor may be a novel endogenous pyrogen in the central nervous system.

    Science.gov (United States)

    Pelá, I R; Ferreira, M E; Melo, M C; Silva, C A; Coelho, M M; Valenzuela, C F

    2000-05-01

    Platelet-derived growth factor (PDGF) exerts neurotrophic and neuromodulatory actions in the mammalian central nervous system (CNS). Like the cytokines, PDGF primarily signals through tyrosine phosphorylation-dependent pathways that activate multiple intracellular molecules including Janus family kinases. We previously showed that microinjection of PDGF-BB into the lateral ventricle induced a febrile response in rats that was reduced by pretreatment with Win 41662, a potent inhibitor of PDGF receptors (Pelá IR, Ferreira MES, Melo MCC, Silva CAA, and Valenzuela CF. Ann NY Acad Sci 856: 289-293, 1998). In this study, we further characterized the role of PDGF-BB in the febrile response in rats. Microinjection of PDGF-BB into the third ventricle produced a dose-dependent increase in colonic temperature that peaked 3-4 h postinjection. Win 41662 attenuated fever induced by intraperitoneal injection of bacterial lipopolysaccharide, suggesting that endogenous PDGF participates in the febrile response to this exogenous pyrogen. Importantly, febrile responses induced by tumor necrosis factor-alpha, interleukin-1beta, and interleukin-6 were unchanged by Win 41662. Both indomethacin and dexamethasone blocked the PDGF-BB-induced increase in colonic temperature, and, therefore, we postulate that PDGF-BB may act via prostaglandin- and/or inducible enzyme-dependent pathways. Thus our findings suggest that PDGF-BB is an endogenous CNS mediator of the febrile response in rats.

  19. Elevated platelet-derived growth factor-BB concentrations in premature neonates who develop chronic lung disease

    Directory of Open Access Journals (Sweden)

    Adcock Kim G

    2004-06-01

    Full Text Available Abstract Background Chronic lung disease (CLD in the preterm newborn is associated with inflammation and fibrosis. Platelet-derived growth factor-BB (PDGF-BB, a potent chemotactic growth factor, may mediate the fibrotic component of CLD. The objectives of this study were to determine if tracheal aspirate (TA concentrations of PDGF-BB increase the first 2 weeks of life in premature neonates undergoing mechanical ventilation for respiratory distress syndrome (RDS, its relationship to the development of CLD, pulmonary hemorrhage (PH and its relationship to airway colonization with Ureaplasma urealyticum (Uu. Methods Infants with a birth weight less than 1500 grams who required mechanical ventilation for RDS were enrolled into this study with parental consent. Tracheal aspirates were collected daily during clinically indicated suctioning. Uu cultures were performed on TA collected in the first week of life. TA supernatants were assayed for PDGF-BB and secretory component of IgA concentrations using ELISA techniques. Results Fifty premature neonates were enrolled into the study. Twenty-eight infants were oxygen dependent at 28 days of life and 16 infants were oxygen dependent at 36 weeks postconceptual age. PDGF-BB concentrations peaked between 4 and 6 days of life. Maximum PDGF-BB concentrations were significantly higher in infants who developed CLD or died from respiratory failure. PH was associated with increased risk of CLD and was associated with higher PDGF-BB concentrations. There was no correlation between maximum PDGF-BB concentrations and Uu isolation from the airway. Conclusions PDGF-BB concentrations increase in TAs of infants who undergo mechanical ventilation for RDS during the first 2 weeks of life and maximal concentrations are greater in those infants who subsequently develop CLD. Elevation in lung PDGF-BB may play a role in the development of CLD.

  20. Platelet-derived growth factor regulates vascular smooth muscle phenotype via mammalian target of rapamycin complex 1

    International Nuclear Information System (INIS)

    Ha, Jung Min; Yun, Sung Ji; Kim, Young Whan; Jin, Seo Yeon; Lee, Hye Sun; Song, Sang Heon; Shin, Hwa Kyoung; Bae, Sun Sik

    2015-01-01

    Mammalian target of rapamycin complex (mTORC) regulates various cellular processes including proliferation, growth, migration and differentiation. In this study, we showed that mTORC1 regulates platelet-derived growth factor (PDGF)-induced phenotypic conversion of vascular smooth muscle cells (VSMCs). Stimulation of contractile VSMCs with PDGF significantly reduced the expression of contractile marker proteins in a time- and dose-dependent manner. In addition, angiotensin II (AngII)-induced contraction of VSMCs was completely blocked by the stimulation of VSMCs with PDGF. PDGF-dependent suppression of VSMC marker gene expression was significantly blocked by inhibition of phosphatidylinositol 3-kinase (PI3K), extracellular signal-regulated kinase (ERK), and mTOR whereas inhibition of p38 MAPK had no effect. In particular, inhibition of mTORC1 by rapamycin or by silencing of Raptor significantly blocked the PDGF-dependent phenotypic change of VSMCs whereas silencing of Rictor had no effect. In addition, loss of AngII-dependent contraction by PDGF was significantly retained by silencing of Raptor. Inhibition of mTORC1 by rapamycin or by silencing of Raptor significantly blocked PDGF-induced proliferation of VSMCs. Taken together, we suggest that mTORC1 plays an essential role in PDGF-dependent phenotypic changes of VSMCs. - Graphical abstract: Regulation of VSMC phenotype by PDGF-dependent activation of mTORC1. - Highlights: • The expression of contractile marker proteins was reduced by PDGF stimulation. • PDGF-dependent phenotypic conversion of VSMCs was blocked by inhibition of mTOR. • PDGF-induced proliferation of VSMCs was attenuated by inhibition of mTORC1. • mTORC1 plays a critical role in PDGF-dependent phenotypic conversion of VSMCs

  1. Platelet-derived growth factor regulates vascular smooth muscle phenotype via mammalian target of rapamycin complex 1

    Energy Technology Data Exchange (ETDEWEB)

    Ha, Jung Min; Yun, Sung Ji; Kim, Young Whan; Jin, Seo Yeon; Lee, Hye Sun [Medical Research Institute, Department of Pharmacology, Pusan National University School of Medicine, Yangsan (Korea, Republic of); Song, Sang Heon [Department of Internal Medicine, Pusan National University Hospital, Busan (Korea, Republic of); Shin, Hwa Kyoung [Department of Anatomy, Pusan National University School of Korean Medicine, Yangsan (Korea, Republic of); Bae, Sun Sik, E-mail: sunsik@pusan.ac.kr [Medical Research Institute, Department of Pharmacology, Pusan National University School of Medicine, Yangsan (Korea, Republic of)

    2015-08-14

    Mammalian target of rapamycin complex (mTORC) regulates various cellular processes including proliferation, growth, migration and differentiation. In this study, we showed that mTORC1 regulates platelet-derived growth factor (PDGF)-induced phenotypic conversion of vascular smooth muscle cells (VSMCs). Stimulation of contractile VSMCs with PDGF significantly reduced the expression of contractile marker proteins in a time- and dose-dependent manner. In addition, angiotensin II (AngII)-induced contraction of VSMCs was completely blocked by the stimulation of VSMCs with PDGF. PDGF-dependent suppression of VSMC marker gene expression was significantly blocked by inhibition of phosphatidylinositol 3-kinase (PI3K), extracellular signal-regulated kinase (ERK), and mTOR whereas inhibition of p38 MAPK had no effect. In particular, inhibition of mTORC1 by rapamycin or by silencing of Raptor significantly blocked the PDGF-dependent phenotypic change of VSMCs whereas silencing of Rictor had no effect. In addition, loss of AngII-dependent contraction by PDGF was significantly retained by silencing of Raptor. Inhibition of mTORC1 by rapamycin or by silencing of Raptor significantly blocked PDGF-induced proliferation of VSMCs. Taken together, we suggest that mTORC1 plays an essential role in PDGF-dependent phenotypic changes of VSMCs. - Graphical abstract: Regulation of VSMC phenotype by PDGF-dependent activation of mTORC1. - Highlights: • The expression of contractile marker proteins was reduced by PDGF stimulation. • PDGF-dependent phenotypic conversion of VSMCs was blocked by inhibition of mTOR. • PDGF-induced proliferation of VSMCs was attenuated by inhibition of mTORC1. • mTORC1 plays a critical role in PDGF-dependent phenotypic conversion of VSMCs.

  2. Platelet-derived growth factor (PDGF) stimulates glycogen synthase activity in 3T3 cells

    International Nuclear Information System (INIS)

    Chan, C.P.; Bowen-Pope, D.F.; Ross, R.; Krebs, E.G.

    1986-01-01

    Hormonal regulation of glycogen synthase, an enzyme that can be phosphorylated on multiple sites, is often associated with changes in its phosphorylation state. Enzyme activation is conventionally monitored by determining the synthase activity ratio [(activity in the absence of glucose 6-P)/(activity in the presence of glucose 6-P)]. Insulin causes an activation of glycogen synthase with a concomitant decrease in its phosphate content. In a previous report, the authors showed that epidermal growth factor (EGF) increases the glycogen synthase activity ratio in Swiss 3T3 cells. The time and dose-dependency of this response was similar to that of insulin. Their recent results indicate that PDGF also stimulates glycogen synthase activity. Enzyme activation was maximal after 30 min. of incubation with PDGF; the time course observed was very similar to that with insulin and EGF. At 1 ng/ml (0.03nM), PDGF caused a maximal stimulation of 4-fold in synthase activity ratio. Half-maximal stimulation was observed at 0.2 ng/ml (6 pM). The time course of changes in enzyme activity ratio closely followed that of 125 I-PDGF binding. The authors data suggest that PDGF, as well as EFG and insulin, may be important in regulating glycogen synthesis through phosphorylation/dephosphorylation mechanisms

  3. Treatment of AVN Using Autologous BM Stem Cells and Activated Platelet-Derived Growth Factor Concentrates.

    Science.gov (United States)

    Nandeesh, Nagaraj H; Janardhan, Kiranmayee; Subramanian, Vignesh; Ashtekar, Abhishek Bhushan; Srikruthi, Nandagiri; Koka, Prasad S; Deb, Kaushik

    Avascular Necrosis (AVN) of hip is a devastating condition seen in younger individuals. It is the ischemic death of the constituents of the bone cartilage of the hip. The femoral head (FH) is the most common site for AVN. It results from interruption of the normal blood flow to the FH that fits into the hip socket. Earlier studies using autologous bone marrow stem cell concentrate injections have shown encouraging results with average success rates. The current study was designed to improve significantly the cartilage regeneration and clinical outcome. Total of 48 patients underwent autologous bone marrow stem cell and activated platelet-rich plasma derived growth factor concentrate (PRP-GFC) therapy for early and advanced stages AVN of femoral head in a single multi-specialty center. The total treatment was divided into three phases. In the phase I, all the clinical diagnostic measurements such as magnetic resonance imaging (MRI), computed tomography (CT) etc. with respect to the AVN patients and bone marrow aspiration from posterior iliac spine from the patients were carried out. In the phase II, isolation of stem cells and preparation from the patients were performed. Subsequently, in phase III, the stem cells and PRP- GFCs were transplanted in the enrolled patients. Ninety three percent of the enrolled AVN patients showed marked enhancement in the hip bone joint space (more than 3mm) after combined stem cells and PRP-GFC treatment as evidenced by comparison of the pre- and post-treatment MRI data thus indicative of regeneration of cartilage. The treated patients showed significant improvement in their motor function, cartilage regrowth (3 to 10mm), and high satisfaction in the two-year follow-up. Combination of stem cell and PRP-GFC therapy has shown promising cartilage regeneration in 45 out of 48 patients of AVN. This study clearly demonstrates the safety and efficacy of this treatment. Larger numbers of patients need to be evaluated to better understand the

  4. Negative feedback regulation of human platelets via autocrine activation of the platelet-derived growth factor alpha-receptor.

    Science.gov (United States)

    Vassbotn, F S; Havnen, O K; Heldin, C H; Holmsen, H

    1994-05-13

    Human platelets contain platelet-derived growth factor (PDGF) in their alpha-granules which is released during platelet exocytosis. We show by immunoprecipitation and 125I-PDGF binding experiments that human platelets have functionally active PDGF alpha-receptors, but not beta-receptors. The PDGF alpha-receptor (PDGFR-alpha) was identified as a 170-kDa glycosylated protein-tyrosine kinase as found in other cell types. Stimulation of platelets with 0.1 unit/ml thrombin resulted in a significant increase (2-5-fold) of the tyrosine phosphorylation of the PDGFR-alpha, as determined by immunoprecipitation with phosphotyrosine antiserum as well as with PDGFR-alpha antiserum. The observed thrombin-induced autophosphorylation of the PDGFR-alpha was inhibited by the addition of a neutralizing monoclonal PDGF antibody. Thus, our results suggest that the platelet PDGFR-alpha is stimulated in an autocrine manner by PDGF secreted during platelet activation. Preincubation of platelets with PDGF inhibited thrombin-induced platelet aggregation and secretion of ATP + ADP and beta-hexosaminidase. Thrombin-induced platelet aggregation was also reversed when PDGF was added 30 s after thrombin stimulation. Inhibition of the autocrine PDGF pathway during platelet activation by the PDGF antibody led to a potentiation of thrombin-induced beta-hexosaminidase secretion. Thus, the PDGFR-alpha takes part in a negative feedback regulation during platelet activation. Our demonstration of PDGF alpha-receptors on human platelets and its inhibitory function during platelet activation identifies a new possible role of PDGF in the regulation of thrombosis.

  5. Expression of platelet-derived growth factor BB, erythropoietin and erythropoietin receptor in canine and feline osteosarcoma.

    Science.gov (United States)

    Meyer, F R L; Steinborn, R; Grausgruber, H; Wolfesberger, B; Walter, I

    2015-10-01

    The discovery of expression of the erythropoietin receptor (EPO-R) on neoplastic cells has led to concerns about the safety of treating anaemic cancer patients with EPO. In addition to its endocrine function, the receptor may play a role in tumour progression through an autocrine mechanism. In this study, the expression of EPO, EPO-R and platelet-derived growth factor BB (PDGF-BB) was analysed in five feline and 13 canine osteosarcomas using immunohistochemistry (IHC) and reverse transcription polymerase chain reaction (RT-PCR). EPO expression was positive in all tumours by IHC, but EPO mRNA was only detected in 38% of the canine and 40% of the feline samples. EPO-R was expressed in all samples by quantitative RT-PCR (RT-qPCR) and IHC. EPO-R mRNA was expressed at higher levels in all feline tumours, tumour cell lines, and kidney when compared to canine tissues. PDGF-BB expression was variable by IHC, but mRNA was detected in all samples. To assess the functionality of the EPO-R on tumour cells, the proliferation of canine and feline osteosarcoma cell lines was evaluated after EPO administration using an alamarBlue assay and Ki67 immunostaining. All primary cell lines responded to EPO treatment in at least one of the performed assays, but the effect on proliferation was very low indicating only a weak responsiveness of EPO-R. In conclusion, since EPO and its receptor are expressed by canine and feline osteosarcomas, an autocrine or paracrine tumour progression mechanism cannot be excluded, although in vitro data suggest a minimal role of EPO-R in osteosarcoma cell proliferation. Copyright © 2015 The Authors. Published by Elsevier Ltd.. All rights reserved.

  6. Assessment of platelet-derived growth factor using A splinted full thickness dermal wound model in bearded dragons (Pogona vitticeps).

    Science.gov (United States)

    Keller, Krista A; Paul-Murphy, Joanne; Weber, E P Scott; Kass, Philip H; Guzman, Sanchez-Migallon David; Park, Shin Ae; Raghunathan, Vijay Krishna; Gustavsen, Kate A; Murphy, Christopher J

    2014-12-01

    Wounds in reptiles are a common reason for presentation to a veterinarian. At this time there is limited information on effective topical medications to aid in wound closure. The objectives of this study were to translate the splinted, full-thickness dermal wound model, validated in mice, to the bearded dragon (Pogona vitticeps) and to determine the effect of topical becaplermin (BP), a platelet-derived growth factor (0.01%), on the rate of wound closure. Ten bearded dragons were anesthetized and two full-thickness cutaneous wounds were made on the dorsum of each lizard. Encircling splints were applied surrounding each wound and subsequently covered by a semi-occlusive dressing. Five lizards had one wound treated with BP and the adjacent wound treated with a vehicle control. Five additional lizards had one wound treated with saline and the second wound treated with a vehicle control. Wounds were imaged daily, and the wound area was measured using digital image analysis. The change in percentage wound closure over 17 days and the time to 50% wound closure was compared among the four treatment groups. There was no significant difference in wound closure rates between BP-treated and saline-treated wounds or in the time to 50% wound closure between any treatments. Vehicle-treated wounds adjacent to saline-treated wounds closed significantly slower than did BP (P dragons. When compared with saline, BP did not have a significant effect on wound closure rates, while the vehicle alone delayed wound closure. Histologic analysis of experimentally created wounds throughout the wound healing process is needed to further evaluate the effects of these treatments on reptile dermal wound healing.

  7. Platelet-derived growth factor receptor beta: a novel urinary biomarker for recurrence of non-muscle-invasive bladder cancer.

    Science.gov (United States)

    Feng, Jiayu; He, Weifeng; Song, Yajun; Wang, Ying; Simpson, Richard J; Zhang, Xiaorong; Luo, Gaoxing; Wu, Jun; Huang, Chibing

    2014-01-01

    Non-muscle-invasive bladder cancer (NMIBC) is one of the most common malignant tumors in the urological system with a high risk of recurrence, and effective non-invasive biomarkers for NMIBC relapse are still needed. The human urinary proteome can reflect the status of the microenvironment of the urinary system and is an ideal source for clinical diagnosis of urinary system diseases. Our previous work used proteomics to identify 1643 high-confidence urinary proteins in the urine from a healthy population. Here, we used bioinformatics to construct a cancer-associated protein-protein interaction (PPI) network comprising 16 high-abundance urinary proteins based on the urinary proteome database. As a result, platelet-derived growth factor receptor beta (PDGFRB) was selected for further validation as a candidate biomarker for NMIBC diagnosis and prognosis. Although the levels of urinary PDGFRB showed no significant difference between patients pre- and post-surgery (n = 185, P>0.05), over 3 years of follow-up, urinary PDGFRB was shown to be significantly higher in relapsed patients (n = 68) than in relapse-free patients (n = 117, P<0.001). The levels of urinary PDGFRB were significantly correlated with the risk of 3-year recurrence of NMIBC, and these levels improved the accuracy of a NMIBC recurrence risk prediction model that included age, tumor size, and tumor number (area under the curve, 0.862; 95% CI, 0.809 to 0.914) compared to PDGFR alone. Therefore, we surmise that urinary PDGFRB could serve as a non-invasive biomarker for predicting NMIBC recurrence.

  8. Bone Morphogenic Protein 4-Smad-Induced Upregulation of Platelet-Derived Growth Factor AA Impairs Endothelial Function.

    Science.gov (United States)

    Hu, Weining; Zhang, Yang; Wang, Li; Lau, Chi Wai; Xu, Jian; Luo, Jiang-Yun; Gou, Lingshan; Yao, Xiaoqiang; Chen, Zhen-Yu; Ma, Ronald Ching Wan; Tian, Xiao Yu; Huang, Yu

    2016-03-01

    Bone morphogenic protein 4 (BMP4) is an important mediator of endothelial dysfunction in cardio-metabolic diseases, whereas platelet-derived growth factors (PDGFs) are major angiogenic and proinflammatory mediator, although the functional link between these 2 factors is unknown. The present study investigated whether PDGF mediates BMP4-induced endothelial dysfunction in diabetes mellitus. We generated Ad-Bmp4 to overexpress Bmp4 and Ad-Pdgfa-shRNA to knockdown Pdgfa in mice through tail intravenous injection. SMAD4-shRNA lentivirus, SMAD1-shRNA, and SMAD5 shRNA adenovirus were used for knockdown in human and mouse endothelial cells. We found that PDGF-AA impaired endothelium-dependent vasodilation in aortas and mesenteric resistance arteries. BMP4 upregulated PDGF-AA in human and mouse endothelial cells, which was abolished by BMP4 antagonist noggin or knockdown of SMAD1/5 or SMAD4. BMP4-impared relaxation in mouse aorta was also ameliorated by PDGF-AA neutralizing antibody. Tail injection of Ad-Pdgfa-shRNA ameliorates endothelial dysfunction induced by Bmp4 overexpression (Ad-Bmp4) in vivo. Serum PDGF-AA was elevated in both diabetic patients and diabetic db/db mice compared with nondiabetic controls. Pdgfa-shRNA or Bmp4-shRNA adenovirus reduced serum PDGF-AA concentration in db/db mice. PDGF-AA neutralizing antibody or tail injection with Pdgfa-shRNA adenovirus improved endothelial function in aortas and mesenteric resistance arteries from db/db mice. The effect of PDGF-AA on endothelial function in mouse aorta was also inhibited by Ad-Pdgfra-shRNA to inhibit PDGFRα. The present study provides novel evidences to show that PDGF-AA impairs endothelium-dependent vasodilation and PDGF-AA mediates BMP4-induced adverse effect on endothelial cell function through SMAD1/5- and SMAD4-dependent mechanisms. Inhibition of PGDF-AA ameliorates vascular dysfunction in diabetic mice. © 2016 American Heart Association, Inc.

  9. An aptamer-based biosensing platform for highly sensitive detection of platelet-derived growth factor via enzyme-mediated direct electrochemistry

    Energy Technology Data Exchange (ETDEWEB)

    Deng Kun; Xiang Yang; Zhang Liqun; Chen Qinghai [Laboratory of the Clinical Experimental Base of Biosensor and Microarray, Center of Molecule and Gene Diagnosis, Southwest Hospital, Third Military Medical University, Chongqing 400042 (China); Fu Weiling, E-mail: weilingfu@yahoo.com [Laboratory of the Clinical Experimental Base of Biosensor and Microarray, Center of Molecule and Gene Diagnosis, Southwest Hospital, Third Military Medical University, Chongqing 400042 (China)

    2013-01-08

    Highlights: Black-Right-Pointing-Pointer Direct electrochemistry of glucose oxidase used for signal generation in aptasensor. Black-Right-Pointing-Pointer Using novel nanocomposite for immobilization and signal amplification. Black-Right-Pointing-Pointer Sensitive electrochemical detection of platelet-derived growth factor. - Abstract: In this work, a new label-free electrochemical aptamer-based sensor (aptasensor) was constructed for detection of platelet-derived growth factor (PDGF) based on the direct electrochemistry of glucose oxidase (GOD). For this proposed aptasensor, poly(diallyldimethylammonium chloride) (PDDA)-protected graphene-gold nanoparticles (P-Gra-GNPs) composite was firstly coated on electrode surface to form the interface with biocompatibility and huge surface area for the adsorption of GOD layer. Subsequently, gold nanoclusters (GNCs) were deposited on the surface of GOD to capture PDGF binding aptamer (PBA). Finally, GOD as a blocking reagent was employed to block the remaining active sites of the GNCs and avoid the nonspecific adsorption. With the direct electron transfer of double layer GOD membranes, the aptasensor showed excellent electrochemical response and the peak current decreased linearly with increasing logarithm of PDGF concentration from 0.005 nM to 60 nM with a relatively low limit of detection of 1.7 pM. The proposed aptasensor exhibited high specificity, good reproducibility and long-term stability, which provided a new promising technique for aptamer-based protein detection.

  10. An aptamer-based biosensing platform for highly sensitive detection of platelet-derived growth factor via enzyme-mediated direct electrochemistry

    International Nuclear Information System (INIS)

    Deng Kun; Xiang Yang; Zhang Liqun; Chen Qinghai; Fu Weiling

    2013-01-01

    Highlights: ► Direct electrochemistry of glucose oxidase used for signal generation in aptasensor. ► Using novel nanocomposite for immobilization and signal amplification. ► Sensitive electrochemical detection of platelet-derived growth factor. - Abstract: In this work, a new label-free electrochemical aptamer-based sensor (aptasensor) was constructed for detection of platelet-derived growth factor (PDGF) based on the direct electrochemistry of glucose oxidase (GOD). For this proposed aptasensor, poly(diallyldimethylammonium chloride) (PDDA)-protected graphene-gold nanoparticles (P-Gra-GNPs) composite was firstly coated on electrode surface to form the interface with biocompatibility and huge surface area for the adsorption of GOD layer. Subsequently, gold nanoclusters (GNCs) were deposited on the surface of GOD to capture PDGF binding aptamer (PBA). Finally, GOD as a blocking reagent was employed to block the remaining active sites of the GNCs and avoid the nonspecific adsorption. With the direct electron transfer of double layer GOD membranes, the aptasensor showed excellent electrochemical response and the peak current decreased linearly with increasing logarithm of PDGF concentration from 0.005 nM to 60 nM with a relatively low limit of detection of 1.7 pM. The proposed aptasensor exhibited high specificity, good reproducibility and long-term stability, which provided a new promising technique for aptamer-based protein detection.

  11. Platelet-derived growth factor (PDGF-BB-mediated induction of monocyte chemoattractant protein 1 in human astrocytes: implications for HIV-associated neuroinflammation

    Directory of Open Access Journals (Sweden)

    Bethel-Brown Crystal

    2012-12-01

    Full Text Available Abstract Chemokine (C-C motif ligand 2, also known as monocyte chemoattractant protein 1 (MCP-1 is an important factor for the pathogenesis of HIV-associated neurocognitive disorders (HAND. The mechanisms of MCP-1-mediated neuropathogenesis, in part, revolve around its neuroinflammatory role and the recruitment of monocytes into the central nervous system (CNS via the disrupted blood-brain barrier (BBB. We have previously demonstrated that HIV-1/HIV-1 Tat upregulate platelet-derived growth factor (PDGF-BB, a known cerebrovascular permeant; subsequently, the present study was aimed at exploring the regulation of MCP-1 by PDGF-BB in astrocytes with implications in HAND. Specifically, the data herein demonstrate that exposure of human astrocytes to HIV-1 LAI elevated PDGF-B and MCP-1 levels. Furthermore, treating astrocytes with the human recombinant PDGF-BB protein significantly increased the production and release of MCP-1 at both the RNA and protein levels. MCP-1 induction was regulated by activation of extracellular-signal-regulated kinase (ERK1/2, c-Jun N-terminal kinase (JNK and p38 mitogen-activated protein (MAP kinases and phosphatidylinositol 3-kinase (PI3K/Akt pathways and the downstream transcription factor, nuclear factor κB (NFκB. Chromatin immunoprecipitation (ChIP assays demonstrated increased binding of NFκB to the human MCP-1 promoter following PDGF-BB exposure. Conditioned media from PDGF-BB-treated astrocytes increased monocyte transmigration through human brain microvascular endothelial cells (HBMECs, an effect that was blocked by STI-571, a tyrosine kinase inhibitor (PDGF receptor (PDGF-R blocker. PDGF-BB-mediated release of MCP-1 was critical for increased permeability in an in vitro BBB model as evidenced by blocking antibody assays. Since MCP-1 is linked to disease severity, understanding its modulation by PDGF-BB could aid in understanding the proinflammatory responses in HAND. These results suggest that astrocyte

  12. Quantification of platelets and platelet derived growth factors from platelet-rich-plasma (PRP) prepared at different centrifugal force (g) and time.

    Science.gov (United States)

    Arora, Satyam; Doda, Veena; Kotwal, Urvershi; Dogra, Mitu

    2016-02-01

    Platelet derived biomaterials represent a key source of cytokines and growth factors extensively used for tissue regeneration; wound healing and tissue repair. Our study was to quantify platelets and growth factors released by PRP when prepared at different centrifugal force (g) and time. Our study was approved by the institutional ethical committee. One hundred millilitres of whole blood (WB) was collected in bag with CPDA as the anticoagulant(AC); (14 mL for 100 mL WB ratio). Nine aliquots of 10 mL each were made from the bag and set of three aliquots were made a group. PRP was prepared at varying centrifugal force (group A: -110 g, group B: -208 g & group C: -440 g) & time (1: -5 min, 2: -10 min & 3: -20 min). Contents of each PRP prepared were analysed. Commercial sandwich ELISA kits were used to quantify the concentrations of CD62P (Diaclone SAS; France), Platelet derived growth factors-AB (Qayee-Bio; China), transforming growth factor-β1 (DRG; Germany) and vascular endothelial growth factor (Boster Immuno Leader; USA) released in each PRP prepared. Eight volunteers were enrolled in the study (24-30 years). The baseline blood counts of all the volunteers were comparable (p ≥ 0.05). Mean ± SD of platelet yield of all nine groups ranged from 17.2 ± 4.2% to 78.7 ± 5.7%. Each PRP was activated with calcified thromboplastin to quantify the growth factors released by them. Significantly higher (p < 0.05) transforming growth factor-β1 and vascular endothelial growth factor were released compared to the baseline. Our study highlights the variation in both force (g) and time results in changes at cellular level and growth factor concentrations. Copyright © 2016 Elsevier Ltd. All rights reserved.

  13. Release kinetics of platelet-derived and plasma-derived growth factors from autologous plasma rich in growth factors.

    Science.gov (United States)

    Anitua, Eduardo; Zalduendo, Mari Mar; Alkhraisat, Mohammad Hamdan; Orive, Gorka

    2013-10-01

    Many studies have evaluated the biological effects of platelet rich plasma reporting the final outcomes on cell and tissues. However, few studies have dealt with the kinetics of growth factor delivery by plasma rich in growth factors. Venous blood was obtained from three healthy volunteers and processed with PRGF-Endoret technology to prepare autologous plasma rich in growth factors. The gel-like fibrin scaffolds were then incubated in triplicate, in a cell culture medium to monitor the release of PDGF-AB, VEGF, HGF and IGF-I during 8 days of incubation. A leukocyte-platelet rich plasma was prepared employing the same technology and the concentrations of growth factors and interleukin-1β were determined after 24h of incubation. After each period, the medium was collected, fibrin clot was destroyed and the supernatants were stored at -80°C until analysis. The growth factor delivery is diffusion controlled with a rapid initial release by 30% of the bioactive content after 1h of incubation and a steady state release when almost 70% of the growth factor content has been delivered. Autologous fibrin matrix retained almost 30% of the amount of the growth factors after 8 days of incubation. The addition of leukocytes to the formula of platelet rich plasma did not increase the concentration of the growth factors, while it drastically increased the presence of pro-inflammatory IL-1β. Further studies employing an in vitro inflammatory model would be interesting to study the difference in growth factors and pro-inflammatory cytokines between leukocyte-free and leukocyte-rich platelet rich plasma. Copyright © 2013 Elsevier GmbH. All rights reserved.

  14. The influence of elevated levels of platelet-derived endothelial cell growth factor/thymidine phosphorylase on tumourigenicity, tumour growth, and oxygenation

    International Nuclear Information System (INIS)

    Griffiths, L.; Stratford, I.J.

    1998-01-01

    Purpose: Investigation of the effect of platelet-derived endothelial cell growth factor/thymidine phosphorylase (PD-ECGF/TP) on various aspects of tumour growth in a xenograft model, including growth rate, tumourigenicity and oxygenation levels. Methods and Materials: MDA 231 breast cancer cells overexpressing PD-ECGF/TP protein were made by retroviral transduction. These cells were grown in vitro and in vivo as xenografts. Direct measurement of tumours was used to record growth parameters, while the comet assay with the bioreductive drug RSU 1069 was used to assess tumour cell oxygenation. Results: We report that MDA 231 breast tumour cell lines expressing an increased range of levels of PD-ECGF/TP have increased tumourigenicity positively related to the level of PD-ECGF/TP when implanted in nude mice. As previously reported, tumours grown from these overexpressing cell lines grew faster than the parental line. These tumours expressed higher levels of TP activity and showed increased immunocytochemical staining for PD-ECGF. In addition, the rate of growth was found to be positively related to the level of PD-ECGF/TP expressed by the tumour cells. When the comet assay was used to compare the oxygenation status of cells between the parental and PD-ECGF/TP overexpressing tumours, the latter were found to have a larger proportion of well oxygenated cells. This is consistent with these tumours having an increased and functionally competent vascular supply in response to the expression of PD-ECGF/TP. Conclusion: PD-ECGF/TP appears to be capable of influencing tumourigenicity, angiogenesis and tumour growth in a proportional manner and can directly influence tumour oxygenation levels via its role in formation of functional vasculature

  15. Activated platelet-derived growth factor β receptor and Ras-mitogen-activated protein kinase pathway in natural bovine urinary bladder carcinomas.

    Science.gov (United States)

    Corteggio, Annunziata; Di Geronimo, Ornella; Roperto, Sante; Roperto, Franco; Borzacchiello, Giuseppe

    2012-03-01

    Bovine papillomavirus types 1 or 2 (BPV-1/2) are involved in the aetiopathogenesis of bovine urinary bladder cancer. BPV-1/2 E5 activates the platelet-derived growth factor β receptor (PDGFβR). The aim of this study was to analyse the Ras/mitogen-activated protein kinase (MAPK) pathway in relation to activation of PDGFβR in natural bovine urinary bladder carcinomas. Co-immunoprecipitation and Western blot analysis demonstrated that recruitment of growth factor receptor bound protein 2 (GRB-2) and Sos-1 to the activated PDGFβR was increased in carcinomas compared to normal tissues. Higher grade bovine urinary bladder carcinomas were associated with activation of Ras, but not with activation of downstream mitogen-activated protein kinase/extracellular signal-regulated kinase (Mek 1/2) or extracellular signal-regulated kinase (Erk 1/2). Copyright © 2011 Elsevier Ltd. All rights reserved.

  16. Post-transcriptional regulation of osteoblastic platelet-derived growth factor receptor-alpha expression by co-cultured primary endothelial cells

    DEFF Research Database (Denmark)

    Finkenzeller, Günter; Mehlhorn, Alexander T; Schmal, Hagen

    2010-01-01

    -alpha downregulation is dependent on time and cell number. This effect was specific to endothelial cells and was not observed when hOBs were co-cultured with human primary chondrocytes or fibroblasts. Likewise, HUVEC-mediated suppression of PDGFR-alpha expression was only seen in hOBs and mesenchymal stem cells......Platelet-derived growth factor receptor (PDGFR) signaling plays an important role in osteoblast function. Inhibition of PDGFR activity leads to a suppression of osteoblast proliferation, whereas mineralized matrix production is enhanced. In previous experiments, we showed that co......-cultivation of human primary endothelial cells and human primary osteoblasts (hOBs) leads to a cell contact-dependent downregulation of PDGFR-alpha expression in the osteoblasts. In this study, we investigated this effect in more detail, revealing that human umbilical vein endothelial cell (HUVEC)-mediated PDGFR...

  17. 15-Deoxy-Δ12,14-prostaglandin J2 and thiazolidinediones transactivate epidermal growth factor and platelet-derived growth factor receptors in vascular smooth muscle cells

    International Nuclear Information System (INIS)

    Ichiki, Toshihiro; Tokunou, Tomotake; Fukuyama, Kae; Iino, Naoko; Masuda, Satoko; Takeshita, Akira

    2004-01-01

    Proliferation of vascular smooth muscle cells (VSMCs) is induced by various mitogens through activation of extracellular signal-regulated protein kinase (ERK) pathway. We recently reported that peroxisome proliferator-activated receptor (PPAR)γ activators such as 15-deoxy-Δ 12,14 -prostaglandin J2 (15-d-PGJ2) and thiazolidinediones (TZDs) activated MEK/ERK pathway through phosphatidylinositol 3-kinase (PI3-K) and induced proliferation of VSMCs. However, the precise mechanisms of PPARγ activators-induced activation of PI3-K/ERK pathway have not been determined. We examined whether transactivation of growth factor receptor is involved in this process. Stimulation of VSMCs with 15-d-PGJ2 or TZDs for 15 min induced phosphorylation of ERK1/2 and Akt. 15-d-PGJ2- or TZDs-induced phosphorylation of ERK1/2 and Akt was inhibited by AG1478, an inhibitor of epidermal growth factor receptor (EGF-R) as well as AG1295, an inhibitor of platelet derived growth factor receptor (PDGF-R). 15-d-PGJ2-induced phosphorylation of both EGF-R and PDGF-R. GM6001, a matrix metalloproteinase inhibitor, and PP2, a Src family protein kinase inhibitor, suppressed 15-d-PGJ2- and TZDs-induced phosphorylation of EGF-R and PDGFβ-R as well as activation of ERK1/2 and Akt. PDGFβ-R was co-immunoprecipitated with EGF-R, regardless of the presence or absence of 15-d-PGJ2. These data suggest that 15-d-PGJ2 and TZDs activate PI3-K/ERK pathway through Src family kinase- and matrix metalloproteinase-dependent transactivation of EGF-R and PDGF-R. Both receptors seemed to associate constitutively. This novel signaling mechanisms may contribute to diverse biological functions of PPARγ activators

  18. Pretreatment with platelet derived growth factor-BB modulates the ability of costochondral resting zone chondrocytes incorporated into PLA/PGA scaffolds to form new cartilage in vivo.

    Science.gov (United States)

    Lohmann, C H; Schwartz, Z; Niederauer, G G; Carnes, D L; Dean, D D; Boyan, B D

    2000-01-01

    Optimal repair of chondral defects is likely to require both a suitable population of chondrogenic cells and a biodegradable matrix to provide a space-filling structural support during the early stages of cartilage formation. This study examined the ability of chondrocytes to support cartilage formation when incorporated into biodegradable scaffolds constructed from copolymers (PLG) of polylactic acid (PLA) and polyglycolic acid (PGA) and implanted in the calf muscle of nude mice. Scaffolds were fabricated to be more hydrophilic (PLG-H) or were reinforced with 10% PGA fibers (PLG-FR), increasing the stiffness of the implant by 20-fold. Confluent primary cultures of rat costochondral resting zone chondrocytes (RC) were loaded into PLG-H foams and implanted intramuscularly. To determine if growth factor pretreatment could modulate the ability of the cells to form new cartilage, RC cells were pretreated with recombinant human platelet derived growth factor-BB IPDGF-BB) for 4 or 24 h prior to implantation. To assess whether scaffold material properties could affect the ability of chondrogenic cells to form cartilage, RC cells were also loaded into PLG-FR scaffolds. To determine if the scaffolds or treatment with PDGF-BB affected the rate of chondrogenesis, tissue at the implant site was harvested at four and eight weeks post-operatively, fixed, decalcified and embedded in paraffin. Sections were obtained along the transverse plane of the lower leg, stained with haematoxylin and eosin, and then assessed by morphometric analysis for area of cartilage, area of residual implant, and area of fibrous connective tissue formation (fibrosis). Whether or not the cartilage contained hypertrophic cells was also assessed. The amount of residual implant did not change with time in any of the implanted tissues. The area occupied by PLG-FR implants was greater than that occupied by PLG-H implants at both time points. All implants were surrounded by fibrous connective tissue, whether

  19. Transforming growth factor β inhibits platelet derived growth factor-induced vascular smooth muscle cell proliferation via Akt-independent, Smad-mediated cyclin D1 downregulation.

    Directory of Open Access Journals (Sweden)

    Abel Martin-Garrido

    Full Text Available In adult tissue, vascular smooth muscle cells (VSMCs exist in a differentiated phenotype, which is defined by the expression of contractile proteins and lack of proliferation. After vascular injury, VSMC adopt a synthetic phenotype associated with proliferation, migration and matrix secretion. The transition between phenotypes is a consequence of the extracellular environment, and in particular, is regulated by agonists such as the pro-differentiating cytokine transforming growth factor β (TGFβ and the pro-proliferative cytokine platelet derived growth factor (PDGF. In this study, we investigated the interplay between TGFβ and PDGF with respect to their ability to regulate VSMC proliferation. Stimulation of human aortic VSMC with TGFβ completely blocked proliferation induced by all isoforms of PDGF, as measured by DNA synthesis and total cell number. Mechanistically, PDGF-induced Cyclin D1 mRNA and protein expression was inhibited by TGFβ. TGFβ had no effect on PDGF activation of its receptor and ERK1/2, but inhibited Akt activation. However, constitutively active Akt did not reverse the inhibitory effect of TGFβ on Cyclin D1 expression even though inhibition of the proteasome blocked the effect of TGFβ. siRNA against Smad4 completely reversed the inhibitory effect of TGFβ on PDGF-induced Cyclin D1 expression and restored proliferation in response to PDGF. Moreover, siRNA against KLF5 prevented Cyclin D1 upregulation by PDGF and overexpression of KLF5 partially reversed TGFβ-induced inhibition of Cyclin D1 expression. Taken together, our results demonstrate that KLF5 is required for PDGF-induced Cyclin D1 expression, which is inhibited by TGFβ via a Smad dependent mechanism, resulting in arrest of VSMCs in the G1 phase of the cell cycle.

  20. Transforming growth factor β inhibits platelet derived growth factor-induced vascular smooth muscle cell proliferation via Akt-independent, Smad-mediated cyclin D1 downregulation.

    Science.gov (United States)

    Martin-Garrido, Abel; Williams, Holly C; Lee, Minyoung; Seidel-Rogol, Bonnie; Ci, Xinpei; Dong, Jin-Tang; Lassègue, Bernard; Martín, Alejandra San; Griendling, Kathy K

    2013-01-01

    In adult tissue, vascular smooth muscle cells (VSMCs) exist in a differentiated phenotype, which is defined by the expression of contractile proteins and lack of proliferation. After vascular injury, VSMC adopt a synthetic phenotype associated with proliferation, migration and matrix secretion. The transition between phenotypes is a consequence of the extracellular environment, and in particular, is regulated by agonists such as the pro-differentiating cytokine transforming growth factor β (TGFβ) and the pro-proliferative cytokine platelet derived growth factor (PDGF). In this study, we investigated the interplay between TGFβ and PDGF with respect to their ability to regulate VSMC proliferation. Stimulation of human aortic VSMC with TGFβ completely blocked proliferation induced by all isoforms of PDGF, as measured by DNA synthesis and total cell number. Mechanistically, PDGF-induced Cyclin D1 mRNA and protein expression was inhibited by TGFβ. TGFβ had no effect on PDGF activation of its receptor and ERK1/2, but inhibited Akt activation. However, constitutively active Akt did not reverse the inhibitory effect of TGFβ on Cyclin D1 expression even though inhibition of the proteasome blocked the effect of TGFβ. siRNA against Smad4 completely reversed the inhibitory effect of TGFβ on PDGF-induced Cyclin D1 expression and restored proliferation in response to PDGF. Moreover, siRNA against KLF5 prevented Cyclin D1 upregulation by PDGF and overexpression of KLF5 partially reversed TGFβ-induced inhibition of Cyclin D1 expression. Taken together, our results demonstrate that KLF5 is required for PDGF-induced Cyclin D1 expression, which is inhibited by TGFβ via a Smad dependent mechanism, resulting in arrest of VSMCs in the G1 phase of the cell cycle.

  1. The Effects of Imatinib Mesylate on Cellular Viability, Platelet Derived Growth Factor and Stem Cell Factor in Mouse Testicular Normal Leydig Cells.

    Science.gov (United States)

    Kheradmand, Fatemeh; Hashemnia, Seyyed Mohammad Reza; Valizadeh, Nasim; Roshan-Milani, Shiva

    2016-01-01

    Growth factors play an essential role in the development of tumor and normal cells like testicular leydig cells. Treatment of cancer with anti-cancer agents like imatinib mesylate may interfere with normal leydig cell activity, growth and fertility through failure in growth factors' production or their signaling pathways. The purpose of the study was to determine cellular viability and the levels of, platelet derived growth factor (PDGF) and stem cell factor (SCF) in normal mouse leydig cells exposed to imatinib, and addressing the effect of imatinib on fertility potential. The mouse TM3 leydig cells were treated with 0 (control), 2.5, 5, 10 and 20 μM imatinib for 2, 4 and 6 days. Each experiment was repeated three times (15 experiments in each day).The cellular viability and growth factors levels were assessed by MTT and ELISA methods, respectively. For statistical analysis, one-way ANOVA with Tukey's post hoc and Kruskal-Wallis test were performed. A p-value less than 0.05 was considered statistically significant. With increasing drug concentration, cellular viability decreased significantly (pcellular viability, PDGF and SCF levels. Imatinib may reduce fertility potential especially at higher concentrations in patients treated with this drug by decreasing cellular viability. The effect of imatinib on leydig cells is associated with PDGF stimulation. Of course future studies can be helpful in exploring the long term effects of this drug.

  2. Chitosan inhibits platelet-mediated clot retraction, increases platelet-derived growth factor release, and increases residence time and bioactivity of platelet-rich plasma in vivo.

    Science.gov (United States)

    Deprés-Tremblay, Gabrielle; Chevrier, Anik; Tran-Khanh, Nicolas; Nelea, Monica; Buschmann, Michael D

    2017-11-10

    Platelet-rich plasma (PRP) has been used to treat different orthopedic conditions, however, the clinical benefits of using PRP remain uncertain. Chitosan (CS)-PRP implants have been shown to improve meniscus, rotator cuff and cartilage repair in pre-clinical models. The purpose of this current study was to investigate in vitro and in vivo mechanisms of action of CS-PRP implants. Freeze-dried formulations containing 1% (w/v) CS (80% degree of deacetylation and number average molar mass 38 kDa), 1% (w/v) trehalose as a lyoprotectant and 42.2 mM calcium chloride as a clot activator were solubilized in PRP. Gravimetric measurements and molecular/cellular imaging studies revealed that clot retraction is inhibited in CS-PRP hybrid clots through physical coating of platelets, blood cells and fibrin strands by chitosan, which interferes with platelet aggregation and platelet-mediated clot retraction. Flow cytometry and ELISA assays revealed that platelets are activated and granules secreted in CS-PRP hybrid clots and that cumulative release of platelet-derived growth factor (PDGF-AB) and epidermal growth factor is higher from CS-PRP hybrid clots compared to PRP clots in vitro. Finally, CS-PRP implants resided for up to 6 weeks in a subcutaneous implantation model and induced cell recruitment and granulation tissue synthesis, confirming greater residency and bioactivity compared to PRP in vivo.

  3. Tyrosine phosphorylation of platelet derived growth factor β receptors in coronary artery lesions: implications for vascular remodelling after directional coronary atherectomy and unstable angina pectoris

    Science.gov (United States)

    Abe, J; Deguchi, J; Takuwa, Y; Hara, K; Ikari, Y; Tamura, T; Ohno, M; Kurokawa, K

    1998-01-01

    Background—Growth factors such as platelet derived growth factor (PDGF) have been postulated to be important mediators of neointimal proliferation observed in atherosclerotic plaques and restenotic lesions following coronary interventions. Binding of PDGF to its receptor results in intrinsic receptor tyrosine kinase activation and subsequent cellular migration, proliferation, and vascular contraction.
Aims—To investigate whether the concentration of PDGF β receptor tyrosine phosphorylation obtained from directional coronary atherectomy (DCA) samples correlate with atherosclerotic plaque burden, the ability of diseased vessels to remodel, coronary risk factors, and clinical events.
Methods—DCA samples from 59 patients and 15 non-atherosclerotic left internal thoracic arteries (LITA) were analysed for PDGF β receptor tyrosine phosphorylation content by receptor immunoprecipitation and antiphosphotyrosine western blot. The amount of PDGF β receptor phosphorylation was analysed in relation to angiographic follow up data and clinical variables.
Results—PDGF β receptor tyrosine phosphorylation in the 59 DCA samples was greater than in the 15 non-atherosclerotic LITA (mean (SD) 0.84 (0.67) v 0.17 (0.08) over a control standard, p atherectomy;  restenosis PMID:9616351

  4. Local administration of platelet-derived growth factor B (PDGFB) improves follicular development and ovarian angiogenesis in a rat model of Polycystic Ovary Syndrome.

    Science.gov (United States)

    Di Pietro, Mariana; Scotti, Leopoldina; Irusta, Griselda; Tesone, Marta; Parborell, Fernanda; Abramovich, Dalhia

    2016-09-15

    Alterations in ovarian angiogenesis are common features in Polycystic Ovary Syndrome (PCOS) patients; the most studied of these alterations is the increase in vascular endothelial growth factor (VEGF) production by ovarian cells. Platelet-derived growth factor B (PDGFB) and D (PDGFD) are decreased in follicular fluid of PCOS patients and in the ovaries of a rat model of PCOS. In the present study, we aimed to analyze the effects of local administration of PDGFB on ovarian angiogenesis, follicular development and ovulation in a DHEA-induced PCOS rat model. Ovarian PDGFB administration to PCOS rats partially restored follicular development, decreased the percentage of cysts, increased the percentage of corpora lutea, and decreased the production of anti-Müllerian hormone. In addition, PDGFB administration improved ovarian angiogenesis by reversing the increase in periendothelial cell area and restoring VEGF levels. Our results shed light into the mechanisms that lead to altered ovarian function in PCOS and provide new data for potential therapeutic strategies. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  5. cDNA cloning and expression of a human platelet-derived growth factor (PDGF) receptor specific for B-chain-containing PDGF molecules

    International Nuclear Information System (INIS)

    Claesson-Welsh, L.; Eriksson, A.; Moren, A.; Severinsson, L.; Ek, B.; Ostman, A.; Betsholtz, C.; Heldin, C.H.

    1988-01-01

    The structure of the human receptor for platelet-derived growth factor (PDGF) has been deduced through cDNA cloning. A 5.45-kilobase-pair cDNA clone predicts a 1,106-amino-acid polypeptide, including the cleavable signal sequence. The overall amino acid sequence similarity with the murine PDGFR receptor is 85%. After transcription of the cDNA and translation in vitro, a PDGR receptor antiserum was used to immunoprecipitate a product of predicted size, which also could be phosphorylated in vitro. Stable introduction of the cDNA into Chinese hamster ovary (CHO) cells led to the expression of a 190-kilodalton component, which was immunoprecipitated by the PDGF receptor antiserum; this most probably represents the mature PDGF receptor. Binding assays with different /sup 125/I-labeled dimeric forms of PDGF A and B chains showed that the PDGFR receptor expressed in CHO cells bound PDGF-BB and, to a lesser extent, PDGF-AB, but not PDGF-AA

  6. UAP56 is an important mediator of Angiotensin II/platelet derived growth factor induced vascular smooth muscle cell DNA synthesis and proliferation

    International Nuclear Information System (INIS)

    Sahni, Abha; Wang, Nadan; Alexis, Jeffrey

    2013-01-01

    Highlights: ► Knockdown of UAP56 inhibits Angiotensin II/PDGF induced vascular smooth muscle cell proliferation. ► UAP56 is a positive regulator of E2F transcriptional activation. ► UAP56 is present in the vessel wall of low flow carotid arteries. -- Abstract: Angiotensin (Ang) II and platelet-derived growth factor (PDGF) are important mediators of pathologic vascular smooth muscle cell (VSMC) proliferation. Identifying downstream mediators of Ang II and PDGF signaling may provide insights for therapies to improve vascular proliferative diseases. We have previously demonstrated that breakpoint cluster region (Bcr) is an important mediator of Ang II/PDGF signaling in VSMC. We have recently reported that the DExD/H box protein UAP56 is an interacting partner of Bcr in regulating VSMC DNA synthesis. We hypothesized that UAP56 itself is an important regulator of VSMC proliferation. In this report we demonstrate that knockdown of UAP56 inhibits Ang II/PDGF induced VSMC DNA synthesis and proliferation, and inhibits E2F transcriptional activity. In addition, we demonstrate that UAP56 is present in the vessel wall of low-flow carotid arteries. These findings suggest that UAP56 is a regulator of VSMC proliferation and identify UAP56 as a target for preventing vascular proliferative disease

  7. Structure activity relationships of quinoxalin-2-one derivatives as platelet-derived growth factor-beta receptor (PDGFbeta R) inhibitors, derived from molecular modeling.

    Science.gov (United States)

    Mori, Yoshikazu; Hirokawa, Takatsugu; Aoki, Katsuyuki; Satomi, Hisanori; Takeda, Shuichi; Aburada, Masaki; Miyamoto, Ken-ichi

    2008-05-01

    We previously reported a quinoxalin-2-one compound (Compound 1) that had inhibitory activity equivalent to existing platelet-derived growth factor-beta receptor (PDGFbeta R) inhibitors. Lead optimization of Compound 1 to increase its activity and selectivity, using structural information regarding PDGFbeta R-ligand interactions, is urgently needed. Here we present models of the PDGFbeta R kinase domain complexed with quinoxalin-2-one derivatives. The models were constructed using comparative modeling, molecular dynamics (MD) and ligand docking. In particular, conformations derived from MD, and ligand binding site information presented by alpha-spheres in the pre-docking processing, allowed us to identify optimal protein structures for docking of target ligands. By carrying out molecular modeling and MD of PDGFbeta R in its inactive state, we obtained two structural models having good Compound 1 binding potentials. In order to distinguish the optimal candidate, we evaluated the structural activity relationships (SAR) between the ligand-binding free energies and inhibitory activity values (IC50 values) for available quinoxalin-2-one derivatives. Consequently, a final model with a high SAR was identified. This model included a molecular interaction between the hydrophobic pocket behind the ATP binding site and the substitution region of the quinoxalin-2-one derivatives. These findings should prove useful in lead optimization of quinoxalin-2-one derivatives as PDGFb R inhibitors.

  8. Enhancement of distribution of dermal multipotent stem cells to bone marrow in rats of total body irradiation by platelet-derived growth factor-AA treatment

    International Nuclear Information System (INIS)

    Zong Zhaowen; Ren Yongchuan; Shen Yue; Chen Yonghua; Ran Xinze; Shi Chunmeng; Cheng Tianmin

    2011-01-01

    Objective: To observe whether dermal multipotent stem cells (dMSCs) treated with platelet-derived growth factor-AA (PDGF-AA) could distribute more frequently to the bone marrow in rats of total body irradiation (TBI). Methods: Male dMSCs were isolated and 10 μg/L PDGF-AA was added to the culture medium and further cultured for 2 h. Then the expression of tenascin-C were examined by Western blot, and the migration ability of dMSCs was assessed in transwell chamber. The pre-treated dMSCs were transplanted by tail vein injection into female rats administered with total body irradiation, and 2 weeks after transplantation, real-time PCR was employed to measure the amount of dMSCs in bone marrow. Non-treated dMSCs served as control.Results PDGF-AA treatment increased the expression of tenascin-C in dMSCs, made (1.79 ± 0.13) × 10 5 cells migrate to the lower chamber under the effect of bone marrow extract, and distributed to bone marrow in TBI rats, significantly more than (1.24 ± 0.09) ×10 5 in non-treated dMSCs (t=8.833, P<0.01). Conclusions: PDGF-AA treatment could enhance the migration ability of dMSCs and increase the amount of dMSCs in bone marrow of TBI rats after transplantation. (authors)

  9. Malignant melanoma of the nasal cavity: a case report with examination of KIT and platelet derived growth factor receptor-α (PDGFRA

    Directory of Open Access Journals (Sweden)

    Tadashi Terada

    2011-10-01

    Full Text Available Although several clinicopathological studies of malignant melanoma of the nasal cavity have been reported, there are no studies of the expression and gene mutation of KIT and platelet derived growth factor receptor-α (PDGFRA in melanoma of the nasal cavity. A 92-year-old Japanese woman consulted to our hospital because of right nasal obstruction and epistaxis. Physical examination and imaging modalities showed a tumor of the right nasal cavity. A biopsy was taken, and it showed malignant epithelioid cells with melanin deposition. Immunohistochemically, the tumor was positive for S100 protein, HMB45, p53, Ki-67 (labeling=20%, KIT and PDGFRA. The tumor was negative for cytokeratins (AE1/3 and CAM5.2. A genetic analysis using PCR-direct sequencing revealed no mutation of KIT gene (exons 9, 11, 13, and 17 or the PDGFRA gene (exons 12 and 18. The pathological diagnosis was primary malignant melanoma of the nasal cavity. The tumor was reduced in size by local resection and chemotherapy (Darthmose regimen: dacarbazine, carmustine, cisplatine, and tamoxifen, and the patient is now alive and free from metastasis 9 months after the first manifestation. In conclusion, the author reported a case of melanoma of the nasal cavity expressing KIT and PDGFRA without gene mutations of KIT and PDGFRA.

  10. Depletion of intracellular calcium stores facilitates the influx of extracellular calcium in platelet derived growth factor stimulated A172 glioblastoma cells.

    Science.gov (United States)

    Vereb, G; Szöllösi, J; Mátyus, L; Balázs, M; Hyun, W C; Feuerstein, B G

    1996-05-01

    Calcium signaling in non-excitable cells is the consequence of calcium release from intracellular stores, at times followed by entry of extracellular calcium through the plasma membrane. To study whether entry of calcium depends upon the level of saturation of intracellular stores, we measured calcium channel opening in the plasma membrane of single confluent A172 glioblastoma cells stimulated with platelet derived growth factor (PDGF) and/or bradykinin (BK). We monitored the entry of extracellular calcium by measuring manganese quenching of Indo-1 fluorescence. PDGF raised intracellular calcium concentration ([Ca2+]i) after a dose-dependent delay (tdel) and then opened calcium channels after a dose-independent delay (tch). At higher doses (> 3 nM), BK increased [Ca2+]i after a tdel approximately 0 s, and tch decreased inversely with both dose and peak [Ca2+]i. Experiments with thapsigargin (TG), BK, and PDGF indicated that BK and PDGF share intracellular Ca2+ pools that are sensitive to TG. When these stores were depleted by treatment with BK and intracellular BAPTA, tdel did not change, but tch fell to almost 0 s in PDGF stimulated cells, indicating that depletion of calcium stores affects calcium channel opening in the plasma membrane. Our data support the capacitative model for calcium channel opening and the steady-state model describing quantal Ca2+ release from intracellular stores.

  11. Expression and developmental control of platelet-derived growth factor A-chain and B-chain/Sis genes in rat aortic smooth muscle cells

    International Nuclear Information System (INIS)

    Majesky, M.W.; Benditt, E.P.; Schwartz, S.M.

    1988-01-01

    Cultured arterial smooth muscle cells (SMC) can produce platelet-derived growth factor (PDGF)-like molecules. This property raises the possibility that SMC-derived PDGFs function as autocrine/paracrine regulators in the formation and maintenance of the artery wall. In this study the authors have asked if levels of mRNAs directing synthesis of PDFG are modulated in aortic SMC during postnatal development. The authors report here that genes encoding PDGF A- and B-chain precursors are expressed at similar low levels in intact aortas from newborn and adult rats. Marked differences in regulation of transcript abundance of these genes were revealed when aortic SMC were grown in cell culture. PDGF B-chain transcripts accumulated in passaged newborn rat SMC but not adult rat SMC, whereas PDGF A-chain RNA was found in comparable amounts in SMC from both age groups. Similarly, SMC from newborn rats secreted at least 60-fold more PDGF-like activity into conditioned medium than did adult rat SMC. These results show that PDGF A- and B-chain genes are transcribed in the normal rat aorta and provide evidence for age-related change in the control of PDGF B-chain gene expression in aortic SMC. Independent regulation of transcript levels in cultured SMC leaves open the possibility that PDGFs of different composition (AA, AB, BB) play different roles in normal function of the artery wall

  12. Changes in Otx2 and Parvalbumin Immunoreactivity in the Superior Colliculus in the Platelet-Derived Growth Factor Receptor-β Knockout Mice

    Directory of Open Access Journals (Sweden)

    Juanjuan Zhao

    2013-01-01

    Full Text Available The superior colliculus (SC, a relay nucleus in the subcortical visual pathways, is implicated in socioemotional behaviors. Homeoprotein Otx2 and β subunit of receptors of platelet-derived growth factor (PDGFR-β have been suggested to play an important role in development of the visual system and development and maturation of GABAergic neurons. Although PDGFR-β-knockout (KO mice displayed socio-emotional deficits associated with parvalbumin (PV-immunoreactive (IR neurons, their anatomical bases in the SC were unknown. In the present study, Otx2 and PV-immunolabeling in the adult mouse SC were investigated in the PDGFR-β KO mice. Although there were no differences in distribution patterns of Otx2 and PV-IR cells between the wild type and PDGFR-β KO mice, the mean numbers of both of the Otx2- and PV-IR cells were significantly reduced in the PDGFR-β KO mice. Furthermore, average diameters of Otx2- and PV-IR cells were significantly reduced in the PDGFR-β KO mice. These findings suggest that PDGFR-β plays a critical role in the functional development of the SC through its effects on Otx2- and PV-IR cells, provided specific roles of Otx2 protein and PV-IR cells in the development of SC neurons and visual information processing, respectively.

  13. Comparative proteome approach demonstrates that platelet-derived growth factor C and D efficiently induce proliferation while maintaining multipotency of hMSCs

    Energy Technology Data Exchange (ETDEWEB)

    Sotoca, Ana M., E-mail: a.sotoca@science.ru.nl [Department of Cell and Applied Biology, Radboud University, Heijendaalseweg 135, 6525 AJ Nijmegen (Netherlands); Roelofs-Hendriks, Jose [Department of Cell and Applied Biology, Radboud University, Heijendaalseweg 135, 6525 AJ Nijmegen (Netherlands); Boeren, Sjef [Laboratory of Biochemistry, Wageningen University, Dreijenlaan 3, 6703 HA Wageningen (Netherlands); Kraan, Peter M. van der [Department of Rheumatology Research and Advanced Therapeutics, Radboud University Nijmegen Medical Centre, Nijmegen (Netherlands); Vervoort, Jacques [Laboratory of Biochemistry, Wageningen University, Dreijenlaan 3, 6703 HA Wageningen (Netherlands); Zoelen, Everardus J.J. van; Piek, Ester [Department of Cell and Applied Biology, Radboud University, Heijendaalseweg 135, 6525 AJ Nijmegen (Netherlands)

    2013-10-15

    This is the first study that comprehensively describes the effects of the platelet-derived growth factor (PDGF) isoforms C and D during in vitro expansion of human mesenchymal stem cells (hMSCs). Our results show that PDGFs can enhance proliferation of hMSCs without affecting their multipotency. It is of great value to culture and expand hMSCs in a safe and effective manner without losing their multipotency for manipulation and further development of cell-based therapies. Moreover, differential effects of PDGF isoforms have been observed on lineage-specific differentiation induced by BMP2 and Vitamin D3. Based on label-free LC-based quantitative proteomics approach we have furthermore identified specific pathways induced by PDGFs during the proliferation process, showing the importance of bioinformatics tools to study cell function. - Highlights: • PDGFs (C and D) significantly increased the number of multipotent undifferentiated hMSCs. • Enhanced proliferation did not impair the ability to undergo lineage-specific differentiation. • Proteomic analysis confirmed the overall signatures of the ‘intact’ cells.

  14. An aptamer-based biosensing platform for highly sensitive detection of platelet-derived growth factor via enzyme-mediated direct electrochemistry.

    Science.gov (United States)

    Deng, Kun; Xiang, Yang; Zhang, Liqun; Chen, Qinghai; Fu, Weiling

    2013-01-08

    In this work, a new label-free electrochemical aptamer-based sensor (aptasensor) was constructed for detection of platelet-derived growth factor (PDGF) based on the direct electrochemistry of glucose oxidase (GOD). For this proposed aptasensor, poly(diallyldimethylammonium chloride) (PDDA)-protected graphene-gold nanoparticles (P-Gra-GNPs) composite was firstly coated on electrode surface to form the interface with biocompatibility and huge surface area for the adsorption of GOD layer. Subsequently, gold nanoclusters (GNCs) were deposited on the surface of GOD to capture PDGF binding aptamer (PBA). Finally, GOD as a blocking reagent was employed to block the remaining active sites of the GNCs and avoid the nonspecific adsorption. With the direct electron transfer of double layer GOD membranes, the aptasensor showed excellent electrochemical response and the peak current decreased linearly with increasing logarithm of PDGF concentration from 0.005 nM to 60 nM with a relatively low limit of detection of 1.7 pM. The proposed aptasensor exhibited high specificity, good reproducibility and long-term stability, which provided a new promising technique for aptamer-based protein detection. Copyright © 2012 Elsevier B.V. All rights reserved.

  15. Two signaling molecules share a phosphotyrosine-containing binding site in the platelet-derived growth factor receptor.

    OpenAIRE

    Nishimura, R; Li, W; Kashishian, A; Mondino, A; Zhou, M; Cooper, J; Schlessinger, J

    1993-01-01

    Autophosphorylation sites of growth factor receptors with tyrosine kinase activity function as specific binding sites for Src homology 2 (SH2) domains of signaling molecules. This interaction appears to be a crucial step in a mechanism by which receptor tyrosine kinases relay signals to downstream signaling pathways. Nck is a widely expressed protein consisting exclusively of SH2 and SH3 domains, the overexpression of which causes cell transformation. It has been shown that various growth fac...

  16. Platelet-derived growth factor (PDGF)-signaling mediates radiation-induced apoptosis in human prostate cancer cells with loss of p53 function

    International Nuclear Information System (INIS)

    Kim, Harold E.; Han, Sue J.; Kasza, Thomas; Han, Richard; Choi, Hyeong-Seon; Palmer, Kenneth C.; Kim, Hyeong-Reh C.

    1997-01-01

    Platelet-derived growth factor (PDGF) signals a diversity of cellular responses in vitro, including cell proliferation, survival, transformation, and chemotaxis. PDGF functions as a 'competence factor' to induce a set of early response genes expressed in G 1 including p21 WAF1/CIP1 , a functional mediator of the tumor suppressor gene p53 in G 1 /S checkpoint. For PDGF-stimulated cells to progress beyond G 1 and transit the cell cycle completely, progression factors in serum such as insulin and IGF-1 are required. We have recently shown a novel role of PDGF in inducing apoptosis in growth-arrested murine fibroblasts. The PDGF-induced apoptosis is rescued by insulin, suggesting that G 1 /S checkpoint is a critical determinant for PDGF-induced apoptosis. Because recent studies suggest that radiation-induced signal transduction pathways interact with growth factor-mediated signaling pathways, we have investigated whether activation of the PDGF-signaling facilitates the radiation-induced apoptosis in the absence of functional p53. For this study we have used the 125-IL cell line, a mutant p53-containing, highly metastatic, and hormone-unresponsive human prostate carcinoma cell line. PDGF signaling is constitutively activated by transfection with a p28 v-sis expression vector, which was previously shown to activate PDGF α- and β- receptors. Although the basal level of p21 WAF1/CIP1 expression and radiation-induced apoptosis were not detectable in control 125-IL cells as would be predicted in mutant p53-containing cells, activation of PDGF-signaling induced expression of p21 WAF1/CIP1 and radiation-induced apoptosis. Our study suggests that the level of 'competence' growth factors including PDGF may be one of the critical determinants for radiation-induced apoptosis, especially in cells with loss of p53 function at the site of radiotherapy in vivo

  17. Effect of plasma-rich in platelet-derived growth factors on peri-implant bone healing: An experimental study in canines

    Science.gov (United States)

    Birang, Reza; Torabi, Alireza; Shahabooei, Mohammad; Rismanchian, Mansour

    2012-01-01

    Background: Tissue engineering principles can be exploited to enhance alveolar and peri-implant bone reconstruction by applying such biological factors as platelet-derived growth factors. The objective of the present study is to investigate the effect of autologous plasma-rich in growth factors (on the healing of peri-implant bone in canine mandible). Materials and Methods: In this prospective experimental animal study, two healthy canines of the Iranian mix breed were selected. Three months after removing their premolar teeth on both sides of the mandible, 12 implants of the Osteo Implant Corporationsystem, 5 mm in diameter and 10 mm in length, were selected to be implanted. Plasma rich in growth factors (PRGF) were applied on six implants while the other six were used as plain implants without the plasma. The implants were installed in osteotomy sites on both sides of the mandible to be removed after 4 weeks with the surrounding bones using a trephine bur. Mesio-distal sections and implant blocks, 50 μ in diameter containing the peri-implant bone, were prepared By basic fuchin toluidine-bluefor histological and histomorphometric evaluation by optical microscope. The data were analyzed using Mann-Whitney Test (PPRGF and control groups had no statistically significant differences (P=0.261, P=0.2) although the parameters showed higher measured values in the PRGF group. However, compared to the control, application of PRGF had significantly increased bone-to-implant contact (P=0.028) Conclusion: Based on the results, it may be concluded that application of PRGF on the surface of implant may enhance bone-to-implant contact. PMID:22363370

  18. Immunohistochemical expression of platelet-derived growth factor receptors in ovarian cancer patients with long-term follow-up

    DEFF Research Database (Denmark)

    Madsen, Christine Vestergaard; Dahl Steffensen, Karina; Waldstrøm, Marianne

    2012-01-01

    relation to histopathological parameters and long-term overall survival. Methods. The immunohistochemical expression of PDGFR-α and PDGFR-β was investigated in tumor and stromal cells in 170 patients with histologically verified epithelial ovarian cancer. Results. Almost half of the tumor specimens showed......Introduction. The well-documented role of the PDGF system in tumor growth and angiogenesis has prompted the development of new biological agents targeting the PDGF system. The aim of the present study was to analyze the expression of the PDGF-receptors in ovarian cancer and to investigate its...... high expression of PDGFR-α and PDGFR-β in tumor cells (43% and 41%) and in stromal compartments (32% and 44%). There was a significant association between high expression of PDGFR-α and high expression of PDGFR-β in both tumor and stromal cells. Coexpression of PDGFR-α and PDGFR-β in stromal cells...

  19. Platelet-derived growth factor-D modulates extracellular matrix homeostasis and remodeling through TIMP-1 induction and attenuation of MMP-2 and MMP-9 gelatinase activities

    International Nuclear Information System (INIS)

    Borkham-Kamphorst, Erawan; Alexi, Pascal; Tihaa, Lidia; Haas, Ute; Weiskirchen, Ralf

    2015-01-01

    Platelet-derived growth factor-D (PDGF-D) is a more recent recognized growth factor involved in the regulation of several cellular processes, including cell proliferation, transformation, invasion, and angiogenesis by binding to and activating its cognate receptor PDGFR-β. After bile duct ligation or in the carbon tetrachloride-induced hepatic fibrosis model , PDGF-D showed upregulation comparable to PDGF-B. Moreover, adenoviral PDGF-D gene transfer induced hepatic stellate cell proliferation and liver fibrosis. We here investigated the molecular mechanism of PDGF-D involvement in liver fibrogenesis. Therefore, the GRX mouse cell line was stimulated with PDGF-D and evaluated for fibrotic markers and PDGF-D signaling pathways in comparison to the other PDGF isoforms. We found that PDGF-D failed to enhance Col I and α-smooth muscle actin (α-SMA) production but has capacity to upregulate expression of the tissue inhibitor of metalloprotease 1 (TIMP-1) resulting in attenuation of MMP-2 and MMP-9 gelatinase activity as indicated by gelatinase zymography. This phenomenon was restored through application of a PDGF-D neutralizing antibody. Unexpectedly, PDGF-D incubation decreased both PDGFR-α and -β in mRNA and protein levels, and PDGF-D phosphorylated typrosines specific for PDGFR-α and -β. We conclude that PDGF-D intensifies fibrogenesis by interfering with the fibrolytic activity of the TIMP-1/MMP system and that PDGF-D signaling is mediated through both PDGF-α and -β receptors. - Highlights: • PDGF-D signals through PDGF receptor type α and β. • PDGF-D modulates extracellular matrix homeostasis and remodeling. • Like PDGF-B, PDGF-D triggers phosphorylation of PLC-γ, Akt/PKB, JNK, ERK1/2, and p38. • PDGF-D induces TIMP-1 expression through ERK and p38 MAPK. • PDGF-D attenuates MMP-2 and MMP-9 gelatinase activities

  20. Platelet-derived growth factor BB and DD and angiopoietin1 are altered in follicular fluid from polycystic ovary syndrome patients.

    Science.gov (United States)

    Scotti, Leopoldina; Parborell, Fernanda; Irusta, Griselda; De Zuñiga, Ignacio; Bisioli, Claudio; Pettorossi, Hernan; Tesone, Marta; Abramovich, Dalhia

    2014-08-01

    Polycystic ovary syndrome (PCOS) is the most common endocrinological pathology among women of reproductive age, and is characterized by abnormalities in ovarian angiogenesis, among other features. Consistent with this association, follicular fluid (FF) concentration and ovarian expression of vascular endothelial growth factor (VEGF) are increased in PCOS patients. In this study, we examined the protein levels of platelet-derived growth factor (PDGF) BB and DD (PDGFBB and PDGFDD), angiopoietin 1 and 2 (ANGPT1 and ANGPT2), and their soluble receptor sTIE2 in FF from PCOS and control patients undergoing assisted reproductive techniques. We also analyzed the effect of FF from PCOS and control patients on tight and adherens junction protein expression in an endothelial cell line. PDGFBB and PDGFDD were significantly lower whereas ANGPT1 concentration was significantly higher in FF from PCOS patients than from control patients. No changes were found in the concentration of ANGPT2 or sTIE2. Expression of claudin-5 was significantly increased in endothelial cells incubated for 24 hr in the presence of FF from PCOS versus from control patients, while vascular-endothelial cadherin, β-catenin, and zonula occludens 1 expression were unchanged. The changes observed in the levels of PDGF isoforms and ANGPT1 may prevent VEGF-induced vascular permeability in the PCOS ovary by regulating endothelial-cell-junction protein levels. Restoring the levels of angiogenic factors may provide new insights into PCOS treatment and the prevention of ovarian hyperstimulation syndrome in affected women. © 2014 Wiley Periodicals, Inc.

  1. Activated platelet-derived growth factor autocrine pathway drives the transformed phenotype of a human glioblastoma cell line.

    Science.gov (United States)

    Vassbotn, F S; Ostman, A; Langeland, N; Holmsen, H; Westermark, B; Heldin, C H; Nistér, M

    1994-02-01

    Human glioblastoma cells (A172) were found to concomitantly express PDGF-BB and PDGF beta-receptors. The receptors were constitutively autophosphorylated in the absence of exogenous ligand, suggesting the presence of an autocrine PDGF pathway. Neutralizing PDGF antibodies as well as suramin inhibited the autonomous PDGF receptor tyrosine kinase activity and resulted in up-regulation of receptor protein. The interruption of the autocrine loop by the PDGF antibodies reversed the transformed phenotype of the glioblastoma cell, as determined by (1) diminished DNA synthesis, (2) inhibition of tumor colony growth, and (3) reversion of the transformed morphology of the tumor cells. The PDGF antibodies showed no effect on the DNA synthesis of another glioblastoma cells line (U-343MGa 31L) or on Ki-ras-transformed fibroblasts. The present study demonstrates an endogenously activated PDGF pathway in a spontaneous human glioblastoma cell line. Furthermore, we provide evidence that the autocrine PDGF pathway drives the transformed phenotype of the tumor cells, a process that can be blocked by extracellular antagonists.

  2. MiR-34a inhibits colon cancer proliferation and metastasis by inhibiting platelet-derived growth factor receptor α.

    Science.gov (United States)

    Li, Chunyan; Wang, Yulin; Lu, Shuming; Zhang, Zhuqing; Meng, Hua; Liang, Lina; Zhang, Yan; Song, Bo

    2015-11-01

    The microRNA (miRNA), miR‑34a is significant in colon cancer progression. In the present study, the role of miR‑34a in colon cancer cell proliferation and metastasis was investigated. It was found that the expression of miR‑34a in colon cancer tissues and cell lines was lower when compared with that of normal tissues and cells. Further research demonstrated that miR‑34a inhibited cell proliferation, induced G1 phase arrest, and suppressed metastasis and epithelial mesenchymal transition in colon cancer cells. Bioinformatic prediction indicated that platelet‑derived growth factor receptor α (PDGFRA) was a potential target gene of miR‑34a and a luciferase assay identified that PDGFRA was a novel direct target gene of miR‑34a. In addition, assays of western blot analyses and quantitative reverse‑transcription polymerase chain reaction confirmed that miR‑34a decreased PDGFRA mRNA expression and protein levels in colon cancer cells. Assessment of cellular function indicated that miR‑34a inhibited colon cancer progression via PDGFRA. These findings demonstrate that miR‑34a may act as a negative regulator in colon cancer by targeting PDGFRA.

  3. Neomycin is a platelet-derived growth factor (PDGF) antagonist that allows discrimination of PDGF alpha- and beta-receptor signals in cells expressing both receptor types.

    Science.gov (United States)

    Vassbotn, F S; Ostman, A; Siegbahn, A; Holmsen, H; Heldin, C H

    1992-08-05

    The aminoglycoside neomycin has recently been found to affect certain platelet-derived growth factor (PDGF) responses in C3H/10T1/2 C18 fibroblasts. Using porcine aortic endothelial cells transfected with PDGF alpha- or beta-receptors, we explored the possibility that neomycin interferes with the interaction between the different PDGF isoforms and their receptors. We found that neomycin (5 mM) inhibited the binding of 125I-PDGF-BB to the alpha-receptor with only partial effect on the binding of 125I-PDGF-AA; in contrast, the binding of 125I-PDGF-BB to the beta-receptor was not affected by the aminoglycoside. Scatchard analyses showed that neomycin (5 mM) decreased the number of binding sites for PDGF-BB on alpha-receptor-expressing cells by 87%. Together with cross-competition studies with 125I-labeled PDGF homodimers, the effect of neomycin indicates that PDGF-AA and PDGF-BB bind to both common and unique structures on the PDGF alpha-receptor. Neomycin specifically inhibited the autophosphorylation of the alpha-receptor by PDGF-BB, with less effect on the phosphorylation induced by PDGF-AA and no effect on the phosphorylation of the beta-receptor by PDGF-BB. Thus, neomycin is a PDGF isoform- and receptor-specific antagonist that provides a possibility to compare the signal transduction pathways of alpha- and beta-receptors in cells expressing both receptor types. This approach was used to show that activation of PDGF beta-receptors by PDGF-BB mediated a chemotactic response in human fibroblasts, whereas activation of alpha-receptors by the same ligand inhibited chemotaxis.

  4. Enrichment of putative pancreatic progenitor cells from mice by sorting for prominin1 (CD133) and platelet-derived growth factor receptor beta.

    Science.gov (United States)

    Hori, Yuichi; Fukumoto, Miki; Kuroda, Yoshikazu

    2008-11-01

    Success in islet transplantation-based therapies for type 1 diabetes mellitus and an extreme shortage of pancreatic islets have motivated recent efforts to develop renewable sources of islet-replacement tissue. Although pancreatic progenitor cells hold a promising potential, only a few attempts have been made at the prospective isolation of pancreatic stem/progenitor cells, because of the lack of specific markers and the development of effective cell culture methods. We found that prominin1 (also known as CD133) recognized the undifferentiated epithelial cells, whereas platelet-derived growth factor receptor beta (PDGFRbeta) was expressed on the mesenchymal cells in the mouse embryonic pancreas. We then developed an isolation method for putative stem/progenitor cells by flow cytometric cell sorting and characterized their potential for differentiation to pancreatic tissue using both in vitro and in vivo protocols. Flow cytometry and the subsequent reverse transcription-polymerase chain reaction and microarray analysis revealed pancreatic epithelial progenitor cells to be highly enriched in the prominin1(high)PDGFRbeta(-) cell population. During in vivo differentiation, these cell populations were able to differentiate into endocrine, exocrine, and ductal tissues, including the formation of an insulin-producing cell cluster. We established the prospective isolation of putative pancreatic epithelial progenitor cells by sorting for prominin1 and PDGFRbeta. Since this strategy is based on the cell surface markers common to human and rodents, these findings may lead to the development of new strategies to derive transplantable islet-replacement tissues from human pancreatic stem/progenitor cells. Disclosure of potential conflicts of interest is found at the end of this article.

  5. Ligustrazine attenuates oxidative stress-induced activation of hepatic stellate cells by interrupting platelet-derived growth factor-β receptor-mediated ERK and p38 pathways

    International Nuclear Information System (INIS)

    Zhang, Feng; Ni, Chunyan; Kong, Desong; Zhang, Xiaoping; Zhu, Xiaojing; Chen, Li; Lu, Yin; Zheng, Shizhong

    2012-01-01

    Hepatic fibrosis represents a frequent event following chronic insult to trigger wound healing reactions with accumulation of extracellular matrix (ECM) in the liver. Activation of hepatic stellate cells (HSCs) is the pivotal event during liver fibrogenesis. Compelling evidence indicates that oxidative stress is concomitant with liver fibrosis irrespective of the underlying etiology. Natural antioxidant ligustrazine exhibits potent antifibrotic activities, but the mechanisms are poorly understood. Our studies were to investigate the ligustrazine effects on HSC activation stimulated by hydrogen peroxide (H 2 O 2 ), an in vitro model mimicking the oxidative stress in liver fibrogenesis, and to elucidate the possible mechanisms. Our results demonstrated that H 2 O 2 at 5 μM significantly stimulated HSC proliferation and expression of marker genes of HSC activation; whereas ligustrazine dose-dependently suppressed proliferation and induced apoptosis in H 2 O 2 -activated HSCs, and attenuated expression of fibrotic marker genes. Mechanistic investigations revealed that ligustrazine reduced platelet-derived growth factor-β receptor (PDGF-βR) expression and blocked the phosphorylation of extracellular regulated protein kinase (ERK) and p38 kinase, two downstream effectors of PDGF-βR. Further molecular evidence suggested that ligustrazine interruption of ERK and p38 pathways was dependent on the blockade of PDGF-βR and might be involved in ligustrazine reduction of fibrotic marker gene expression under H 2 O 2 stimulation. Furthermore, ligustrazine modulated some proteins critical for HSC activation and ECM homeostasis in H 2 O 2 -stimulated HSCs. These data collectively indicated that ligustrazine could attenuate HSC activation caused by oxidative stress, providing novel insights into ligustrazine as a therapeutic option for hepatic fibrosis. Highlights: ► Ligustrazine inhibits oxidative stress-induced HSC activation. ► Ligustrazine reduces fibrotic marker genes

  6. The inhibitory effect of dexamethasone on platelet-derived growth factor-induced vascular smooth muscle cell migration through up-regulating PGC-1α expression

    International Nuclear Information System (INIS)

    Xu, Wei; Guo, Ting; Zhang, Yan; Jiang, Xiaohong; Zhang, Yongxian; Zen, Ke; Yu, Bo; Zhang, Chen-Yu

    2011-01-01

    Dexamethasone has been shown to inhibit vascular smooth muscle cell (VSMC) migration, which is required for preventing restenosis. However, the mechanism underlying effect of dexamethasone remains unknown. We have previously demonstrated that peroxisome proliferator-activated receptor gamma (PPARγ) coactivator-1 alpha (PGC-1α) can inhibit VSMC migration and proliferation. Here, we investigated the role of PGC-1α in dexamethasone-reduced VSMC migration and explored the possible mechanism. We first examined PGC-1α expression in cultured rat aortic VSMCs. The results revealed that incubation of VSMCs with dexamethasone could significantly elevate PGC-1α mRNA expression. In contrast, platelet-derived growth factor (PDGF) decreased PGC-1α expression while stimulating VSMC migration. Mechanistic study showed that suppression of PGC-1α by small interfering RNA strongly abrogated the inhibitory effect of dexamethasone on VSMC migration, whereas overexpression of PGC-1α had the opposite effect. Furthermore, an analysis of MAPK signal pathways showed that dexamethasone inhibited ERK and p38 MAPK phosphorylation in VSMCs. Overexpression of PGC-1α decreased both basal and PDGF-induced p38 MAPK phosphorylation, but it had no effect on ERK phosphorylation. Finally, inhibition of PPARγ activation by a PPARγ antagonist GW9662 abolished the suppressive effects of PGC-1α on p38 MAPK phosphorylation and VSMC migration. These effects of PGC-1α were enhanced by a PPARγ agonist troglitazone. Collectively, our data indicated for the first time that one of the anti-migrated mechanisms of dexamethasone is due to the induction of PGC-1α expression. PGC-1α suppresses PDGF-induced VSMC migration through PPARγ coactivation and, consequently, p38 MAPK inhibition.

  7. Platelet-derived growth factor receptor-β, carrying the activating mutation D849N, accelerates the establishment of B16 melanoma

    International Nuclear Information System (INIS)

    Suzuki, Shioto; Heldin, Carl-Henrik; Heuchel, Rainer Lothar

    2007-01-01

    Platelet-derived growth factor (PDGF)-BB and PDGF receptor (PDGFR)-β are mainly expressed in the developing vasculature, where PDGF-BB is produced by endothelial cells and PDGFR-β is expressed by mural cells, including pericytes. PDGF-BB is produced by most types of solid tumors, and PDGF receptor signaling participates in various processes, including autocrine stimulation of tumor cell growth, recruitment of tumor stroma fibroblasts, and stimulation of tumor angiogenesis. Furthermore, PDGF-BB-producing tumors are characterized by increased pericyte abundance and accelerated tumor growth. Thus, there is a growing interest in the development of tumor treatment strategies by blocking PDGF/PDGFR function. We have recently generated a mouse model carrying an activated PDGFR-β by replacing the highly conserved aspartic acid residue (D) 849 in the activating loop with asparagine (N). This allowed us to investigate, in an orthotopic tumor model, the role of increased stromal PDGFR-β signaling in tumor-stroma interactions. B16 melanoma cells lacking PDGFR-β expression and either mock-transfected or engineered to express PDGF-BB, were injected alone or in combination with matrigel into mice carrying the activated PDGFR-β (D849N) and into wild type mice. The tumor growth rate was followed and the vessel status of tumors, i.e. total vessel area/tumor, average vessel surface and pericyte density of vessels, was analyzed after resection. Tumors grown in mice carrying an activated PDGFR-β were established earlier than those in wild-type mice. In this early phase, the total vessel area and the average vessel surface were higher in tumors grown in mice carrying the activated PDGFR-β (D849N) compared to wild-type mice, whereas we did not find a significant difference in the number of tumor vessels and the pericyte abundance around tumor vessels between wild type and mutant mice. At later phases of tumor progression, no significant difference in tumor growth rate was

  8. Platelet-derived growth factor-D modulates extracellular matrix homeostasis and remodeling through TIMP-1 induction and attenuation of MMP-2 and MMP-9 gelatinase activities

    Energy Technology Data Exchange (ETDEWEB)

    Borkham-Kamphorst, Erawan, E-mail: ekamphorst@ukaachen.de; Alexi, Pascal; Tihaa, Lidia; Haas, Ute; Weiskirchen, Ralf, E-mail: rweiskirchen@ukaachen.de

    2015-02-13

    Platelet-derived growth factor-D (PDGF-D) is a more recent recognized growth factor involved in the regulation of several cellular processes, including cell proliferation, transformation, invasion, and angiogenesis by binding to and activating its cognate receptor PDGFR-β. After bile duct ligation or in the carbon tetrachloride-induced hepatic fibrosis model{sub ,} PDGF-D showed upregulation comparable to PDGF-B. Moreover, adenoviral PDGF-D gene transfer induced hepatic stellate cell proliferation and liver fibrosis. We here investigated the molecular mechanism of PDGF-D involvement in liver fibrogenesis. Therefore, the GRX mouse cell line was stimulated with PDGF-D and evaluated for fibrotic markers and PDGF-D signaling pathways in comparison to the other PDGF isoforms. We found that PDGF-D failed to enhance Col I and α-smooth muscle actin (α-SMA) production but has capacity to upregulate expression of the tissue inhibitor of metalloprotease 1 (TIMP-1) resulting in attenuation of MMP-2 and MMP-9 gelatinase activity as indicated by gelatinase zymography. This phenomenon was restored through application of a PDGF-D neutralizing antibody. Unexpectedly, PDGF-D incubation decreased both PDGFR-α and -β in mRNA and protein levels, and PDGF-D phosphorylated typrosines specific for PDGFR-α and -β. We conclude that PDGF-D intensifies fibrogenesis by interfering with the fibrolytic activity of the TIMP-1/MMP system and that PDGF-D signaling is mediated through both PDGF-α and -β receptors. - Highlights: • PDGF-D signals through PDGF receptor type α and β. • PDGF-D modulates extracellular matrix homeostasis and remodeling. • Like PDGF-B, PDGF-D triggers phosphorylation of PLC-γ, Akt/PKB, JNK, ERK1/2, and p38. • PDGF-D induces TIMP-1 expression through ERK and p38 MAPK. • PDGF-D attenuates MMP-2 and MMP-9 gelatinase activities.

  9. The platelet-derived growth factor signaling system in snapping turtle embryos, Chelydra serpentina: potential role in temperature-dependent sex determination and testis development.

    Science.gov (United States)

    Rhen, Turk; Jangula, Adam; Schroeder, Anthony; Woodward-Bosh, Rikki

    2009-05-01

    The platelet-derived growth factor (Pdgf) signaling system is known to play a significant role during embryonic and postnatal development of testes in mammals and birds. In contrast, genes that comprise the Pdgf system in reptiles have never been cloned or studied in any tissue, let alone developing gonads. To explore the potential role of PDGF ligands and their receptors during embryogenesis, we cloned cDNA fragments of Pdgf-A, Pdgf-B, and receptors PdgfR-alpha and PdgfR-beta in the snapping turtle, a reptile with temperature-dependent sex determination (TSD). We then compared gene expression profiles in gonads from embryos incubated at a male-producing temperature to those from embryos at a female-producing temperature, as well as between hatchling testes and ovaries. Expression of Pdgf-B mRNA in embryonic gonads was significantly higher at a male temperature than at a female temperature, but there was no difference between hatchling testes and ovaries. This developmental pattern was reversed for Pdgf-A and PdgfR-alpha mRNA: expression of these genes did not differ in embryos, but diverged in hatchling testes and ovaries. Levels of PdgfR-beta mRNA in embryonic gonads were not affected by temperature and did not differ between testes and ovaries. However, expression of both receptors increased at least an order of magnitude from the embryonic to the post-hatching period. Finally, we characterized expression of these genes in several other embryonic tissues. The brain, heart, and liver displayed unique expression patterns that distinguished these tissues from each other and from intestine, lung, and muscle. Incubation temperature had a significant effect on expression of PdgfR-alpha and PdgfR-beta in the heart but not other tissues. Together, these findings demonstrate that temperature has tissue specific effects on the Pdgf system and suggest that Pdgf signaling is involved in sex determination and the ensuing differentiation of testes in the snapping turtle.

  10. Local Application of Gelatin Hydrogel Sheets Impregnated With Platelet-Derived Growth Factor BB Promotes Tendon-to-Bone Healing After Rotator Cuff Repair in Rats.

    Science.gov (United States)

    Tokunaga, Takuya; Ide, Junji; Arimura, Hitoshi; Nakamura, Takayuki; Uehara, Yusuke; Sakamoto, Hidetoshi; Mizuta, Hiroshi

    2015-08-01

    To determine whether the local application of platelet-derived growth factor BB (PDGF-BB) in hydrogel sheets would promote healing and improve histologic characteristics and biomechanical strength after rotator cuff (RC) repair in rats. To assess the effect of PDGF-BB on tendon-to-bone healing we divided 36 adult male Sprague-Dawley rats treated with bilateral surgery to repair the supraspinatus tendon at its insertion site into 3 groups: group 1 = suture-only group; group 2 = suture and gelatin hydrogel sheets impregnated with phosphate-buffered saline (PBS); and group 3 = suture and gelatin hydrogel sheets impregnated with PDGF-BB (0.5 μg). Semiquantitative histologic evaluation was carried out 2, 6, and 12 weeks later; cell proliferation was assessed 2 and 6 weeks postoperatively by immunostaining for proliferating cell nuclear antigen (PCNA), and biomechanical testing, including ultimate load to failure, stiffness, and ultimate stress to failure, was performed 12 weeks after the operation. At 2 weeks, the average percentage of PCNA-positive cells at the insertion site was significantly higher in group 3 (40.5% ± 2.4%) than in group 1 (32.1% ± 6.9%; P = .03) and group 2 (31.9% ± 3.7%; P = .02). At 2 and 6 weeks, the histologic scores were similar among the 3 groups. At 12 weeks, the histologic score was significantly higher in group 3 (10.3 ± 0.8) than in group 1 (8.5 ± 0.5; P = .002) or group 2 (8.8 ± 0.8; P = .009), whereas ultimate load to failure, stiffness, and ultimate load to stress (normal control population, 44.73 ± 9.75 N, 27.59 ± 4.32 N/mm, and 21.33 ± 4.65 N/mm(2), respectively) were significantly higher in group 3 (28.28 ± 6.28 N, 11.05 ± 2.37 N/mm, and 7.99 ± 2.13 N/mm(2), respectively) than in group 1 (10.44 ± 1.98 N, 4.74 ± 1.31 N/mm, and 3.28 ± 1.27 N/mm(2), respectively; all P repair in humans. Copyright © 2015 Arthroscopy Association of North America. Published by Elsevier Inc. All rights reserved.

  11. Potential of pomegranate fruit extract (Punica granatum Linn.) to increase vascular endothelial growth factor and platelet-derived growth factor expressions on the post-tooth extraction wound of Cavia cobaya.

    Science.gov (United States)

    Nirwana, Intan; Rachmadi, Priyawan; Rianti, Devi

    2017-08-01

    Pomegranates fruit extracts have several activities, among others, anti-inflammatory, antibacterial, and antioxidants that have the main content punicalagin and ellagic acid. Pomegranate has the ability of various therapies through different mechanisms. Vascular endothelial growth factor (VEGF) function was to form new blood vessels produced by various cells one of them was macrophages. Platelet-derived growth factor (PDGF) was a growth factor proven chemotactic, increased fibroblast proliferation and collagen matrix production. In addition, VEGF and PDGF synergize in their ability to vascularize tissues. The PDGF function was to stabilize and regulate maturation of new blood vessels. Activities of pomegranate fruit extract were observed by measuring the increased of VEGF and PDGF expression as a marker of wound healing process. To investigate the potential of pomegranate extracts on the tooth extraction wound to increase the expression of VEGF and PDGF on the 4 th day of wound healing process. This study used 12 Cavia cobaya , which were divided into two groups, namely, the provision of 3% sodium carboxymethyl cellulose and pomegranate extract. The 12 C. cobaya would be executed on the 4 th day, the lower jaw of experimental animals was taken, decalcified about 30 days. The expression of VEGF and PDGF was examined using immunohistochemical techniques. The differences of VEGF and PDGF expression were evaluated statistically using t-test. Statistically analysis showed that there were significant differences between control and treatment groups (p<0.05). Pomegranate fruit extract administration increased VEGF and PDGF expression on post-tooth extraction wound.

  12. Mammary tumors that become independent of the type I insulin-like growth factor receptor express elevated levels of platelet-derived growth factor receptors

    Directory of Open Access Journals (Sweden)

    Campbell Craig I

    2011-11-01

    Full Text Available Abstract Background Targeted therapies are becoming an essential part of breast cancer treatment and agents targeting the type I insulin-like growth factor receptor (IGF-IR are currently being investigated in clinical trials. One of the limitations of targeted therapies is the development of resistant variants and these variants typically present with unique gene expression patterns and characteristics compared to the original tumor. Results MTB-IGFIR transgenic mice, with inducible overexpression of the IGF-IR were used to model mammary tumors that develop resistance to IGF-IR targeting agents. IGF-IR independent mammary tumors, previously shown to possess characteristics associated with EMT, were found to express elevated levels of PDGFRα and PDGFRβ. Furthermore, these receptors were shown to be inversely expressed with the IGF-IR in this model. Using cell lines derived from IGF-IR-independent mammary tumors (from MTB-IGFIR mice, it was demonstrated that PDGFRα and to a lesser extent PDGFRβ was important for cell migration and invasion as RNAi knockdown of PDGFRα alone or PDGFRα and PDGFRβ in combination, significantly decreased tumor cell migration in Boyden chamber assays and suppressed cell migration in scratch wound assays. Somewhat surprisingly, concomitant knockdown of PDGFRα and PDGFRβ resulted in a modest increase in cell proliferation and a decrease in apoptosis. Conclusion During IGF-IR independence, PDGFRs are upregulated and function to enhance tumor cell motility. These results demonstrate a novel interaction between the IGF-IR and PDGFRs and highlight an important, therapeutically relevant pathway, for tumor cell migration and invasion.

  13. Injury induces in vivo expression of platelet-derived growth factor (PDGF) and PDGF receptor mRNAs in skin epithelial cells and PDGF mRNA in connective tissue fibroblasts

    International Nuclear Information System (INIS)

    Antoniades, H.N.; Galanopoulos, T.; Neville-Golden, J.; Kiritsy, C.P.; Lynch, S.E.

    1991-01-01

    Platelet-derived growth factor (PDGF) stimulates many of the processes important in tissue repair, including proliferation of fibroblasts and synthesis of extracellular matrices. In this study, the authors have demonstrated with in situ hydridization and immunocytochemistry the reversible expression of 3-sis/PDGF-2 and PDGF receptor (PDGF-R) b mRNAs and their respective protein products in epithelial cells and fibroblasts following cutaneous injury in pigs. Epithelial cells in control, unwounded skin did not express c-sis and PDGF-R mRNAs, and fibroblasts expressed only PDGF-R mRNA. The expression levels in the injured site were correlated with the stage of tissue repair, being highest during the initial stages of the repair process and declining at the time of complete re-epithelialization and tissue remodeling. These studies provide a mulecular basis for understanding the mechanisms contributing to normal tissue repair. They suggest the possibility that a defect in these mechanisms may be associated with defective wound healing. It is also conceivable that chronic injury may induce irreversible gene expression leading to pathologic, unregulated cell growth

  14. cDNA for the human β2-adrenergic receptor: a protein with multiple membrane-spanning domains and encoded by a gene whose chromosomal location is shared with that of the receptor for platelet-derived growth factor

    International Nuclear Information System (INIS)

    Kobilka, B.K.; Dixon, R.A.F.; Frielle, T.

    1987-01-01

    The authors have isolated and sequenced a cDNA encoding the human β 2 -adrenergic receptor. The deduced amino acid sequence (413 residues) is that of a protein containing seven clusters of hydrophobic amino acids suggestive of membrane-spanning domains. While the protein is 87% identical overall with the previously cloned hamster β 2 -adrenergic receptor, the most highly conserved regions are the putative transmembrane helices (95% identical) and cytoplasmic loops (93% identical), suggesting that these regions of the molecule harbor important functional domains. Several of the transmembrane helices also share lesser degrees of identity with comparable regions of select members of the opsin family of visual pigments. They have localized the gene for the β 2 -adrenergic receptor to q31-q32 on chromosome 5. This is the same position recently determined for the gene encoding the receptor for platelet-derived growth factor and is adjacent to that for the FMS protooncogene, which encodes the receptor for the macrophage colony-stimulating factor

  15. The C-terminal SH2 domain of p85 accounts for the high affinity and specificity of the binding of phosphatidylinositol 3-kinase to phosphorylated platelet-derived growth factor beta receptor.

    Science.gov (United States)

    Klippel, A; Escobedo, J A; Fantl, W J; Williams, L T

    1992-01-01

    Upon stimulation by its ligand, the platelet-derived growth factor (PDGF) receptor associates with the 85-kDa subunit of phosphatidylinositol (PI) 3-kinase. The 85-kDa protein (p85) contains two Src homology 2 (SH2) domains and one SH3 domain. To define the part of p85 that interacts with the PDGF receptor, a series of truncated p85 mutants was analyzed for association with immobilized PDGF receptor in vitro. We found that a fragment of p85 that contains a single Src homology domain, the C-terminal SH2 domain (SH2-C), was sufficient for directing the high-affinity interaction with the receptor. Half-maximal binding of SH2-C to the receptor was observed at an SH2-C concentration of 0.06 nM. SH2-C, like full-length p85, was able to distinguish between wild-type PDGF receptor and a mutant receptor lacking the PI 3-kinase binding site. An excess of SH2-C blocked binding of full-length p85 and PI 3-kinase to the receptor but did not interfere with the binding of two other SH2-containing proteins, phospholipase C-gamma and GTPase-activating protein. These results demonstrate that a region of p85 containing a single SH2 domain accounts both for the high affinity and specificity of binding of PI 3-kinase to the PDGF receptor. Images PMID:1312663

  16. Analysis of receptor signaling pathways by mass spectrometry: identification of vav-2 as a substrate of the epidermal and platelet-derived growth factor receptors

    DEFF Research Database (Denmark)

    Pandey, A; Podtelejnikov, A V; Blagoev, B

    2000-01-01

    Oligomerization of receptor protein tyrosine kinases such as the epidermal growth factor receptor (EGFR) by their cognate ligands leads to activation of the receptor. Transphosphorylation of the receptor subunits is followed by the recruitment of signaling molecules containing src homology 2 (SH2...

  17. Overexpression of microRNA-375 impedes platelet-derived growth factor-induced proliferation and migration of human fetal airway smooth muscle cells by targeting Janus kinase 2.

    Science.gov (United States)

    Ji, Yamei; Yang, Xin; Su, Huixia

    2018-02-01

    The abnormal proliferation and migration of airway smooth muscle (ASM) cells play a critical role in airway remodeling during the development of asthma. MicroRNAs (miRNAs) have emerged as critical regulators of ASM cell proliferation and migration in airway remodeling. In this study, we aimed to investigate the potential role of miR-375 in the regulation of platelet-derived growth factor (PDGF)-induced fetal ASM cell proliferation and migration. Our results showed that miR-375 expression was significantly decreased in fetal ASM cells that were treated with PDGF. Functional data showed that overexpression of miR-375 inhibited the proliferation and migration of fetal ASM cells, whereas inhibition of miR-375 enhanced the proliferation and migration of fetal ASM cells. The results of bioinformatics analysis and a dual-luciferase reporter assay showed that miR-375 binds directly to the 3'-untranslated region of Janus kinase 2 (JAK2). Further data confirmed that miR-375 negatively regulates the expression of JAK2 in fetal ASM cells. Moreover, miR-375 also impeded the PDGF-induced activation of signal transducer and activator of transcription 3 (STAT3) in fetal ASM cells. However, restoration of JAK2 expression partially reversed the inhibitory effect of miR-375 on fetal ASM cell proliferation and migration. Overall, our results demonstrate that miR-375 inhibits fetal ASM cell proliferation and migration by targeting JAK2/STAT3 signaling. Our study provides a potential therapeutic target for the development of novel treatment strategies for pediatric asthma. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  18. MicroRNA-20b-5p inhibits platelet-derived growth factor-induced proliferation of human fetal airway smooth muscle cells by targeting signal transducer and activator of transcription 3.

    Science.gov (United States)

    Tang, Jin; Luo, Lingying

    2018-06-01

    Pediatric asthma is still a health threat to the pediatric population in recent years. The airway remodeling induced by abnormal airway smooth muscle (ASM) cell proliferation is an important cause of asthma. MicroRNAs (miRNAs) are important regulators of ASM cell proliferation. Numerous studies have reported that miR-20b-5p is a critical regulator for cell proliferation. However, whether miR-20b-5p is involved in regulating ASM cell proliferation remains unknown. In this study, we aimed to investigate the potential role of miR-20b-5p in regulating the proliferation of fetal ASM cell induced by platelet-derived growth factor (PDGF). Here, we showed that miR-20b-5p was significantly decreased in fetal ASM cells treated with PDGF. Biological experiments showed that the overexpression of miR-20b-5p inhibited the proliferation while miR-20b-5p inhibition markedly promoted the proliferation of fetal ASM cells. Bioinformatics analysis and luciferase reporter assay showed that miR-20b-5p directly targeted the 3'-UTR of signal transducer and activator of transcription 3 (STAT3). Further data showed that miR-20b-5p negatively regulated the expression of STAT3 in fetal ASM cells. Moreover, miR-20b-5p regulates the transcriptional activity of STAT3 in fetal ASM cells. Overexpression of STAT3 reversed the inhibitory effect of miR-20b-5p overexpression on fetal ASM cell proliferation while the knockdown of STAT3 abrogated the promoted effect of miR-20b-5p inhibition on fetal ASM cell proliferation. Overall, our results show that miR-20b-5p impedes PDGF-induced proliferation of fetal ASM cells through targeting STAT3. Our study suggests that miR-20b-5p may play an important role in airway remodeling during asthma and suggests that miR-20b-5p may serve as a potential therapeutic target for pediatric asthma. Copyright © 2018 Elsevier Masson SAS. All rights reserved.

  19. Platelet-Derived Microvesicles in Cardiovascular Diseases

    Directory of Open Access Journals (Sweden)

    Maria T. K. Zaldivia

    2017-11-01

    Full Text Available Microvesicles (MVs circulating in the blood are small vesicles (100–1,000 nm in diameter derived from membrane blebs of cells such as activated platelets, endothelial cells, and leukocytes. A growing body of evidence now supports the concept that platelet-derived microvesicles (PMVs, the most abundant MVs in the circulation, are important regulators of hemostasis, inflammation, and angiogenesis. Compared with healthy individuals, a large increase of circulating PMVs has been observed, particularly in patients with cardiovascular diseases. As observed in MVs from other parent cells, PMVs exert their biological effects in multiple ways, such as triggering various intercellular signaling cascades and by participating in transcellular communication by the transfer of their “cargo” of cytoplasmic components and surface receptors to other cell types. This review describes our current understanding of the potential role of PMVs in mediating hemostasis, inflammation, and angiogenesis and their consequences on the pathogenesis of cardiovascular diseases, such as atherosclerosis, myocardial infarction, and venous thrombosis. Furthermore, new developments of the therapeutic potential of PMVs for the treatment of cardiovascular diseases will be discussed.

  20. The influence of platelet- derived products on angiogenesis and tissue repair: a concise update

    Directory of Open Access Journals (Sweden)

    Constanza E Martínez

    2015-10-01

    Full Text Available Platelet degranulation allows the release of a large amount of soluble mediators, is an essential step for wound healing initiation, and stimulates clotting and angiogenesis. The latter process is one of the most critical biological events observed during tissue repair,increasing the growth of blood vessels in the maturing wound. Angiogenesis requires the action of a variety of growth factors that act in an appropriate physiological ratio to assure functional blood vessel restoration. Platelets release main regulators of angiogenesis: Vascular Endothelial Growth Factors (VEGFs, basic fibroblast growth factor (FGF-2, and Platelet derived growth factors (PDGFs, among others. In order to stimulate tissue repair, platelet derived fractions have been used as an autologous source of growth factors and biomolecules, namely Platelet Rich Plasma (PRP, Platelet Poor Plasma (PPP and Platelet Rich Fibrin(PRF. The continuous release of these growth factors has been proposed to promote angiogenesis both in vitro and in vivo. Considering the existence of clinical trials currently evaluating the efficacy of autologous PRP, the present review analyses fundamental questions regarding the putative role of platelet derived fractions as regulators of angiogenesis and evaluates the possible clinical implications of these formulations.

  1. Incorporation of Human-Platelet-Derived Growth Factor-BB Encapsulated Poly(lactic-co-glycolic acid) Microspheres into 3D CORAGRAF Enhances Osteogenic Differentiation of Mesenchymal Stromal Cells

    DEFF Research Database (Denmark)

    Mohan, Saktiswaren; Raghavendran, Hanumantharao Balaji; Karunanithi, Puvanan

    2017-01-01

    Tissue engineering aims to generate or facilitate regrowth or healing of damaged tissues by applying a combination of biomaterials, cells, and bioactive signaling molecules. In this regard, growth factors clearly play important roles in regulating cellular fate. However, uncontrolled release...... was noted to support rapid cell expansion and differentiation of stromal cells into osteogenic cells in vitro for bone tissue engineering applications....

  2. Association assessment of platelet derived growth factor B gene ...

    African Journals Online (AJOL)

    Shayesteh Rezayani

    2017-04-21

    Apr 21, 2017 ... susceptibility and environmental risk factors and their interactions. [1] and starts .... Germany) as internal control, and 30 lM of each specific primer. (Eurofins .... thank the Arya Tina Gene company for recruiting study subjects.

  3. Platelet-derived growth factor mediates interleukin-13-induced ...

    Indian Academy of Sciences (India)

    2014-07-01

    4), September 2014, 693–700, * Indian Academy of Sciences. 693. Keywords. ... ence, France) were dissolved in 100% dimethyl sulfox- ide (DMSO) at ..... of China (81070045), the Key Clinical Project for Affiliated. Hospital of ...

  4. Using growth factors in human extraction sockets: a histologic and histomorphometric evaluation of short-term healing

    NARCIS (Netherlands)

    Geurs, N.; Ntounis, A.; Vassilopoulos, P.; van der Velden, U.; Loos, B.G.; Reddy, M.

    2014-01-01

    Purpose: Ridge preservation protocols reduce crestal remodeling after tooth extraction. There is insufficient evidence on bone grafting in combination with platelet-rich plasma (PRP) or recombinant human platelet-derived growth factor (rhPDGF-BB). The aim of this study is to evaluate healing of

  5. Platelet-derived microparticles regulates thrombin generation via phophatidylserine in abdominal sepsis.

    Science.gov (United States)

    Wang, Yongzhi; Zhang, Su; Luo, Lingtao; Norström, Eva; Braun, Oscar Ö; Mörgelin, Matthias; Thorlacius, Henrik

    2018-02-01

    Sepsis is associated with dysfunctional coagulation. Recent data suggest that platelets play a role in sepsis by promoting neutrophil accumulation. Herein, we show that cecal ligation and puncture (CLP) triggered systemic inflammation, which is characterized by formation of IL-6 and CXC chemokines as well as neutrophil accumulation in the lung. Platelet depletion decreased neutrophil accumulation, IL-6, and CXC chemokines formation in septic lungs. Depletion of platelets increased peak thrombin formation and total thrombin generation (TG) in plasma from septic animals. CLP elevated circulating levels of platelet-derived microparticles (PMPs). In vitro generated PMPs were a potent inducer of TG. Interestingly, in vitro wild-type recombinant annexin V abolished PMP-induced thrombin formation whereas a mutant annexin V protein, which does not bind to phosphatidylserine (PS), had no effect. Administration of wild-type, but not mutant annexin V, significantly inhibited thrombin formation in septic animals. Moreover, CLP-induced formation of thrombin-antithrombin complexes were reduced in platelet-depleted mice and in animals pretreated with annexin V. PMP-induced TG attenuated in FXII- and FVII-deficient plasma. These findings suggest that sepsis-induced TG is dependent on platelets. Moreover, PMPs formed in sepsis are a potent inducer of TG via PS exposure, and activation of both the intrinsic and extrinsic pathway of coagulation. In conclusion, these observations suggest that PMPs and PS play an important role in dysfunctional coagulation in abdominal sepsis. © 2017 Wiley Periodicals, Inc.

  6. Binding properties of beetal recombinant caprine growth hormone to ...

    African Journals Online (AJOL)

    SAM

    2014-07-23

    Jul 23, 2014 ... The aim of the study was to illustrate the radio-receptor assay of beetal recombinant caprine growth hormone (rcGH) ... interaction with microsomal membrane that shall be beneficial to study hormone receptor interactions of other Bovidae .... adding 1 ml of ice cold assay buffer, followed by 1 ml of 25% (w/v).

  7. Binding properties of beetal recombinant caprine growth hormone to ...

    African Journals Online (AJOL)

    The aim of the study was to illustrate the radio-receptor assay of beetal recombinant caprine growth hormone (rcGH). Tracer (125I-rcGH) was prepared by iodinating beetal rcGH with iodine-125 and its biological activity was analyzed by rabbit anti-rcGH antibodies. Liver microsomal membranes of the Bovidae species ...

  8. Recombinant-derived chicken growth hormone used for radioimmunoassay

    International Nuclear Information System (INIS)

    Proudman, J.A.

    1984-01-01

    The use of recombinant-derived chicken growth hormone (rcGH) in an avian growth hormone (GH) radioimmunoassay (RIA) procedure is described. Antiserum to turkey GH bound 125 I-labeled rcGH, and unlabeled rcGH or turkey GH displaced binding in a dose-related manner. The dose-response curves of sera and pituitary extract from chickens and turkeys were parallel to the rcGH standard curve. Sera from hypophysectomized (hypox) chickens and turkeys produced no dose-response and did not inhibit binding of labeled rcGH. Recovery of rcGH added to hypox sera was quantitative. Modification of the homologous turkey GH RIA protocol of Proudman and Wentworth (1) to use rcGH made possible either an increase in assay sensitivity or a 3-day reduction in incubation time

  9. Recombiner

    International Nuclear Information System (INIS)

    Kikuchi, Nobuo.

    1983-01-01

    Purpose: To shorten the pre-heating time for a recombiner and obtain a uniform temperature distribution for the charged catalyst layer in a BWR type reactor. Constitution: A pre-heating heater is disposed to the outer periphery of a vessel for a recombiner packed with catalysts for recombining hydrogen and oxygen in gases flowing through a radioactive gaseous wastes processing system. Heat pipes for transmitting the heat applied to said container to the catalyst are disposed vertically and horizontally within the container. Different length of the heat pipes are combined. In this way, pre-heating time for the recombiner before the operation start and before the system switching can be shortened and the uniform pre-heating for the inside of the recombiner is also made possible. Further, heater control in the pre-heating can be carried out effectively and with ease. (Moriyama, K.)

  10. Effect of cigarette smoke on monocyte procoagulant activity: Focus on platelet-derived brain-derived neurotrophic factor (BDNF).

    Science.gov (United States)

    Amadio, Patrizia; Baldassarre, Damiano; Sandrini, Leonardo; Weksler, Babette B; Tremoli, Elena; Barbieri, Silvia S

    2017-01-01

    Cigarette smoke (CS) activates platelets, promotes vascular dysfunction, and enhances Tissue Factor (TF) expression in blood monocytes favoring pro-thrombotic states. Brain-derived neurotrophic factor (BDNF), a member of the family of neurotrophins involved in survival, growth, and maturation of neurons, is released by activated platelets (APLTs) and plays a role in the cardiovascular system. The effect of CS on circulating levels of BDNF is controversial and the function of circulating BDNF in atherothrombosis is not fully understood. Here, we have shown that human platelets, treated with an aqueous extract of CS (CSE), released BDNF in a dose-dependent manner. In addition, incubation of human monocytes with BDNF or with the supernatant of platelets activated with CSE increased TF activity by a Tropomyosin receptor kinase B (TrkB)-dependent mechanism. Finally, comparing serum and plasma samples of 12 male never smokers (NS) and 29 male active smokers (AS) we observed a significant increase in microparticle-associated TF activity (MP-TF) as well as BDNF in AS, while in serum, BDNF behaved oppositely. Taken together these findings suggest that platelet-derived BDNF is involved in the regulation of TF activity and that CS plays a role in this pathway by favoring a pro-atherothrombotic state.

  11. Short-term effect of recombinant human growth hormone in patients with alcoholic cirrhosis

    DEFF Research Database (Denmark)

    Møller, S; Becker, U; Grønbaek, M

    1994-01-01

    As growth hormone possesses anabolic properties that are active on protein metabolism, and thus of potential benefit to patients with chronic liver disease, we determined the metabolic effects of recombinant human growth hormone on insulin-like growth factor-I (IGF-I) its specific binding proteins......, and liver function. Twenty consecutive patients with cirrhosis were randomized to recombinant human growth hormone (Norditropin, 4 I.U. twice daily) subcutaneously for 6 weeks (n = 10) or conventional medical treatment (n = 10). The serum concentrations of insulin-like growth factor-I in the recombinant...... patients as well as in controls, whereas no change in insulin-like growth factor binding protein-1 concentrations was found. No significant changes were seen in the area under the curve for biochemical liver function tests. We conclude that administration of recombinant human growth hormone induces...

  12. Recombiner

    International Nuclear Information System (INIS)

    Osumi, Morimichi.

    1979-01-01

    Purpose: To provide a recombiner which is capable of converting hydrogen gas into water by use of high-frequency heating at comparatively low temperatures and is safe and cheap in cost. Constitution: Hydrogen gas is introduced from an outer pipeline to the main structure of a recombiner, and when it passes through the vicinity of the central part of the recombiner, it is reacted with copper oxide (CuO 2 ) heated to a temperature more than 300 0 C by a high-frequency heater, and converted gently into water by reduction operation (2H 2 + CuO 2 → Cu + 2H 2 O). The thus prepared water is exhausted through the outer pipeline to a suppression pool. A part of hydrogen gas which has not been converted completely into water by the reaction and is remaining as hydrogen is recovered through exhaust nozzles and again introduced into the main structure of the recombiner. (Yoshino, Y.)

  13. Platelet-Derived MRP-14 Induces Monocyte Activation in Patients With Symptomatic Peripheral Artery Disease.

    Science.gov (United States)

    Dann, Rebecca; Hadi, Tarik; Montenont, Emilie; Boytard, Ludovic; Alebrahim, Dornaszadat; Feinstein, Jordyn; Allen, Nicole; Simon, Russell; Barone, Krista; Uryu, Kunihiro; Guo, Yu; Rockman, Caron; Ramkhelawon, Bhama; Berger, Jeffrey S

    2018-01-02

    Peripheral artery disease (PAD), a diffuse manifestation of atherothrombosis, is a major cardiovascular threat. Although platelets are primary mediators of atherothrombosis, their role in the pathogenesis of PAD remains unclear. The authors sought to investigate the role of platelets in a cohort of symptomatic PAD. The authors profiled platelet activity, mRNA, and effector roles in patients with symptomatic PAD and in healthy controls. Patients with PAD and carotid artery stenosis were recruited into ongoing studies (NCT02106429 and NCT01897103) investigating platelet activity, platelet RNA, and cardiovascular disease. Platelet RNA sequence profiling mapped a robust up-regulation of myeloid-related protein (MRP)-14 mRNA, a potent calcium binding protein heterodimer, in PAD. Circulating activated platelets were enriched with MRP-14 protein, which augmented the expression of the adhesion mediator, P-selectin, thereby promoting monocyte-platelet aggregates. Electron microscopy confirmed the firm interaction of platelets with monocytes in vitro and colocalization of macrophages with MRP-14 confirmed their cross talk in atherosclerotic manifestations of PAD in vivo. Platelet-derived MRP-14 was channeled to monocytes, thereby fueling their expression of key PAD lesional hallmarks and increasing their directed locomotion, which were both suppressed in the presence of antibody-mediated blockade. Circulating MRP-14 was heightened in the setting of PAD, significantly correlated with PAD severity, and was associated with incident limb events. The authors identified a heightened platelet activity profile and unraveled a novel immunomodulatory effector role of platelet-derived MRP-14 in reprograming monocyte activation in symptomatic PAD. (Platelet Activity in Vascular Surgery and Cardiovascular Events [PACE]; NCT02106429; and Platelet Activity in Vascular Surgery for Thrombosis and Bleeding [PIVOTAL]; NCT01897103). Copyright © 2018 American College of Cardiology Foundation

  14. Short-term effect of recombinant human growth hormone in patients with alcoholic cirrhosis

    DEFF Research Database (Denmark)

    Møller, S; Becker, U; Grønbaek, M

    1994-01-01

    As growth hormone possesses anabolic properties that are active on protein metabolism, and thus of potential benefit to patients with chronic liver disease, we determined the metabolic effects of recombinant human growth hormone on insulin-like growth factor-I (IGF-I) its specific binding proteins...

  15. Recombiner

    International Nuclear Information System (INIS)

    Saalfrank, H.

    1985-01-01

    Air containing hydrogen can be oxidized by heating in a container called a recombiner, in order to avoid the collection of hydrogen. The container is long and a large number of straight heating bars are arranged in parallel in it and they are flanged to a lid. The heating bars are surrounded by tubes, in order to obtain good heat transfer by a narrow annular gap. (orig.) [de

  16. Cortical bone growth and maturational changes in dwarf rats induced by recombinant human growth hormone

    Science.gov (United States)

    Martinez, D. A.; Orth, M. W.; Carr, K. E.; Vanderby, R. Jr; Vailas, A. C.

    1996-01-01

    The growth hormone (GH)-deficient dwarf rat was used to investigate recombinant human (rh) GH-induced bone formation and to determine whether rhGH facilitates simultaneous increases in bone formation and bone maturation during rapid growth. Twenty dwarf rats, 37 days of age, were randomly assigned to dwarf plus rhGH (GH; n = 10) and dwarf plus vehicle (n = 10) groups. The GH group received 1.25 mg rhGH/kg body wt two times daily for 14 days. Biochemical, morphological, and X-ray diffraction measurements were performed on the femur middiaphysis. rhGH stimulated new bone growth in the GH group, as demonstrated by significant increases (P < 0.05) in longitudinal bone length (6%), middiaphyseal cross-sectional area (20%), and the amount of newly accreted bone collagen (28%) in the total pool of middiaphyseal bone collagen. Cortical bone density, mean hydroxyapatite crystal size, and the calcium and collagen contents (microgram/mm3) were significantly smaller in the GH group (P < 0.05). Our findings suggest that the processes regulating new collagen accretion, bone collagen maturation, and mean hydroxyapatite crystal size may be independently regulated during rapid growth.

  17. Cortical bone growth and maturational changes in dwarf rats induced by recombinant human growth hormone

    Science.gov (United States)

    Martinez, D. A.; Orth, M. W.; Carr, K. E.; Vanderby, R. Jr; Vailas, A. C.

    1996-01-01

    The growth hormone (GH)-deficient dwarf rat was used to investigate recombinant human (rh) GH-induced bone formation and to determine whether rhGH facilitates simultaneous increases in bone formation and bone maturation during rapid growth. Twenty dwarf rats, 37 days of age, were randomly assigned to dwarf plus rhGH (GH; n = 10) and dwarf plus vehicle (n = 10) groups. The GH group received 1.25 mg rhGH/kg body wt two times daily for 14 days. Biochemical, morphological, and X-ray diffraction measurements were performed on the femur middiaphysis. rhGH stimulated new bone growth in the GH group, as demonstrated by significant increases (P bone length (6%), middiaphyseal cross-sectional area (20%), and the amount of newly accreted bone collagen (28%) in the total pool of middiaphyseal bone collagen. Cortical bone density, mean hydroxyapatite crystal size, and the calcium and collagen contents (microgram/mm3) were significantly smaller in the GH group (P bone collagen maturation, and mean hydroxyapatite crystal size may be independently regulated during rapid growth.

  18. The Role of Platelet-Derived Growth Factor C and Its Splice Variant in Breast Cancer

    Science.gov (United States)

    2012-02-01

    epithelial-mesenchymal transition leading instead to apoptosis (8). Furthermore, inhibition of PDGFR signaling by the PDGFR inhibitor STI571...PDGFRs are expressed in many different cell types including endothelial, epithelial and neural cells and are necessary for embryological development (as

  19. Graves’ Ophthalmopathy: A Comprehensive Role For Platelet-Derived Growth Factors

    NARCIS (Netherlands)

    L. van Steensel (Leendert)

    2012-01-01

    textabstractAutoimmune thyroid disorders include Graves’ disease (GD), which leads to hyperthyroidism, and Hashimoto’s thyroiditis, which leads to hypothyroidism. The incidence of autoimmune hypothyroidism is around five times higher than the incidence of autoimmune hyperthyroidism, but together

  20. Platelet-Derived Short-Chain Polyphosphates Enhance the Inactivation of Tissue Factor Pathway Inhibitor by Activated Coagulation Factor XI.

    Directory of Open Access Journals (Sweden)

    Cristina Puy

    Full Text Available Factor (F XI supports both normal human hemostasis and pathological thrombosis. Activated FXI (FXIa promotes thrombin generation by enzymatic activation of FXI, FIX, FX, and FV, and inactivation of alpha tissue factor pathway inhibitor (TFPIα, in vitro. Some of these reactions are now known to be enhanced by short-chain polyphosphates (SCP derived from activated platelets. These SCPs act as a cofactor for the activation of FXI and FV by thrombin and FXIa, respectively. Since SCPs have been shown to inhibit the anticoagulant function of TFPIα, we herein investigated whether SCPs could serve as cofactors for the proteolytic inactivation of TFPIα by FXIa, further promoting the efficiency of the extrinsic pathway of coagulation to generate thrombin.Purified soluble SCP was prepared by size-fractionation of sodium polyphosphate. TFPIα proteolysis was analyzed by western blot. TFPIα activity was measured as inhibition of FX activation and activity in coagulation and chromogenic assays. SCPs significantly accelerated the rate of inactivation of TFPIα by FXIa in both purified systems and in recalcified plasma. Moreover, platelet-derived SCP accelerated the rate of inactivation of platelet-derived TFPIα by FXIa. TFPIα activity was not affected by SCP in recalcified FXI-depleted plasma.Our data suggest that SCP is a cofactor for TFPIα inactivation by FXIa, thus, expanding the range of hemostatic FXIa substrates that may be affected by the cofactor functions of platelet-derived SCP.

  1. Apheresis platelet concentrates contain platelet-derived and endothelial cell-derived microparticles.

    Science.gov (United States)

    Rank, A; Nieuwland, R; Liebhardt, S; Iberer, M; Grützner, S; Toth, B; Pihusch, R

    2011-02-01

    Microparticles (MP) are membrane vesicles with thrombogenic and immunomodulatory properties. We determined MP subgroups from resting platelets, activated platelets and endothelial cells in donors and apheresis platelet concentrates (PC). MP were double stained with annexin V and CD61 (platelet-derived MP; PMP), P-selectin or CD63 (MP from activated platelets) and CD144 plus E-selectin (endothelial cell-derived MP; EMP) and detected by flow cytometry in platelet donors (n=36) and apheresis PC (n=11; Trima™). PC contained MP, mainly from resting platelets [93% (90-95)], and minor fractions of PMP from activated platelets [P-selectin(+) or CD63(+); 4·8% (3·2-7·7) and 2·6% (2·0-4·0)]. Compared to donors, levels of annexin V+ MP, PMP, P-selectin(+) and CD63(+) MP were 1·7-, 2·3-, 8·6- and 3·1-fold higher in PC (all P<0·05). During storage (1-5 days), levels of annexin V+ MP and PMP did not increase, although small increases in the fraction of P-selectin(+) or CD63(+) MP occurred (both P<0·05). PC also contained EMP, which were 2·6- to 3·7-fold enriched in PC compared to donors (P<0·05). Transfusion of apheresis PC also results in transfusion of HLA-carrying PMP and EMP. This might counteract the aim of reducing transfused HLA load by leucodepletion. The increases in PMP exposing P-selectin or CD63 reflect mild platelet activation during storage. We conclude that in leucodepleted platelet apheresis using fluidized particle bed technology, MP are harvested mainly from the donor by apheresis. Improvement in apheresis technology might reduce MP load. © 2010 The Author(s). Vox Sanguinis © 2010 International Society of Blood Transfusion.

  2. Intraarticular Sprifermin (Recombinant Human Fibroblast Growth Factor 18) in Knee Osteoarthritis

    DEFF Research Database (Denmark)

    Lohmander, L. S.; Hellot, S.; Dreher, D.

    2014-01-01

    Objective. To evaluate the efficacy and safety of intraarticular sprifermin (recombinant human fibroblast growth factor 18) in the treatment of symptomatic knee osteoarthritis (OA). Methods. The study was a randomized, double-blind, placebo-controlled, proof-of-concept trial. Intraarticular sprif...

  3. Short-term effects of recombinant human growth hormone and feeding on gluconeogenesis in humans

    Science.gov (United States)

    After a short-term fast, lactating women have increased rates of glucose production but not gluconeogenesis (GNG) despite relative hypoinsulinemia. We explored the effects of non-insulin-dependent increase in glucose utilization and recombinant human growth hormone (rhGH) on glucose production, glyc...

  4. A Child with Local Lipohypertrophy following Recombinant Human Growth Hormone Administration

    NARCIS (Netherlands)

    Koppen, Ilan J. N.; Bakx, Roel; de Kruiff, Chris C.; van Trotsenburg, A. S. Paul

    2016-01-01

    Local lipohypertrophy due to recombinant human growth hormone (rhGH) administration is a rare phenomenon. Here, we report a case of an 11-year-old girl who presented with a paraumbilical swelling, approximately one year after the start of rhGH treatment for short stature due to the presumed

  5. Preservation of Biological Activity of Plasma and Platelet-Derived Eye Drops After Their Different Time and Temperature Conditions of Storage.

    Science.gov (United States)

    Anitua, Eduardo; de la Fuente, María; Riestra, Ana; Merayo-Lloves, Jesús; Muruzábal, Francisco; Orive, Gorka

    2015-09-01

    To analyze whether plasma rich in growth factors (PRGF) eye drops preserve their biological characteristics and activity after storage for 3 and 6 months at -20°C, at 4°C, and at room temperature for 72 hours, compared with fresh samples (t0). Blood from 6 healthy donors was harvested and centrifuged to obtain PRGF free of leukocytes. Resulting PRGF eye drops were stored for 3 and 6 months at -20°C. At each time, 2 aliquots were maintained at room temperature or at 4°C for 72 hours. Platelet-derived growth factor-AB, transforming growth factor-β1, vascular endothelial growth factor, epidermal growth factor, insulin-like growth factor-1, angiopoietin-1, and thrombospondin-1 were quantified at each time and temperature of storage. Also, the effect of PRGF eye drops on proliferation of primary human keratocytes was evaluated. All the analyzed growth factor levels remained constant at each time and storage condition. No differences were observed in the proliferative activity of keratocytes after treatment with PRGF eye drops at any studied time or temperature. Finally, there was no microbial contamination in any of the PRGF eye drops. The preservation of the PRGF eye drops at -20°C for up to 3 and 6 months does not mean reduction of the main growth factors and proteins implicated in ocular surface wound healing. Eye drop characteristics and in vitro biological activity were not affected by their usage and conservation for 72 hours at 4°C or at room temperature.

  6. Correlation of mutations and recombination with growth kinetics of poliovirus vaccine strains.

    Science.gov (United States)

    Pliaka, V; Kyriakopoulou, Z; Tsakogiannis, D; Ruether, I G A; Gartzonika, C; Levidiotou-Stefanou, S; Krikelis, A; Markoulatos, P

    2010-12-01

    Attenuated strains of Sabin poliovirus vaccine replicate in the human gut and, in rare cases, may cause vaccine-associated paralytic poliomyelitis (VAPP). The genetic instability of Sabin strains constitutes one of the main causes of VAPP, a disease that is most frequently associated with type 3 and type 2 Sabin strains, and more rarely with type 1 Sabin strains. In the present study, the growth phenotype of eight oral poliovirus vaccine (OPV) isolates (two non-recombinants and six recombinants), as well as of Sabin vaccine strains, was evaluated using two different assays, the reproductive capacity at different temperatures (Rct) test and the one-step growth curve test in Hep-2 cells at two different temperatures (37°C and 40°C). The growth phenotype of isolates was correlated with genomic modifications in order to identify the determinants and mechanisms of reversion towards neurovirulence. All of the recombinant OPV isolates showed a thermoresistant phenotype in the Rct test. Moreover, both recombinant Sabin-3 isolates showed significantly higher viral yield than Sabin 3 vaccine strain at 37°C and 40°C in the one-step growth curve test. All of the OPV isolates displayed mutations at specific sites of the viral genome, which are associated with the attenuated and temperature-sensitive phenotype of Sabin strains. The results showed that both mutations and recombination events could affect the phenotype traits of Sabin derivatives and may lead to the reversion of vaccinal strains to neurovirulent ones. The use of phenotypic markers along with the genomic analysis may shed additional light on the molecular determinants of the reversed neurovirulent phenotype of Sabin derivatives.

  7. Homologous recombination occurs in Entamoeba and is enhanced during growth stress and stage conversion.

    Directory of Open Access Journals (Sweden)

    Nishant Singh

    Full Text Available Homologous recombination (HR has not been demonstrated in the parasitic protists Entamoeba histolytica or Entamoeba invadens, as no convenient method is available to measure it. However, HR must exist to ensure genome integrity, and possible genetic exchange, especially during stage conversion from trophozoite to cyst. Here we show the up regulation of mitotic and meiotic HR genes in Entamoeba during serum starvation, and encystation. To directly demonstrate HR we use a simple PCR-based method involving inverted repeats, which gives a reliable read out, as the recombination junctions can be determined by sequencing the amplicons. Using this read out, we demonstrate enhanced HR under growth stress in E. histolytica, and during encystation in E. invadens. We also demonstrate recombination between chromosomal inverted repeats. This is the first experimental demonstration of HR in Entamoeba and will help future investigations into this process, and to explore the possibility of meiosis in Entamoeba.

  8. Exogenous recombinant human growth hormone effects during suboptimal energy and zinc intake

    OpenAIRE

    Rising, Russell; Scaglia, Julio F; Cole, Conrad; Tverskaya, Rozalia; Duro, Debora; Lifshitz, Fima

    2005-01-01

    Abstract Background Energy and Zinc (Zn) deficiencies have been associated with nutritional related growth retardation as well as growth hormone (GH) resistance. In this study, the relationship between suboptimal energy and/or Zn intake and growth in rats and their response to immunoreactive exogenous recombinant human GH (GHi), was determined. Results Rats treated with GHi and fed ad-libitum energy and Zn (100/100) had increased IGFBP-3 (p < 0.05) as compared with NSS (215 ± 23 vs. 185 ± 17 ...

  9. The effects of genetic polymorphism on treatment response of recombinant human growth hormone.

    Science.gov (United States)

    Chen, Shi; You, Hanxiao; Pan, Hui; Zhu, Huijuan; Yang, Hongbo; Gong, Fengying; Wang, Linjie; Jiang, Yu; Yan, Chengsheng

    2017-12-06

    Recombinant human growth hormone (rhGH) has been widely used in clinical treatment of growth hormone deficiency (GHD) or non GHD since 1985 and technology have achieved a great development in different long-acting formulations. Although the mathematical models for predicting the growth hormone response could help clinicians get to an individual personalized growth dose, many patients just can't reach the target height and the growth hormone responses differed.Genetic polymorphisms may play a role in the varies of individual responses in this treatment process.This article gives an overview of the genetic polymorphisms research of growth hormone in recent years, in order to give some potential suggestion and guide for the dose titration during treatment. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  10. Treatment of dwarfism with recombinant human insulin-like growth factor-1.

    Science.gov (United States)

    Ranke, Michael B; Wölfle, Joachim; Schnabel, Dirk; Bettendorf, Markus

    2009-10-01

    The growth hormone-IGF (insulin-like growth factor) system plays a central role in hormonal growth regulation. Recombinant human (rh) growth hormone (GH) has been available since the late 1980s for replacement therapy in GH-deficient patients and for the stimulation of growth in patients with short stature of various causes. Growth promotion by GH occurs in part indirectly through the induction of IGF-1 synthesis. In primary disturbances of IGF-1 production, short stature can only be treated with recombinant human IGF-1 (rhIGF-1). rhIGF-1 was recently approved for this indication but can also be used to treat other conditions. Selective review of the literature on IGF-1 therapy, based on a PubMed search. In children with severe primary IGF-1 deficiency (a rare condition whose prevalence is less than 1:10,000), the prognosis for final height is very poor (ca. 130 cm), and IGF-1 therapy is the appropriate form of pathophysiologically based treatment. There is no alternative treatment at present. The subcutaneous administration of IGF-1 twice daily in doses of 80 to 120 microg/kg accelerates growth and increases final height by 12 to 15 cm, according to current data. There is, however, a risk of hypoglycemia, as IGF-1 has an insulin-like effect. As treatment with IGF-1 is complex, this new medication should only be prescribed, for the time being, by experienced pediatric endocrinologists and diabetologists.

  11. Recombinant Human Acidic Fibroblast Growth Factor (aFGF) Expressed in Nicotiana benthamiana Potentially Inhibits Skin Photoaging.

    Science.gov (United States)

    Ha, Jang-Ho; Kim, Ha-Neul; Moon, Ki-Beom; Jeon, Jae-Heung; Jung, Dai-Hyun; Kim, Su-Jung; Mason, Hugh S; Shin, Seo-Yeon; Kim, Hyun-Soon; Park, Kyung-Mok

    2017-07-01

    Responding to the need for recombinant acidic fibroblast growth factor in the pharmaceutical and cosmetic industries, we established a scalable expression system for recombinant human aFGF using transient and a DNA replicon vector expression in Nicotiana benthamiana . Recombinant human-acidic fibroblast growth factor was recovered following Agrobacterium infiltration of N. benthamiana . The optimal time point at which to harvest recombinant human acidic fibroblast growth factor expressing leaves was found to be 4 days post-infiltration, before necrosis was evident. Commassie-stained SDS-PAGE gels of His-tag column eluates, concentrated using a 10 000 molecular weight cut-off column, showed an intense band at the expected molecular weight for recombinant human acidic fibroblast growth factor. An immunoblot confirmed that this band was recombinant human acidic fibroblast growth factor. Up to 10 µg recombinant human-acidic fibroblast growth factor/g of fresh leaves were achieved by a simple affinity purification protocol using protein extract from the leaves of agroinfiltrated N. benthamiana . The purified recombinant human acidic fibroblast growth factor improved the survival rate of UVB-irradiated HaCaT and CCD-986sk cells approximately 89 and 81 %, respectively. N. benthamiana -derived recombinant human acidic fibroblast growth factor showed similar effects on skin cell proliferation and UVB protection compared to those of Escherichia coli -derived recombinant human acidic fibroblast growth factor. Additionally, N. benthamiana- derived recombinant human acidic fibroblast growth factor increased type 1 procollagen synthesis up to 30 % as well as reduced UVB-induced intracellular reactive oxygen species generation in fibroblast (CCD-986sk) cells.UVB is a well-known factor that causes various types of skin damage and premature aging. Therefore, the present study demonstrated that N. benthamiana -derived recombinant human acidic fibroblast growth factor

  12. Angiogenic Type I Collagen Extracellular Matrix Integrated with Recombinant Bacteriophages Displaying Vascular Endothelial Growth Factors.

    Science.gov (United States)

    Yoon, Junghyo; Korkmaz Zirpel, Nuriye; Park, Hyun-Ji; Han, Sewoon; Hwang, Kyung Hoon; Shin, Jisoo; Cho, Seung-Woo; Nam, Chang-Hoon; Chung, Seok

    2016-01-21

    Here, a growth-factor-integrated natural extracellular matrix of type I collagen is presented that induces angiogenesis. The developed matrix adapts type I collagen nanofibers integrated with synthetic colloidal particles of recombinant bacteriophages that display vascular endothelial growth factor (VEGF). The integration is achieved during or after gelation of the type I collagen and the matrix enables spatial delivery of VEGF into a desired region. Endothelial cells that contact the VEGF are found to invade into the matrix to form tube-like structures both in vitro and in vivo, proving the angiogenic potential of the matrix. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  13. Enhancement of bone formation in rabbits by recombinant human growth hormone

    International Nuclear Information System (INIS)

    Ehrnberg, A.; Brosjoe, O.; Laaftman, P.; Nilsson, O.; Stroemberg, L.

    1993-01-01

    We studied the effect of human recombinant growth hormone on diaphyseal bone in 40 adult rabbits. The diaphyseal periosteum of one femur in each animal was mechanically stimulated by a nylon cerclage band. The bands induced an increase in bone formation, bone mineral content, and maximum torque capacity of the diaphyseal bone at 1 and 2 months. Growth hormone enhanced the anabolic effect of the cerclage bands on bone metabolism, evidenced by a further increase in torsional strength of the femurs. (au) (32 refs.)

  14. Exogenous recombinant human growth hormone effects during suboptimal energy and zinc intake

    Directory of Open Access Journals (Sweden)

    Duro Debora

    2005-04-01

    Full Text Available Abstract Background Energy and Zinc (Zn deficiencies have been associated with nutritional related growth retardation as well as growth hormone (GH resistance. In this study, the relationship between suboptimal energy and/or Zn intake and growth in rats and their response to immunoreactive exogenous recombinant human GH (GHi, was determined. Results Rats treated with GHi and fed ad-libitum energy and Zn (100/100 had increased IGFBP-3 (p Conclusion These results suggest that GHi enhances weight gain in rats with suboptimal energy and Zn intake but does not modify energy expenditure or physical activity index. Suboptimal Zn intake did not exacerbate the reduced growth or decrease in energy expenditure observed with energy restriction.

  15. GROWTH RESPONSE OF CLOWN LOACH (Chromobotia macracanthus Bleeker 1852 JUVENILES IMMERSED IN WATER CONTAINING RECOMBINANT GROWTH HORMONE

    Directory of Open Access Journals (Sweden)

    Asep Permana

    2015-12-01

    Full Text Available The main problem in the culture of clown loach (Chromobotia macracanthus is the slow growth rate, which takes about six months to reach its market size (two inches total body length. Slow growth eventually cause a long production time and increase the production costs. An alternative solution can be proposed in order to enhance the growth is by using recombinant growth hormone. The aim of this study was to determine the immersion dose of recombinant Epinephelus lanceolatus growth hormone (rElGH which can generate the highest growth in clown loach. Larvae at seven day after hatching were hyperosmotic treated with NaCl 2.0% for one minute, then immersed for one hour in water containing 0.3% NaCl, 0.01% bovine serum albumin (BSA, and different doses of rElGH, namely: 0.12 (treatment A, 1.2 (B, 12 (C, and 120 mg/L (D. As control, fish were immersed in water without rElGH and NaCl (control-1, water containing 0.3% NaCl and 0.01% BSA (control-2, and 0.3% NaCl water (control-3. Each treatment was replicated three times. The results showed that clown loach juveniles in treatment B, C, and D had longer total body length (P0.05. In addition, the percentage of large size juveniles increased approximately 5% in treatment B, almost the same as in the medium size, while the small size were decrease compared to the control-1. Thus, the best immersion dose of rElGH was 1.2 mg/L water.

  16. A Child with Local Lipohypertrophy following Recombinant Human Growth Hormone Administration

    Directory of Open Access Journals (Sweden)

    Ilan J. N. Koppen

    2016-01-01

    Full Text Available Local lipohypertrophy due to recombinant human growth hormone (rhGH administration is a rare phenomenon. Here, we report a case of an 11-year-old girl who presented with a paraumbilical swelling, approximately one year after the start of rhGH treatment for short stature due to the presumed diagnosis of partial growth hormone insensitivity. Ultrasound imaging revealed an asymmetric distribution of subcutaneous fat tissue at the rhGH administration site, indicating local lipohypertrophy. After sparing her routine injection site and alternating other sites, the swelling disappeared within 6 months. Although the precise cause of local lipohypertrophy resulting from rhGH administration is still unclear, it might be related to the presumed diagnosis of partial growth hormone insensitivity.

  17. Atelocollagen sponge and recombinant basic fibroblast growth factor combination therapy for resistant wounds with deep cavities.

    Science.gov (United States)

    Nakanishi, Asako; Hakamada, Arata; Isoda, Ken-ichi; Mizutani, Hitoshi

    2005-05-01

    Recent advances in bioengineering have introduced materials that enhance wound healing. Even with such new tools, some deep ulcers surrounded by avascular tissues, including bone, tendon, and fascia, are resistant to various therapies and easily form deep cavities with loss of subcutaneous tissue. Atelocollagen sponges have been used as an artificial dermis to cover full-thickness skin defects. Topical recombinant human basic fibroblast growth factor has been introduced as a growth factor to induce fibroblast proliferation in skin ulcers. We applied these materials in combination in two patients with deep resistant wounds: one with a cavity reaching the mediastinum through a divided sternum and one with deep necrotic wounds caused by electric burns. These wounds did not respond to the topical basic fibroblast growth factor alone. In contrast, the combination therapy closed the wounds rapidly without further surgical treatment. This combination therapy is a potent treatment for resistant wounds with deep cavities.

  18. The role and future challenges for recombinant growth hormone therapy to promote growth in children after renal transplantation.

    Science.gov (United States)

    Janjua, Halima S; Mahan, John D

    2011-01-01

    Chronic kidney disease can severely impair linear growth in children. For many children, growth improves after renal transplantation, but for some, growth velocity remains low and for others, catch-up growth is insufficient to compensate for the deficit imparted by renal disease in the preceding years. Inadequate final adult height after renal transplant is multifactorial and can adversely affect the quality of life (QOL), psychosocial development and long term prospects for these children as they grow into adulthood. Growth failure after renal transplant requires thorough evaluation and its management in renal transplant recipients can involve improved nutritional intake, correction of metabolic acidosis, treatment of secondary hyperparathyroidism, steroid-sparing immunosuppression and/or use of recombinant human growth hormone (rGH). Treatment with rGH after renal transplant has been evaluated by a limited number of clinical trials suggesting efficacy and safety for this treatment strategy. Several important clinical questions regarding rGH use in children post-renal transplant remain unanswered. © 2011 John Wiley & Sons A/S.

  19. Brief study about the distribution of recombinant human Epidermic Growth Factor (rh-EGF)

    International Nuclear Information System (INIS)

    Rodriguez Garcia, J.C.; De Dios D Espaux, R.; Bello Garciga, J.L.

    1997-01-01

    This report describes results of the study about biodistribution of I-125 recombinant human Epidermic Growth Factor (rhEGF). The radiolabelled product was administrated to Sprague Dawley rats in three different ways: intramuscular, subcutaneous and epidermic; the highest concentration of EGF in blood was found 4 hours after rhEGF administration, with a greater distribution in the plasma with regard to cellular pellet. The slowest plasma clearance corresponded to the intramuscular administration. The highest concentration of radiolabelled rhEGF was found in liver, kidney and intestine. It was found that radiolabelled EGF is excreted mainly throughout urine and faeces although other excretion pathways could exist

  20. A recombinant endogenous retrovirus amplified in a mouse neuroblastoma is involved in tumor growth in vivo.

    Science.gov (United States)

    Pothlichet, Julien; Heidmann, Thierry; Mangeney, Marianne

    2006-08-15

    The theory of immunoediting postulates that tumor cells exhibit a reduced immunogenicity to escape eradication by the host immune system. It has been proposed that endogenous retroviruses--provided that they are active--could play a role in this process, via the immunosuppressive domain carried by their envelope protein. Here, we demonstrate that the Neuro-2a tumor cell line--originating from a spontaneous A/J mouse neuroblastoma--produces an infectious retrovirus that most probably results from a recombination event between 2 mouse endogenous retroviral elements. This Neuro-2a-associated recombinant retrovirus derives from the unique ecotropic provirus located at the Emv-1 locus, but with a gag sequence conferring B-tropism, thus allowing its high-level amplification in Neuro-2a cells. We show that knocking down -by RNA interference- this endogenous retrovirus in Neuro-2a cells has no effect on the transformed phenotype of the cells, but results in delayed tumor growth and prolonged animal survival, following engraftment of the cells into immunocompetent mice. Recombination between endogenous retroviruses, amplification of the resulting element and high-level expression of its immunosuppressive activity are therefore likely steps of an immunoediting process, leading to an invading tumor. Copyright 2006 Wiley-Liss, Inc.

  1. Nerve growth factor, clinical applications and production of the recombinant protei

    Directory of Open Access Journals (Sweden)

    M. Zangi

    2017-01-01

    Full Text Available The mammalian neurotrophin family proteins, nerve growth factor (NGF, brain-derived neurotrophic factor (BDNF, neurotrophin-3 (NT-3 and neurotrophin-4/5 (NT-4/5 are known as neuronal survival factors. NGF, one of the most important cytokines, is composed of 118 amino acids. NGF is involved in the growth and differentiation of neural cells of the vertebrate peripheral sympathetic nerve as well as basal forebrain cholinergic neurons which degenerate in Alzheimer’s disease. In addition, it is implicated in the regulation of synaptic transmission and synaptogenesis in the central nervous system. NGF is produced by a variety of immune cells, including B cells, T cells, monocytes and mast cells as well as nervous system and binds through two distinct receptors, TrkA and p75NTR which signaling through them leads to the neuronal differentiation and cell death respectively. Considering the importance of this protein as a drug, NGF has been proposed for the treatment of neuron degenerative diseases such as Alzheimer's, Parkinson's and multiple sclerosis. To produce enough protein for research and clinical applications, genetic engineering techniques are used to produce recombinant forms. To date, there are no reports about the systems for production of the recombinant human NGF in an effective, low cost, with industrial production. Plants as a safe host generally offer major advantages such as free of animal pathogens, low costs, the ability to produce a protein similar to natural protein, and industrial production in large scale. Then they are suitable for the production of recombinant human NGF.

  2. Translational mixed-effects PKPD modelling of recombinant human growth hormone - from hypophysectomized rat to patients

    DEFF Research Database (Denmark)

    Thorsted, A; Thygesen, P; Agersø, H

    2016-01-01

    BACKGROUND AND PURPOSE: We aimed to develop a mechanistic mixed-effects pharmacokinetic (PK)-pharmacodynamic (PD) (PKPD) model for recombinant human growth hormone (rhGH) in hypophysectomized rats and to predict the human PKPD relationship. EXPERIMENTAL APPROACH: A non-linear mixed-effects model...... was developed from experimental PKPD studies of rhGH and effects of long-term treatment as measured by insulin-like growth factor 1 (IGF-1) and bodyweight gain in rats. Modelled parameter values were scaled to human values using the allometric approach with fixed exponents for PKs and unscaled for PDs...... s.c. administration was over predicted. After correction of the human s.c. absorption model, the induction model for IGF-1 well described the human PKPD data. CONCLUSIONS: A translational mechanistic PKPD model for rhGH was successfully developed from experimental rat data. The model links...

  3. Optimization of the purification methods for recovery of recombinant growth hormone from Paralichthys olivaceus

    Science.gov (United States)

    Zang, Xiaonan; Zhang, Xuecheng; Mu, Xiaosheng; Liu, Bin

    2013-03-01

    This study aimed to optimize the purification of recombinant growth hormone from Paralichthys olivaceus. Recombinant flounder growth hormone (r-fGH) was expressed by Escherichia coli in form of inclusion body or as soluble protein under different inducing conditions. The inclusion body was renatured using two recovery methods, i.e., dilution and dialysis. Thereafter, the refolded protein was purified by Glutathione Sepharase 4B affinity chromatography and r-fGH was obtained by cleavage of thrombin. For soluble products, r-fGH was directly purified from the lysates by Glutathione Sepharase 4B affinity chromatography. ELISA-receptor assay demonstrated that despite its low receptor binding activity, the r-fGH purified from refolded inclusion body had a higher yield (2.605 mg L-1) than that from soluble protein (1.964 mg L-1). Of the tested recovery methods, addition of renaturing buffer (pH 8.5) into denatured inclusion body yielded the best recovery rate (17.9%). This work provided an optimized purification method for high recovery of r-fGH, thus contributing to the application of r-fGH to aquaculture.

  4. Platelet-derived stromal cell-derived factor-1 is required for the transformation of circulating monocytes into multipotential cells.

    Directory of Open Access Journals (Sweden)

    Noriyuki Seta

    Full Text Available BACKGROUND: We previously described a primitive cell population derived from human circulating CD14(+ monocytes, named monocyte-derived multipotential cells (MOMCs, which are capable of differentiating into mesenchymal and endothelial lineages. To generate MOMCs in vitro, monocytes are required to bind to fibronectin and be exposed to soluble factor(s derived from circulating CD14(- cells. The present study was conducted to identify factors that induce MOMC differentiation. METHODS: We cultured CD14(+ monocytes on fibronectin in the presence or absence of platelets, CD14(- peripheral blood mononuclear cells, platelet-conditioned medium, or candidate MOMC differentiation factors. The transformation of monocytes into MOMCs was assessed by the presence of spindle-shaped adherent cells, CD34 expression, and the potential to differentiate in vitro into mesenchymal and endothelial lineages. RESULTS: The presence of platelets or platelet-conditioned medium was required to generate MOMCs from monocytes. A screening of candidate platelet-derived soluble factors identified stromal cell-derived factor (SDF-1 as a requirement for generating MOMCs. Blocking an interaction between SDF-1 and its receptor CXCR4 inhibited MOMC generation, further confirming SDF-1's critical role in this process. Finally, circulating MOMC precursors were found to reside in the CD14(+CXCR4(high cell population. CONCLUSION: The interaction of SDF-1 with CXCR4 is essential for the transformation of circulating monocytes into MOMCs.

  5. Production of functional human insulin-like growth factor binding proteins (IGFBPs) using recombinant expression in HEK293 cells

    DEFF Research Database (Denmark)

    Wanscher, Anne Sofie Molsted; Williamson, Michael; Ebersole, Tasja Wainani

    2015-01-01

    on human proteins with therapeutic relevance is needed to design and process the next generation of protein therapeutics. In order to conduct structural and functional investigations large quantities of recombinant proteins are needed. However, finding a suitable recombinant production system for proteins...... and the final protein yields were between 1 and 12mg protein per liter culture media. The recombinant IGFBPs contained PTMs and exhibited high-affinity interactions with their natural ligands IGF-1 and IGF-2.......Insulin-like growth factor binding proteins (IGFBPs) display many functions in humans including regulation of the insulin-like growth factor (IGF) signaling pathway. The various roles of human IGFBPs make them attractive protein candidates in drug discovery. Structural and functional knowledge...

  6. In situ expression of platelet-derived growth factor (PDGF-beta) during chronic rejection is abolished by retransplantation.

    Science.gov (United States)

    Yang, H C; McElroy, R J; Kreider, J W; Marshall, R L; Martinie, J B; Diamond, J

    1995-07-01

    We have shown that Fischer 344-->Lewis renal allografts (ALLO) develop chronic rejection which is not detected in Lewis-->Lewis isografts (ISO). The progression of chronic rejection in ALLO can be reversed by retransplantation (RE-TX) of kidneys from ALLO back into syngeneic Fischer 344 recipients. The purpose of this study was to assess the in situ expression of PDGF-beta, a cytokine associated with wound injury, in ISO, ALLO, and RE-TX. In situ PDGF-beta mRNA expression in kidney sections was assessed early (8 weeks) and late (16 weeks) during the development of chronic rejection. Kidneys from ALLO were transplanted back into syngeneic Fischer recipients at 12 weeks and evaluated for PDGF-beta expression 12 weeks later. Differences in glomerular staining were graded quantitatively on a minimum of 25 glomeruli per section with grade 0, no positive cells in the glomerulus; grade 1, 1 or 2 positive cells; grade 2, 3 or more positive cells in a segmental distribution; and grade 3, > 4 positive cells of moderate intensity in a diffuse distribution. According to this grading system, glomerular PDGF-beta mRNA expression in isografts (N = 6) at 8 and 16 weeks after transplantation was 0.09 +/- 0.03 and 0.2 +/- 0.04, respectively. In allografts (N = 6), PDGF-beta mRNA was significantly higher (P < .001) for the same time periods, 1.28 +/- 0.6 and 1.89 +/- 0.08, respectively. In situ expression of PDGF in retransplants (N = 6) at 24 weeks, 0.07 +/- 0.02, was significantly diminished (P < .001) at 24 weeks compared to allografts at 8 or 16 weeks.(ABSTRACT TRUNCATED AT 250 WORDS)

  7. Bacteria-induced release of white cell--and platelet-derived vascular endothelial growth factor in vitro

    DEFF Research Database (Denmark)

    Nielsen, Hans Jørgen; Werther, K; Mynster, T

    2001-01-01

    of buffy-coat-depleted red cell (SAGM) blood were donated by healthy blood donors. Subsequently, half of every unit was leucocyte depleted by filtration, and all 32 half-units were stored under standard conditions for 35 days. Just after storage, and on days 7, 21 and 35 during storage, aliquots...

  8. Influence of a ras oncogene on platelet-derived growth factor (PDGF)-stimulated phosphoinositide hydrolysis in murine fibroblasts

    International Nuclear Information System (INIS)

    Parries, G.; Racker, E.

    1986-01-01

    The authors have examined the effects of transfection of rat-1 fibroblasts with the ras oncogene on the metabolism of phosphatidylinositol (PI). Incubation of [ 3 H]inositol-labeled rat-1 cells with PDGF resulted in a 2- to 3-fold increase in [ 3 H]IP3 levels within 90 s. In the presence of 25 mM Li+, [ 3 H]IP1 levels were increased 8-fold after 30 min. In contrast, incubation of ras-transfected fibroblasts (EJ-2 line) with PDGF had little or no effect on the level of either [ 3 H]IP3 or [ 3 H]IP1. Similar stimulations by PDGF were observed in NIH 3T3 cells, but not in Kirsten virus-transformed or Harvey ras-transfected cell lines. On the other hand, NIH 3T3 cells transfected with v-src responded to PDGF by stimulation of PI turnover similar to the parent cell line. In NIH 3T3 cells transfected with an expression vector containing the v-Ha-ras gene under transcriptional control of the glucocorticoid-inducible mouse mammary tumor virus promoter, the PDGF stimulation of [ 3 H]inositol incorporation into PI was reduced from 10-fold in the absence of dexamethasone to 1.8-fold when the cells were pretreated for 26 h with 2 μM dexamethasone. In the parental 3T3 cells PDGF stimulation was reduced by about 40% in the presence of dexamethasone. In the absence of PDGF the rate of PI turnover (i.e., the kinetics of [ 3 H]IP1 accumulation in the presence of Li+) in EJ-2 cells was similar to that in rat-1 cells. Thus, in the presence of PDGF, the rate of PI turnover in rat-1 cells was several fold higher than in the transfected cells. These results suggest that the ras gene product (p21) may exert an inhibitory effect on PDGF-stimulated phosphoinositide metabolism

  9. Growth regulation on human acute myeloid leukemia effects of five recombinant hematopoietic factors in a serum-free culture system

    NARCIS (Netherlands)

    Delwel, E.; Salem, M.; Pellens, C.; Dorssers, L.; Wagemaker, G.; Clark, S.; Loewenberg, B

    1988-01-01

    The response of human acute myeloid leukemia (AML) cells to the distinct hematopoietic growth factors (HGFs), ie, recombinant interleukin-3 (IL-3), granulocyte-macrophage-CSF (GM-CSF), granulocyte-CSF (G-CSF), macrophage-CSF (M-CSF), and erythropoietin (Epo) was investigated under well-defined

  10. Development of a NanoBioAnalytical platform for "on-chip" qualification and quantification of platelet-derived microparticles.

    Science.gov (United States)

    Obeid, Sameh; Ceroi, Adam; Mourey, Guillaume; Saas, Philippe; Elie-Caille, Celine; Boireau, Wilfrid

    2017-07-15

    Blood microparticles (MPs) are small membrane vesicles (50-1000nm), derived from different cell types. They are known to play important roles in various biological processes and also recognized as potential biomarkers of various health disorders. Different methods are currently used for the detection and characterization of MPs, but none of these methods is capable to quantify and qualify total MPs at the same time, hence, there is a need to develop a new approach for simultaneous detection, characterization and quantification of microparticles. Here we show the potential of surface plasmon resonance (SPR) method coupled to atomic force microscopy (AFM) to quantify and qualify platelet-derived microparticles (PMPs), on the whole nano-to micro-meter scale. The different subpopulations of microparticles could be determined via their capture onto the surface using specific ligands. In order to verify the correlation between the capture level and the microparticles concentration in solution, two calibration standards were used: Virus-Like Particles (VLPs) and synthetic beads with a mean diameter of 53nm and 920nm respectively. The AFM analysis of the biochip surface allowed metrological analysis of captured PMPs and revealed that more than 95% of PMPs were smaller than 300nm. Our results suggest that our NanoBioAnalytical platform, combining SPR and AFM, is a suitable method for a sensitive, reproducible, label-free characterization and quantification of MPs over a wide concentration range (≈10 7 to 10 12 particles/mL; with a limit of detection (LOD) in the lowest ng/µL range) which matches with their typical concentrations in blood. Copyright © 2016 Elsevier B.V. All rights reserved.

  11. Height Outcome of Recombinant Human Growth Hormone Treatment in Achondroplasia Children: A Meta-Analysis.

    Science.gov (United States)

    Miccoli, Mario; Bertelloni, Silvano; Massart, Francesco

    2016-01-01

    Although recombinant human growth hormone (rhGH) is not approved to treat short stature of achondroplasia (ACH), some studies suggested growth improvement during short-term rhGH treatment. A meta-analysis of rhGH therapy efficacy in ACH children was performed. From 12 English-language studies, 558 (54.0% males) rhGH-treated ACH children were enrolled. Administration of rhGH (median dosage 0.21 mg/kg/ week; range 0.16-0.42 mg/kg/week) improved height (Ht) from baseline [-5.069 standard deviation score (SDS; 95% CI -5.109 to -5.029); p < 0.0001] to 12 [-4.325 SDS (95% CI -4.363 to -4.287); p < 0.0001] and 24 months [-4.073 SDS (95% CI -4.128 to -4.019); p < 0.0001]. Then, Ht remained approximately constant up to 5 years [-3.941 SDS (95% CI -4.671 to -3.212); p < 0.0001]. In ACH children, rhGH treatment increased Ht from -5.0 to -4.0 SDS during 5 years, but insufficient data are available on both the adult Ht and the changes of body proportions. © 2016 S. Karger AG, Basel.

  12. Expression and purification of recombinant truncated human keratinocyte growth factor-1

    International Nuclear Information System (INIS)

    Deng Lin; Ma Jisheng; Liu Xiaoju; Wang Xiaojie; Li Xiaokun; Gong Shouliang; Wang Huiyan; Tian Haishan

    2010-01-01

    Objective: To construct the genetic engineering bacteria highly expressing 23 amino acids human keratinocyte growth factor-1 (rhKGF1 dest23 ) missing N terminal, and provide experimental data for development of new drug for treatment of oral mucositis after radiotherapy and chemotherapy. Methods: PCR was used to synthese 23 amino acids rhKGF1 dest23 missing N terminal and sumo gene fragments, and construct four kinds of recombinant prokaryotic expression vectors: pET22b-rhKGF1 dest23 , pET22b-sumo-rhKGF1 dest23 , pET3c-rhKGF1 dest23 and pET3c-sumo-rhKGF1 dest23 , then they were transformed into prokaryotic expression host bacteria: Rosetta (DE3) plysS, BL21 (DE3), BL21 (DE3) Star plysS, origima(DE3) and BL21AI, the best expression combination of plasmid and host strain of rhKGF1 dest23 protein was screened and purified by CM ion-exchange and heparin affinity chromatography and identified with Western blotting. Results: pET22b-rhKGF1 dest23 plasmid and the BL21AI host bacteria was the best combination of expression, after induced by IPTG and arabinose, the majority of recombinant protein was expressed in soluble form, accounting for about 12% of the total bacterial proteins. Its purity reached to more than 95% of the protein after two steps chromatography, then conformed with Western blotting. Conclusion: Human genetic engineering bacteria of KGF1 dest23 is successfully constructed and induced by IPTG and arabinose, then after CM weak cation exchange and heparin affinity chromatography, the purified rhKGF1 dest23 protein is obtained. (authors)

  13. Long-term response to recombinant human growth hormone treatment: a new predictive mathematical method.

    Science.gov (United States)

    Migliaretti, G; Ditaranto, S; Guiot, C; Vannelli, S; Matarazzo, P; Cappello, N; Stura, I; Cavallo, F

    2018-07-01

    Recombinant GH has been offered to GH-deficient (GHD) subjects for more than 30 years, in order to improve height and growth velocity in children and to enhance metabolic effects in adults. The aim of our work is to describe the long-term effect of rhGH treatment in GHD pediatric patients, suggesting a growth prediction model. A homogeneous database is defined for diagnosis and treatment modalities, based on GHD patients afferent to Hospital Regina Margherita in Turin (Italy). In this study, 232 GHD patients are selected (204 idiopathic GHD and 28 organic GHD). Each measure is shown in terms of mean with relative standard deviations (SD) and 95% confidence interval (95% CI). To estimate the final height of each patient on the basis of few measures, a mathematical growth prediction model [based on Gompertzian function and a mixed method based on the radial basis functions (RBFs) and the particle swarm optimization (PSO) models] was performed. The results seem to highlight the benefits of an early start of treatment, further confirming what is suggested by the literature. Generally, the RBF-PSO method shows a good reliability in the prediction of the final height. Indeed, RMSE is always lower than 4, i.e., in average the forecast will differ at most of 4 cm to the real value. In conclusion, the large and accurate database of Italian GHD patients allowed us to assess the rhGH treatment efficacy and compare the results with those obtained in other Countries. Moreover, we proposed and validated a new mathematical model forecasting the expected final height after therapy which was validated on our cohort.

  14. The preventive effect of recombinant human growth factor (rhEGF) on the recurrence of radiodermatitis

    International Nuclear Information System (INIS)

    Ryu, Seung-Hee; Kim, Yeun-Hwa; Lee, Sang-Wook; Hong, Joon-Pio

    2010-01-01

    The effects of topical application of recombinant human epidermal growth factor (rhEGF) on wound healing and the recurrence of radiodermatitis were assessed in the irradiated skin of BALB/c Nu/Nu mice. Mice irradiated with 45 Gy of radiation were divided into 5 groups and treated with 10, 50, and 100 μg/g rhEGF ointment, vehicle alone, or no treatment (control) for 6 months. Wounds were observed initially in all groups and complete healing time (HT 100 ) for initial wound repair did not differ significantly among groups. However, the rate of recurrence over 6 months was significantly lower in the EGF-treated groups than in the control group (p<0.05). Histological examination showed that treatment with the optimum dose of EGF (50 μg/g) accelerated normal wound healing when compared with the higher dose of EGF (100 μg/g), vehicle alone, or no treatment, with the latter group showing irregular epidermal thickness, poor definition of epidermis and dermis, and unstable dermal structure. Collagen distribution was also significantly increased in mice treated with 50 μg/g rhEGF (p<0.05) compared with the control or vehicle-treated group. Taken together, these results indicate that treatment with exogenous EGF (50 μg/g dose) can enhance radiation-induced wound repair while preserving structural tissue stability and preventing the recurrence of radiodermatitis. (author)

  15. Recombinant human laminin isoforms can support the undifferentiated growth of human embryonic stem cells

    International Nuclear Information System (INIS)

    Miyazaki, Takamichi; Futaki, Sugiko; Hasegawa, Kouichi; Kawasaki, Miwa; Sanzen, Noriko; Hayashi, Maria; Kawase, Eihachiro; Sekiguchi, Kiyotoshi; Nakatsuji, Norio; Suemori, Hirofumi

    2008-01-01

    Human embryonic stem cells (hESCs) are thought to be a promising cell source for cell transplantation therapy. For such a clinical application, the hESCs should be manipulated using appropriate and qualified materials. In this study, we examined the efficacy of recombinant human laminin (rhLM) isoforms on the undifferentiated growth of hESCs. We first determined the major integrins expressed on the hESCs to reveal the preference of the hESCs for rhLMs, and found that the hESCs mainly expressed integrin α6β1, which binds predominantly to laminin-111, -332 and -511/-521. When the hESCs were seeded onto rhLMs, the cells indeed adhered markedly to rhLM-332, and to rhLM-511 and rhLM-111 to a lesser extent. The hESCs proliferated on these three rhLMs for several passages while preserving their pluripotency. These results show that rhLM-111, -332, and -511 are good substrates to expand undifferentiated hESCs due to their high affinity to integrin α6β1 expressed on hESCs

  16. Recombinant human growth hormone treatment in short children with renal disease: Our first experience

    Directory of Open Access Journals (Sweden)

    Spasojević-Dimitrijeva Brankica

    2010-01-01

    Full Text Available Introduction. Growth retardation is a hallmark of chronic illnesses such as chronic kidney disease in children, and it is associated with increased morbidity and mortality. The growth hormone (GH resistance observed in uraemia can be overcome by supraphysiological doses of exogenous GH. Objective. We would like to present our first results of recombinant human growth hormone (rhGH treatment, mainly in children on haemodialysis. Methods. Sixteen children, aged 4.5-17.1 years (mean age 11.25±3.57 with height below -2.0 standard deviation score (SDS for age or height velocity below -2.0 SDS for age, were selected to receive rhGH therapy at our Nephrology and Haemodialysis Department. Most of them were on haemodialysis (14 children with mean spent time 2.88±2.68 years (0-9 years before the initiation of rhGH therapy. One half of patients were prepubertal (8 children and the second half were in early puberty (testicular volume between 4 and 8 ml for boys and breast development B2 or B3 in girls. All patients received 28-30IU/m² rhGH per week by daily subcutaneous injection. The year before rhGH therapy served as a control period. Results. During the first year of treatment, mean height velocity in haemodialysis patients increased from 2.25 cm/year to 6.59 cm/year (p<0.0001 and in the second year it was 5.25 cm/ year (p=0.004. The mean height SDS in haemodialysis children did not improve significantly during the first year of rhGH treatment (from -3.01 SDS to -2.77 SDS, p=0.063. Neither weight nor the body mass index varied compared with the pretreatment period. Two patients developed worsened secondary hyperparathyroidism and were excluded from the study, but the relationship with rhGH remains uncertain. Conclusion. Mean height velocity significantly improved during rhGH therapy in haemodialysis patients. No significant side-effects were observed in children during three-year treatment with GH.

  17. [Effect of recombinant human growth hormone therapy on metabolic parameters in patients with craniopharyngioma].

    Science.gov (United States)

    Mao, J F; Wang, X; Xiong, S Y; Zheng, J J; Yu, B Q; Nie, M; Wu, X Y; Qi, S T

    2017-11-14

    Objective: To investigate the effects of recombinant human growth hormone (rhGH) on metabolic parameters in patients with craniopharyngioma surgeries. Methods: Totallys 30 patients with craniopharyngioma were included in this retrospective study. They were divided into growth hormone (GH) group and control group according to whether they received rhGH therapy or not. The following parameters, including body mass index (BMI), weight, waist circumstance, transaminase, fasting blood glucose, lipid profile and high-sensitivity C-reactive protein (hsCRP) were compared after rhGH therapy for 4-6 months. Results: In GH group, patients were 18-46 (30.0±8.8) years old. The duration after craniopharyngioma surgery was (12.9±5.4) years. Before rhGH therapy, they had got sufficient thyroid and glucocorticoid hormone replacement. After rhGH therapy, the body weight decreased from (92.3±20.1) to (87.6 ±14.6) kg ( P =0.190), with a reduction of BMI from (30.1±5.9) to (28.2±3.7) kg/m(2) ( P =0.120). The waist circumference decreased from (104.4±9.4) cm to (98.8±10.6) cm ( P =0.002). Alanine aminotransferase (ALT) decreased from (52±34) to (28±19) U/L ( P =0.029), with a reduction of aspartate transaminase (AST) from (46±21) to (33±18) U/L ( P =0.035) and γ-glutamyl transpeptadase (GGT) from (59±42) to (29±15) U/L ( P =0.02). hsCRP decreased from (5.3±4.9) to (2.3±2.8) mg/L ( P =0.006) and triglyceride (TG) decreased from (1.8±0.7) to (1.5±0.6) mmol/L ( P =0.028). Fasting blood glucose, low density lipoprotein cholesterol (LDL-C), high density lipoprotein cholesterol (HDL-C) and free fat acid (FFA) were not significantly changed(all P >0.05). In the control group, the above mentioned parameters did not changed significantly during 4-6 months of observational period(all P >0.05). Conclusion: rhGH therapy improves metabolic parameters in patients after craniopharyngioma surgery by decreasing body weight, waist circumstance and fat deposit in liver, as well as

  18. Anti-aggregatory effect of cyclodextrins in the refolding process of recombinant growth hormones from Escherichia coli inclusion bodies

    DEFF Research Database (Denmark)

    Bajorunaite, Egle; Cirkovas, Andrejus; Radzevicius, Kostas

    2009-01-01

    Cyclodextrins with different ring size and ring substituents were tested for recombinant mink and porcine growth hormones aggregation suppression in the refolding process from Escherichia coli inclusion bodies. Methyl-β-cyclodextrin and 2-hydroxypropyl-β-cyclodextrin show a positive effect...... on the aggregation suppression of both proteins. The influence of different methyl-β-cyclodextrin and 2-hydroxypropyl-β-cyclodextrin concentrations on the renaturation yield of both growth hormones was investigated. Moreover, methyl-β-cyclodextrin and 2-hydroxypropyl-β-cyclodextrin suppress not only folding...

  19. Data regarding the growth of Lactobacillus acidophilus NCFM on different carbohydrates and recombinant production of elongation factor G and pyruvate kinase

    DEFF Research Database (Denmark)

    Celebioglu, Hasan Ufuk; Olesen, Sita Vaag; Prehn, Kennie

    2017-01-01

    The present study describes the growth of the very well-known probiotic bacterium Lactobacillus acidophilus NCFM on different carbohydrates. Furthermore, recombinant production of putative moonlighting proteins elongation factor G and pyruvate kinase from this bacterium is described. For further...

  20. Role of Platelet-derived Microvesicles as Crosstalk Mediators in Atherothrombosis and Future Pharmacology Targets: a Link between Inflammation, Atherosclerosis and Thrombosis

    Directory of Open Access Journals (Sweden)

    Lina Badimon

    2016-08-01

    Full Text Available Reports in the last decade have suggested that the role of platelets in atherosclerosis and its thrombotic complications may be mediated, in part, by local secretion of platelet-derived microvesicles (pMVs, small cell blebs released during the platelet activation process. MVs are the most abundant cell-derived microvesicle subtype in the circulation. High concentrations of circulating MVs have been reported in patients with atherosclerosis, acute vascular syndromes, and/or diabetes mellitus, suggesting a potential correlation between the quantity of microvesicles and the clinical severity of the atherosclerotic disease. pMVs are considered to be biomarkers of disease but new information indicates that pMVs are also involved in signaling functions. pMVs evoke or promote haemostatic and inflammatory responses, neovascularization, cell survival and apoptosis, processes involved in the pathophysiology of cardiovascular disease. This review is focused on the complex cross-talk between platelet-derived microvesicles, inflammatory cells and vascular elements and their relevance in the development of the atherosclerotic disease and its clinical outcomes, providing an updated state-of-the art of pMV involvement in atherothrombosis and pMV potential use as therapeutic agent influencing cardiovascular biomedicine in the future.

  1. Production of functional human insulin-like growth factor binding proteins (IGFBPs) using recombinant expression in HEK293 cells.

    Science.gov (United States)

    Wanscher, Anne Sofie Molsted; Williamson, Michael; Ebersole, Tasja Wainani; Streicher, Werner; Wikström, Mats; Cazzamali, Giuseppe

    2015-04-01

    Insulin-like growth factor binding proteins (IGFBPs) display many functions in humans including regulation of the insulin-like growth factor (IGF) signaling pathway. The various roles of human IGFBPs make them attractive protein candidates in drug discovery. Structural and functional knowledge on human proteins with therapeutic relevance is needed to design and process the next generation of protein therapeutics. In order to conduct structural and functional investigations large quantities of recombinant proteins are needed. However, finding a suitable recombinant production system for proteins such as full-length human IGFBPs, still remains a challenge. Here we present a mammalian HEK293 expression method suitable for over-expression of secretory full-length human IGFBP-1 to -7. Protein purification of full-length human IGFBP-1, -2, -3 and -5 was conducted using a two-step chromatography procedure and the final protein yields were between 1 and 12mg protein per liter culture media. The recombinant IGFBPs contained PTMs and exhibited high-affinity interactions with their natural ligands IGF-1 and IGF-2. Copyright © 2014 Elsevier Inc. All rights reserved.

  2. Height outcome of the recombinant human growth hormone treatment in Turner syndrome: a meta-analysis

    Directory of Open Access Journals (Sweden)

    Ping Li

    2018-04-01

    Full Text Available Objective: This study sought to determine the effect of the recombinant human growth hormone (rhGH treatment of Turner syndrome (TS on height outcome. Methods: We searched in MEDLINE, EMBASE and Cochrane Central Register of Controlled Trials and Cochrane Database of Systematic Reviews. A literature search identified 640 records. After screening and full-text assessment, 11 records were included in the systematic review. Methodological quality was assessed using the Cochrane Risk of Bias tool. RevMan 5.3 software was used for meta-analysis. We also assessed the quality of evidence with the GRADE system. Results: Compared with controls, rhGH therapy led to increased final height (MD = 7.22 cm, 95% CI 5.27–9.18, P < 0.001, I2 = 4%; P = 0.18, height standard deviation (HtSDS (SMD = 1.22, 95% CI 0.88–1.56, P < 0.001, I2 = 49%; P = 0.14 and height velocity (HV (MD 2.68 cm/year; 95% CI 2.34, 3.02; P < 0.001, I2 = 0%; P = 0.72. There was a small increase in bone age (SMD 0.32 years; 95% CI 0.1, 0.54; P = 0.004, I2 = 73%; P = 0.02 after rhGH therapy for 12 months. What is more, the rhGH/oxandrolone combination therapy suggested greater final height (MD 2.46 cm; 95% CI 0.73, 4.18; P = 0.005, I2 = 32%; P = 0.22, increase and faster HV (SMD 1.67 cm/year; 95% CI 1.03, 2.31; P < 0.03, I2 = 80%; P < 0.001, with no significant increase in HtSDS and bone maturation compared with rhGH therapy alone. Conclusions: For TS patients, rhGH alone or with concomitant use of oxandrolone treatment had advantages on final height.

  3. Growth Response and Feed Utilization of Giant Gourami (Osphronemus goramy Juvenile Feeding Different Protein Levels of the Diets Supplemented with Recombinant Growth Hormone

    Directory of Open Access Journals (Sweden)

    DARMAWAN SETIA BUDI

    2015-01-01

    Full Text Available The purpose of this study was to examine the effect of dietary supplementation with recombinant growth hormone (rGH on the growth and dietary utility of juvenile giant gourami. The rGH was mixed with chicken egg yolk and sprayed on to artificial feed with different protein levels (34, 28, and 21%; isoenergy. Each treatment group of gourami was paired with a control group that received feed of the same protein level, but without rGH supplementation. Juvenile of giant gourami (weight 15.83 ± 0.13 g were fed diets containing rGH, to apparent satiation, 2 times a week. Fish were reared from less than 2 months old for 42 days in 100 L glass aquaria at an initial density of 10 fish per aquarium. At the end of this period, the biomass and daily growth rate (SGR of the fish were measured and the feed conversion ratio calculated and compared. Our data showed that fish fed rGH-supplemented diets experienced higher growth than fish in the control groups and showed that fish with higher protein diets experienced higher growth than the groups with less protein diets. The group with the highest biomass gain, SGR, and lowest feed conversion ratio (FCR was the group fed a 34% protein diet supplemented with rGH. Furthermore, biomass gain, SGR, and FCR in the rGH treatment group with a 28% protein diet matched the measurements of the non-rGH control group receiving a 34% protein diet (P > 0.05. We conclude that giant juvenile gourami given feed supplemented with recombinant growth hormone will experience increased growth and dietary utility compared with gourami given the same feed without supplementation.

  4. Recombinant human interleukin 2 acts as a B cell growth and differentiation promoting factor

    OpenAIRE

    Emmrich, F.; Moll, Heidrun; Simon, Markus M.

    2009-01-01

    Human B cells appropriately activated by a B cell mitogen are rendered susceptible to human Interleukin 2 (IL-2) as demonstrated with recombinant human IL-2 (rec. h IL-2). They show increased proliferation and drastically enhanced immunoglobulin secretion. Susceptibility to IL-2 is accompanied with the expression of the IL-2 receptor (Tac antigen) on B cells. The data suggest that IL-2 is one of the lymphokines directly involved in the activation of B lymphocytes.

  5. The important role of von Willebrand factor in platelet-derived FVIII gene therapy for murine hemophilia A in the presence of inhibitory antibodies.

    Science.gov (United States)

    Shi, Q; Schroeder, J A; Kuether, E L; Montgomery, R R

    2015-07-01

    Our previous studies have demonstrated that targeting FVIII expression to platelets results in FVIII storage together with von Willebrand factor (VWF) in platelet α-granules and that platelet-derived FVIII (2bF8) corrects the murine hemophilia A phenotype even in the presence of high-titer anti-FVIII inhibitory antibodies (inhibitors). To explore how VWF has an impact on platelet gene therapy for hemophilia A with inhibitors. 2bF8 transgenic mice in the FVIII(-/-) background (2bF8(tg+/-) F8(-/-) ) with varying VWF phenotypes were used in this study. Animals were analyzed by VWF ELISA, FVIII activity assay, Bethesda assay and tail clip survival test. Only 18% of 2bF8(tg+/-) F8(-/-) VWF(-/-) animals, in which VWF was deficient, survived the tail clip challenge with inhibitor titers of 3-8000 BU mL(-1) . In contrast, 82% of 2bF8(tg+/-) F8(-/-) VWF(+/+) mice, which had normal VWF levels, survived tail clipping with inhibitor titers of 10-50,000 BU mL(-1) . All 2bF8(tg+/-) F8(-/-) VWF(-/-) mice without inhibitors survived tail clipping and no VWF(-/-) F8(-/-) mice survived this challenge. Because VWF is synthesized by endothelial cells and megakaryocytes and is distributed in both plasma and platelets in peripheral blood, we further investigated the effect of each compartment of VWF on platelet-FVIII gene therapy for hemophilia A with inhibitors. In the presence of inhibitors, 42% of animals survived tail clipping in the group with plasma-VWF and 50% survived in the platelet-VWF group. VWF is essential for platelet gene therapy for hemophilia A with inhibitors. Both platelet-VWF and plasma-VWF are required for optimal platelet-derived FVIII gene therapy for hemophilia A in the presence of inhibitors. © 2015 International Society on Thrombosis and Haemostasis.

  6. Height outcome of the recombinant human growth hormone treatment in patients with SHOX gene haploinsufficiency: a meta-analysis.

    Science.gov (United States)

    Massart, Francesco; Bizzi, Martina; Baggiani, Angelo; Miccoli, Mario

    2013-04-01

    Patients with mutations or deletions of the SHOX gene present variable growth impairment, with or without mesomelic skeletal dysplasia. If untreated, short patients with SHOX haplodeficiency (SHOXD) remain short into adulthood. Although recombinant human growth hormone (rhGH) treatment improves short-term linear growth, there are episodic data on the final height of treated SHOXD subjects. After a thorough search of the published literature for pertinent studies, we undertook a meta-analysis evaluation of the efficacy and safety of rhGH treatment in SHOXD patients. In SHOXD patients, administration of rhGH progressively improved the height deficit from baseline to 24 months, although the major catch-up growth was detected after 12 months. The rhGH-induced growth appeared constant until final height. Our meta-analysis suggested rhGH therapy improves height outcome of SHOXD patients, though future studies using carefully titrated rhGH protocols are needed. Original submitted 29 October 2012; Revision submitted 22 February 2013.

  7. Growth Hormone Safety Workshop Position Paper: a critical appraisal of recombinant human growth hormone therapy in children and adults

    DEFF Research Database (Denmark)

    Allen, David B; Backeljauw, Philippe; Bidlingmaier, Martin

    2015-01-01

    Recombinant human GH (rhGH) has been in use for 30 years, and over that time its safety and efficacy in children and adults has been subject to considerable scrutiny. In 2001, a statement from the GH Research Society (GRS) concluded that 'for approved indications, GH is safe'; however, the statem...... (PES) convened a meeting to reappraise the safety of rhGH. The ouput of the meeting is a concise position statement.......Recombinant human GH (rhGH) has been in use for 30 years, and over that time its safety and efficacy in children and adults has been subject to considerable scrutiny. In 2001, a statement from the GH Research Society (GRS) concluded that 'for approved indications, GH is safe'; however......, the statement highlighted a number of areas for on-going surveillance of long-term safety, including cancer risk, impact on glucose homeostasis, and use of high dose pharmacological rhGH treatment. Over the intervening years, there have been a number of publications addressing the safety of rhGH with regard...

  8. Therapeutic Recombinant Monoclonal Antibodies

    Science.gov (United States)

    Bakhtiar, Ray

    2012-01-01

    During the last two decades, the rapid growth of biotechnology-derived techniques has led to a myriad of therapeutic recombinant monoclonal antibodies with significant clinical benefits. Recombinant monoclonal antibodies can be obtained from a number of natural sources such as animal cell cultures using recombinant DNA engineering. In contrast to…

  9. The Effect of Recombinant Growth Hormone Treatment in Children with Idiopathic Short Stature and Low Insulin-Like Growth Factor-1 Levels.

    Science.gov (United States)

    Şıklar, Zeynep; Kocaay, Pınar; Çamtosun, Emine; İsakoca, Mehmet; Hacıhamdioğlu, Bülent; Savaş Erdeve, Şenay; Berberoğlu, Merih

    2015-12-01

    Idiopathic short stature (ISS) constitutes a heterogeneous group of short stature which is not associated with an endocrine or other identifiable cause. Some ISS patients may have varying degrees of insulin-like growth factor-1 (IGF-1) deficiency. Recombinant growth hormone (rGH) treatment has been used by some authors with variable results. Reports on long-term rGH treatment are limited. In this study, 21 slowly growing, non-GH-deficient ISS children who received rGH treatment for 3.62±0.92 years were evaluated at the end of a 5.42±1.67-year follow-up period. The study group included patients with low IGF-1 levels who also responded well to an IGF generation test. The patients were divided into two groups as good responders [height increment >1 standard deviation (SD)] and poor responders (height increment deficit and almost 40% of patients may reach their target height.

  10. Transposon mutagenesis to improve the growth of recombinant Saccharomyces cerevisiae on D-xylose

    Science.gov (United States)

    Haiying Ni; Jose M. Laplaza; Thomas W. Jeffries

    2007-01-01

    Saccharomyces cerevisiae L2612 transformed with genes for xylose reductase and xylitol dehydrogenase (XYL1 and XYL2) grows well on glucose but very poorly on D-xylose. When a gene for D-xylulokinase (XYL3 or XKS1) is overexpressed, growth on glucose is unaffected, but growth on xylose is blocked. Spontaneous or chemically induced mutants of this engineered yeast that...

  11. Quantitative trait local analysis of growth-related traits in a new Arabidopsis recombinant inbred population

    NARCIS (Netherlands)

    El-Lithy, M.E.M.; Clerkx, E.J.M.; Ruijs, G.J.; Koornneef, M.; Vreugdenhil, D.

    2004-01-01

    Arabidopsis natural variation was used to analyze the genetics of plant growth rate. Screening of 22 accessions revealed a large variation for seed weight, plant dry weight and relative growth rate but not for water content. A positive correlation was observed between seed weight and plant area 10 d

  12. Effects of growth hormone deficiency and recombinant growth hormone therapy on postprandial gallbladder motility and cholecystokinin release.

    NARCIS (Netherlands)

    Moschetta, A.; Twickler, M.; Rehfeld, J.F.; Ooteghem, N.A. van; Castro Cabezas, M.; Portincasa, P.; Berge-Henegouwen, G.P. van; Erpecum, K.J. van

    2004-01-01

    In addition to cholecystokinin, other hormones have been suggested to be involved in regulation of postprandial gallbladder contraction. We aimed to evaluate effects of growth hormone (GH) on gallbladder contractility and cholecystokinin release. Gallbladder and gastric emptying (by ultrasound) and

  13. The effect of recombinant growth hormone on intestinal anastomotic wound healing in rats with obstructive jaundice.

    Science.gov (United States)

    Cağlikülekçi, Mehmet; Ozçay, Necdet; Oruğ, Taner; Aydoğ, Gülden; Renda, Nurten; Atalay, Fuat

    2002-03-01

    Several clinical and experimental studies have shown that obstructive jaundice delays wound healing. Growth hormone may prevent delayed wound healing, since it has effects on the release of mediators in jaundice, as well as increasing the protein synthesis. Forty male Wistar rats were allocated to four groups: Group I (n=10): intestinal anastomosis to normal small bowel, Group II (n=10): intestinal anastomosis to normal small bowel followed by growth hormone therapy (2mg/kg/day, subcutaneously), Group III (n=10): intestinal anastomosis to obstructive jaundice rat's small bowel, Group IV (n=10): intestinal anastomosis to obstructive jaundice rat's small bowel followed by growth hormone therapy at the same dosage The animals were observed for seven days then killed. Intraabdominal adhesions, anastomotic complications and anastomotic bursting pressures were recorded and tissue samples from the anastomotic site were obtained to measure hydroxyproline levels and for histopathologic examination. Growth hormone had a beneficial effect on the healing of intestinal anastomosis in both jaundiced and non-jaundiced rats. This was demonstrated by clinical and mechanical parameters such as a significant increase in anastomotic bursting pressure, hydroxyproline content and histopathological scores. Growth hormone reverses the adverse effects of obstructive jaundice on small bowel anastomotic healing. It can be hypothesized that this effect is due to augmentation of insulin-like growth factors, protection of hepatocytes, enhancement of intestinal epithelization, and reversal of the resultant malnutritional state caused by growth hormone in obstructive jaundice.

  14. Heterologous expression of a truncated form of human recombinant vascular endothelial growth factor-A and its biological activity in wound healing.

    Science.gov (United States)

    Khaki, Mohsen; Salmanian, Ali Hatef; Mosayebi, Ghasem; Baazm, Maryam; Babaei, Saeed; Molaee, Neda; Abtahi, Hamid

    2017-07-01

    Vascular endothelial growth factor (VEGF) is one of the most effective proteins in angiogenesis, mesenchymal stem cells (MSCs) differentiation and wound healing. These abilities are therapeutic potential of VEGF in diabetic retinopathy, nephropathy and other tissue damage circumstances. In this study, recombinant VEGF was produced in Escherichia coli ( E. coli ) system and then biological activity of this protein was evaluated in animal wound healing. E. coli BL21 (DE3) competent cells were transformed with pET32a-VEGF clone and induced by isopropyl-β-D-thio-galactoside (IPTG). The recombinant protein was purified by affinity chromatography. Recombinant VEGF-A-based ointment (VEGF/Vaseline 0.8 mg/100 w/w) was used for external wound (25×15mm thickness) healing in animal model. In vivo activity of ointment was evaluated by clinical evidences and cytological microscopic assessment. The recombinant protein with molecular weight of 45 kilodaltons (kDa) and concentration of 0.8 mg/ml was produced. Immunoblotting data showed that the antigenic region of VEGF can be expressed in E. coli and the recombinant protein has similar epitopes with close antigenic properties to the natural form. Macroscopic findings and microscopic data showed that the recombinant VEGF-A ointment was effective on excisional wound healing. Recombinant VEGF-A produced by pET32a in E. coli , possesses acceptable structure and has wound healing capability.

  15. Heterologous expression of a truncated form of human recombinant vascular endothelial growth factor-A and its biological activity in wound healing

    Directory of Open Access Journals (Sweden)

    Mohsen Khaki

    2017-07-01

    Full Text Available Objective(s: Vascular endothelial growth factor (VEGF is one of the most effective proteins in angiogenesis, mesenchymal stem cells (MSCs differentiation and wound healing. These abilities are therapeutic potential of VEGF in diabetic retinopathy, nephropathy and other tissue damage circumstances. In this study, recombinant VEGF was produced in Escherichia coli (E. coli system and then biological activity of this protein was evaluated in animal wound healing. Materials and Methods: E. coli BL21 (DE3 competent cells were transformed with pET32a-VEGF clone and induced by isopropyl-β-D-thio-galactoside (IPTG. The recombinant protein was purified byaffinity chromatography. Recombinant VEGF-A-based ointment (VEGF/Vaseline 0.8 mg/100 w/w was used for external wound (25×15mm thickness healing in animal model. In vivo activity of ointment was evaluated by clinical evidences and cytological microscopic assessment. Results: The recombinant protein with molecular weight of 45 kilodaltons (kDa and concentration of 0.8 mg/ml was produced.Immunoblotting data showed that the antigenic region of VEGF can be expressed in E. coli and the recombinant protein has similar epitopes with close antigenic properties to the natural form. Macroscopic findings and microscopic data showed that the recombinant VEGF-A ointment was effective on excisional wound healing. Conclusion: Recombinant VEGF-A produced by pET32a in E. coli, possesses acceptable structure and has wound healing capability.

  16. Epidermal growth factor receptor antibody plus recombinant human endostatin in treatment of hepatic metastases after remnant gastric cancer resection

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    We report a 55-year-old male who developed advanced hepatic metastasis and peritoneal carcinomatosis after resection of remnant gastric cancer resection 3 mo ago. The patient only received epidermal growth factor (EGF) receptor antibody (Cetuximab) plus recombinant human endostatin (Endostar).Anti-tumor activity was assessed by 18F-fluorodeoxyglucose (18F-FDG)positron emission tomography/computer tomography (PET/CT) at baseline and then every 4 wk. The case illustrates that 18FDG-PET/CT could make an early prediction of the response to Cetuximab plus Endostar in such clinical situations. 18FDG-PET/CT is a useful molecular imaging modality to evaluate the biological response advanced hepatic metastasis and peritoneal carcinomatosis to Cetuximab plus Endostar in patients after remnant gastric cancer resection.

  17. Decreased prothrombotic effects of pegylated recombinant human megakaryocyte growth and development factor in thrombocytopenic state in a rat thrombosis model.

    Science.gov (United States)

    Nishiyama, U; Kuwaki, T; Akahori, H; Kato, T; Ikeda, Y; Miyazaki, H

    2005-02-01

    Previous in vitro studies demonstrated that thrombopoietin (TPO) acts on platelets to activate a variety of intracellular signaling pathways and to enhance platelet sensitivity to multiple agonists. Little is known, however, about whether TPO exerts prothrombotic effects in vivo. The aim of this study was to examine the effects of pegylated recombinant human megakaryocyte growth and development factor (PEG-rHuMGDF), a pegylated N-terminal domain of human TPO, in a rat model of venous thrombosis. A microthrombus was photochemically induced on the vessel wall of a mesenteric venule, but the vessel was not occluded by it. A single intravenous injection of PEG-rHuMGDF (3 microg kg(-1)) after the thrombus generation into normal rats enhanced the thrombus size, resulting in transient thrombotic occlusion in the majority of rats. Stimulatory effects on thrombus growth were also observed following administration of glycosylated recombinant human full-length TPO (6 microg kg(-1)). In rats rendered thrombocytopenic by total body irradiation, however, PEG-rHuMGDF, even at 300 microg kg(-1), did not induce a significant increase in thrombus size or thrombotic occlusion. Platelets from thrombocytopenic rats had decreased surface levels of c-Mpl and decreased sensitivity to PEG-rHuMGDF in an in vitro aggregation response. Thus, decreased prothrombotic effects of PEG-rHuMGDF in thrombocytopenic rats might be the result not only of low platelet counts but also of decreased platelet reactivity to PEG-rHuMGDF. These results indicate that PEG-rHuMGDF has little effect on venous thrombus formation in thrombocytopenic states associated with high endogenous TPO levels.

  18. GrowthHormone Research Society workshop summary: consensus guidelines for recombinant human growth hormone therapy in Prader-Willi syndrome.

    Science.gov (United States)

    Deal, Cheri L; Tony, Michèle; Höybye, Charlotte; Allen, David B; Tauber, Maïthé; Christiansen, Jens Sandahl

    2013-06-01

    Recombinant human GH (rhGH) therapy in Prader-Willi syndrome (PWS) has been used by the medical community and advocated by parental support groups since its approval in the United States in 2000 and in Europe in 2001. Its use in PWS represents a unique therapeutic challenge that includes treating individuals with cognitive disability, varied therapeutic goals that are not focused exclusively on increased height, and concerns about potential life-threatening adverse events. The aim of the study was to formulate recommendations for the use of rhGH in children and adult patients with PWS. We performed a systematic review of the clinical evidence in the pediatric population, including randomized controlled trials, comparative observational studies, and long-term studies (>3.5 y). Adult studies included randomized controlled trials of rhGH treatment for ≥ 6 months and uncontrolled trials. Safety data were obtained from case reports, clinical trials, and pharmaceutical registries. Forty-three international experts and stakeholders followed clinical practice guideline development recommendations outlined by the AGREE Collaboration (www.agreetrust.org). Evidence was synthesized and graded using a comprehensive multicriteria methodology (EVIDEM) (http://bit.ly.PWGHIN). Following a multidisciplinary evaluation, preferably by experts, rhGH treatment should be considered for patients with genetically confirmed PWS in conjunction with dietary, environmental, and lifestyle interventions. Cognitive impairment should not be a barrier to treatment, and informed consent/assent should include benefit/risk information. Exclusion criteria should include severe obesity, uncontrolled diabetes mellitus, untreated severe obstructive sleep apnea, active cancer, or psychosis. Clinical outcome priorities should vary depending upon age and the presence of physical, mental, and social disability, and treatment should be continued for as long as demonstrated benefits outweigh the risks.

  19. Recombinant bovine growth hormone (rBGH) enhances somatic growth by regulating the GH-IGF axis in fingerlings of gilthead sea bream (Sparus aurata).

    Science.gov (United States)

    Vélez, Emilio J; Perelló, Miquel; Azizi, Sheida; Moya, Alberto; Lutfi, Esmail; Pérez-Sánchez, Jaume; Calduch-Giner, Josep A; Navarro, Isabel; Blasco, Josefina; Fernández-Borràs, Jaume; Capilla, Encarnación; Gutiérrez, Joaquim

    2018-02-01

    The growth hormone (GH)/insulin-like growth factors (IGFs) endocrine axis is the main growth-regulator system in vertebrates. Some authors have demonstrated the positive effects on growth of a sustained-release formulation of a recombinant bovine GH (rBGH) in different fish species. The aim of this work was to characterize the effects of a single injection of rBGH in fingerlings of gilthead sea bream on growth, GH-IGF axis, and both myogenic and osteogenic processes. Thus, body weight and specific growth rate were significantly increased in rBGH-treated fish respect to control fish at 6weeks post-injection, whereas the hepatosomatic index was decreased and the condition factor and mesenteric fat index were unchanged, altogether indicating enhanced somatic growth. Moreover, rBGH injection increased the plasma IGF-I levels in parallel with a rise of hepatic mRNA from total IGF-I, IGF-Ic and IGF-II, the binding proteins IGFBP-1a and IGFBP-2b, and also the receptors IGF-IRb, GHR-I and GHR-II. In skeletal muscle, the expression of IGF-Ib and GHR-I was significantly increased but that of IGF-IRb was reduced; the mRNA levels of myogenic regulatory factors, proliferation and differentiation markers (PCNA and MHC, respectively), or that of different molecules of the signaling pathway (TOR/AKT) were unaltered. Besides, the growth inhibitor myostatin (MSTN1 and MSTN2) and the hypertrophic marker (MLC2B) expression resulted significantly enhanced, suggesting altogether that the muscle is in a non-proliferative stage of development. Contrarily in bone, although the expression of most molecules of the GH/IGF axis was decreased, the mRNA levels of several osteogenic genes were increased. The histology analysis showed a GH induced lipolytic effect with a clear decrease in the subcutaneous fat layer. Overall, these results reveal that a better growth potential can be achieved on this species and supports the possibility to improve growth and quality through the optimization of its

  20. Taguchi Experimental Design for Optimization of Recombinant Human Growth Hormone Production in CHO Cell Lines and Comparing its Biological Activity with Prokaryotic Growth Hormone.

    Science.gov (United States)

    Aghili, Zahra Sadat; Zarkesh-Esfahani, Sayyed Hamid

    2018-02-01

    Growth hormone deficiency results in growth retardation in children and the GH deficiency syndrome in adults and they need to receive recombinant-GH in order to rectify the GH deficiency symptoms. Mammalian cells have become the favorite system for production of recombinant proteins for clinical application compared to prokaryotic systems because of their capability for appropriate protein folding, assembly, post-translational modification and proper signal. However, production level in mammalian cells is generally low compared to prokaryotic hosts. Taguchi has established orthogonal arrays to describe a large number of experimental situations mainly to reduce experimental errors and to enhance the efficiency and reproducibility of laboratory experiments.In the present study, rhGH was produced in CHO cells and production of rhGH was assessed using Dot blotting, western blotting and Elisa assay. For optimization of rhGH production in CHO cells using Taguchi method An M16 orthogonal experimental design was used to investigate four different culture components. The biological activity of rhGH was assessed using LHRE-TK-Luciferase reporter gene system in HEK-293 and compared to the biological activity of prokaryotic rhGH.A maximal productivity of rhGH was reached in the conditions of 1%DMSO, 1%glycerol, 25 µM ZnSO 4 and 0 mM NaBu. Our findings indicate that control of culture conditions such as the addition of chemical components helps to develop an efficient large-scale and industrial process for the production of rhGH in CHO cells. Results of bioassay indicated that rhGH produced by CHO cells is able to induce GH-mediated intracellular cell signaling and showed higher bioactivity when compared to prokaryotic GH at the same concentrations. © Georg Thieme Verlag KG Stuttgart · New York.

  1. Recombinant β-1,3-1,4-glucanase from Theobroma cacao impairs Moniliophthora perniciosa mycelial growth.

    Science.gov (United States)

    Britto, Dahyana Santos; Pirovani, Carlos Priminho; Andrade, Bruno Silva; Dos Santos, Tassiara Pereira; Pungartnik, Cristina; Cascardo, Júlio Cezar M; Micheli, Fabienne; Gesteira, Abelmon S

    2013-09-01

    In this work, we identified a gene from Theobroma cacao L. genome and cDNA libraries, named TcGlu2, that encodes a β-1,3-1,4-glucanase. The TcGlu2 ORF was 720 bp in length and encoded a polypeptide of 239 amino acids with a molecular mass of 25.58 kDa. TcGlu2 contains a conserved domain characteristic of β-1,3-1,4-glucanases and presented high protein identity with β-1,3-1,4-glucanases from other plant species. Molecular modeling of TcGlu2 showed an active site of 13 amino acids typical of glucanase with β-1,3 and 1,4 action mode. The recombinant cDNA TcGlu2 obtained by heterologous expression in Escherichia coli and whose sequence was confirmed by mass spectrometry, has a molecular mass of about 22 kDa (with His-Tag) and showed antifungal activity against the fungus Moniliophthora perniciosa, causal agent of the witches' broom disease in cacao. The integrity of the hyphae membranes of M. perniciosa, incubated with protein TcGlu2, was analyzed with propidium iodide. After 1 h of incubation, a strong fluorescence emitted by the hyphae indicating the hydrolysis of the membrane by TcGlu2, was observed. To our knowledge, this is the first study of a cacao β-1,3-1,4-glucanase expression in heterologous system and the first analysis showing the antifungal activity of a β-1,3-1,4-glucanase, in particular against M. perniciosa.

  2. Influence of recombinant interferon alpha on nutritional status and growth pattern in children with chronic viral hepatitis.

    Science.gov (United States)

    Gottrand, F; Michaud, L; Guimber, D; Ategbo, S; Dubar, G; Turck, D; Farriaux, J P

    1996-12-01

    Anorexia and weight loss are frequently reported as adverse effects during recombinant interferon alpha (rIFN-alpha) treatment. The aim of the present study was to assess both nutritional status and growth of children and adolescents treated with rIFN-alpha for chronic viral hepatitis. Eleven patients aged 4-16 years with histologically proven chronic active hepatitis (hepatitis B, n = 9; hepatitis C, n = 2) receiving rIFN-alpha subcutaneously thrice a week for 6 months were studied. Weight and height increments were assessed during the 6 months before starting rIFN-alpha. Weight and height were measured every 3 months (M0, M3, M6) during the 6 months of rIFN-alpha treatment, then every 6 months during the follow up period (6-36 months). Weight decreased in every child during rIFN-alpha treatment (weight loss varies from 0.5 to 2.6 kg after 3 months of treatment). Weight/age Z-score decreased from 0.12 at M0 to -0.69 at M3 (P Nutritional status was significantly impaired during treatment (Z-score for weight/height decreased from 0.18 at M0 to -0.74 at M3, P nutritional status are encountered constantly at the beginning of rIFN-alpha therapy without any deleterious effect on growth. Information of the families and nutritional intervention during treatment should be required, in order to limit the importance of weight loss.

  3. Health-Related Quality of Life of Young Adults Treated with Recombinant Human Growth Hormone during Childhood.

    Directory of Open Access Journals (Sweden)

    Grit Sommer

    Full Text Available Since recombinant human growth hormone (rhGH became available in 1985, the spectrum of indications has broadened and the number of treated patients increased. However, long-term health-related quality of life (HRQoL after childhood rhGH treatment has rarely been documented. We assessed HRQoL and its determinants in young adults treated with rhGH during childhood.For this study, we retrospectively identified former rhGH patients in 11 centers of paediatric endocrinology, including university hospitals and private practices. We sent a questionnaire to all patients treated with rhGH for any diagnosis, who were older than 18 years, and who resided in Switzerland at time of the survey. Three hundred participants (58% of 514 eligible returned the questionnaire. Mean age was 23 years; 56% were women; 43% had isolated growth hormone deficiency, or idiopathic short stature; 43% had associated diseases or syndromes, and 14% had growth hormone deficiency after childhood cancer. Swiss siblings of childhood cancer survivors and the German norm population served as comparison groups. HRQoL was assessed using the Short Form-36. We found that the Physical Component Summary of healthy patients with isolated growth hormone deficiency or idiopathic short stature resembled that of the control group (53.8 vs. 54.9. Patients with associated diseases or syndromes scored slightly lower (52.5, and former cancer patients scored lowest (42.6. The Mental Component Summary was similar for all groups. Lower Physical Component Summary was associated with lower educational level (coeff. -1.9. Final height was not associated with HRQoL.In conclusion, HRQoL after treatment with rhGH in childhood depended mainly on the underlying indication for rhGH treatment. Patients with isolated growth hormone deficiency/idiopathic short stature or patients with associated diseases or syndromes had HRQoL comparable to peers. Patients with growth hormone deficiency after childhood cancer were

  4. A radioimmunoassay of chicken growth hormone using growth hormone produced by recombinant DNA technology: validation and observations of plasma hormone variations in genetically fat and lean chickens

    International Nuclear Information System (INIS)

    Picaper, G.; Leclercq, B.; Saadoun, A.; Mongin, P.

    1986-01-01

    A radioimmunoassay (RIA) of chicken growth hormone (c-GH) has been developed using growth hormone produced by recombinant DNA technology. The best rabbit antiserum was used at 1/300,000 final dilution. Hormone labelling by iodine-125, achieved by chloramine T, allowed a specific activity of 3.7 MBq/μg. The equilibrium curves show that optimal conditions of incubation were reached at room temperature for 24h. This RIA used a second sheep antibody which precipitated the whole c-GH bound to the first antibody in the presence of polyethylene glycol solution (6%) at room temperature for 30 min. In our conditions, sensitivity was about 30 pg of c-GH per tube. Coefficient of variation was around 10%. No cross reaction was found with avian LH and prolactin. Thyrotrophin-releasing hormone (TRH) injection to young chickens induced 20-fold higher plasma c-GH concentrations. Simultaneous injection of somatostatin and TRH slightly reduced these concentrations. Hypoglycemia induced by insulin led to a drop of the plasma c-GH concentration. Conversely, refeeding or glucose load induced slight increases of the c-GH level. Genetically fat chickens tended to exhibit higher plasma c-GH concentrations than lean chickens

  5. Translational mixed-effects PKPD modelling of recombinant human growth hormone - from hypophysectomized rat to patients

    DEFF Research Database (Denmark)

    Thorsted, Anders; Thygesen, Peter; Agersø, Henrik

    2016-01-01

    was developed from experimental PKPD studies of rhGH and effects of long-term treatment as measured by insulin-like growth factor 1 (IGF-1) and bodyweight gain in rats. Modelled parameter values were scaled to human values using the allometric approach with fixed exponents for PKs and unscaled for PDs...... and validated through simulations relative to patient data. KEY RESULTS: The final model described rhGH PK as a two compartmental model with parallel linear and non-linear elimination terms, parallel first-order absorption with a total s.c. bioavailability of 87% in rats. Induction of IGF-1 was described...... by an indirect response model with stimulation of kin and related to rhGH exposure through an Emax relationship. Increase in bodyweight was directly linked to individual concentrations of IGF-1 by a linear relation. The scaled model provided robust predictions of human systemic PK of rhGH, but exposure following...

  6. A recombinant lentiviral PDGF-driven mouse model of proneural glioblastoma.

    Science.gov (United States)

    Rahme, Gilbert J; Luikart, Bryan W; Cheng, Chao; Israel, Mark A

    2018-02-19

    Mouse models of glioblastoma (GBM), the most aggressive primary brain tumor, are critical for understanding GBM pathology and can contribute to the preclinical evaluation of therapeutic agents. Platelet-derived growth factor (PDGF) signaling has been implicated in the development and pathogenesis of GBM, specifically the proneural subtype. Although multiple mouse models of PDGF-driven glioma have been described, they require transgenic mice engineered to activate PDGF signaling and/or impair tumor suppressor genes and typically represent lower-grade glioma. We designed recombinant lentiviruses expressing both PDGFB and a short hairpin RNA targeting Cdkn2a to induce gliomagenesis following stereotactic injection into the dentate gyrus of adult immunocompetent mice. We engineered these viruses to coexpress CreERT2 with PDGFB, allowing for deletion of floxed genes specifically in transduced cells, and designed another version of this recombinant lentivirus in which enhanced green fluorescent protein was coexpressed with PDGFB and CreERT2 to visualize transduced cells. The dentate gyrus of injected mice showed hypercellularity one week post-injection and subsequently developed bona fide tumors with the pathologic hallmarks of GBM leading to a median survival of 77 days post-injection. Transcriptomic analysis of these tumors revealed a proneural gene expression signature. Informed by the genetic alterations observed in human GBM, we engineered a novel mouse model of proneural GBM. While reflecting many of the advantages of transgenic mice, this model allows for the facile in vivo testing of gene function in tumor cells and makes possible the rapid production of large numbers of immunocompetent tumor-bearing mice for preclinical testing of therapeutics. © The Author(s) 2017. Published by Oxford University Press on behalf of the Society for Neuro-Oncology. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com

  7. Recombinant growth differentiation factor 11 influences short-term memory and enhances Sox2 expression in middle-aged mice.

    Science.gov (United States)

    Zhang, Min; Jadavji, Nafisa M; Yoo, Hyung-Suk; Smith, Patrice D

    2018-04-02

    Previous evidence suggests that a significant decline in cognitive ability begins during middle-age and continues to deteriorate with increase in age. Recent work has demonstrated the potential rejuvenation impact of growth differentiation factor-11 (GDF-11) in aged mice. We carried out experiments to evaluate the impact of a single dose of recombinant (rGDF-11) on short-term visual and spatial memory in middle-aged male mice. On the novel object recognition task, we observed middle-aged mice treated rGDF-11 showed improved performance on the novel object recognition task. However, middle-aged mice did not show increased expression of phosphorylated-Smad2/3, a downstream effector of GDF-11. We noted however that the expression of the transcription factor, Sox2 was increased within the dentate gyrus. Our data suggest that a single injection of rGDF-11 contributes to improvements in cognitive function of middle-aged animals, which may be critical in the preservation of short-term memory capacity in old age. Copyright © 2017 Elsevier B.V. All rights reserved.

  8. Efficacy of recombinant bovine epidermal growth factor in the treatment of experimental subclinical Staphylococcus aureus mastitis in a ewe model.

    Science.gov (United States)

    Gabadage, Kamal; Chirino-Trejo, Manuel; Campbell, John; Luby, Christopher

    2017-01-01

    Staphylococcus aureus is the most common contagious mastitis pathogen of dairy cattle. Antimicrobial treatment of infected cattle results in variable cure rates. Epidermal growth factor (EGF) plays an important role in the modulation of host innate immune responses and the regulation of mammary epithelial regeneration, indicating that EGF may be useful as a treatment for mastitis. A pilot study was conducted to evaluate the efficacy of recombinant bovine EGF (rbEGF) for the treatment of S aureus intramammary infection (IMI) using an ovine model. Each ewe was experimentally infected with S aureus in both udder halves. One udder half of each ewe received one of two treatments: EGF (n=13) or pirlimycin (n=13). The contralateral udder half of each ewe received sterile saline as a control. The bacteriological cure rate following rbEGF was significantly lower (15 per cent) than that attained with pirlimycin hydrochloride (61 per cent) and did not differ from that following treatment with sterile saline. Cure rates following treatment with rbEGF were not significantly different to those following sterile saline. Given that EGF is associated with modulation of host immunity and wound healing, future studies into EGF should not focus on whether EGF increases cure rates of S aureus IMI.

  9. Absolute hypoxic exercise training enhances in vitro thrombin generation by increasing procoagulant platelet-derived microparticles under high shear stress in sedentary men.

    Science.gov (United States)

    Chen, Yu-Wen; Chen, Yi-Ching; Wang, Jong-Shyan

    2013-05-01

    HS (high shear) stress associated with artery stenosis facilitates TG (thrombin generation) by increasing the release of procoagulant PDMPs (platelet-derived microparticles). Physical exercise and hypoxia may paradoxically modulate vascular thrombotic risks. The aim of the present study was to investigate how exercise training with/without hypoxia affected TG mediated by PDMPs under physio-pathological shear flows. A total of 75 sedentary males were randomly divided into five groups (n=15 in each group): 21% O2 [NC (normoxic control)] or 15% O2 [HC (hypoxic control)] at rest or were trained at 50% of peak work rate under 21% O2 [NT (normoxic training)] or 15% O2 [HAT (hypoxic-absolute training)], or 50% of HR (heart rate) reserve under 15% O2 [HRT (hypoxic-relative training)] for 30 min/day, 5 days/week for 4 weeks. The PDMP characteristics and dynamic TG were measured by flow cytometry and thrombinography respectively. Before the intervention, strenuous exercise markedly increased the PDMP count (14.8%) and TG rate (19.5%) in PDMP-rich plasma at 100 dynes/cm2 of shear stress (Pexercise. Conversely, HAT notably promoted the PDMP count (37.3%) and TG rate (38.9%) induced by HS (Pexercise. We conclude that both HRT and NT depress similarly HS-mediated TG during exercise, but HAT accelerates the prothrombotic response to vigorous exercise. These findings provide new insights into how exercise training under a hypoxic condition influences the risk of thrombosis associated with stenotic arteries.

  10. A new pen device for injection of recombinant human growth hormone: a convenience, functionality and usability evaluation study

    Directory of Open Access Journals (Sweden)

    Sauer M

    2017-12-01

    Full Text Available   Maritta Sauer,1 Carole Abbotts2 1Global Strategic Insights, Merck KGaA, Darmstadt, Germany; 2Pharmaceutical Market Research Consultant, London, UK Purpose: Adherence to recombinant human growth hormone (r-hGH is critical to growth and other outcomes in patients with growth disorders, but the requirement for daily injections means that ease of use is an important factor. This study assessed the perceived ease of use and functionality of the prototype of a reusable pen injector (pen device for r-hGH that incorporates several advanced features. Participants and methods: Semi-structured 60-minute qualitative interviews were conducted in 5 countries with 57 health care professionals (HCPs and 30 patients with GH deficiency/caregivers administering r-hGH to patients, including children. HCPs had to be responsible for training in the use of r-hGH pen devices and to see ≥4 r-hGH patients/caregivers per month. Patients/caregivers had to have experience with r-hGH administration for at least 6 months.Results: Thirty-seven (65% of HCPs described the pen device as “simple” or “easy” to use. The aluminum body was generally perceived as attractive, high quality and comfortable to hold and operate. The ease of preparation and use made it suitable for both children and adults. The ability to dial back the r-hGH dose, if entered incorrectly, was mentioned as a major benefit, because other devices need several user steps to reset. Patients/caregivers felt the pen device was easy to use and the injection-feedback features reassured them that the full dose had been given. Overall, 40 (70% HCPs and 16 (52% patients/caregivers were likely to recommend or request the pen device. Moreover, patients/caregivers rated the pen device higher than their current reusable pens and almost equal to the leading disposable device for ease of learning, preparation, administration and ease of use.Conclusion: The prototype pen device successfully met its design

  11. [HPV DNA vaccines expressing recombinant CRT/HPV6bE7 fusion protein inhibit tumor growth and angiogenic activity].

    Science.gov (United States)

    Xu, Yan; Cheng, Hao; Zhao, Ke-Jia; Zhu, Ke-Jian; Zhang, Xing

    2007-11-01

    This paper was to study the angiogenic inhibitory effect and the potential antitumor effect of the constructed recombinant DNA vaccine CRT/HPV6bE7 in vivo. The C57BL/6 mice were vaccinated respectively with recombinant CRT/HPV6bE7 DNA plamids. The inhibitory effects on angiogenesis of generated vaccines in vivo were evaluated by a bFGF-induced angiogenesis assay using the Matrigel kit. To investigate the potential antitumor effect, the mean tumor weights, sizes and tumor appearing times were measured in C57BL/6 mice treated with HPV6bE7-expressing B16 cells. The results indicated that the recombinants CRT180/HPV6bE7 and CRT180 showed strong anti-angiogenic effects in bFGF-induced angiogenesis in vivo. Moreover, CRT180/HPV6bE7 and CRT180 DNA vaccines could significantly inhibit the tumor growth in tumor challenge experiment, and CRT180/HPV6bE7 was superior to other vaccines in delaying tumor formation time, limiting tumor size and weight in tumor protection experiment. In conclusion, recombinant CRT180/HPV6bE7 DNA could elicit a most efficient anti-angiogenic effect and inhibit tumor growth in mice inoculated with DNA vaccines. The antiangiogenic activity of CRT were suggested residing in a domain between CRT 120-180 aa.

  12. Long-term efficacy of recombinant human growth hormone therapy in short-statured patients with Noonan syndrome

    Directory of Open Access Journals (Sweden)

    Insook Jeong

    2016-03-01

    Full Text Available PurposeNoonan syndrome (NS is characterized by short stature, heart anomalies, developmental delays, dysmorphic features, cryptorchidism, and coagulation defects. Several studies reported the short-term effects of recombinant human growth hormone (rhGH treatment on the improvement of height. This study was performed to evaluate the long-term efficacy of rhGH in children with NS in Korea.MethodsThis study included 15 prepubertal NS children who received rhGH subcutaneously at a dose of 50–75 µg/kg/day for 6 days a week for at least >3 years. Preand posttreatment data, such as height, weight, bone age, insulin-like growth factor 1 (IGF-1, and IGF binding protein 3 (IGFBP-3 levels, were collected every 6 months.ResultsChronologic age and bone age at the start of treatment were 7.97±1.81 and 5.09±2.12 years, respectively. Height standard deviation score (SDS was increased from –2.64±0.64 to –1.54±1.24 years after 3 years (P<0.001. Serum IGF-1 SDS levels were elevated from –1.28±1.03 to –0.10±0.94 (P<0.001. Height SDS was more increased in subjects without PTPN11 mutations compared to those with mutations after 3 years (P=0.012. However, the other parameters, including bone age, IGF-1 SDS, and IGFBP-3 SDS, were not significantly different between patients with and without PTPN11 mutations.ConclusionAlthough this study included a relatively small number of patients, long-term rhGH therapy in NS patients was safe and effective at improving height, growth velocity, and serum IGF-1 levels, in accordance with previous studies. However, the meticulous monitoring of potential adverse events is still needed because of high dose of rhGH and preexisting hyperactivity of RAS-MAPK pathway. Patients with PTPN11 mutations demonstrated a decreased response to rhGH therapy compared to those without mutations.

  13. Suppression of Homologous Recombination by insulin-like growth factor-1 inhibition sensitizes cancer cells to PARP inhibitors

    International Nuclear Information System (INIS)

    Amin, Oreekha; Beauchamp, Marie-Claude; Nader, Paul Abou; Laskov, Ido; Iqbal, Sanaa; Philip, Charles-André; Yasmeen, Amber; Gotlieb, Walter H.

    2015-01-01

    Impairment of homologous recombination (HR) is found in close to 50 % of ovarian and breast cancer. Tumors with BRCA1 mutations show increased expression of the Insulin-like growth factor type 1 receptor (IGF-1R). We previously have shown that inhibition of IGF-1R results in growth inhibition and apoptosis of ovarian tumor cells. In the current study, we aimed to investigate the correlation between HR and sensitivity to IGF-1R inhibition. Further, we hypothesized that IGF-1R inhibition might sensitize HR proficient cancers to Poly ADP ribose polymerase (PARP) inhibitors. Using ovarian and breast cancer cellular models with known BRCA1 status, we evaluated their HR functionality by RAD51 foci formation assay. The 50 % lethal concentration (LC50) of Insulin-like growth factor type 1 receptor kinase inhibitor (IGF-1Rki) in these cells was assessed, and western immunoblotting was performed to determine the expression of proteins involved in the IGF-1R pathway. Moreover, IGF-1R inhibitors were added on HR proficient cell lines to assess mRNA and protein expression of RAD51 by qPCR and western blot. Also, we explored the interaction between RAD51 and Insulin receptor substance 1 (IRS-1) by immunoprecipitation. Next, combination effect of IGF-1R and PARP inhibitors was evaluated by clonogenic assay. Cells with mutated/methylated BRCA1 showed an impaired HR function, and had an overactivation of the IGF-1R pathway. These cells were more sensitive to IGF-1R inhibition compared to HR proficient cells. In addition, the IGF-IR inhibitor reduced RAD51 expression at mRNA and protein levels in HR proficient cells, and sensitized these cells to PARP inhibitor. Targeting IGF-1R might lead to improved personalized therapeutic approaches in cancer patients with HR deficiency. Targeting both PARP and IGF-1R might increase the clinical efficacy in HR deficient patients and increase the population of patients who may benefit from PARP inhibitors

  14. Use of a small molecule cell cycle inhibitor to control cell growth and improve specific productivity and product quality of recombinant proteins in CHO cell cultures.

    Science.gov (United States)

    Du, Zhimei; Treiber, David; McCarter, John D; Fomina-Yadlin, Dina; Saleem, Ramsey A; McCoy, Rebecca E; Zhang, Yuling; Tharmalingam, Tharmala; Leith, Matthew; Follstad, Brian D; Dell, Brad; Grisim, Brent; Zupke, Craig; Heath, Carole; Morris, Arvia E; Reddy, Pranhitha

    2015-01-01

    The continued need to improve therapeutic recombinant protein productivity has led to ongoing assessment of appropriate strategies in the biopharmaceutical industry to establish robust processes with optimized critical variables, that is, viable cell density (VCD) and specific productivity (product per cell, qP). Even though high VCD is a positive factor for titer, uncontrolled proliferation beyond a certain cell mass is also undesirable. To enable efficient process development to achieve consistent and predictable growth arrest while maintaining VCD, as well as improving qP, without negative impacts on product quality from clone to clone, we identified an approach that directly targets the cell cycle G1-checkpoint by selectively inhibiting the function of cyclin dependent kinases (CDK) 4/6 with a small molecule compound. Results from studies on multiple recombinant Chinese hamster ovary (CHO) cell lines demonstrate that the selective inhibitor can mediate a complete and sustained G0/G1 arrest without impacting G2/M phase. Cell proliferation is consistently and rapidly controlled in all recombinant cell lines at one concentration of this inhibitor throughout the production processes with specific productivities increased up to 110 pg/cell/day. Additionally, the product quality attributes of the mAb, with regard to high molecular weight (HMW) and glycan profile, are not negatively impacted. In fact, high mannose is decreased after treatment, which is in contrast to other established growth control methods such as reducing culture temperature. Microarray analysis showed major differences in expression of regulatory genes of the glycosylation and cell cycle signaling pathways between these different growth control methods. Overall, our observations showed that cell cycle arrest by directly targeting CDK4/6 using selective inhibitor compound can be utilized consistently and rapidly to optimize process parameters, such as cell growth, qP, and glycosylation profile in

  15. Data regarding the growth of Lactobacillus acidophilus NCFM on different carbohydrates and recombinant production of elongation factor G and pyruvate kinase

    Directory of Open Access Journals (Sweden)

    Hasan Ufuk Celebioglu

    2017-10-01

    Full Text Available The present study describes the growth of the very well-known probiotic bacterium Lactobacillus acidophilus NCFM on different carbohydrates. Furthermore, recombinant production of putative moonlighting proteins elongation factor G and pyruvate kinase from this bacterium is described. For further and detailed interpretation of the data presented here, please see the research article “Mucin- and carbohydrate-stimulated adhesion and subproteome changes of the probiotic bacterium Lactobacillus acidophilus NCFM” (Celebioglu et al., 2017 [1].

  16. Camptothecin inhibits platelet-derived growth factor-BB-induced proliferation of rat aortic vascular smooth muscle cells through inhibition of PI3K/Akt signaling pathway

    Energy Technology Data Exchange (ETDEWEB)

    Park, Eun-Seok [Department of Applied Biochemistry, Division of Life Science, College of Health and Biomedical Science, Konkuk University, Chungju, Chungbuk (Korea, Republic of); Kang, Shin-il [College of Pharmacy Medical Research Center, Chungbuk National University, Cheongju (Korea, Republic of); Yoo, Kyu-dong [Hazardous Substances Analysis Division, Gwangju Regional Food and Drug Administration, Gwangju (Korea, Republic of); Lee, Mi-Yea [Department of Nursing Kyungbok University, Pocheon (Korea, Republic of); Yoo, Hwan-Soo; Hong, Jin-Tae [College of Pharmacy Medical Research Center, Chungbuk National University, Cheongju (Korea, Republic of); Shin, Hwa-Sup [Department of Applied Biochemistry, Division of Life Science, College of Health and Biomedical Science, Konkuk University, Chungju, Chungbuk (Korea, Republic of); Kim, Bokyung [Department of Physiology, Konkuk Medical School, Konkuk University, Chungju, Chungbuk (Korea, Republic of); Yun, Yeo-Pyo, E-mail: ypyun@chungbuk.ac.kr [College of Pharmacy Medical Research Center, Chungbuk National University, Cheongju (Korea, Republic of)

    2013-04-15

    The abnormal proliferation of vascular smooth muscle cells (VSMCs) in arterial wall is a major cause of vascular disorders such as atherosclerosis and restenosis after angioplasty. In this study, we investigated not only the inhibitory effects of camptothecin (CPT) on PDGF-BB-induced VSMC proliferation, but also its molecular mechanism of this inhibition. CPT significantly inhibited proliferation with IC50 value of 0.58 μM and the DNA synthesis of PDGF-BB-stimulated VSMCs in a dose-dependent manner (0.5–2 μM ) without any cytotoxicity. CPT induced the cell cycle arrest at G0/G1 phase. Also, CPT decreased the expressions of G0/G1-specific regulatory proteins including cyclin-dependent kinase (CDK)2, cyclin D1 and PCNA in PDGF-BB-stimulated VSMCs. Pre-incubation of VSMCs with CPT significantly inhibited PDGF-BB-induced Akt activation, whereas CPT did not affect PDGF-receptor beta phosphorylation, extracellular signal-regulated kinase (ERK) 1/2 phosphorylation and phospholipase C (PLC)-γ1 phosphorylation in PDGF-BB signaling pathway. Our data showed that CPT pre-treatment inhibited VSMC proliferation, and that the inhibitory effect of CPT was enhanced by LY294002, a PI3K inhibitor, on PDGF-BB-induced VSMC proliferation. In addition, inhibiting the PI3K/Akt pathway by LY294002 significantly enhanced the suppression of PCNA expression and Akt activation by CPT. These results suggest that the anti-proliferative activity of CPT is mediated in part by downregulating the PI3K/Akt signaling pathway. - Highlights: ► CPT inhibits proliferation of PDGF-BB-induced VSMC without cytotoxicity. ► CPT arrests the cell cycle in G0/G1 phase by downregulation of cyclin D1 and CDK2. ► CPT significantly attenuates Akt phosphorylation in PDGF-BB signaling pathway. ► LY294002 enhanced the inhibitory effect of CPT on VSMC proliferation. ► Thus, CPT is mediated by downregulating the PI3K/Akt signaling pathway.

  17. Camptothecin inhibits platelet-derived growth factor-BB-induced proliferation of rat aortic vascular smooth muscle cells through inhibition of PI3K/Akt signaling pathway

    International Nuclear Information System (INIS)

    Park, Eun-Seok; Kang, Shin-il; Yoo, Kyu-dong; Lee, Mi-Yea; Yoo, Hwan-Soo; Hong, Jin-Tae; Shin, Hwa-Sup; Kim, Bokyung; Yun, Yeo-Pyo

    2013-01-01

    The abnormal proliferation of vascular smooth muscle cells (VSMCs) in arterial wall is a major cause of vascular disorders such as atherosclerosis and restenosis after angioplasty. In this study, we investigated not only the inhibitory effects of camptothecin (CPT) on PDGF-BB-induced VSMC proliferation, but also its molecular mechanism of this inhibition. CPT significantly inhibited proliferation with IC50 value of 0.58 μM and the DNA synthesis of PDGF-BB-stimulated VSMCs in a dose-dependent manner (0.5–2 μM ) without any cytotoxicity. CPT induced the cell cycle arrest at G0/G1 phase. Also, CPT decreased the expressions of G0/G1-specific regulatory proteins including cyclin-dependent kinase (CDK)2, cyclin D1 and PCNA in PDGF-BB-stimulated VSMCs. Pre-incubation of VSMCs with CPT significantly inhibited PDGF-BB-induced Akt activation, whereas CPT did not affect PDGF-receptor beta phosphorylation, extracellular signal-regulated kinase (ERK) 1/2 phosphorylation and phospholipase C (PLC)-γ1 phosphorylation in PDGF-BB signaling pathway. Our data showed that CPT pre-treatment inhibited VSMC proliferation, and that the inhibitory effect of CPT was enhanced by LY294002, a PI3K inhibitor, on PDGF-BB-induced VSMC proliferation. In addition, inhibiting the PI3K/Akt pathway by LY294002 significantly enhanced the suppression of PCNA expression and Akt activation by CPT. These results suggest that the anti-proliferative activity of CPT is mediated in part by downregulating the PI3K/Akt signaling pathway. - Highlights: ► CPT inhibits proliferation of PDGF-BB-induced VSMC without cytotoxicity. ► CPT arrests the cell cycle in G0/G1 phase by downregulation of cyclin D1 and CDK2. ► CPT significantly attenuates Akt phosphorylation in PDGF-BB signaling pathway. ► LY294002 enhanced the inhibitory effect of CPT on VSMC proliferation. ► Thus, CPT is mediated by downregulating the PI3K/Akt signaling pathway

  18. Optimization of Platelet-Derived Growth Factor Receptor (PDGFR) Inhibitors for Duration of Action, as an Inhaled Therapy for Lung Remodeling in Pulmonary Arterial Hypertension.

    Science.gov (United States)

    Shaw, Duncan E; Baig, Ferheen; Bruce, Ian; Chamoin, Sylvie; Collingwood, Stephen P; Cross, Sarah; Dayal, Satish; Drückes, Peter; Furet, Pascal; Furminger, Vikki; Haggart, Deborah; Hussey, Martin; Konstantinova, Irena; Loren, Jon C; Molteni, Valentina; Roberts, Sonia; Reilly, John; Saunders, Alex M; Stringer, Rowan; Sviridenko, Lilya; Thomas, Matthew; Thomson, Christopher G; Tomlins, Christine; Wen, Ben; Yeh, Vince; Pearce, Andrew C

    2016-09-08

    A series of potent PDGFR inhibitors has been identified. The series was optimized for duration of action in the lung. A novel kinase occupancy assay was used to directly measure target occupancy after i.t. dosing. Compound 25 shows 24 h occupancy of the PDGFR kinase domain, after a single i.t. dose and has efficacy at 0.03 mg/kg, in the rat moncrotaline model of pulmonary arterial hypertension. Examination of PK/PD data from the optimization effort has revealed in vitro:in vivo correlations which link duration of action in vivo with low permeability and high basicity and demonstrate that nonspecific binding to lung tissue increases with lipophilicity.

  19. Platelet-derived Growth-factor-releasing Aligned Collagen-nanoparticle Fibers Promote the Proliferation and Tenogenic Differentiation of Adipose-derived Stem Cells

    Science.gov (United States)

    2013-11-27

    among the most common ortho- pedic injuries to soldiers due to repeated exercise , heavy-duty work and battlefield injuries [1,2]. Tendon /ligament...longer, sustained effect on cells. This is highly desirable because tendons /ligaments tend to need a longer time to healing due to the lack of...C). A rat Achilles tendon can sustain a maximum force of 32 N. Depending on the outcome of mechanical evaluation, the aligned collagen fiber scaffold

  20. A Novel Roscovitine Derivative Potently Induces G(1)-Phase Arrest in Platelet-Derived Growth Factor-BB-Activated Vascular Smooth Muscle Cells

    Czech Academy of Sciences Publication Activity Database

    Sroka, I. M.; Heiss, E.H.; Havlíček, Libor; Totzke, F.; Aristei, Y.; Pechan, P.; Kubbutat, M.H.G.; Strnad, Miroslav; Dirsch, V.M.

    2010-01-01

    Roč. 77, č. 2 (2010), s. 255-261 ISSN 0026-895X R&D Projects: GA ČR GA301/08/1649 Grant - others:_(XE) LSHB-CT-2004-503467 Institutional research plan: CEZ:AV0Z50380511 Keywords : CYCLIN-DEPENDENT KINASES * DRUG-ELUTING STENTS * CYC202 R-ROSCOVITINE Subject RIV: FD - Oncology ; Hematology Impact factor: 4.725, year: 2010

  1. Platelet-derived growth factor (PDGF) B-chain gene expression by activated blood monocytes precedes the expression of the PDGF A-chain gene

    International Nuclear Information System (INIS)

    Martinet, Y.; Jaffe, H.A.; Yamauchi, K.; Betsholtz, C.; Westermark, B.; Heldin, C.H.; Crystal, R.G.

    1987-01-01

    When activated, normal human blood monocytes are known to express the c-sis proto-oncogene coding for PDGF B-chain. Since normal human platelet PDGF molecules are dimers of A and B chains and platelets and monocytes are derived from the same marrow precursors, activated blood monocytes were simultaneously evaluated for their expression of PDGF A and B chain genes. Human blood monocytes were purified by adherence, cultured with or without activation by lipopolysaccharide and poly(A)+ RNA evaluated using Northern analysis and 32 P-labeled A-chain and B-chain (human c-sis) probes. Unstimulated blood monocytes did not express either A-chain or B-chain genes. In contrast, activated monocytes expressed a 4.2 kb mRNA B-chain transcript at 4 hr, but the B-chain mRNA levels declined significantly over the next 18 hr. In comparison, activated monocytes expressed very little A-chain mRNA at 4 hr, but at 12 hr 1.9, 2.3, and 2.8 kb transcripts were observed and persisted through 24 hr. Thus, activation of blood monocytes is followed by PDGF B-chain gene expression preceding PDGF A-chain gene expression, suggesting a difference in the regulation of the expression of the genes for these two chains by these cells

  2. Platelet-derived growth factor (PDGF) in oncogenesis: development of a vascular connective tissue stroma in xenotransplanted human melanoma producing PDGF-BB.

    OpenAIRE

    Forsberg, K; Valyi-Nagy, I; Heldin, C H; Herlyn, M; Westermark, B

    1993-01-01

    Human WM9 melanoma cells, previously shown to be devoid of PDGF expression, were stably transfected with a PDGF-B cDNA under the transcriptional control of a cytomegalovirus promoter. Northern blot analysis revealed high expression of an mRNA of the expected size in the PDGF-B-transfected cells. Synthesis and secretion of PDGF-BB was confirmed by immunoprecipitation. Furthermore, conditioned medium from PDGF-B-transfected cells contained a mitogenic activity for fibroblasts. For analysis of t...

  3. Overactivation of phospholipase C-gamma1 renders platelet-derived growth factor beta-receptor-expressing cells independent of the phosphatidylinositol 3-kinase pathway for chemotaxis

    DEFF Research Database (Denmark)

    Rönnstrand, L; Siegbahn, A; Rorsman, C

    1999-01-01

    ., Siegbahn, A. , Rorsman, C., Engström, U., Wernstedt, C., Heldin, C.-H., and Rönnstrand, L. (1996) EMBO J. 15, 5299-5313). Here we show that the increased chemotaxis correlates with increased activation of phospholipase C-gamma1 (PLC-gamma1), measured as inositol-1,4, 5-trisphosphate release. By two......-dimensional phosphopeptide mapping, the increase in phosphorylation of PLC-gamma1 was shown not to be selective for any site, rather a general increase in phosphorylation of PLC-gamma1 was seen. Specific inhibitors of protein kinase C, bisindolylmaleimide (GF109203X), and phosphatidylinositol 3-kinase (PI3-kinase), LY294002......, did not affect the activation of PLC-gamma1. To assess whether increased activation of PLC-gamma1 is the cause of the hyperchemotactic behavior of the Y934F mutant cell line, we constructed cell lines expressing either wild-type or a catalytically compromised version of PLC-gamma1 under a tetracycline...

  4. Platelet-derived growth factor B retention is essential for development of normal structure and function of conduit vessels and capillaries

    DEFF Research Database (Denmark)

    Nyström, Henrik C.; Lindblom, Per; Wickman, Anna

    2006-01-01

    . This outward eutrophic remodelling of the aorta was accompanied by endothelial dysfunction. RetKO showed decreased capillary density in skeletal muscle and signs of a defective delivery of capillary oxygen to skeletal muscle, as shown by a decreased physical performance. In RetKO mice, echocardiography...

  5. Biopharmaceutical characterisation of insulin and recombinant human growth hormone loaded lipid submicron particles produced by supercritical gas micro-atomisation.

    Science.gov (United States)

    Salmaso, Stefano; Bersani, Sara; Elvassore, Nicola; Bertucco, Alberto; Caliceti, Paolo

    2009-09-08

    Homogeneous dispersions of insulin and recombinant human growth hormone (rh-GH) in tristearin/phosphatidylcholine/PEG mixtures (1.3:1.3:0.25:0.15 w/w ratio) were processed by supercritical carbon dioxide gas micro-atomisation to produce protein-loaded lipid particles. The process yielded spherical particles, with a 197+/-94 nm mean diameter, and the insulin and rh-GH recovery in the final product was 57+/-8% and 48+/-5%, respectively. In vitro, the proteins were slowly released for about 70-80 h according to a diffusive mechanism. In vivo, the insulin and glucose profiles in plasma obtained by subcutaneous administration of a dose of particles containing 2 microg insulin to diabetic mice overlapped that obtained with 2 microg of insulin in solution. Administration of a dose of particles containing 5 microg insulin resulted in faster and longer glycaemia reduction. Oral administration of 20 and 50 microg insulin equivalent particles produced a significant hypoglycaemic effect. The glucose levels decreased since 2h after administration, reaching about 50% and 70% glucose reduction in 1-2h with the lower and higher dose, respectively. As compared to subcutaneous administration, the relative pharmacological bioavailability obtained with 20 and 50 microg equivalent insulin particles was 7.7% and 6.7%, respectively. Daily subcutaneous administration of 40 microg of rh-GH-loaded particles to hypophysectomised rats induced similar body weight increase as 40 microg rh-GH in solution. The daily oral administration of 400 microg rh-GH equivalent particles elicited a slight body weight increase, which corresponded to a relative pharmacological bioavailability of 3.4% compared to subcutaneous administration.

  6. Effects of recombinant human epidermal growth factor (rhEGF) on experimental radiation-induced oral mucositis in rats

    International Nuclear Information System (INIS)

    Jung, Kwon Il; Kim, Sun Hee; Moon, Soo Young; Kim, Yeon Wha; Hong, Joon Pio; Lee, Sang Wook; Kim, Hyun Sook

    2006-01-01

    Oral mucositis is a common toxicity of radiation or chemotherapy, which is used a treatment for head and neck cancer. We investigated effects of recombinant human epidermal growth factor (rhEGF) on radiation-induced oral mucositis in rat model. Spraque-Dawley rats (7 per group) exposed to a single dose of 25 Gy (day 0) on their head, except for one group, were randomly divided into un-treated, vehicle-treated, and two rhEGF-treated groups. Rats were topically applied with rhEGF (15 or 30 μ g/oral cavity/day) or vehicle to their oral mucosa. Survival rate of rats, weight changes, and food intakes were examined from day 0 to 18 after radiation. Histology study was performed from oral mucosa of rats at day 7 and 18 after radiation. rhEGF-treated groups (15 or 30 μ g/day) showed all survival rate 33%, whereas un-treated and vehicle-treated groups showed all survival rate 0% at the end of experiment. rhEGF-treated groups statistically had less weight loss compared to vehicle-treated group from day 2 to 7 after radiation. Food intake of rats with rhEGF treatment turned to increase at day 14 after radiation. At 7 day after radiation, un-treated and vehicle-treated groups showed severe pseudomembraneous of ulcerative oral mucositis. On the other hand, rhEGF-treated groups had no more than cellular swelling and degeneration of epidermal cells in oral mucosa of rats. These results suggest that rhEGF has significantly positive effects on radiation-induced oral mucositis in rats. rhEGF display a therapeutic potential on a clinical level

  7. A better anti-diabetic recombinant human fibroblast growth factor 21 (rhFGF21 modified with polyethylene glycol.

    Directory of Open Access Journals (Sweden)

    Zhifeng Huang

    Full Text Available As one of fibroblast growth factor (FGF family members, FGF21 has been extensively investigated for its potential as a drug candidate to combat metabolic diseases. In the present study, recombinant human FGF21 (rhFGF21 was modified with polyethylene glycol (PEGylation in order to increase its in vivo biostabilities and therapeutic potency. At N-terminal residue rhFGF21 was site-selectively PEGylated with mPEG20 kDa-butyraldehyde. The PEGylated rhFGF21 was purified to near homogeneity by Q Sepharose anion-exchange chromatography. The general structural and biochemical features as well as anti-diabetic effects of PEGylated rhFGF21 in a type 2 diabetic rat model were evaluated. By N-terminal sequencing and MALDI-TOF mass spectrometry, we confirmed that PEG molecule was conjugated only to the N-terminus of rhFGF21. The mono-PEGylated rhFGF21 retained the secondary structure, consistent with the native rhFGF21, but its biostabilities, including the resistance to physiological temperature and trypsinization, were significantly enhanced. The in vivo immunogenicity of PEGylated rhFGF21 was significantly decreased, and in vivo half-life time was significantly elongated. Compared to the native form, the PEGylated rhFGF21 had a similar capacity of stimulating glucose uptake in 3T3-L1 cells in vitro, but afforded a significantly long effect on reducing blood glucose and triglyceride levels in the type 2 diabetic animals. These results suggest that the PEGylated rhFGF21 is a better and more effective anti-diabetic drug candidate than the native rhFGF21 currently available. Therefore, the PEGylated rhFGF21 may be potentially applied in clinics to improve the metabolic syndrome for type 2 diabetic patients.

  8. A Single-Center Pilot Prospective Study of Topical Application of Platelet-Derived Eye Drops for Patients with Ocular Chronic Graft-versus-Host Disease.

    Science.gov (United States)

    Zallio, Francesco; Mazzucco, Laura; Monaco, Federico; Astori, Maria Rosa; Passera, Roberto; Drago, Giovanna; Tamiazzo, Stefania; Rapetti, Manuela; Dolcino, Daniela; Guaschino, Roberto; Pini, Massimo; Ladetto, Marco

    2016-09-01

    Ocular involvement of chronic graft-versus-host disease (cGVHD) is a complication that occurs in up to 60% of patients after allogeneic hematopoietic stem cell transplantation. Conventional therapeutic options include medical and surgical procedures that are administered depending on the severity of the condition, but most of them have provided unsatisfactory results and, to date, there is no consensus about treatment. We considered that topical application of a platelet lysate, administered as eye drops, might be considered an alternative worthwhile of investigation to treat ocular surface disorders in patients suffering from cGVHD. Therefore, we conducted a single-center prospective pilot study to assess the efficacy and safety of using eye drops made from reconstituted lysed platelet concentrate. Twenty-six patients with ocular cGVHD were eligible for the study; all but 2 completed their scheduled 1-year treatment and complied with the hematologic and ophthalmic regimen. At their first assessment interviews, after 30 days of treatment, 91% of patients reported an improvement in their symptoms and for 32%, substantive objective differences were measured. Remission of corneal damage was seen for 86% of our cohort, and improved National Institutes of Health scores for 73%, of whom 8% achieved the best score of 0 (ie, non-dry eye). Similar results were seen at later time points. Comparing outcomes for our patient cohort to those determined retrospectively for patients in our institutional database revealed a 5-year overall survival (OS) of 65%. This OS is comparable to patients with limited cGVHD (75%) and is superior to that of patients with nonocular extensive cGVHD or without cGVHD (30% and 59%, respectively) (P = .013). Our results suggest that platelet-derived eye drops are a safe, practical, and well-tolerated therapeutic option that offers substantial benefits for most patients affected by ocular cGVHD at onset. The favorable OS of our patient cohort

  9. Differences in levels of platelet-derived microparticles in platelet components prepared using the platelet rich plasma, buffy coat, and apheresis procedures.

    Science.gov (United States)

    Noulsri, Egarit; Udomwinijsilp, Prapaporn; Lerdwana, Surada; Chongkolwatana, Viroje; Permpikul, Parichart

    2017-04-01

    There has been an increased interest in platelet-derived microparticles (PMPs) in transfusion medicine. Little is known about PMP status during the preparation of platelet concentrates for transfusion. The aim of this study is to compare the PMP levels in platelet components prepared using the buffy coat (BC), platelet-rich plasma platelet concentrate (PRP-PC), and apheresis (AP) processes. Platelet components were prepared using the PRP-PC and BC processes. Apheresis platelets were prepared using the Trima Accel and Amicus instruments. The samples were incubated with annexin A5-FITC, CD41-PE, and CD62P-APC. At day 1 after processing, the PMPs and activated platelets were determined using flow cytometry. Both the percentage and number of PMPs were higher in platelet components prepared using the Amicus instrument (2.6±1.8, 32802±19036 particles/μL) than in platelet components prepared using the Trima Accel instrument (0.5±0.4, 7568±5298 particles/μL), BC (1.2±0.6, 12,920±6426 particles/μL), and PRP-PC (0.9±0.6, 10731±5514 particles/μL). Both the percentage and number of activated platelets were higher in platelet components prepared using the Amicus instrument (33.2±13.9, 427553±196965 cells/μL) than in platelet components prepared using the Trima Accel instrument (16.2±6.1, 211209±87706 cells/μL), BC (12.9±3.2, 140624±41003 cells/μL), and PRP-PC (21.1±6.3, 265210±86257 cells/μL). The study suggests high variability of PMPs and activated platelets in platelet components prepared using different processes. This result may be important in validating the instruments involved in platelet blood collection and processing. Copyright © 2016 Elsevier Ltd. All rights reserved.

  10. Time-dependent inhibitory effects of cGMP-analogues on thrombin-induced platelet-derived microparticles formation, platelet aggregation, and P-selectin expression

    International Nuclear Information System (INIS)

    Nygaard, Gyrid; Herfindal, Lars; Kopperud, Reidun; Aragay, Anna M.; Holmsen, Holm; Døskeland, Stein Ove; Kleppe, Rune; Selheim, Frode

    2014-01-01

    Highlights: • We investigated the impact of cyclic nucleotide analogues on platelet activation. • Different time dependence were found for inhibition of platelet activation. • Additive effect was found using PKA- and PKG-activating analogues. • Our results may explain some of the discrepancies reported for cNMP signalling. - Abstract: In platelets, nitric oxide (NO) activates cGMP/PKG signalling, whereas prostaglandins and adenosine signal through cAMP/PKA. Cyclic nucleotide signalling has been considered to play an inhibitory role in platelets. However, an early stimulatory effect of NO and cGMP-PKG signalling in low dose agonist-induced platelet activation have recently been suggested. Here, we investigated whether different experimental conditions could explain some of the discrepancy reported for platelet cGMP-PKG-signalling. We treated gel-filtered human platelets with cGMP and cAMP analogues, and used flow cytometric assays to detect low dose thrombin-induced formation of small platelet aggregates, single platelet disappearance (SPD), platelet-derived microparticles (PMP) and thrombin receptor agonist peptide (TRAP)-induced P-selectin expression. All four agonist-induced platelet activation phases were blocked when platelets were costimulated with the PKG activators 8-Br-PET-cGMP or 8-pCPT-cGMP and low-doses of thrombin or TRAP. However, extended incubation with 8-Br-PET-cGMP decreased its inhibition of TRAP-induced P-selectin expression in a time-dependent manner. This effect did not involve desensitisation of PKG or PKA activity, measured as site-specific VASP phosphorylation. Moreover, PKG activators in combination with the PKA activator Sp-5,6-DCL-cBIMPS revealed additive inhibitory effect on TRAP-induced P-selectin expression. Taken together, we found no evidence for a stimulatory role of cGMP/PKG in platelets activation and conclude rather that cGMP/PKG signalling has an important inhibitory function in human platelet activation

  11. Recombinant Programming

    OpenAIRE

    Pawlak , Renaud; Cuesta , Carlos; Younessi , Houman

    2004-01-01

    This research report presents a promising new approach to computation called Recombinant Programming. The novelty of our approach is that it separates the program into two layers of computation: the recombination and the interpretation layer. The recombination layer takes sequences as inputs and allows the programmer to recombine these sequences through the definition of cohesive code units called extensions. The output of such recombination is a mesh that can be used by the interpretation la...

  12. Construction and growth properties of bovine herpesvirus type 5 recombinants defective in the glycoprotein E or thymidine kinase gene or both

    Directory of Open Access Journals (Sweden)

    M.C.S. Brum

    2010-02-01

    Full Text Available Bovine herpesvirus type 5 (BoHV-5 is an important pathogen of cattle in South America. We describe here the construction and characterization of deletion mutants defective in the glycoprotein E (gE or thymidine kinase (TK gene or both (gE/TK from a highly neurovirulent and well-characterized Brazilian BoHV-5 strain (SV507/99. A gE-deleted recombinant virus (BoHV-5 gE∆ was first generated in which the entire gE open reading frame was replaced with a chimeric green fluorescent protein gene. A TK-deleted recombinant virus (BoHV-5 TK∆ was then generated in which most of the TK open reading frame sequences were deleted and replaced with a chimeric β-galactosidase gene. Subsequently, using the BoHV-5 gE∆ virus as backbone, a double gene-deleted (TK plus gE BoHV-5 recombinant (BoHV-5 gE/TK∆ was generated. The deletion of the gE and TK genes was confirmed by immunoblotting and PCR, respectively. In Madin Darby bovine kidney (MDBK cells, the mutants lacking gE (BoHV-5 gE∆ and TK + gE (BoHV-5 gE/TK∆ produced small plaques while the TK-deleted BoHV-5 produced wild-type-sized plaques. The growth kinetics and virus yields in MDBK cells for all three recombinants (BoHV-5 gE∆, BoHV-5 TK∆ and BoHV-5 gE/TK∆ were similar to those of the parental virus. It is our belief that the dual gene-deleted recombinant (BoHV-5 gE/TK∆ produced on the background of a highly neurovirulent Brazilian BoHV-5 strain may have potential application in a vaccine against BoHV-5.

  13. Impact of alg3 gene deletion on growth, development, pigment production, protein secretion, and functions of recombinant Trichoderma reesei cellobiohydrolases in Aspergillus niger

    Energy Technology Data Exchange (ETDEWEB)

    Dai, Ziyu; Aryal, Uma K.; Shukla, Anil; Qian, Wei-Jun; Smith, Richard D.; Magnuson, Jon K.; Adney, William S.; Beckham, Gregg T.; Brunecky, Roman; Himmel, Michael E.; Decker, Stephen R.; Ju, Xiaohui; Zhang, Xiao; Baker, Scott E.

    2013-12-01

    ALG3 is a Family 58 glycosyltransferase enzyme involved in early N-linked glycan synthesis. Here, we investigated the effect of the alg3 gene disruption on growth, development, metabolism, and protein secretion in Aspergillus niger. The alg3 gene deletion resulted in a significant reduction of growth on complete (CM) and potato dextrose agar (PDA) media and a substantial reduction of spore production on CM. It also delayed spore germination in the liquid cultures of both CM and PDA media, but led to a significant accumulation of red pigment on both CM and liquid modified minimal medium (MM) supplemented with yeast extract. The relative abundance of 55 proteins of the total 190 proteins identified in the secretome was significantly different as a result of alg3 gene deletion. Comparison of a Trichoderma reesei cellobiohydrolase (Cel7A) heterologously expressed in A. niger parental and Δalg3 strains showed that the recombinant Cel7A expressed in the mutant background was smaller in size than that from the parental strains. This study suggests that ALG3 is critical for growth and development, pigment production, and protein secretion in A. niger. Functional analysis of recombinant Cel7A with aberrant glycosylation demonstrates the feasibility of this alternative approach to evaluate the role of N-linked glycosylation in glycoprotein secretion and function.

  14. Production and characterization of recombinantly derived peptides and antibodies for accurate determinations of somatolactin, growth hormone and insulin-like growth factor-I in European sea bass (Dicentrarchus labrax).

    Science.gov (United States)

    de Celis, S Vega-Rubín; Gómez-Requeni, P; Pérez-Sánchez, J

    2004-12-01

    A specific radioimmunoassay (RIA) for European sea bass (Dicentrarchus labrax) growth hormone (GH) was developed and validated. For this purpose, a stable source of GH was produced by means of recombinant DNA technology in a bacteria system. The identity of the purified protein (ion exchange chromatography) was demonstrated by Western blot and a specific GH antiserum was raised in rabbit. In Western blot and RIA system, this antiserum recognized specifically native and recombinant GH, and it did not cross-react with fish prolactin (PRL) and somatolactin (SL). In a similar way, a specific polyclonal antiserum against the now available recombinant European sea bass SL was raised and used in the RIA system to a sensitivity of 0.3 ng/ml (90% of binding of tracer). Further, European sea bass insulin-like growth factor-I (IGF-I) was cloned and sequenced, and its high degree of identity with IGF-I peptides of barramundi, tuna, and sparid fish allowed the use of a commercial IGF-I RIA based on barramundi IGF-I antiserum. These assay tools assisted for the first time accurate determinations of SL and GH-IGF-I axis activity in a fish species of the Moronidae family. Data values were compared to those found with gilthead sea bream (Sparus aurata), which is currently used as a Mediterranean fish model for growth endocrinology studies. As a characteristic feature, the average concentration year round of circulating GH in growing mature males of European sea bass was higher than in gilthead sea bream. By contrast, the average concentration of circulating SL was lower. Concerning to circulating concentration of IGF-I, the measured plasma values for a given growth rate were also lower in European sea bass. These findings are discussed on the basis of a different energy status that might allowed a reduced but more continuous growth in European sea bass.

  15. Recombinant Newcastle disease virus (NDV/Anh-IL-2 expressing human IL-2 as a potential candidate for suppresses growth of hepatoma therapy

    Directory of Open Access Journals (Sweden)

    Yunzhou Wu

    2016-09-01

    Full Text Available Newcastle disease virus (NDV have shown oncolytic therapeutic efficacy in preclinical study and are currently approved for clinical trials. NDV Anhinga strain which is a mesogenic strain should be classified as lytic strain and has a therapeutic efficacy in hepatocellular cancer. In this study, we evaluated the capacity of NDV Anhinga strain to elicit immune reaction in vivo and the possibility for using as a vaccine vector for expressing tumor therapeutic factors. Interleukin-2 (IL-2 could boost the immune response against the tumor cells. Therefore, we use NDV Anhinga strain as backbone to construct a recombinant virus (NDV/Anh-IL-2 expressing IL-2. The virus growth curve showed that the production of recombinant NDV/Anh-IL-2 was slightly delayed compared to the wild type. The NDV/Anh-IL-2 strain could express soluble IL-2 and effectively inhibit the growth of hepatocellular carcinoma in vivo. 60 days post-treatment, mice which were completely cured by previous treatment were well protected when rechallenged with the same tumor cell. From the H&E-stained sections, intense infiltration of lymphocyte was observed in the NDV Anhinga strain treated group, especially in NDV/Anh-IL-2 group. The NDV Anhinga strain could not only kill the tumor directly, but could also elicit immune reaction and a potent immunological memory when killing tumor in vivo. In conclusion, the Anhinga strain could be an effective vector for tumor therapy; the recombinant NDV/Anh-IL-2 strain expressing soluble IL-2 is a promising candidate for hepatoma therapy.

  16. Validation and ease of use of a new pen device for self-administration of recombinant human growth hormone: results from a two-center usability study

    Directory of Open Access Journals (Sweden)

    Rapaport R

    2013-09-01

    Full Text Available Robert Rapaport,1 Paul Saenger,2 Heinrich Schmidt,3 Yukihiro Hasegawa,4 Michel Colle,5 Sandro Loche,6 Sandra Marcantonio,7 Walter Bonfig,8 Markus Zabransky,9 Fima Lifshitz10 1Division of Pediatric Endocrinology and Diabetes, Mount Sinai School of Medicine, 2Winthrop University Hospital, Mineola, NY, USA; 3University Children's Hospital, Division of Endocrinology and Diabetology, Munich, Germany; 4Department of Endocrinology and Metabolism, Tokyo Metropolitan Children's Medical Center, Tokyo, Japan; 525 rue Boudet, Bordeaux, France; 6Servizio di Endocrinologia Pediatrica, Ospedale Microcitemico ASL Cagliari, Cagliari, Italy; 7Clinica de Endocrinologia Pediátrica, Londrina, Brazil; 8Division of Pediatric Endocrinology, Technical University München, Munich, Germany; 9Sandoz International GmbH, Holzkirchen, Germany; 10Pediatric Sunshine Academics, Inc, Santa Barbara, CA, USA Abstract: Close adherence to the recommended treatment regimen is important for the success of recombinant human growth hormone therapy, although nonadherence can be common. Ease of use and safety during use/storage are among several important factors in the design of a growth hormone injection device intended for long-term use. This study was performed to validate the usability and assess the ease of use of a new pen device (SurePal™ that has been developed to support daily administration of the recombinant human growth hormone product, Omnitrope® (somatropin. The primary objectives of the study were to assess if study participants, representing intended users of the pen in clinical practice, were able to perform an injection procedure into an injection pad effectively and safely and disassemble the pen without receiving a needlestick injury. A total of 106 participants (61 adults and 45 children/adolescents were enrolled at two study centers (one in the US, one in Germany. Results for both primary usability tasks met the predefined acceptance criteria, with >85% of

  17. Angiopoietin-2 impairs collateral artery growth associated with the suppression of the infiltration of macrophages in mouse hindlimb ischaemia

    Directory of Open Access Journals (Sweden)

    Xiaoyong Tan

    2016-10-01

    Full Text Available Abstract Background Angiopoietin-2 (Ang-2, a ligand of the Tie-2 receptor, plays an important role in maintaining endothelial cells and in destabilizing blood vessels. Collateral artery growth (arteriogenesis is a key adaptive response to arterial occlusion. It is unknown whether the destabilization of blood vessels by Ang-2 can affect arteriogenesis and modulate mononuclear cell function. This study aimed to investigate the effects of Ang-2 on collateral artery growth. Methods Hindlimb ischaemia model was produced in C57BL/6 mice by femoral artery ligation. Blood flow perfusion was measured using a laser Doppler perfusion imager quantitative RT-PCR analysis was applied to identify the level of angiogenic factors. Results After the induction of hindlimb ischaemia, blood flow recovery was impaired in mice treated with recombinant Ang-2 protein; this was accompanied by a reduction of peri-collateral macrophage infiltration. In addition, quantitative RT-PCR analysis revealed that Ang-2 treatment decreased monocyte chemotactic protein-1 (MCP-1, platelet-derived growth factor-BB (PDGF-BB mRNA levels in ischaemic adductor muscles. Ang-2 can lead to macrophage M1/M2 polarization shift inhibition in the ischaemic muscles. Furthermore, Ang-2 reduced the in vitro inflammatory response in macrophages and vascular cells involved in arteriogenesis. Conclusions Our results demonstrate that Ang-2 is essential for efficient arteriogenesis, which controls macrophage infiltration.

  18. Long-term auxological and pubertal outcome of patients with hereditary insulin-like growth factor-I deficiency (Laron and growth hormone-gene deletion syndrome) treated with recombinant human insulin-like growth factor-I.

    Science.gov (United States)

    Messina, M F; Arrigo, T; Valenzise, M; Ghizzoni, L; Caruso-Nicoletti, M; Zucchini, S; Chiabotto, P; Crisafulli, G; Zirilli, G; De Luca, F

    2011-04-01

    GH-IGF-I axis is mainly involved in the complex process of somatic growth but emerging evidence suggests that it also influences hypothalamic-pituitary-gonadal (HPG) function. We report some data regarding long-term auxological and pubertal outcome of five female patients with hereditary forms of GH-IGF-I deficiency (Laron and GH-gene deletion syndrome) and a mean age of 23.4±5.3 yr (range 19-32). All the patients received recombinant human IGF-I (rhIGF-I, Pharmacia and Upjohn, Stockholm, Sweden, and rhIGF-I, Genentech, San Francisco, CA, USA) from a mean age of 8.6 yr (range 3.2-14.2) up to the final height. Final height was very disappointing (≤ -5.0 SD scores) and lower than target height in all the patients. Pubertal onset was delayed in most of them but menarche occurred spontaneously in all the patients. Median age at menarche was 15.1 yr. Menstrual cycles were regular for several years. Median duration of gynecological follow- up was 8.3 yr with the longest span of 17.2 yr. We can assert that GH-IGF-I axis has an essential role in promoting linear growth in humans and its physiological action cannot be replaced by pharmacological treatment in most patients with hereditary forms of IGF-I insufficiency as demonstrated by their subnormal final height. Our clinical observations can also support an essential role of IGF-I in genitalia growth but not in the function of HPG axis as demonstrated by the maintenance of regular menstrual cycles in the presence of subnormal levels of IGF-I after treatment discontinuation.

  19. Effects of recombinant human growth hormone in the treatment of dwarfism and relationship between IGF-1, IGFBP-3 and thyroid hormone.

    Science.gov (United States)

    Ren, Shanxiang; Nie, Yuxiang; Wang, Aihong

    2016-12-01

    The effects of recombinant human growth hormone (rhGH) in the treatment of dwarfism and the relationship between insulin-like growth factor (IGF)-1, IGF-binding protein (IGFBP)-3 and thyroid hormone were examined in the present study. For this purpose, 66 patients diagnosed with dwarfism were selected retrospectively, with 36 cases of growth hormone deficiency (GHD) and 30 cases of idiopathic short stature (ISS). The therapeutic dose of GHD 0.10 IU/kg·day and ISS 0.15 IU/kg·day were injected subcutaneously every night before sleep until adulthood. The average follow-up was 5 years, and the results were evaluated and measured every 3 months, including height, BA, secondary test of growth hormone (GH peak), IGF-1, IGFBP-3 and thyroid hormone (FT3, FT4 and TSH). After treatment, the height, BA, GH peak, IGF-A and IGFBP-3 of the GHD group were all increased, and the differences were statistically significant (P0.05). The results of the Pearson-related analysis suggested that GH peak of the GHD group, IGF-1 and IGFBP-3 were positively associated with height (P0.05). rhGH was effective for GHD and ISS, with the GHD effect being positively associated with the GH peak, IGF-1 and IGFBP-3. ISS had no obvious relationship with GH peak, IGF-1 and IGFBP-3 although other influencing factors may be involved.

  20. Controlled chondrogenesis from adipose-derived stem cells by recombinant transforming growth factor-β3 fusion protein in peptide scaffolds.

    Science.gov (United States)

    Zheng, Dong; Dan, Yang; Yang, Shu-hua; Liu, Guo-hui; Shao, Zeng-wu; Yang, Cao; Xiao, Bao-jun; Liu, Xiangmei; Wu, Shuilin; Zhang, Tainjin; Chu, Paul K

    2015-01-01

    Adipose-derived stem cells (ADSCs) are promising for cartilage repair due to their easy accessibility and chondrogenic potential. Although chondrogenesis of transforming growth factor-β (TGF-β) mediated mesenchymal stem cells (MSCs) is well established in vitro, clinical tissue engineering requires effective and controlled delivery of TGF-β in vivo. In this work, a self-assembled peptide scaffold was employed to construct cartilages in vivo through the chondrogenesis from ADSCs controlled by recombinant fusion protein LAP-MMP-mTGF-β3 that was transfected by lentiviral vectors. During this course, the addition of matrix metalloproteinases (MMPs) can trigger the release of mTGF-β3 from the recombinant fusion protein of LAP-MMP-mTGF-β3 in the combined scaffolds, thus stimulating the differentiation of ADSCs into chondrogenesis. The specific expression of cartilage genes was analyzed by real-time polymerase chain reaction and Western blot. The expression of chondrocytic markers was obviously upregulated to a higher level compared to the one by commonly used TGF-β3 alone. After 3 weeks of in vitro culturing, the hybrids with differentiated chondrogenesis were then injected subcutaneously into nude mice and retrieved after 4 weeks of culturing in vivo. Histological analysis also confirmed that the recombinant fusion protein was more effective for the formation of cartilage matrix than the cases either with TGF-β3 alone or without LAP-MMP-mTGF-β3 (P<0.05). This study demonstrates that controlled local delivery of the LAP-MMP-mTGF-β3 constructs can accelerate differentiation of ADSCs into the cartilage in vivo, which indicates the great potential of this hybrid in rapid therapy of osteoarthritis. Copyright © 2014 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

  1. Genetic Recombination

    Science.gov (United States)

    Whitehouse, H. L. K.

    1973-01-01

    Discusses the mechanisms of genetic recombination with particular emphasis on the study of the fungus Sordaria brevicollis. The study of recombination is facilitated by the use of mutants of this fungus in which the color of the ascospores is affected. (JR)

  2. Recombinant epidermal growth factor-like domain-1 from coagulation factor VII functionalized iron oxide nanoparticles for targeted glioma magnetic resonance imaging.

    Science.gov (United States)

    Liu, Heng; Chen, Xiao; Xue, Wei; Chu, Chengchao; Liu, Yu; Tong, Haipeng; Du, Xuesong; Xie, Tian; Liu, Gang; Zhang, Weiguo

    The highly infiltrative and invasive nature of glioma cells often leads to blurred tumor margins, resulting in incomplete tumor resection and tumor recurrence. Accurate detection and precise delineation of glioma help in preoperative delineation, surgical planning and survival prediction. In this study, recombinant epidermal growth factor-like domain-1, derived from human coagulation factor VII, was conjugated to iron oxide nanoparticles (IONPs) for targeted glioma magnetic resonance (MR) imaging. The synthesized EGF1-EGFP-IONPs exhibited excellent targeting ability toward tissue factor (TF)-positive U87MG cells and human umbilical vein endothelial cells in vitro, and demonstrated persistent and efficient MR contrast enhancement up to 12 h for preclinical glioma models with high targeting specificity in vivo. They hold great potential for clinical translation and developing targeted theranostics against brain glioma.

  3. Effect of Recombinant Human Growth Hormone and Rosiglitazone for HIV-Associated Abdominal Fat Accumulation on Adiponectin and other Markers of Inflammation

    Science.gov (United States)

    Leung, Vivien; Chiu, Ya-Lin; Kotler, Donald P.; Albu, Jeanine; Zhu, Yuan-Shan; Ham, Kirsis; Engelson, Ellen S.; Hammad, Hoda; Christos, Paul; Donovan, Daniel S.; Ginsberg, Henry N.; Glesby, Marshall J.

    2016-01-01

    Background/Objective In a previous report of HIV-infected patients with fat redistribution, we found that recombinant human growth hormone (rhGH) therapy reduced visceral adipose tissue (VAT) but increased insulin resistance, and that the addition of rosiglitazone reversed the negative effects of rhGH on insulin sensitivity. In this study, we sought to determine the effects of recombinant human growth hormone (rhGH) and rosiglitazone therapy on an array of inflammatory and fibrinolytic markers. Methods 72 patients with HIV-associated abdominal obesity and insulin resistance were randomized to treatment with rhGH, rosiglitazone, the combination of rhGH and rosiglitazone, or placebo for 12 weeks. Subjects with plasma and serum samples available at weeks 0 (n = 63) and 12 (n = 46-48) were assessed for adiponectin, C-reactive protein (CRP), homocysteine, interleukin-1 (IL-1), interleukin-6 (IL-6), tumor necrosis factor alpha (TNF-α), interferon gamma (IFN-γ), fibrinogen, plasminogen activator inhibitor-1 (PAI-1) antigen, and tissue plasminogen activator (tPA) antigen. Results Treatment with both rosiglitazone alone and the combination of rosiglitazone and rhGH for 12 weeks resulted in significant increases in adiponectin levels from baseline. Adiponectin levels did not change significantly in the rhGH alone arm. There were no significant changes in the other biomarkers amongst the different treatment groups. Discussion In this study of HIV-infected patients with altered fat distribution, treatment with rosiglitazone had beneficial effects on adiponectin concentrations, an effect that was also seen with combination rosiglitazone and rhGH. RhGH administration alone, however, did not demonstrate any significant impact on adiponectin levels despite reductions in VAT. PMID:27077672

  4. Recombinant tumor necrosis factor alpha inhibits growth of methylcholanthrene-induced sarcoma and enhances natural killer activity of tumor-infiltrating lymphocytes in aging rats

    International Nuclear Information System (INIS)

    Ziolkowska, Maria; Nowak Joanna, J.; Janiak, Marek; Ryzewska, Alicja

    1994-01-01

    The effect of recombinant human tumor necrosis factors alpha (rHuTNF-α) on the growth of immunogenic, methylcholanthrene-induced sarcoma (MC-Sa) and natural killer (NK) cell activity of tumor-infiltrating lymphocytes (TIL) in adult and aging rats was investigated. In both groups of animals the growth of transplantable MC-Sa was markedly and similarly inhibited by multiple intratumoral (i.t.) injections of rHuTF-α. This effect was accompanied by stimulation of NK activity of tumor-infiltrating lymphocytes in adult as well as in aging rats. Studies ''in vitro'' demonstrated additionally that rHuTNF-α was a potent stimulator of NK but not of ADCC (antibody-dependent cellular cytotoxicity) activity of spleen lymphocytes from healthy animals. Our results indicate that the antitumor effect of TNF-α is comparable in adult and in aging rats bearing immunogenic MC-Sa. The inhibition of MC-Sa growth may be attributed not only to the TNF-α-induced necrosis of the neoplastic tissue but also to the ''in vivo'' stimulatory effect of this cytokine upon the NK-type function of lymphocytes infiltrating the tumor mass. (author). 31 refs, 5 figs, 2 tabs

  5. Effects of recombinant human epidermal growth factor on the proliferation and radiation survival of human fibroblast cell lines in vitro

    International Nuclear Information System (INIS)

    Kim, Hyun Sook; Kang, Ki Mun; Na, Jae Boem; Chai, Gyu Young; Lee, Sang Wook

    2006-01-01

    To explore the effect of recombinant human EGF on the proliferation and survival of human fibroblast cell lines following irradiation. Fibroblast was originated human skin and primary cultured. The trypan blue stain assay and MTT assay were used to study the proliferative effects of EGF on human fibroblast cell lines in vitro. An incubation of fibroblasts with rhEGF for 24 hours immediately after irradiation was counted everyday. Cell cycle distributions were analyzed by FACS analysis. Number of fibroblast was significant more increased rhEGF (1.0 nM, 10 nM, 100 nM, 1,000 nM) treated cell than control after 8 Gy irradiation. Most effective dose of rhEGF was at 160 nM. These survival differences were maintained at 1 week later. Proportion of S phase was significantly increased on rhEGF treated cells. rhEGF cause increased fibroblast proliferation following irradiation. We expect that rhEGF was effective for radiation induced wound healing

  6. Minimally invasive esthetic ridge preservation with growth-factor enhanced bone matrix.

    Science.gov (United States)

    Nevins, Marc L; Said, Sherif

    2017-12-28

    Extraction socket preservation procedures are critical to successful esthetic implant therapy. Conventional surgical approaches are technique sensitive and often result in alteration of the soft tissue architecture, which then requires additional corrective surgical procedures. This case series report presents the ability of flapless surgical techniques combined with a growth factor-enhanced bone matrix to provide esthetic ridge preservation at the time of extraction for compromised sockets. When considering esthetic dental implant therapy, preservation, or further enhancement of the available tissue support at the time of tooth extraction may provide an improved esthetic outcome with reduced postoperative sequelae and decreased treatment duration. Advances in minimally invasive surgical techniques combined with recombinant growth factor technology offer an alternative for bone reconstruction while maintaining the gingival architecture for enhanced esthetic outcome. The combination of freeze-dried bone allograft (FDBA) and rhPDGF-BB (platelet-derived growth factor-BB) provides a growth-factor enhanced matrix to induce bone and soft tissue healing. The use of a growth-factor enhanced matrix is an option for minimally invasive ridge preservation procedures for sites with advanced bone loss. Further studies including randomized clinical trials are needed to better understand the extent and limits of these procedures. The use of minimally invasive techniques with growth factors for esthetic ridge preservation reduces patient morbidity associated with more invasive approaches and increases the predictability for enhanced patient outcomes. By reducing the need for autogenous bone grafts the use of this technology is favorable for patient acceptance and ease of treatment process for esthetic dental implant therapy. © 2017 Wiley Periodicals, Inc.

  7. Expression and Purification of Recombinant Human Basic Fibroblast Growth Factor Fusion Proteins and Their Uses in Human Stem Cell Culture.

    Science.gov (United States)

    Imsoonthornruksa, Sumeth; Pruksananonda, Kamthorn; Parnpai, Rangsun; Rungsiwiwut, Ruttachuk; Ketudat-Cairns, Mariena

    2015-01-01

    To reduce the cost of cytokines and growth factors in stem cell research, a simple method for the production of soluble and biological active human basic fibroblast growth factor (hbFGF) fusion protein in Escherichia coli was established. Under optimal conditions, approximately 60-80 mg of >95% pure hbFGF fusion proteins (Trx-6xHis-hbFGF and 6xHis-hbFGF) were obtained from 1 liter of culture broth. The purified hbFGF proteins, both with and without the fusion tags, were biologically active, which was confirmed by their ability to stimulate proliferation of NIH3T3 cells. The fusion proteins also have the ability to support several culture passages of undifferentiated human embryonic stem cells and induce pluripotent stem cells. This paper describes a low-cost and uncomplicated method for the production and purification of biologically active hbFGF fusion proteins. © 2015 S. Karger AG, Basel.

  8. Safety and PK/PD correlation of TV-1106, a recombinant fused human albumin-growth hormone, following repeat dose administration to monkeys.

    Science.gov (United States)

    Ashkenazi, Nurit; Rosenstock, Moti; Hallak, Hussein; Bassan, Merav; Rasamoelisolo, Michele; Leuschner, Jost; Shinar, Doron

    TV-1106 is a recombinant human albumin genetically fused to growth hormone which is intended to reduce the frequency of injections for GH therapy users. We report the safety, tolerability, pharmacokinetics and pharmacodynamics of repeated subcutaneous injections of TV-1106 in Cynomolgus monkeys. Cynomolgus monkeys received four weekly subcutaneous injections of 0, 5, 10 or 20mg/kg TV-1106 and were monitored for safety signals throughout the study. Serum levels of TV-1106 and insulin-like growth factor 1 (IGF-1) were assayed. Treated animals showed no adverse effects or histopathological changes. TV-1106 serum concentrations showed sustained exposure to the drug. Exposure increased in a dose-dependent manner with peak concentrations at approximately 24h post-dosing and elimination half-lives in the range of 12 to 24h. IGF-1 serum concentrations were elevated throughout the entire study duration, indicative of the pharmacological response. There was a clear correlation between change in IGF-1 levels and dose or exposure to TV-1106. The safety, pharmacokinetic and pharmacodynamic findings support the further development of TV-1106 as a once-weekly administered treatment for patients with GHD. Copyright © 2016 Elsevier Ltd. All rights reserved.

  9. Recombinant FIP-gat, a Fungal Immunomodulatory Protein from Ganoderma atrum, Induces Growth Inhibition and Cell Death in Breast Cancer Cells.

    Science.gov (United States)

    Xu, Hui; Kong, Ying-Yu; Chen, Xin; Guo, Meng-Yuan; Bai, Xiao-Hui; Lu, Yu-Jia; Li, Wei; Zhou, Xuan-Wei

    2016-04-06

    FIP-gat, an immunomodulatory protein isolated from Ganoderma atrum, is a new member of the FIP family. Little is known, however, about its expressional properties and antitumor activities. It was availably expressed in Escherichia coli with a total yield of 29.75 mg/L. The migration of recombinant FIP-gat (rFIP-gat) on SDS-PAGE corresponded to the predicted molecular mass, and the band was correctly detected by a specific antibody. To characterize the direct effects of rFIP-gat on MDA-MB-231 breast cancer cells, MDA-MB-231 cells were treated with different concentrations of rFIP-gat in vitro; the results showed that this protein could reduce cell viability dose-dependently with a median inhibitory concentration (IC50) of 9.96 μg/mL and agglutinate the MDA-MB-231 cells at a concentration as low as 5 μg/mL. Furthermore, FIP-gat at a concentration of 10 μg/mL can induce significant growth inhibition and cell death in MDA-MB-231 cells. Notably, FIP-gat treatment triggers significant cell cycle arrest at the G1/S transition and pronounced increase in apoptotic cell population. Molecular assays based on microarray and real-time PCR further revealed the potential mechanisms encompassing growth arrest, apoptosis, and autophagy underlying the phenotypic effects.

  10. A phase I/II exploratory clinical trial for intracordal injection of recombinant hepatocyte growth factor for vocal fold scar and sulcus.

    Science.gov (United States)

    Hirano, Shigeru; Kawamoto, Atsuhiko; Tateya, Ichiro; Mizuta, Masanobu; Kishimoto, Yo; Hiwatashi, Nao; Kawai, Yoshitaka; Tsuji, Takuya; Suzuki, Ryo; Kaneko, Mami; Naito, Yasushi; Kagimura, Tatsuo; Nakamura, Tatsuo; Kanemaru, Shin-Ichi

    2018-04-01

    Vocal fold scar and sulcus are intractable diseases with no effective established treatments. Hepatocyte growth factor (HGF) has preclinically proven to have potent antifibrotic and regenerative effects on vocal fold scar. The current Phase I/II clinical trial aims to examine the safety and effectiveness of intracordal injection of a recombinant human HGF drug for patients with vocal fold scar or sulcus. This is an open-label, dose-escalating, first-in-human clinical trial. Eighteen patients with bilateral vocal fold scar or sulcus were enrolled and divided into three groups: Step I received 1 μg of HGF per vocal fold; Step II received 3 μg of HGF; and Step III received 10 μg of HGF. Injections were administered once weekly for 4 weeks. The protocol treatment was performed starting with Step I and escalating to Step III. Patients were followed for 6 months post-treatment. Local and systemic safety aspects were examined as primary endpoints, and therapeutic effects were assessed as secondary endpoints using voice handicap index-10; maximum phonation time; vocal fold vibratory amplitude; grade, rough, breathy, asthenic, strained scale; and jitter. The results indicated no serious drug-related adverse events in either the systemic or local examinations. In whole-subject analysis, voice handicap index-10, vocal fold vibratory amplitude, and grade, rough, breathy, asthenic, strained scale were significantly improved at 6 months, whereas maximum phonation time and jitter varied. There were no significant differences in phonatory data between the step groups. In conclusion, intracordal injection of a recombinant human HGF drug was safe, feasible, and potentially effective for human patients with vocal fold scar or sulcus. Copyright © 2017 John Wiley & Sons, Ltd.

  11. Increased levels of specific leukocyte- and platelet-derived substances during normal anti-tetanus antibody synthesis in patients with inactive Crohn disease

    DEFF Research Database (Denmark)

    Nielsen, Hans Jørgen; Mortensen, T; Holten-Andersen, M

    2001-01-01

    , vascular endothelial growth factor (VEGF), tissue inhibitor of metalloproteinases-1 (TIMP-1), plasminogen activator inhibitor type-1 (PAI-1) and myeloperoxidase (MPO) were determined in serum or plasma obtained on the same days. RESULTS: After inoculation anti-tetanus antibody levels were equally raised...... immunization in patients with Crohn disease and the subsequent release of various inflammatory mediators and growth factors in blood. METHODS: Ten patients with inactive disease and no concurrent medication and 12 age-and gender-matched healthy volunteers with anti-tetanus antibody levels less than 0.1 IU...... range and IL-6, TNF-alpha, MPO and histamine levels were unchanged in patients and volunteers during the study period. The levels of VEGF, TIMP-1 and PAI-1 were unchanged in the healthy volunteers during the study period, but were significantly (P

  12. Evaluation of a multiple-cycle, recombinant virus, growth competition assay that uses flow cytometry to measure replication efficiency of human immunodeficiency virus type 1 in cell culture.

    Science.gov (United States)

    Dykes, Carrie; Wang, Jiong; Jin, Xia; Planelles, Vicente; An, Dong Sung; Tallo, Amanda; Huang, Yangxin; Wu, Hulin; Demeter, Lisa M

    2006-06-01

    Human immunodeficiency virus type 1 (HIV-1) replication efficiency or fitness, as measured in cell culture, has been postulated to correlate with clinical outcome of HIV infection, although this is still controversial. One limitation is the lack of high-throughput assays that can measure replication efficiency over multiple rounds of replication. We have developed a multiple-cycle growth competition assay to measure HIV-1 replication efficiency that uses flow cytometry to determine the relative proportions of test and reference viruses, each of which expresses a different reporter gene in place of nef. The reporter genes are expressed on the surface of infected cells and are detected by commercially available fluorescence-labeled antibodies. This method is less labor-intensive than those that require isolation and amplification of nucleic acids. The two reporter gene products are detected with similar specificity and sensitivity, and the proportion of infected cells in culture correlates with the amount of viral p24 antigen produced in the culture supernatant. HIV replication efficiencies of six different drug-resistant site-directed mutants were reproducibly quantified and were similar to those obtained with a growth competition assay in which the relative proportion of each variant was measured by sequence analysis, indicating that recombination between the pol and reporter genes was negligible. This assay also reproducibly quantified the relative fitness conferred by protease and reverse transcriptase sequences containing multiple drug resistance mutations, amplified from patient plasma. This flow cytometry-based growth competition assay offers advantages over current assays for HIV replication efficiency and should prove useful for the evaluation of patient samples in clinical trials.

  13. In vivo delivery of recombinant human growth hormone from genetically engineered human fibroblasts implanted within Baxter immunoisolation devices.

    Science.gov (United States)

    Josephs, S F; Loudovaris, T; Dixit, A; Young, S K; Johnson, R C

    1999-01-01

    Continuous delivery of therapeutic peptide to the systemic circulation would be the optimal treatment for a variety of diseases. The Baxter TheraCyte system is a membrane encapsulation system developed for implantation of tissues, cells such as endocrine cells or cell lines genetically engineered for therapeutic peptide delivery in vivo. To demonstrate the utility of this system, cell lines were developed which expressed human growth hormone (hGH) at levels exceeding 1 microgram per million cells per day. These were loaded into devices which were then implanted into juvenile nude rats. Significant levels of hGH of up to 2.5 ng/ml were detected in plasma throughout the six month duration of the study. In contrast, animals implanted with free cells showed peak plasma levels of 0.5 to 1.2 ng four days after implantation with no detectable hGH beyond 10 days. Histological examination of explanted devices showed they were vascularized and contained cells that were viable and morphologically healthy. After removal of the implants, no hGH could be detected which confirmed that the source of hGH was from cells contained within the device. The long term expression of human growth hormone as a model peptide has implications for the peptide therapies for a variety of human diseases using membrane encapsulated cells.

  14. Increased levels of specific leukocyte- and platelet-derived substances during normal anti-tetanus antibody synthesis in patients with inactive Crohn disease

    DEFF Research Database (Denmark)

    Nielsen, Hans Jørgen; Mortensen, T; Holten-Andersen, M

    2001-01-01

    /ml were inoculated with 1 ml (6 Lf units) of tetanus toxoid vaccine. The anti-tetanus antibody levels were determined in serum obtained before inoculation and after 7, 14 and 28 days, respectively. C-reactive protein (CRP), interleukin-6 (IL-6), tumour necrosis factor-alpha (TNF-alpha), histamine......, vascular endothelial growth factor (VEGF), tissue inhibitor of metalloproteinases-1 (TIMP-1), plasminogen activator inhibitor type-1 (PAI-1) and myeloperoxidase (MPO) were determined in serum or plasma obtained on the same days. RESULTS: After inoculation anti-tetanus antibody levels were equally raised...

  15. Effects of recombinant eel growth hormone on the uptake of ( sup 35 S)sulfate by ceratobranchial cartilages of the Japanese eel, Anguilla japonica

    Energy Technology Data Exchange (ETDEWEB)

    Duan, C.M.; Inui, Y. (Univ. of Tokyo (Japan))

    1990-08-01

    Effects of growth hormone (GH) on the synthesis of mucopolysaccharide by ceratobranchial cartilages of the Japanese eel, Anguilla japonica, were examined by monitoring the in vitro uptake of ({sup 35}S)sulfate. The ({sup 35}S)sulfate uptake decreased rapidly to one-third of the initial level during the first 3 days after hypophysectomy, and decreased gradually thereafter. When hypophysectomized eels were injected intramuscularly with recombinant eel GH (2 micrograms/g), the plasma GH concentrations increased maximally after 6 hr, and declined rapidly thereafter. On the other hand, the sulfate uptake increased significantly after 12 hr, and high levels were maintained until 48 hr. The stimulating effect of GH was dose dependent (0.02-2 micrograms/g). However, the addition of eel GH (0.05-5 micrograms/ml) to the culture medium did not affect the sulfate uptake by hypophysectomized eel cartilages, suggesting that the stimulative action of GH on the sulfate uptake by the cartilages is indirect.

  16. Effect of recombinant adenovirus encoding human p53 tumor suppressor gene (rAd-p53) on the growth and radiotherapeutic sensitivity of human lymphoma cell lines

    International Nuclear Information System (INIS)

    Yu Zeyang; Fan Wo; Li Dongqing; Zhu Ran; Wang Yongqing; Wu Jinchang

    2008-01-01

    Objective: To explore the inhibitory effect and radiation sensitization of recombinant adenovirus encoding human p53 tumor suppressor gene (rAd-p53) on human lymphoma cell lines. Methods: Human lymphoma cell lines Raji and Daudi were treated with rAd-p53, radiation therapy and combined treatment, respectively. The cell growth inhibition was assessed by MTT. The p53 protein expression was detected by Western blotting, and p53 mRNA was detected by BT-PCB. Results: The MTT results showed that the inhibitory effect and radiosensitivity enhancement of rAd-p53 on human lymphoma cell lines were not obvious [Raji: (27.5±4.1)%; Daudi: (28.1±1.6)%]. The results of Western blotting and BT-PCB showed that extrinsic p53 protein and p53 mRNA were expressed to some degree, but not at high-level. In addition, the results didn't demonstrate obvious radiosensitivity enhancement. Conclusions: The role of inhibition and radiosensitivity enhancement of rAd-p53 was not significant on human lymphoma cell lines. (authors)

  17. Immunoradiometric assay for the determination of E. coli proteins in recombinant dna derived human growth hormone produced at IPEN-CNEN/SP

    International Nuclear Information System (INIS)

    Soares, Carlos R.J.

    1995-01-01

    An immunoradiometric assay (IRMA) for the determination of multiple antigens was set up in order to quantify E. coli (ECP) in lots of purified recombinant human growth hormone (rec-hGH). SDS-PAGE and Western Blotting techniques were carried out, in parallel, to confirm the results obtained by IRMA and to provide more information about the contaminants. Anti-ECP antibodies were obtained by rabbit immunization with ECP, which were submitted to the same purification process utilized for rec-hGH with the exception of the last step. A strain-process-specific assay was thus set up. The antiserum obtained was purified through an affinity column prepared with the same ECP used for immunization, this provided an highly sensitive assay (0,03 ng ECP/mL). This IRMA was shown to be specific, not presenting any cross reaction with hGH and studies carried out on precision, accuracy and linearity of response with dilution confirmed its validity as one of the fundamental purity tests for rec-hGH produced at IPEN-CNEN/SP, whose principles can be easily extended to the analysis of other similar products. These studies have also shown that the utilization of an affinity column, prepared with the described anti-ECP antiserum was very effective, providing rec-hGH lots with less then 10 parts per million (0,001%) of contaminating proteins. (author). 45 refs., 15 figs., 11 tabs

  18. [Effect of concomitant use of dental drug on the properties of recombinant human basic fibroblast growth factor formulation for periodontal disease].

    Science.gov (United States)

    Sato, Yasuhiko; Oba, Takuma; Danjo, Kazumi

    2013-01-01

    We have discussed the essential property for periodontal disease medication using protein, such as recombinant human basic fibroblast growth factor (rhbFGF). In our previous study, the criteria of thickener for the medication, viscosity, flowability etc., were set. The aim of this study was to evaluate the physical and chemical effect of concomitant use of general dental drug or device on thickener properties for the clinical use of viscous rhbFGF formulation. Viscous formulation was prepared with six cellulose derivatives, two types hydroxy propyl cellulose (HPC), three types hydroxy ethyl cellulose (HEC) and methyl cellulose (MC). Antibiotic ointment, local anesthetic, bone graft substitute, agent for gargle and mouthwashes, were chosen as general dental drug and device. These drugs and device were mixed with the viscous formulations and the change of viscosity and flowability, the remaining ratio of rhbFGF were evaluated. When the various thickener solutions were mixed with the liquid drugs, viscosity and flowability did not changed much. However, in the case of MC solution, viscous property declined greatly when MC solution was mixed with cationic surfactant for gargle. The flowabilities of thickener solutions were declined with insoluble bone graft. The stabilities of rhbFGF in thickener solutions were no problem for 24 hours even in the case of mixing with dental drug or device. Our findings suggested that the viscous rhbFGF formulations prepared in this research were not substantially affected by the concomitant use of dental drug or device, especially the formulation with HPC or HEC was useful.

  19. Recombinant human growth hormone treatment, using two dose regimens in children with chronic renal failure--a report on linear growth and adverse effects

    DEFF Research Database (Denmark)

    Hertel, Niels Thomas; Holmberg, Christer; Rönnholm, Kai A R

    2002-01-01

    The aim of this study was to study the efficiency and the adverse effects of 2 or 4 IU/m2/day of growth hormone (GH) in the first year and 4 IU/m2/day in the second. Of 29 growth-retarded children with chronic renal failure (CRF) (aged 3.4-15.1 years), 23 completed the first year of therapy, and 16...... completed the second year. Height velocity SDS (HVSDS) increased in the first year in the low-dose group with 3.0, and 3.8 in the high-dose group. In the second year, HVSDS increased by 1.3 in the low-dose group and by 2.1 in high-dose group (p 3 ratio rose identically during...... the first year (p year of therapy in both groups. HbA1c, levels did not change. The number of adverse events was highest in the low-dose group, in which one patient developed...

  20. The effect of recombinant human growth hormone with or without rosiglitazone on hepatic fat content in HIV-1-infected individuals: a randomized clinical trial.

    Science.gov (United States)

    Kotler, Donald P; He, Qing; Engelson, Ellen S; Albu, Jeanine B; Glesby, Marshall J

    2016-01-01

    Hepatic fat is related to insulin resistance (IR) and visceral adipose tissue (VAT) in HIV+ and uninfected individuals. Growth hormone (GH) reduces VAT but increases IR. We evaluated the effects of recombinant human GH (rhGH) and rosiglitazone (Rosi) on hepatic fat in a substudy of a randomized controlled trial. HIV+ subjects with abdominal obesity and IR (QUICKI≤0.33) were randomized to rhGH 3 mg daily, Rosi 4 mg twice daily, the combination or double placebo. Hepatic fat was measured by magnetic resonance spectroscopy, visceral fat by MRI and IR by frequently sampled intravenous glucose tolerance tests at baseline and week 12. 31 subjects were studied at both time points. Significant correlations between hepatic fat and VAT (r=0.41; P=0.02) and QUICKI (r=0.39; P<0.05) were seen at baseline. IR rose with rhGH but not Rosi. When rhGH treatment groups were combined, hepatic fat expressed as percentage change decreased significantly (P<0.05) but did not change in Rosi (P=0.71). There were no correlations between changes in hepatic fat and VAT (P=0.4) or QUICKI (P=0.6). In a substudy of 21 subjects, a trend was noticed between changes in hepatic fat and serum insulin-like growth factor-1 (IGF-1; P=0.09). Hepatic fat correlates significantly with both VAT and IR, but changes in hepatic fat do not correlate with changes in VAT and glucose metabolism. Hepatic fat content is reduced by rhGH but Rosi has no effect. These results suggest an independent effect of GH or IGF-1 on hepatic fat. The study was registered at Clinicaltrials.gov (NCT00130286).

  1. The effect of recombinant human growth hormone with or without rosiglitazone on hepatic fat content in HIV-1 infected individuals; a randomized clinical trial

    Science.gov (United States)

    Kotler, Donald P; He, Qing; Engelson, Ellen S; Albu, Jeanine B; Glesby, Marshall J

    2016-01-01

    Background Hepatic fat is related to insulin resistance (IR) and visceral adipose tissue (VAT) in HIV+ and uninfected individuals. Growth hormone (GH) reduces VAT but increases IR. We evaluated the effects of recombinant human GH (rhGH) and rosiglitazone (Rosi) on hepatic fat in a substudy of a randomized controlled trial. Methods HIV+ subjects with abdominal obesity and IR (QUICKI ≤ 0.33) were randomized to rhGH 3 mg daily, Rosi 4 mg twice daily, the combination, or double placebo. Hepatic fat was measured by magnetic resonance spectroscopy (MRS), visceral fat by MRI, and IR by frequently sampled IV glucose tolerance tests at baseline and week 12. Results 31 subjects were studied at both time points. Significant correlations between hepatic fat and VAT (r = 0.41, p=0.02) and QUICKI (r = 0.39, p<0.05) were seen at baseline. Insulin resistance rose with rhGH but not Rosi. When rhGH treatment groups were combined, hepatic fat expressed as percent change decreased significantly (p<0.05) but did not change in Rosi (p=0.71). There were no correlations between changes in hepatic fat and VAT (p=0.4) or QUICKI (p=0.6). In a substudy of 21 subjects, a trend was noticed between changes in hepatic fat and serum IGF-1 (p=0.09). Conclusions Hepatic fat correlates significantly with both VAT and IR, but changes in hepatic fat do not correlate with changes in VAT and glucose metabolism. Hepatic fat content is reduced by rhGH but Rosi has no effect. These results suggest an independent effect of growth hormone or IGF-1 on hepatic fat. The study was registered at Clinicaltrials.gov (NCT00130286). PMID:25536669

  2. Industrial case study: evaluation of a mixed-mode resin for selective capture of a human growth factor recombinantly expressed in E. coli.

    Science.gov (United States)

    Kaleas, Kimberly A; Schmelzer, Charles H; Pizarro, Shelly A

    2010-01-08

    Mixed-mode chromatography resins are gaining popularity as effective purification tools for challenging feedstocks. This study presents the development of an industrial application to selectively capture recombinant human vascular endothelial growth factor (rhVEGF) on Capto MMC from an alkaline feedstock. Capto MMC resin contains a ligand that has the potential to participate in ionic, hydrophobic, and hydrogen boding interactions with proteins and is coupled to a highly cross-linked agarose bead matrix. VEGF is a key growth factor involved in angiogenesis and has therapeutic applications for wound healing. In this process, it is expressed in Escherichia coli as inclusion bodies. Solids are harvested from the cell lysate, and the rhVEGF is solubilized and refolded at pH 9.8 in the presence of urea and redox reagents. The unique mixed-mode characteristics of Capto MMC enabled capture of this basic protein with minimal load conditioning and delivered a concentrated pool for downstream processing with >95% yields while reducing host cell protein content to study explores the impact of loading conditions and residence time on the dynamic binding capacity as well as the development of elution conditions for optimal purification performance. After evaluating various elution buffers, l-arginine HCl was shown to be an effective eluting agent for rhVEGF desorption from the Capto MMC mixed-mode resin since it successfully disrupted the multiple interactions between the resin and rhVEGF. The lab scale effort produced a robust chromatography step that was successfully implemented at commercial manufacturing scale. Copyright 2009 Elsevier B.V. All rights reserved.

  3. [Antitumor effect of recombinant Xenopus laevis vascular endothelial growth factor (VEGF) as a vaccine combined with adriamycin on EL4 lymphoma in mice].

    Science.gov (United States)

    Niu, Ting; Liu, Ting; Jia, Yong-Qian; Liu, Ji-Yan; Wu, Yang; Hu, Bing; Tian, Ling; Yang, Li; Kan, Bing; Wei, Yu-Quan

    2005-09-01

    To explore the antitumor effect of immunotherapy with recombinant Xenopus laevis vascular endothelial growth factor (xVEGF) as a vaccine combined with adriamycin on lymphoma model in mice. EL4 lymphoma model was established in C57BL/6 mice. Mice were randomized into four groups: combination therapy, adriamycin alone, xVEGF alone and normal saline (NS) groups, and then were given relevant treatments. The growth of tumor, the survival rate of tumor-bearing mice, and the potential toxicity of regimens above were observed. Anti-VEGF antibody-producing B cells (APBCs) were detected by enzyme-linked immunospot (ELISPOT) assay. In addition, microvessel density (MVD) of tumor was detected by immunohistochemistry, and tumor cell apoptosis was also detected by TUNEL staining. The tumor volumes of mice were significantly smaller in combination group than those in other three groups (P < 0.05). Complete regression of tumor was observed in 3 of 10 mice in combination group. Forty-eight days after inoculation of tumor cells, the survival rate of mice was significantly higher in combination group than in NS group (P < 0.01). The anti-VEGF APBC count in combination group or xVEGF group was significantly higher, compared with that in adriamycin group or NS group (P < 0.01). MVD in tumor tissues was significantly lower in combination group than those in other three groups (P < 0.05). Moreover, tumor cell apoptosis was significantly higher in combination group than those in other three groups (P < 0.05). In this experimental study, the use of xVEGF vaccine and adriamycin as a combination of immunotherapy with chemotherapy has sucessfully produced synergistic antitumor effect on lymphoma in mice.

  4. Spectrum Recombination.

    Science.gov (United States)

    Greenslade, Thomas B., Jr.

    1984-01-01

    Describes several methods of executing lecture demonstrations involving the recombination of the spectrum. Groups the techniques into two general classes: bringing selected portions of the spectrum together using lenses or mirrors and blurring the colors by rapid movement or foreshortening. (JM)

  5. Effect of recombinant insulin-like growth factor-1 treatment on short-term linear growth in a child with Majewski osteodysplastic primordial dwarfism type II and hepatic insufficiency.

    Science.gov (United States)

    Faienza, Maria Felicia; Acquafredda, Angelo; D'Aniello, Mariangela; Soldano, Lucia; Marzano, Flaviana; Ventura, Annamaria; Cavallo, Luciano

    2013-01-01

    We report the case of a boy affected by severe intrauterine and postnatal growth retardation, microcephaly, facial dysmorphisms and postnecrotic cirrhosis, diagnosed at birth as having Seckel syndrome, and subsequently confirmed as Majewski osteodysplastic primordial dwarfism type II (MOPD II) on the basis of clinical and radiological features of skeletal dysplasia. At our observation (6 years 7 months) he presented height -10.3 standard deviation score (SDS), weight -22.1 SDS, head circumference -8 SDS, delayed bone age of 4 years with respect to chronological age. In consideration of the low levels of insulin-like growth factor-1 (IGF-1) as well as of hepatic insufficiency, we started the treatment with recombinant human IGF-1 (rhIGF-1) at the dose of 0.04 mg/kg in 2 doses/day, with an increase of 0.04 mg/kg after 1 week until the maximum dose of 0.12 mg/kg. We observed an early response to rhIGF-1 treatment, with a shift of height velocity from 1.8 cm/year (-4.6 SDS) at 4 cm/year (-1.9 SDS), and an increase in bone age of 1.5 years during the first 6 months. rhIGF-1 treatment does not seem to be able to replace the physiological action of IGF-1 in patients with MOPD II and hepatic insufficiency, however, it seems to preserve the typical growth pattern of MOPD II patients, avoiding a further widening of the growth deficiency in these subjects.

  6. Impact of alg3 gene deletion on growth, development, pigment production, protein secretion, and functions of recombinant Trichoderma reesei cellobiohydrolases in Aspergillus niger.

    Science.gov (United States)

    Dai, Ziyu; Aryal, Uma K; Shukla, Anil; Qian, Wei-Jun; Smith, Richard D; Magnuson, Jon K; Adney, William S; Beckham, Gregg T; Brunecky, Roman; Himmel, Michael E; Decker, Stephen R; Ju, Xiaohui; Zhang, Xiao; Baker, Scott E

    2013-12-01

    Dolichyl-P-Man:Man(5)GlcNAc(2)-PP-dolichyl α-1,3-mannosyltransferase (also known as "asparagine-linked glycosylation 3", or ALG3) is involved in early N-linked glycan synthesis and thus is essential for formation of N-linked protein glycosylation. In this study, we examined the effects of alg3 gene deletion (alg3Δ) on growth, development, pigment production, protein secretion and recombinant Trichoderma reesei cellobiohydrolase (rCel7A) expressed in Aspergillus niger. The alg3Δ delayed spore germination in liquid cultures of complete medium (CM), potato dextrose (PD), minimal medium (MM) and CM with addition of cAMP (CM+cAMP), and resulted in significant reduction of hyphal growth on CM, potato dextrose agar (PDA), and CM+cAMP and spore production on CM. The alg3Δ also led to a significant accumulation of red pigment on both liquid and solid CM cultures. The relative abundances of 54 of the total 215 proteins identified in the secretome were significantly altered as a result of alg3Δ, 63% of which were secreted at higher levels in alg3Δ strain than the parent. The rCel7A expressed in the alg3Δ mutant was smaller in size than that expressed in both wild-type and parental strains, but still larger than T. reesei Cel7A. The circular dichroism (CD)-melt scans indicated that change in glycosylation of rCel7A does not appear to impact the secondary structure or folding. Enzyme assays of Cel7A and rCel7A on nanocrystalline cellulose and bleached kraft pulp demonstrated that the rCel7As have improved activities on hydrolyzing the nanocrystalline cellulose. Overall, the results suggest that alg3 is critical for growth, sporulation, pigment production, and protein secretion in A. niger, and demonstrate the feasibility of this alternative approach to evaluate the roles of N-linked glycosylation in glycoprotein secretion and function. Copyright © 2013 Elsevier Inc. All rights reserved.

  7. Development and Characterization of Sodium Hyaluronate Microparticle-Based Sustained Release Formulation of Recombinant Human Growth Hormone Prepared by Spray-Drying.

    Science.gov (United States)

    Kim, Sun J; Kim, Chan W

    2016-02-01

    The purpose of this study was to develop and characterize a sodium hyaluronate microparticle-based sustained release formulation of recombinant human growth hormone (SR-rhGH) prepared by spray-drying. Compared to freeze-drying, spray-dried SR-rhGH showed not only prolonged release profiles but also better particle property and injectability. The results of size-exclusion high-performance liquid chromatography showed that no aggregate was detected, and dimer was just about 2% and also did not increase with increase of inlet temperature up to 150 °C. Meanwhile, the results of reversed-phase high-performance liquid chromatography revealed that related proteins increased slightly from 4.6% at 100 °C to 6.3% at 150 °C. Thermal mapping test proved that product temperature did not become high to cause protein degradation during spray-drying because thermal energy was used for the evaporation of surface moisture of droplets. The structural characterization by peptide mapping, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, and circular dichroism revealed that the primary, secondary, and tertiary structures of rhGH in SR-rhGH were highly comparable to those of reference somatropin materials. The biological characterization by rat weight gain and cell proliferation assays provided that bioactivity of SR-rhGH was equivalent to that of native hGH. These data establish that spray-dried SR-rhGH is highly stable by preserving intact rhGH and hyaluronate microparticle-based formulation by spray-drying can be an alternative delivery system for proteins. Copyright © 2016 American Pharmacists Association®. Published by Elsevier Inc. All rights reserved.

  8. A pilot study of regenerative therapy using controlled release of recombinant human fibroblast growth factor for patients with pre-collapse osteonecrosis of the femoral head.

    Science.gov (United States)

    Kuroda, Yutaka; Asada, Ryuta; So, Kazutaka; Yonezawa, Atsushi; Nankaku, Manabu; Mukai, Kumi; Ito-Ihara, Toshiko; Tada, Harue; Yamamoto, Michio; Murayama, Toshinori; Morita, Satoshi; Tabata, Yasuhiko; Yokode, Masayuki; Shimizu, Akira; Matsuda, Shuichi; Akiyama, Haruhiko

    2016-08-01

    We evaluated the safety and clinical outcomes of a single local administration of gelatin hydrogel impregnated with recombinant human fibroblast growth factor (rhFGF)-2 for the treatment of the precollapse stage of osteonecrosis of the femoral head (ONFH). Patients with ONFH (precollapse stage ≤2) received a single local administration of 800 μg of rhFGF-2-impregnated gelatin hydrogel and were followed up for one year. The surgery was performed using a minimally invasive technique involving a 1-cm skin incision, and walking was allowed from day one postoperatively. The primary outcomes included occurrence of adverse events and complications. The secondary outcomes included changes in the Harris hip scores, visual analog scale for pain scores, University of California, Los Angeles (UCLA) activity scores, and radiological images. We included ten patients, of which five experienced 14 adverse events, including one complication from spinal anesthesia. However, patients completely recovered from all adverse events. The mean clinical scores significantly improved by one year postoperatively compared with the pre-operative scores (before vs. after: visual analog score for pain, 21.2 vs. 5.3 mm; UCLA activity score, 5.5 vs. 6.6; Harris hip score, 81.0 vs. 96.9 points). There was only one case of femoral head collapse; however, this occurred in a hip with extensive necrosis. Stage progression and collapse did not occur in the other nine cases. Computed tomography confirmed bone regeneration in the femoral heads. Clinical application of rhFGF-2-impregnated gelatin hydrogel for patients with precollapse ONFH was feasible and safe.

  9. Chimeric Recombinant Human Metapneumoviruses with the Nucleoprotein or Phosphoprotein Open Reading Frame Replaced by That of Avian Metapneumovirus Exhibit Improved Growth In Vitro and Attenuation In Vivo

    Science.gov (United States)

    Pham, Quynh N.; Biacchesi, Stéphane; Skiadopoulos, Mario H.; Murphy, Brian R.; Collins, Peter L.; Buchholz, Ursula J.

    2005-01-01

    Chimeric versions of recombinant human metapneumovirus (HMPV) were generated by replacing the nucleoprotein (N) or phosphoprotein (P) open reading frame with its counterpart from the closely related avian metapneumovirus (AMPV) subgroup C. In Vero cells, AMPV replicated to an approximately 100-fold-higher titer than HMPV. Surprisingly, the N and P chimeric viruses replicated to a peak titer that was 11- and 25-fold higher, respectively, than that of parental HMPV. The basis for this effect is not known but was not due to obvious changes in the efficiency of gene expression. AMPV and the N and P chimeras were evaluated for replication, immunogenicity, and protective efficacy in hamsters. AMPV was attenuated compared to HMPV in this mammalian host on day 5 postinfection, but not on day 3, and only in the nasal turbinates. In contrast, the N and P chimeras were reduced approximately 100-fold in both the upper and lower respiratory tract on day 3 postinfection, although there was little difference by day 5. The N and P chimeras induced a high level of neutralizing serum antibodies and protective efficacy against HMPV; AMPV was only weakly immunogenic and protective against HMPV challenge, reflecting antigenic differences. In African green monkeys immunized intranasally and intratracheally, the mean peak titer of the P chimera was reduced 100- and 1,000-fold in the upper and lower respiratory tracts, whereas the N chimera was reduced only 10-fold in the lower respiratory tract. Both chimeras were comparable to wild-type HMPV in immunogenicity and protective efficacy. Thus, the P chimera is a promising live HMPV vaccine candidate that paradoxically combines improved growth in vitro with attenuation in vivo. PMID:16306583

  10. Effects of up to 15 years of recombinant human GH (rhGH) replacement on bone metabolism in adults with growth hormone deficiency (GHD): the Leiden Cohort Study.

    Science.gov (United States)

    Appelman-Dijkstra, Natasha M; Claessen, Kim M J A; Hamdy, Neveen A T; Pereira, Alberto M; Biermasz, Nienke R

    2014-11-01

    Growth hormone deficiency (GHD) in adulthood may be associated with a decreased bone mineral density (BMD), a decreased bone mineral content (BMC) and an increased fracture risk. Recombinant human GH (rhGH) replacement induces a progressive increase in BMD for up to 5-7 years of treatment. Data on longer follow-up are, however, scarce. Two hundred and thirty-adult GHD patients (mean age 47·1 years, 52·6% female), of whom 88% patients had adult-onset (AO) GHD, receiving rhGH replacement for ≥5 years were included in the study. Most patients had multiple pituitary hormone deficiencies. Bone turnover markers, BMC and BMD and T-scores at the lumbar spine and femoral neck were evaluated at baseline, and after 5, 10 and 15 years of rhGH replacement. In addition, clinical fracture incidence was assessed. Mean lumbar spine BMD, lumbar spine BMC and T-scores gradually increased during the first 10 years of rhGH replacement and remained stable thereafter. Largest effects of rhGH supplementation were found in men. In the small subset of patients using bisphosphonates, use of bisphosphonates did not impact additional beneficial effects in the long term. Low baseline BMD positively affected the change in BMD and BMC over time, but there was a negative effect of high GH dose at 1 year on the change in BMD and BMC over time. Clinical fracture incidence during long-term rhGH replacement was 20.1/1000 py. Fifteen years of rhGH replacement in GHD adults resulted in a sustained increase in BMD values at the lumbar spine, particularly in men, and stabilization of BMD values at the femoral neck. Clinical fracture incidence was suggested not to be increased during long-term rhGH replacement. © 2014 John Wiley & Sons Ltd.

  11. Ridge augmentation using recombinant human fibroblast growth factor-2 with biodegradable gelatin sponges incorporating β-tricalcium phosphate: a preclinical study in dogs.

    Science.gov (United States)

    Hoshi, S; Akizuki, T; Matsuura, T; Ikawa, T; Kinoshita, A; Oda, S; Tabata, Y; Matsui, M; Izumi, Y

    2016-02-01

    Fibroblast growth factor-2 (FGF-2) regulates the proliferation and differentiation of osteogenic cells, resulting in the promotion of bone formation. Biodegradable gelatin sponges incorporating β-tricalcium phosphate (β-TCP) have been reported as a scaffold, which has the ability to control growth factor release, offering sufficient mechanical strength and efficient migration of mesenchymal cells. In this study, we evaluated the effects of the combined use of recombinant human FGF-2 (rhFGF-2) and gelatin/β-TCP sponge on ridge augmentation in dogs. Six male beagle dogs were used in this study. Twelve wk after tooth extraction, bilateral 10 × 5 mm (width × depth) saddle-type defects were created 3 mm apart from the mesial side of the maxillary canine. At the experimental sites, the defects were filled with gelatin/β-TCP sponge infiltrated with 0.3% rhFGF-2, whereas gelatin/β-TCP sponge infiltrated with saline was applied to the control sites. Eight wk after surgery, qualitative and quantitative analyses were performed. There were no signs of clinical inflammation at 8 wk after surgery. Histometric measurements revealed that new bone height at the experimental sites (2.98 ± 0.65 mm) was significantly greater than that at the control sites (1.56 ± 0.66 mm; p = 0.004). The total tissue height was greater at the experimental sites (6.62 ± 0.66 mm) than that at the control sites (5.95 ± 0.74 mm), although there was no statistical significant difference (p = 0.051). Cast model measurements revealed that the residual defect height at the experimental sites (2.31 ± 0.50 mm) was significantly smaller than that at the control sites (3.51 ± 0.78 mm; p = 0.012). The combined use of rhFGF-2 and gelatin/β-TCP sponge promotes ridge augmentation in canine saddle-type bone defects. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  12. Genetic blockade of insulin-like growth factor-1 receptor via recombinant adenovirus in lung cancer can be enhanced by the histone deacetylase inhibitor, vorinostat.

    Science.gov (United States)

    Park, Mi-Young; Kim, Dal Rae; Eo, Eun Young; Lim, Hyo Jeong; Park, Jong Sun; Cho, Young-Jae; Yoon, Ho-Il; Lee, Jae Ho; Lee, Choon-Taek

    2013-01-01

    Many approaches have been suggested as anti-tumor therapy for targeting insulin-like growth factor 1 receptor (IGF-1R), such as monoclonal antibodies and tyrosine kinase inhibitor. We introduced recombinant adenoviruses expressing antisense, dominant negative or short hairpin RNA to IGF-1R. Moreover, we demonstrated that histone deacetylase inhibitor (vorinostat) can increase the transduction efficiency of adenoviruses by increasing CAR-induced transduction and by enhancing the transcription of the adenoviral transgene. In the present study, we showed that the combination of ad-sh (short hairpin) IGF-1R with vorinostat leads to a synergistic enhancement of IGF-1R blockade. We measured the change in IGF-1R upon cotreatment with vorinostat and ad-shIGF-1R. Changes in transduction efficiency of ad-shIGF-1R were measured by fluorescent microscopy. Changes in apoptotic proportion and cell survival after the cotreatment were measured by the sub-G1 assay and cell counts. The effect of nuclear factor (NF)-κB activation was also measured by NF-κB p65 activation enzyme-linked immunosorbent assay. Drug interactions were analyzed upon cotreatment with ad-shIGF-1R, vorinostat and cisplatin. Combined treatment of ad-shIGF-1R and vorinostat synergistically suppressed the IGF-1R expression in lung cancer cell lines and also increased the transduction efficiency of ad-shIGF-1R. Ad-shIGF-1R and vorinostat cotreatment increased apoptotic cell death and synergistically suppressed cell growth compared to ad-shIGF-1R or vorinostat treatment alone. Vorinostat suppressed NF-κB activation, which was activated by ad-shIGF-1R. Moreover, triple combination of ad-shIGF-1R, vorinostat and cisplatin demonstrated synergistic cytotoxicity on lung cancer cells. Vorinostat enhanced the blocking capability of ad-shIGF-1R. The combined treatment of vorinostat and ad-sh-IGF-1R appears to have promising potential as a new therapeutic approach for lung cancer. Copyright © 2013 John Wiley & Sons, Ltd.

  13. A prospective, randomized pilot study on the safety and efficacy of recombinant human growth and differentiation factor-5 coated onto β-tricalcium phosphate for sinus lift augmentation.

    Science.gov (United States)

    Koch, Felix P; Becker, Jürgen; Terheyden, Hendrik; Capsius, Björn; Wagner, Wilfried

    2010-11-01

    The aim of this prospective, randomized clinical trial was to investigate the potential of recombinant human growth and differentiation factor-5 (rhGDF-5) coated onto β-tricalcium phosphate (β-TCP) (rhGDF-5/β-TCP) to support bone formation after sinus lift augmentation. In total, 31 patients participated in this multicenter clinical trial. They required a two-stage unilateral maxillary sinus floor augmentation (residual bone height augmentation with rhGDF-5/β-TCP and a 3-month healing period, (b) augmentation with rhGDF-5/β-TCP and a 4-month healing period and (c) medical device β-TCP mixed with autologous bone and a 4-month healing period. The primary study objective was the area of newly formed bone within the augmented area as assessed by histomorphometric evaluation of trephine bur biopsies. The osseous regeneration was similar in each treatment group; the amount of newly formed bone ranged between 28% (± 15.5%) and 31.8% (± 17.9%). Detailed analysis of histological data will be published elsewhere. As secondary efficacy variables, the augmentation height at the surgery site was measured by radiography. The largest augmentation was radiologically achieved in the rhGDF-5/β-TCP - 3-month and the rhGDF-5/β-TCP - 4-month treatment groups. As safety parameters, adverse events were recorded and anti-drug antibody levels were evaluated. Most of the adverse events were judged as unrelated to the study medication. Four out of 47 (8.5%) implants failed in patients treated with rhGDF-5/β-TCP, a result that is in agreement with the general implant failure rate of 5-15%. Transiently very low amounts of anti-rhGDF-5 antibodies were detected in some patients who received rhGDF-5, which was not related to the bone formation outcome. rhGDF-5/β-TCP was found to be effective and safe as the control treatment with autologous bone mixed β-TCP in sinus floor augmentation. Thus, further investigation regarding efficacy and safety will be carried out in larger patient

  14. Production of recombinant AAV vectors encoding insulin-like growth factor I is enhanced by interaction among AAV rep regulatory sequences

    Directory of Open Access Journals (Sweden)

    Dilley Robert

    2009-01-01

    Full Text Available Abstract Background Adeno-associated virus (AAV vectors are promising tools for gene therapy. Currently, their potential is limited by difficulties in producing high vector yields with which to generate transgene protein product. AAV vector production depends in part upon the replication (Rep proteins required for viral replication. We tested the hypothesis that mutations in the start codon and upstream regulatory elements of Rep78/68 in AAV helper plasmids can regulate recombinant AAV (rAAV vector production. We further tested whether the resulting rAAV vector preparation augments the production of the potentially therapeutic transgene, insulin-like growth factor I (IGF-I. Results We constructed a series of AAV helper plasmids containing different Rep78/68 start codon in combination with different gene regulatory sequences. rAAV vectors carrying the human IGF-I gene were prepared with these vectors and the vector preparations used to transduce HT1080 target cells. We found that the substitution of ATG by ACG in the Rep78/68 start codon in an AAV helper plasmid (pAAV-RC eliminated Rep78/68 translation, rAAV and IGF-I production. Replacement of the heterologous sequence upstream of Rep78/68 in pAAV-RC with the AAV2 endogenous p5 promoter restored translational activity to the ACG mutant, and restored rAAV and IGF-I production. Insertion of the AAV2 p19 promoter sequence into pAAV-RC in front of the heterologous sequence also enabled ACG to function as a start codon for Rep78/68 translation. The data further indicate that the function of the AAV helper construct (pAAV-RC, that is in current widespread use for rAAV production, may be improved by replacement of its AAV2 unrelated heterologous sequence with the native AAV2 p5 promoter. Conclusion Taken together, the data demonstrate an interplay between the start codon and upstream regulatory sequences in the regulation of Rep78/68 and indicate that selective mutations in Rep78/68 regulatory elements

  15. Recombinant human growth hormone and rosiglitazone for abdominal fat accumulation in HIV-infected patients with insulin resistance: a randomized, double-blind, placebo-controlled, factorial trial.

    Directory of Open Access Journals (Sweden)

    Marshall J Glesby

    Full Text Available Recombinant human growth hormone (rhGH reduces visceral adipose tissue (VAT volume in HIV-infected patients but can worsen glucose homeostasis and lipoatrophy. We aimed to determine if adding rosiglitazone to rhGH would abrogate the adverse effects of rhGH on insulin sensitivity (SI and subcutaneous adipose tissue (SAT volume.Randomized, double-blind, placebo-controlled, multicenter trial using a 2×2 factorial design in which HIV-infected subjects with abdominal obesity and insulin resistance were randomized to rhGH 3 mg daily, rosiglitazone 4 mg twice daily, combination rhGH + rosiglitazone, or double placebo (control for 12 weeks. The primary endpoint was change in SI by frequently sampled intravenous glucose tolerance test from entry to week 12. Body composition was assessed by whole body magnetic resonance imaging (MRI and dual Xray absorptiometry (DEXA. Seventy-seven subjects were randomized of whom 72 initiated study drugs. Change in SI from entry to week 12 differed across the 4 arms by 1-way ANCOVA (P = 0.02; by pair-wise comparisons, only rhGH (decreasing SI; P = 0.03 differed significantly from control. Changes from entry to week 12 in fasting glucose and glucose area under the curve on 2-hour oral glucose tolerance test differed across arms (1-way ANCOVA P = 0.004, increasing in the rhGH arm relative to control. VAT decreased significantly in the rhGH arms (-17.5% in rhGH/rosiglitazone and -22.7% in rhGH but not in the rosiglitazone alone (-2.5% or control arms (-1.9%. SAT did not change significantly in any arm. DEXA results were consistent with the MRI data. There was no significant rhGH x rosiglitazone interaction for any body composition parameter.The addition of rosiglitazone abrogated the adverse effects of rhGH on insulin sensitivity and glucose tolerance while not significantly modifying the lowering effect of rhGH on VAT.Clinicaltrials.gov NCT00130286.

  16. Recombinant human growth hormone and rosiglitazone for abdominal fat accumulation in HIV-infected patients with insulin resistance: a randomized, double-blind, placebo-controlled, factorial trial.

    Science.gov (United States)

    Glesby, Marshall J; Albu, Jeanine; Chiu, Ya-Lin; Ham, Kirsis; Engelson, Ellen; He, Qing; Muthukrishnan, Varalakshmi; Ginsberg, Henry N; Donovan, Daniel; Ernst, Jerry; Lesser, Martin; Kotler, Donald P

    2013-01-01

    Recombinant human growth hormone (rhGH) reduces visceral adipose tissue (VAT) volume in HIV-infected patients but can worsen glucose homeostasis and lipoatrophy. We aimed to determine if adding rosiglitazone to rhGH would abrogate the adverse effects of rhGH on insulin sensitivity (SI) and subcutaneous adipose tissue (SAT) volume. Randomized, double-blind, placebo-controlled, multicenter trial using a 2×2 factorial design in which HIV-infected subjects with abdominal obesity and insulin resistance were randomized to rhGH 3 mg daily, rosiglitazone 4 mg twice daily, combination rhGH + rosiglitazone, or double placebo (control) for 12 weeks. The primary endpoint was change in SI by frequently sampled intravenous glucose tolerance test from entry to week 12. Body composition was assessed by whole body magnetic resonance imaging (MRI) and dual Xray absorptiometry (DEXA). Seventy-seven subjects were randomized of whom 72 initiated study drugs. Change in SI from entry to week 12 differed across the 4 arms by 1-way ANCOVA (P = 0.02); by pair-wise comparisons, only rhGH (decreasing SI; P = 0.03) differed significantly from control. Changes from entry to week 12 in fasting glucose and glucose area under the curve on 2-hour oral glucose tolerance test differed across arms (1-way ANCOVA P = 0.004), increasing in the rhGH arm relative to control. VAT decreased significantly in the rhGH arms (-17.5% in rhGH/rosiglitazone and -22.7% in rhGH) but not in the rosiglitazone alone (-2.5%) or control arms (-1.9%). SAT did not change significantly in any arm. DEXA results were consistent with the MRI data. There was no significant rhGH x rosiglitazone interaction for any body composition parameter. The addition of rosiglitazone abrogated the adverse effects of rhGH on insulin sensitivity and glucose tolerance while not significantly modifying the lowering effect of rhGH on VAT. Clinicaltrials.gov NCT00130286.

  17. Curative Metatarsal Bone Surgery Combined with Intralesional Administration of Recombinant Human Epidermal Growth Factor in Diabetic Neuropathic Ulceration of the Forefoot: A Prospective, Open, Uncontrolled, Nonrandomized, Observational Study

    Directory of Open Access Journals (Sweden)

    Aristides L. Garcia Herrera, MD, PhD

    2017-01-01

    Conclusions: The combination of curative metatarsal bone surgery with intralesional administration of recombinant human EGF resulted in a significant reduction in the re-epithelization time, recidivism, and development of new diabetic lesions. The safety profile was appropriate. However, more randomized, triple-blind, and placebo trials are needed to evaluate the efficacy and safety of this new therapy.

  18. Targeted recombinant fusion proteins of IFNγ and mimetic IFNγ with PDGFβR bicyclic peptide inhibits liver fibrogenesis in vivo.

    Directory of Open Access Journals (Sweden)

    Ruchi Bansal

    Full Text Available Hepatic stellate cells (HSCs, following transdifferentiation to myofibroblasts plays a key role in liver fibrosis. Therefore, attempts to attenuate this myofibroblastic phenotype would be a promising therapeutic approach. Interferon gamma (IFNγ is a potent anti-fibrotic cytokine, but its pleiotropic receptor expression leading to severe adverse effects has limited its clinical application. Since, activated HSC express high-level of platelet derived growth factor beta receptor (PDGFβR, we investigated the potential of PDGFβR-specific targeting of IFNγ and its signaling peptide that lacks IFNγR binding site (mimetic IFNγ or mimIFNγ in liver fibrosis. We prepared DNA constructs expressing IFNγ, mimIFNγ or BiPPB (PDGFβR-specific bicyclic peptide-IFNγ, BiPPB-mimIFNγ fusion proteins. Both chimeric proteins alongwith IFNγ and mimIFNγ were produced in E.coli. The expressed proteins were purified and analyzed for PDGFβR-specific binding and in vitro effects. Subsequently, these recombinant proteins were investigated for the liver uptake (pSTAT1α signaling pathway, for anti-fibrotic effects and adverse effects (platelet counts in CCl4-induced liver fibrogenesis in mice. The purified HSC-targeted IFNγ and mimIFNγ fusion proteins showed PDGFβR-specific binding and significantly reduced TGFβ-induced collagen-I expression in human HSC (LX2 cells, while mouse IFNγ and mimIFNγ did not show any effect. Conversely, mouse IFNγ and BiPPB-IFNγ induced activation and dose-dependent nitric oxide release in mouse macrophages (express IFNγR while lack PDGFβR, which was not observed with mimIFNγ and BiPPB-mimIFNγ, due to the lack of IFNγR binding sites. In vivo, targeted BiPPB-IFNγ and BiPPB-mimIFNγ significantly activated intrahepatic IFNγ-signaling pathway compared to IFNγ and mimIFNγ suggesting increased liver accumulation. Furthermore, the targeted fusion proteins ameliorated liver fibrogenesis in mice by significantly reducing

  19. Recombinant Collagenlike Proteins

    Science.gov (United States)

    Fertala, Andzej

    2007-01-01

    A group of collagenlike recombinant proteins containing high densities of biologically active sites has been invented. The method used to express these proteins is similar to a method of expressing recombinant procollagens and collagens described in U. S. Patent 5,593,859, "Synthesis of human procollagens and collagens in recombinant DNA systems." Customized collagenous proteins are needed for biomedical applications. In particular, fibrillar collagens are attractive for production of matrices needed for tissue engineering and drug delivery. Prior to this invention, there was no way of producing customized collagenous proteins for these and other applications. Heretofore, collagenous proteins have been produced by use of such biological systems as yeasts, bacteria, and transgenic animals and plants. These products are normal collagens that can also be extracted from such sources as tendons, bones, and hides. These products cannot be made to consist only of biologically active, specific amino acid sequences that may be needed for specific applications. Prior to this invention, it had been established that fibrillar collagens consist of domains that are responsible for such processes as interaction with cells, binding of growth factors, and interaction with a number of structural proteins present in the extracellular matrix. A normal collagen consists of a sequence of domains that can be represented by a corresponding sequence of labels, e.g., D1D2D3D4. A collagenlike protein of the present invention contains regions of collagen II that contain multiples of a single domain (e.g., D1D1D1D1 or D4D4D4D4) chosen for its specific biological activity. By virtue of the multiplicity of the chosen domain, the density of sites having that specific biological activity is greater than it is in a normal collagen. A collagenlike protein according to this invention can thus be made to have properties that are necessary for tissue engineering.

  20. Growth

    Science.gov (United States)

    John R. Jones; George A. Schier

    1985-01-01

    This chapter considers aspen growth as a process, and discusses some characteristics of the growth and development of trees and stands. For the most part, factors affecting growth are discussed elsewhere, particularly in the GENETICS AND VARIATION chapter and in chapters in PART 11. ECOLOGY. Aspen growth as it relates to wood production is examined in the WOOD RESOURCE...

  1. Enhanced direct-gap light emission from Si-capped n+-Ge epitaxial layers on Si after post-growth rapid cyclic annealing: impact of non-radiative interface recombination toward Ge/Si double heterostructure lasers.

    Science.gov (United States)

    Higashitarumizu, Naoki; Ishikawa, Yasuhiko

    2017-09-04

    Enhanced direct-gap light emission is reported for Si-capped n + -Ge layers on Si after post-growth rapid cyclic annealing (RCA), and impact of non-radiative recombination (NRR) at the Ge/Si interface is discussed toward Ge/Si double heterostructure (DH) lasers. P-doped n + -Ge layer (1 × 10 19 cm -3 , 400 nm) is grown on Si by ultra-high vacuum chemical vapor deposition, followed by a growth of Si capping layer (5 nm) to form a Si/Ge/Si DH structure. Post-growth RCA to eliminate defects in Ge is performed in N 2 at temperatures between 900°C and 780°C, where the annealing time is minimized to be 5 s in each RCA cycle to prevent an out-diffusion of P dopants from the Ge surface. Direct-gap photoluminescence (PL) intensity at 1.6 µm increases with the RCA cycles up to 40, although the threading dislocation density in Ge is not reduced after 3 cycles in the present condition. The PL enhancement is ascribed to the suppression of NRR at the Ge/Si interface, where an intermixed SiGe alloy is formed. For Ge/Si DH lasers, NRR at the Ge/Si interface is found to have a significant impact on the threshold current density Jth. In order to achieve Jth on the order of 1 kA/cm 2 , similar to III-V lasers, the interface recombination velocity S is required below 10 3 cm/s in spite of S as large as 10 5 cm/s at the ordinary defect-rich Ge/Si interface.

  2. Minoxidil Promotes Hair Growth through Stimulation of Growth Factor Release from Adipose-Derived Stem Cells

    Directory of Open Access Journals (Sweden)

    Nahyun Choi

    2018-02-01

    Full Text Available Minoxidil directly promotes hair growth via the stimulation of dermal papilla (DP and epithelial cells. Alternatively, there is little evidence for indirect promotion of hair growth via stimulation of adipose-derived stem cells (ASCs. We investigated whether minoxidil stimulates ASCs and if increased growth factor secretion by ASCs facilitates minoxidil-induced hair growth. Telogen-to-anagen induction was examined in mice. Cultured DP cells and vibrissae hair follicle organ cultures were used to further examine the underlying mechanisms. Subcutaneous injection of minoxidil-treated ASCs accelerated telogen-to-anagen transition in mice, and increased hair weight at day 14 post-injection. Minoxidil did not alter ASC proliferation, but increased migration and tube formation. Minoxidil also increased the secretion of growth factors from ASCs, including chemokine (C-X-C motif ligand 1 (CXCL1, platelet-derived endothelial cell growth factor (PD-ECGF, and platelet-derived growth factor-C (PDGF-C. Minoxidil increased extracellular signal–regulated kinases 1/2 (ERK1/2 phosphorylation, and concomitant upregulation of PD-ECGF and PDGF-C mRNA levels were attenuated by an ERK inhibitor. Subcutaneous injection of CXCL1, PD-ECGF, or PDGF-C enhanced anagen induction in mice, and both CXCL1 and PDGF-C increased hair length in ex vivo organ culture. Treatment with CXCL1, PD-ECGF, or PDGF-C also increased the proliferation index in DP cells. Finally, topical application of CXCL1, PD-ECGF, or PDGF-C with 2% minoxidil enhanced anagen induction when compared to minoxidil alone. Minoxidil stimulates ASC motility and increases paracrine growth factor signaling. Minoxidil-stimulated secretion of growth factors by ASCs may enhance hair growth by promoting DP proliferation. Therefore, minoxidil can be used as an ASC preconditioning agent for hair regeneration.

  3. Minoxidil Promotes Hair Growth through Stimulation of Growth Factor Release from Adipose-Derived Stem Cells

    Science.gov (United States)

    Choi, Nahyun; Shin, Soyoung; Song, Sun U.; Sung, Jong-Hyuk

    2018-01-01

    Minoxidil directly promotes hair growth via the stimulation of dermal papilla (DP) and epithelial cells. Alternatively, there is little evidence for indirect promotion of hair growth via stimulation of adipose-derived stem cells (ASCs). We investigated whether minoxidil stimulates ASCs and if increased growth factor secretion by ASCs facilitates minoxidil-induced hair growth. Telogen-to-anagen induction was examined in mice. Cultured DP cells and vibrissae hair follicle organ cultures were used to further examine the underlying mechanisms. Subcutaneous injection of minoxidil-treated ASCs accelerated telogen-to-anagen transition in mice, and increased hair weight at day 14 post-injection. Minoxidil did not alter ASC proliferation, but increased migration and tube formation. Minoxidil also increased the secretion of growth factors from ASCs, including chemokine (C-X-C motif) ligand 1 (CXCL1), platelet-derived endothelial cell growth factor (PD-ECGF), and platelet-derived growth factor-C (PDGF-C). Minoxidil increased extracellular signal–regulated kinases 1/2 (ERK1/2) phosphorylation, and concomitant upregulation of PD-ECGF and PDGF-C mRNA levels were attenuated by an ERK inhibitor. Subcutaneous injection of CXCL1, PD-ECGF, or PDGF-C enhanced anagen induction in mice, and both CXCL1 and PDGF-C increased hair length in ex vivo organ culture. Treatment with CXCL1, PD-ECGF, or PDGF-C also increased the proliferation index in DP cells. Finally, topical application of CXCL1, PD-ECGF, or PDGF-C with 2% minoxidil enhanced anagen induction when compared to minoxidil alone. Minoxidil stimulates ASC motility and increases paracrine growth factor signaling. Minoxidil-stimulated secretion of growth factors by ASCs may enhance hair growth by promoting DP proliferation. Therefore, minoxidil can be used as an ASC preconditioning agent for hair regeneration. PMID:29495622

  4. Growth factor effects on costal chondrocytes for tissue engineering fibrocartilage

    Science.gov (United States)

    Johns, D.E.; Athanasiou, K.A.

    2010-01-01

    Tissue engineered fibrocartilage could become a feasible option for replacing tissues like the knee meniscus or temporomandibular joint disc. This study employed five growth factors insulin-like growth factor-I, transforming growth factor-β1, epidermal growth factor, platelet-derived growth factor-BB, and basic fibroblast growth factor in a scaffoldless approach with costal chondrocytes, attempting to improve biochemical and mechanical properties of engineered constructs. Samples were quantitatively assessed for total collagen, glycosaminoglycans, collagen type I, collagen type II, cells, compressive properties, and tensile properties at two time points. Most treated constructs were worse than the no growth factor control, suggesting a detrimental effect, but the IGF treatment tended to improve the constructs. Additionally, the 6wk time point was consistently better than 3wks, with total collagen, glycosaminoglycans, and aggregate modulus doubling during this time. Further optimization of the time in culture and exogenous stimuli will be important in making a more functional replacement tissue. PMID:18597118

  5. RNAi and heterochromatin repress centromeric meiotic recombination

    DEFF Research Database (Denmark)

    Ellermeier, Chad; Higuchi, Emily C; Phadnis, Naina

    2010-01-01

    During meiosis, the formation of viable haploid gametes from diploid precursors requires that each homologous chromosome pair be properly segregated to produce an exact haploid set of chromosomes. Genetic recombination, which provides a physical connection between homologous chromosomes, is essen......During meiosis, the formation of viable haploid gametes from diploid precursors requires that each homologous chromosome pair be properly segregated to produce an exact haploid set of chromosomes. Genetic recombination, which provides a physical connection between homologous chromosomes....... Surprisingly, one mutant derepressed for recombination in the heterochromatic mating-type region during meiosis and several mutants derepressed for centromeric gene expression during mitotic growth are not derepressed for centromeric recombination during meiosis. These results reveal a complex relation between...... types of repression by heterochromatin. Our results also reveal a previously undemonstrated role for RNAi and heterochromatin in the repression of meiotic centromeric recombination and, potentially, in the prevention of birth defects by maintenance of proper chromosome segregation during meiosis....

  6. BIOTECHNOLOGY OF RECOMBINANT HORMONES IN DOPING

    Directory of Open Access Journals (Sweden)

    Biljana Vitošević

    2011-09-01

    Full Text Available Recombinant DNA technology has allowed rapid progress in creating biosynthetic gene products for the treatment of many diseases. In this way it can produce large amounts of hormone, which is intended for the treatment of many pathological conditions. Recombinant hormones that are commonly used are insulin, growth hormone and erythropoietin. Precisely because of the availability of these recombinant hormones, it started their abuse by athletes. Experiments in animal models confirmed the potential effects of some of these hormones in increasing physical abilities, which attracted the attention of athletes who push the limits of their competitive capability by such manipulation. The risks of the use of recombinant hormones in doping include serious consequences for the health of athletes. Methods of detection of endogenous hormones from recombined based on the use of a monoclonal antibodies, capillary zone electrophoresis and protein biomarkers

  7. 重组人表皮生长因子对植皮创面成活的影响%Effect of recombinant human epithelium growth factor on livability of skin graft

    Institute of Scientific and Technical Information of China (English)

    龙剑虹; 张明华; 谢庭鸿; 杨兴华; 黄晓元; 周捷

    2003-01-01

    AIM: To investigate the effects of recombinant human epithelium growth factor (rhEGF) applied to skin graft. METHODS: 96 cases between February 2000 and December 2001, were treated. During the operation, After scar removed and skin grafted, the rhEGF was injected under the skin graft. 80 cases without injection of rhEGF were made as contrast. Ten days later, the area of survived skin was measured and the livability of skin was calculated. RESULTS: The skin livability of cases with injection of rhEGF was (90.67 ± 10.02)% and the skin livability of contrast cases was(76. 85 ± 8.35)%. There axisted evident differences between them( P < 0. 01) . CONCLUSION: The rhEGF was an effective method for increasing livability of skin graft.

  8. Effect of recombinant human insulin-like growth factor-I on progression of ALS. A placebo-controlled study. The North America ALS/IGF-I Study Group.

    Science.gov (United States)

    Lai, E C; Felice, K J; Festoff, B W; Gawel, M J; Gelinas, D F; Kratz, R; Murphy, M F; Natter, H M; Norris, F H; Rudnicki, S A

    1997-12-01

    The objective of this study was to investigate the safety and efficacy of recombinant human insulinlike growth factor-I (rhIGF-I) in the treatment of sporadic ALS. A double-blind, placebo-controlled, randomized study of 266 patients was conducted at eight centers in North America. Placebo or rhIGF-I (0.05 mg/kg/day or 0.10 mg/kg/day) was administered for 9 months. The primary outcome measure was disease symptom progression, assessed by the rate of change (per patient slope) in the Appel ALS rating scale total score. The Sickness Impact Profile (SIP), a patient-perceived, health-related quality of life assessment, was a secondary outcome variable. Progression of functional impairment in patients receiving high-dose (0.10 mg/kg/day) rhIGF-I was 26% slower than in patients receiving placebo (p = 0.01). The high-dose treatment group was less likely to terminate the study due to protocol-defined markers of disease symptom progression, and members in this group exhibited a slower decline in quality of life, as assessed by the SIP. Patients receiving 0.05 mg/kg/day of rhIGF-I exhibited trends similar to those associated with high-dose treatment, suggesting a dose-dependent response. The incidence of clinically significant adverse experiences was comparable among the three treatment groups. Recombinant human insulin-like growth factor-I slowed the progression of functional impairment and the decline in health-related quality of life in patients with ALS with no medically important adverse effects.

  9. Inhibition of human MCF-7 breast cancer cells and HT-29 colon cancer cells by rice-produced recombinant human insulin-like growth binding protein-3 (rhIGFBP-3.

    Directory of Open Access Journals (Sweden)

    Stanley C K Cheung

    Full Text Available BACKGROUND: Insulin-like growth factor binding protein-3 (IGFBP-3 is a multifunctional molecule which is closely related to cell growth, apoptosis, angiogenesis, metabolism and senescence. It combines with insulin-like growth factor-I (IGF-I to form a complex (IGF-I/IGFBP-3 that can treat growth hormone insensitivity syndrome (GHIS and reduce insulin requirement in patients with diabetes. IGFBP-3 alone has been shown to have anti-proliferation effect on numerous cancer cells. METHODOLOGY/PRINCIPAL FINDINGS: We reported here an expression method to produce functional recombinant human IGFBP-3 (rhIGFBP-3 in transgenic rice grains. Protein sorting sequences, signal peptide and endoplasmic reticulum retention tetrapeptide (KDEL were included in constructs for enhancing rhIGFBP-3 expression. Western blot analysis showed that only the constructs with signal peptide were successfully expressed in transgenic rice grains. Both rhIGFBP-3 proteins, with or without KDEL sorting sequence inhibited the growth of MCF-7 human breast cancer cells (65.76 ± 1.72% vs 45.00 ± 0.86%, p < 0.05; 50.84 ± 1.97% vs 45.00 ± 0.86%, p < 0.01 respectively and HT-29 colon cancer cells (65.14 ± 3.84% vs 18.01 ± 13.81%, p < 0.05 and 54.7 ± 9.44% vs 18.01 ± 13.81%, p < 0.05 respectively when compared with wild type rice. CONCLUSION/SIGNIFICANCE: These findings demonstrated the feasibility of producing biological active rhIGFBP-3 in rice using a transgenic approach, which will definitely encourage more research on the therapeutic use of hIGFBP-3 in future.

  10. The dwarf phenotype in GH240B mice, haploinsufficient for the autism candidate gene Neurobeachin, is caused by ectopic expression of recombinant human growth hormone.

    Science.gov (United States)

    Nuytens, Kim; Tuand, Krizia; Fu, Quili; Stijnen, Pieter; Pruniau, Vincent; Meulemans, Sandra; Vankelecom, Hugo; Creemers, John W M

    2014-01-01

    Two knockout mouse models for the autism candidate gene Neurobeachin (Nbea) have been generated independently. Although both models have similar phenotypes, one striking difference is the dwarf phenotype observed in the heterozygous configuration of the GH240B model that is generated by the serendipitous insertion of a promoterless human growth hormone (hGH) genomic fragment in the Nbea gene. In order to elucidate this discrepancy, the dwarfism present in this Nbea mouse model was investigated in detail. The growth deficiency in Nbea+/- mice coincided with an increased percentage of fat mass and a decrease in bone mineral density. Low but detectable levels of hGH were detected in the pituitary and hypothalamus of Nbea+/- mice but not in liver, hippocampus nor in serum. As a consequence, several members of the mouse growth hormone (mGH) signaling cascade showed altered mRNA levels, including a reduction in growth hormone-releasing hormone mRNA in the hypothalamus. Moreover, somatotrope cells were less numerous in the pituitary of Nbea+/- mice and both contained and secreted significantly less mGH resulting in reduced levels of circulating insulin-like growth factor 1. These findings demonstrate that the random integration of the hGH transgene in this mouse model has not only inactivated Nbea but has also resulted in the tissue-specific expression of hGH causing a negative feedback loop, mGH hyposecretion and dwarfism.

  11. The dwarf phenotype in GH240B mice, haploinsufficient for the autism candidate gene Neurobeachin, is caused by ectopic expression of recombinant human growth hormone.

    Directory of Open Access Journals (Sweden)

    Kim Nuytens

    Full Text Available Two knockout mouse models for the autism candidate gene Neurobeachin (Nbea have been generated independently. Although both models have similar phenotypes, one striking difference is the dwarf phenotype observed in the heterozygous configuration of the GH240B model that is generated by the serendipitous insertion of a promoterless human growth hormone (hGH genomic fragment in the Nbea gene. In order to elucidate this discrepancy, the dwarfism present in this Nbea mouse model was investigated in detail. The growth deficiency in Nbea+/- mice coincided with an increased percentage of fat mass and a decrease in bone mineral density. Low but detectable levels of hGH were detected in the pituitary and hypothalamus of Nbea+/- mice but not in liver, hippocampus nor in serum. As a consequence, several members of the mouse growth hormone (mGH signaling cascade showed altered mRNA levels, including a reduction in growth hormone-releasing hormone mRNA in the hypothalamus. Moreover, somatotrope cells were less numerous in the pituitary of Nbea+/- mice and both contained and secreted significantly less mGH resulting in reduced levels of circulating insulin-like growth factor 1. These findings demonstrate that the random integration of the hGH transgene in this mouse model has not only inactivated Nbea but has also resulted in the tissue-specific expression of hGH causing a negative feedback loop, mGH hyposecretion and dwarfism.

  12. Photoionization and Recombination

    Science.gov (United States)

    Nahar, Sultana N.

    2000-01-01

    Theoretically self-consistent calculations for photoionization and (e + ion) recombination are described. The same eigenfunction expansion for the ion is employed in coupled channel calculations for both processes, thus ensuring consistency between cross sections and rates. The theoretical treatment of (e + ion) recombination subsumes both the non-resonant recombination ("radiative recombination"), and the resonant recombination ("di-electronic recombination") processes in a unified scheme. In addition to the total, unified recombination rates, level-specific recombination rates and photoionization cross sections are obtained for a large number of atomic levels. Both relativistic Breit-Pauli, and non-relativistic LS coupling, calculations are carried out in the close coupling approximation using the R-matrix method. Although the calculations are computationally intensive, they yield nearly all photoionization and recombination parameters needed for astrophysical photoionization models with higher precision than hitherto possible, estimated at about 10-20% from comparison with experimentally available data (including experimentally derived DR rates). Results are electronically available for over 40 atoms and ions. Photoionization and recombination of He-, and Li-like C and Fe are described for X-ray modeling. The unified method yields total and complete (e+ion) recombination rate coefficients, that can not otherwise be obtained theoretically or experimentally.

  13. Inhibitory effects of recombinant plasmid pshuttle-Egr1-shTRAIL transfection in combination with X-irradiation on growth of liver cancer cells SMMC7721

    International Nuclear Information System (INIS)

    Chen Zhiyong; Liu Min; Dong Lihua; Gong Shouliang

    2011-01-01

    Objective: To investigate the effect of recombinant plasmid pshuttle-Egr1-shTRAIL stable transfection in combination with X-ray irradiation on the TRAIL protein expression and the apoptosis in human SMMC7721 hepatoma cells. Methods: The pshuttle-Egr1-shTRAIL packaged with liposome was stably transfected into SMMC7721 cells in vitro. The shTRAIL protein expression were measured with ELISA assay, Annexin V-FITC kit was adopted to measure the apoptosis of pshuttle-Egr1-shTRAIL cells, and the changes in survival rate of SMMC7721 cells measured with cell cloning assay. Results: The TRAIL protein expressions in pshuttle-Egr1-shTRAIL plus different doses of irradiation groups were significantly increased compared with 0 Gy group (P<0.001). The percentage of apoptotic cells was significantly higher than that in 0 Gy group (P<0.05 or P<0.001), and the survival rate of SMMC7721 cells was decreased significantly (P<0.05 or P<0.001). Conclusion: The pshuttle-Egr1-shTRAIL stable transfection in combination with irradiation can significantly induce the apoptosis of SMMC7721 tumor cells and inhibit the cell proliferation. (authors)

  14. RECOMBINANT HUMAN MAST-CELL GROWTH-FACTOR SUPPORTS ERYTHROID COLONY FORMATION IN POLYCYTHEMIA-VERA IN THE PRESENCE AND ABSENCE OF ERYTHROPOIETIN AND SERUM

    NARCIS (Netherlands)

    MULLER, EW; DEWOLF, JTM; HENDRIKS, DW; ESSELINK, MT; HALIE, MR; VELLENGA, E

    The effect of mast cell growth factor (MGF) was studied on erythropoietin (Epo)-dependent and Epo-independent (''spontaneous'') erythroid colony formation in patients with polycythemia vera (PV). MGF stimulated both Epo-dependent and Epo-independent erythroid colony formation from PV peripheral

  15. Low FT4 Concentrations around the Start of Recombinant Human Growth Hormone Treatment: Predictor of Congenital Structural Hypothalamic-Pituitary Abnormalities?

    NARCIS (Netherlands)

    van Iersel, L. (Laura); H.M. Van Santen (Hanneke M.); van Zandwijken, G.R.J. (Gladys R.J.); N. Zwaveling-Soonawala (Nitash); A.C.S. Hokken-Koelega (Anita); A.S.P. van Trotsenburg (Paul)

    2018-01-01

    textabstractBackground: Growth hormone (GH) treatment may unmask central hypothyroidism (CeH). This was first observed in children with GH deficiency (GHD), later also in adults with GHD due to acquired “organic” pituitary disease. We hypothesized that newly diagnosed CeH in children after starting

  16. Low FT4 Concentrations around the Start of Recombinant Human Growth Hormone Treatment: Predictor of Congenital Structural Hypothalamic-Pituitary Abnormalities?

    NARCIS (Netherlands)

    van Iersel, Laura; van Santen, Hanneke M.; Zandwijken, Gladys R. J.; Zwaveling-Soonawala, Nitash; Hokken-Koelega, Anita C. S.; van Trotsenburg, A. S. Paul

    2018-01-01

    Growth hormone (GH) treatment may unmask central hypothyroidism (CeH). This was first observed in children with GH deficiency (GHD), later also in adults with GHD due to acquired "organic" pituitary disease. We hypothesized that newly diagnosed CeH in children after starting GH treatment for

  17. Clinical assessment of bone quality of human extraction sockets after conversion with growth factors.

    Science.gov (United States)

    Ntounis, Athanasios; Geurs, Nico; Vassilopoulos, Philip; Reddy, Michael

    2015-01-01

    The study was conducted to evaluate the effect of mineralized freeze-dried bone allograft (FDBA), alone or in combination with growth factors in extraction sockets, on subjective assessment of bone quality during implant placement. Forty-one patients whose treatment plan involved extraction of anterior or premolar teeth were randomized into four groups: Group 1, collagen plug (control); Group 2, FDBA/β-tricalcium phosphate (β-TCP)/collagen plug; Group 3, FDBA/β-TCP/platelet-rich plasma (PRP)/collagen plug; Group 4, FDBA/β-TCP/recombinant human platelet-derived growth factor BB (rhPDGF-BB)/collagen plug. After 8 weeks of healing, implants were placed. The clinicians assessed bone quality according to the Misch classification. A benchtop calibration exercise test was conducted to evaluate agreement and accuracy of operators in recognizing different bone qualities. Differences were analyzed using one-way analysis of variance (ANOVA) or chi-square tests for continuous and categorical data. Pairwise comparisons were tested using least squares means (LS means). Spearman correlation coefficients were used to evaluate the relationship of bone growth with potential confounders. P .05). Inclusion of bone grafting is associated with a shift from D4 quality to D3 quality bone. Inclusion of PRP in bone grafting eliminates the incidence of D4 bone, establishing D3 and D2 quality bone as prevalent (56% vs. 42%, respectively). Inclusion of rhPDGF-BB and β-TCP in combination with the bone grafting has the same effect, although D2 quality is less prevalent. When compared to sockets grafted with FDBA/β-TCP/collagen plug alone, the sockets with growth factors demonstrated fewer residual bone graft particles. (1) Inclusion of bone grafting enhanced bone quality as assessed during implant placement. (2) Overall inclusion of PRP and rhPDGF-BB enhanced subjective bone quality, eliminating incidence of D4 quality in human extraction sockets. (3) The use of PRP or rhPDGF-BB may

  18. Growth factors and new periodontology

    Directory of Open Access Journals (Sweden)

    Paknejad M

    1999-06-01

    Full Text Available Growth factors are biological mediators that have a key roll in proliferation, chemotaxy and"ndifferentiation by acting on specific receptors on the surface of cells and regulating events in wound"nhealing.They can be considered hormones that are not released in to the blood stream but have one a"nlocal action. Some of these factors can regulate premature change in GO to Gl phase in cell devesion"ncycle and even may stimulate synthesis of DNA in suitable cells, Growth substances, primarily secreted"nby fibroblasts, endothelia! cells, macrophages and platelet, include platelet derived growth factor"n(PDGF, insulin like growth factor (IGF transforming growth factor (TGFa and (3 and bone"nmorphogenetic proteins BMPs that approximately are the most important of them. (BMPs could be"nused to control events during periodontal, craniofacial and implant wound healing through favoring bone"nformation"nAccording toLynch, combination of PGDF and IGF1 would be effective in promoting growth of all the"ncomponents of the periodontium."nThe aim of this study was to characterize growth factor and review the literature to determine the"nmechanism of their function, classification and application in implant and periodontal treatment.

  19. Effects of quantum well growth temperature on the recombination efficiency of InGaN/GaN multiple quantum wells that emit in the green and blue spectral regions

    Energy Technology Data Exchange (ETDEWEB)

    Hammersley, S.; Dawson, P. [School of Physics and Astronomy, Photon Science Institute, University of Manchester, Manchester M13 9PL (United Kingdom); Kappers, M. J.; Massabuau, F. C.-P.; Sahonta, S.-L.; Oliver, R. A.; Humphreys, C. J. [Department of Materials Science and Metallurgy, University of Cambridge, 27 Charles Babbage Road, Cambridge CB3 0FS (United Kingdom)

    2015-09-28

    InGaN-based light emitting diodes and multiple quantum wells designed to emit in the green spectral region exhibit, in general, lower internal quantum efficiencies than their blue-emitting counter parts, a phenomenon referred to as the “green gap.” One of the main differences between green-emitting and blue-emitting samples is that the quantum well growth temperature is lower for structures designed to emit at longer wavelengths, in order to reduce the effects of In desorption. In this paper, we report on the impact of the quantum well growth temperature on the optical properties of InGaN/GaN multiple quantum wells designed to emit at 460 nm and 530 nm. It was found that for both sets of samples increasing the temperature at which the InGaN quantum well was grown, while maintaining the same indium composition, led to an increase in the internal quantum efficiency measured at 300 K. These increases in internal quantum efficiency are shown to be due reductions in the non-radiative recombination rate which we attribute to reductions in point defect incorporation.

  20. Effects of quantum well growth temperature on the recombination efficiency of InGaN/GaN multiple quantum wells that emit in the green and blue spectral regions

    International Nuclear Information System (INIS)

    Hammersley, S.; Dawson, P.; Kappers, M. J.; Massabuau, F. C.-P.; Sahonta, S.-L.; Oliver, R. A.; Humphreys, C. J.

    2015-01-01

    InGaN-based light emitting diodes and multiple quantum wells designed to emit in the green spectral region exhibit, in general, lower internal quantum efficiencies than their blue-emitting counter parts, a phenomenon referred to as the “green gap.” One of the main differences between green-emitting and blue-emitting samples is that the quantum well growth temperature is lower for structures designed to emit at longer wavelengths, in order to reduce the effects of In desorption. In this paper, we report on the impact of the quantum well growth temperature on the optical properties of InGaN/GaN multiple quantum wells designed to emit at 460 nm and 530 nm. It was found that for both sets of samples increasing the temperature at which the InGaN quantum well was grown, while maintaining the same indium composition, led to an increase in the internal quantum efficiency measured at 300 K. These increases in internal quantum efficiency are shown to be due reductions in the non-radiative recombination rate which we attribute to reductions in point defect incorporation

  1. Recombination of cluster ions

    Science.gov (United States)

    Johnsen, Rainer

    1993-01-01

    Some of our recent work on molecular band emissions from recombination of molecular dimer ions (N4(+) and CO(+) CO) is discussed. Much of the experimental work was done by Y. S. Cao; the results on N4(+) recombination have been published. A brief progress report is given on our ongoing measurements of neutral products of recombination using the flowing-afterglow Langmuir-probe technique in conjunction with laser-induced fluorescence.

  2. Integrated Bottom-Up and Top-Down Liquid Chromatography-Mass Spectrometry for Characterization of Recombinant Human Growth Hormone Degradation Products.

    Science.gov (United States)

    Wang, Yu Annie; Wu, Di; Auclair, Jared R; Salisbury, Joseph P; Sarin, Richa; Tang, Yang; Mozdzierz, Nicholas J; Shah, Kartik; Zhang, Anna Fan; Wu, Shiaw-Lin; Agar, Jeffery N; Love, J Christopher; Love, Kerry R; Hancock, William S

    2017-12-05

    With the advent of biosimilars to the U.S. market, it is important to have better analytical tools to ensure product quality from batch to batch. In addition, the recent popularity of using a continuous process for production of biopharmaceuticals, the traditional bottom-up method, alone for product characterization and quality analysis is no longer sufficient. Bottom-up method requires large amounts of material for analysis and is labor-intensive and time-consuming. Additionally, in this analysis, digestion of the protein with enzymes such as trypsin could induce artifacts and modifications which would increase the complexity of the analysis. On the other hand, a top-down method requires a minimum amount of sample and allows for analysis of the intact protein mass and sequence generated from fragmentation within the instrument. However, fragmentation usually occurs at the N-terminal and C-terminal ends of the protein with less internal fragmentation. Herein, we combine the use of the complementary techniques, a top-down and bottom-up method, for the characterization of human growth hormone degradation products. Notably, our approach required small amounts of sample, which is a requirement due to the sample constraints of small scale manufacturing. Using this approach, we were able to characterize various protein variants, including post-translational modifications such as oxidation and deamidation, residual leader sequence, and proteolytic cleavage. Thus, we were able to highlight the complementarity of top-down and bottom-up approaches, which achieved the characterization of a wide range of product variants in samples of human growth hormone secreted from Pichia pastoris.

  3. Health-related quality of life of young adults with Turner syndrome following a long-term randomized controlled trial of recombinant human growth hormone

    Directory of Open Access Journals (Sweden)

    Van Vliet Guy

    2011-05-01

    Full Text Available Abstract Background There are limited long-term randomized controlled trials of growth hormone (GH supplementation to adult height and few published reports of the health-related quality of life (HRQOL following treatment. The present follow-up study of young adults from a long-term controlled trial of GH treatment in patients with Turner syndrome (TS yielded data to examine whether GH supplementation resulted in a higher HRQOL (either due to taller stature or from the knowledge that active treatment and not placebo had been received or alternatively a lower HRQOL (due to medicalization from years of injections. Methods The original trial randomized 154 Canadian girls with TS aged 7-13 years from 13 centres to receive either long-term GH injections at the pharmacologic dose of 0.3 mg/kg/week or to receive no injections; estrogen prescription for induction of puberty was standardized. Patients were eligible for the follow-up study if they were at least 16 years old at the time of follow-up. The instrument used to study HRQOL was the SF-36, summarized into physical and mental component scales (PCS and MCS; higher scores indicate better HRQOL. Results Thirty-four of the 48 eligible participants (71% consented to participate; data were missing for one patient. Both groups (GH and no treatment had normal HRQOL at this post-treatment assessment. The GH group had a (mean ± SD PCS score of 56 ± 5; the untreated group 58 ± 4; mean score for 16-24 year old females in the general population 53.5 ± 6.9. The GH group had a mean MCS score of 52 ± 6; the untreated group 49 ± 13; mean score for 16-24 year old females in the general population 49.6 ± 9.8. Secondary analyses showed no relationship between HRQOL and height. Conclusions We found no benefit or adverse effect on HRQOL either from receiving or not receiving growth hormone injections in a long-term randomized controlled trial, confirming larger observational studies. We suggest that it remains

  4. Periodontal wound healing/regeneration following implantation of recombinant human growth/differentiation factor-5 (rhGDF-5) in an absorbable collagen sponge carrier into one-wall intrabony defects in dogs: a dose-range study.

    Science.gov (United States)

    Kim, Tae-Gyun; Wikesjö, Ulf M E; Cho, Kyoo-Sung; Chai, Jung-Kiu; Pippig, Susanne D; Siedler, Michael; Kim, Chong-Kwan

    2009-07-01

    Recombinant human growth/differentiation factor-5 (rhGDF-5) is being evaluated as a candidate therapy in support of periodontal regeneration. The objective of this study was to evaluate cementum and alveolar bone formation, and aberrant healing events following surgical implantation of rhGDF-5 in an absorbable collagen sponge (ACS) carrier using an established periodontal defect model. Bilateral 4 x 5 mm (width x depth), one-wall, critical-size, intrabony periodontal defects were surgically created at the mandibular second and fourth pre-molar teeth in 15 Beagle dogs. Five animals received 1 microg/defect and five animals 20 microg/defect rhGDF-5 in unilateral defect sites. Contralateral sites received treatments reported elsewhere. Five animals received rhGDF-5/ACS with 0 (buffer control) and 100 microg/defect rhGDF-5 in contralateral defect sites. The animals were euthanized at 8 weeks post-surgery for histologic and histometric evaluation. Surgical implantation of rhGDF-5 stimulated significant periodontal regeneration. Cementum formation was significantly enhanced in sites implanted with rhGDF-5 (1 and 100 microg) compared with control (phealing/regeneration in intrabony periodontal defects without complications.

  5. Scale-up of Escherichia coli growth and recombinant protein expression conditions from microwell to laboratory and pilot scale based on matched k(L)a.

    Science.gov (United States)

    Islam, R S; Tisi, D; Levy, M S; Lye, G J

    2008-04-01

    Fermentation optimization experiments are ideally performed at small scale to reduce time, cost and resource requirements. Currently microwell plates (MWPs) are under investigation for this purpose as the format is ideally suited to automated high-throughput experimentation. In order to translate an optimized small-scale fermentation process to laboratory and pilot scale stirred-tank reactors (STRs) it is necessary to characterize key engineering parameters at both scales given the differences in geometry and the mechanisms of aeration and agitation. In this study oxygen mass transfer coefficients are determined in three MWP formats and in 7.5 L and 75 L STRs. k(L)a values were determined in cell-free media using the dynamic gassing-out technique over a range of agitation conditions. Previously optimized culture conditions at the MWP scale were then scaled up to the larger STR scales on the basis of matched k(L)a values. The accurate reproduction of MWP (3 mL) E. coli BL21 (DE3) culture kinetics at the two larger scales was shown in terms of cell growth, protein expression, and substrate utilization for k(L)a values that provided effective mixing and gas-liquid distribution at each scale. This work suggests that k(L)a provides a useful initial scale-up criterion for MWP culture conditions which enabled a 15,000-fold scale translation in this particular case. This work complements our earlier studies on the application of DoE techniques to MWP fermentation optimization and in so doing provides a generic framework for the generation of large quantities of soluble protein in a rapid and cost-effective manner.

  6. Growth Factor Mediated Signaling in Pancreatic Pathogenesis

    Energy Technology Data Exchange (ETDEWEB)

    Nandy, Debashis; Mukhopadhyay, Debabrata, E-mail: mukhopadhyay.debabrata@mayo.edu [Department of Biochemistry and Molecular Biology, College of Medicine, Mayo Clinic, 200 First Street SW, Guggenheim 1321C, Rochester, MN 55905 (United States)

    2011-02-24

    Functionally, the pancreas consists of two types of tissues: exocrine and endocrine. Exocrine pancreatic disorders mainly involve acute and chronic pancreatitis. Acute pancreatitis typically is benign, while chronic pancreatitis is considered a risk factor for developing pancreatic cancer. Pancreatic carcinoma is the fourth leading cause of cancer related deaths worldwide. Most pancreatic cancers develop in the exocrine tissues. Endocrine pancreatic tumors are more uncommon, and typically are less aggressive than exocrine tumors. However, the endocrine pancreatic disorder, diabetes, is a dominant cause of morbidity and mortality. Importantly, different growth factors and their receptors play critical roles in pancreatic pathogenesis. Hence, an improved understanding of how various growth factors affect pancreatitis and pancreatic carcinoma is necessary to determine appropriate treatment. This chapter describes the role of different growth factors such as vascular endothelial growth factor (VEGF), insulin-like growth factor (IGF), platelet derived growth factor (PDGF), fibroblast growth factor (FGF), epidermal growth factor (EGF), and transforming growth factor (TGF) in various pancreatic pathophysiologies. Finally, the crosstalk between different growth factor axes and their respective signaling mechanisms, which are involved in pancreatitis and pancreatic carcinoma, are also discussed.

  7. Growth Factor Mediated Signaling in Pancreatic Pathogenesis

    International Nuclear Information System (INIS)

    Nandy, Debashis; Mukhopadhyay, Debabrata

    2011-01-01

    Functionally, the pancreas consists of two types of tissues: exocrine and endocrine. Exocrine pancreatic disorders mainly involve acute and chronic pancreatitis. Acute pancreatitis typically is benign, while chronic pancreatitis is considered a risk factor for developing pancreatic cancer. Pancreatic carcinoma is the fourth leading cause of cancer related deaths worldwide. Most pancreatic cancers develop in the exocrine tissues. Endocrine pancreatic tumors are more uncommon, and typically are less aggressive than exocrine tumors. However, the endocrine pancreatic disorder, diabetes, is a dominant cause of morbidity and mortality. Importantly, different growth factors and their receptors play critical roles in pancreatic pathogenesis. Hence, an improved understanding of how various growth factors affect pancreatitis and pancreatic carcinoma is necessary to determine appropriate treatment. This chapter describes the role of different growth factors such as vascular endothelial growth factor (VEGF), insulin-like growth factor (IGF), platelet derived growth factor (PDGF), fibroblast growth factor (FGF), epidermal growth factor (EGF), and transforming growth factor (TGF) in various pancreatic pathophysiologies. Finally, the crosstalk between different growth factor axes and their respective signaling mechanisms, which are involved in pancreatitis and pancreatic carcinoma, are also discussed

  8. Production, purification and characterization of two recombinant ...

    African Journals Online (AJOL)

    STORAGESEVER

    2008-06-17

    Jun 17, 2008 ... Two recombinant DNA-derived variants of ovine growth hormone were produced, purified, characterized and compared with the authentic pituitary derived GH. The variants oGH3 and oGH5 were isolated by differential centrifugation method and were purified after refolding by ion-exchange.

  9. Immunoassays for the measurement of IGF-II, IGFBP-2 and -3, and ICTP as indirect biomarkers of recombinant human growth hormone misuse in sport. Values in selected population of athletes.

    Science.gov (United States)

    Abellan, Rosario; Ventura, Rosa; Palmi, Ilaria; di Carlo, Simonetta; Bacosi, Antonella; Bellver, Montse; Olive, Ramon; Pascual, Jose Antonio; Pacifici, Roberta; Segura, Jordi; Zuccaro, Piergiorgio; Pichini, Simona

    2008-11-04

    Insulin-like growth factor-II (IGF-II), insulin-like growth factor binding proteins (IGFBPs) -2 and -3 and C-terminal telopeptide of type I collagen (ICTP) have been proposed, among others, as indirect biomarkers of the recombinant human growth hormone misuse in sport. An extended intra- and inter-laboratory validation of commercially available immunoassays for biomarkers detection was performed. ELISA assays for total IGF-II, IGFBP-2 and IGFBP-3 (IGF-II/ELISA1: DSLabs, IGFBP-2/ELISA2: Biosource, and IGFBP-3/ELISA3: BioSource) and an EIA assay for ICTP (ICTP/EIA: Orion Diagnostica) were evaluated. The inter- and intra-laboratory precision values were acceptable for all evaluated assays (maximum imprecision of 30% and 66% were found only for the lowest quality control samples of IGF-II and IGFBP-3). Correct accuracy was obtained for all inter-laboratory immunoassays and for IGFBP-2 intra-laboratory immunoassay. The range of concentrations found in serum samples under investigation was always covered by the calibration curves of the studied immunoassays. However, 11% and 15% of the samples felt below the estimated LOQ for IGF-II and ICTP, respectively, in the zone where lower precision was obtained. Although the majority of evaluated assays showed an overall reliability not always suitable for antidoping control analysis, relatively high concordances between laboratory results were obtained for all assays. Evaluated immunoassays were used to measure serum concentrations of IGF-II, IGFBP-2 and -3 and ICTP in elite athletes of various sport disciplines at different moments of the training season; in recreational athletes at baseline conditions and finally in sedentary individuals. Serum IGF-II was statistically higher both in recreational and elite athletes compared to sedentary individuals. Elite athletes showed lower IGFBP-2 and higher IGFBP-3 concentration with respect to recreational athletes and sedentary people. Among elite athletes, serum IGFBP-3 (synchronized

  10. Activated recombinant adenovirus proteinases

    Science.gov (United States)

    Anderson, Carl W.; Mangel, Walter F.

    1999-08-10

    This application describes methods and expression constructs for producing activatable recombinant adenovirus proteinases. Purified activatable recombinant adenovirus proteinases and methods of purification are described. Activated adenovirus proteinases and methods for obtaining activated adenovirus proteinases are further included. Isolated peptide cofactors of adenovirus proteinase activity, methods of purifying and identifying said peptide cofactors are also described. Antibodies immunoreactive with adenovirus proteinases, immunospecific antibodies, and methods for preparing them are also described. Other related methods and materials are also described.

  11. Hadron correlations from recombination

    Energy Technology Data Exchange (ETDEWEB)

    Fries, Rainer J [School of Physics and Astronomy, University of Minnesota, Minneapolis, MN 55455 (United States)

    2005-01-01

    Quark recombination is a successful model to describe the hadronization of a deconfined quark gluon plasma. Jet-like dihadron correlations measured at RHIC provide a challenge for this picture. We discuss how correlations between hadrons can arise from correlations between partons before hadronization. An enhancement of correlations through the recombination process, similar to the enhancement of elliptic flow is found. Hot spots from completely or partially quenched jets are a likely source of such parton correlations.

  12. Extracellular matrix organization modulates fibroblast growth and growth factor responsiveness.

    Science.gov (United States)

    Nakagawa, S; Pawelek, P; Grinnell, F

    1989-06-01

    To learn more about the relationship between extracellular matrix organization, cell shape, and cell growth control, we studied DNA synthesis by fibroblasts in collagen gels that were either attached to culture dishes or floating in culture medium during gel contraction. After 4 days of contraction, the collagen density (initially 1.5 mg/ml) reached 22 mg/ml in attached gels and 55 mg/ml in floating gels. After contraction, attached collagen gels were well organized; collagen fibrils were aligned in the plane of cell spreading; and fibroblasts had an elongated, bipolar morphology. Floating collagen gels, however, were unorganized; collagen fibrils were arranged randomly; and fibroblasts had a stellate morphology. DNA synthesis by fibroblasts in contracted collagen gels was suppressed if the gels were floating in medium but not if the gels were attached, and inhibition was independent of the extent of gel contraction. Therefore, growth of fibroblasts in contracted collagen gels could be regulated by differences in extracellular matrix organization and cell shape independently of extracellular matrix density. We also compared the responses of fibroblasts in contracted collagen gels and monolayer culture to peptide growth factors including fibroblast growth factor, platelet-derived growth factor, transforming growth factor-beta, and interleukin 1. Cells in floating collagen gels were generally unresponsive to any of the growth factors. Cells in attached collagen gels and monolayer culture were affected similarly by fibroblast growth factor but not by the others. Our results indicate that extracellular matrix organization influenced not only cell growth, but also fibroblast responsiveness to peptide growth factors.

  13. Regulation of Meiotic Recombination

    Energy Technology Data Exchange (ETDEWEB)

    Gregory p. Copenhaver

    2011-11-09

    Meiotic recombination results in the heritable rearrangement of DNA, primarily through reciprocal exchange between homologous chromosome or gene conversion. In plants these events are critical for ensuring proper chromosome segregation, facilitating DNA repair and providing a basis for genetic diversity. Understanding this fundamental biological mechanism will directly facilitate trait mapping, conventional plant breeding, and development of genetic engineering techniques that will help support the responsible production and conversion of renewable resources for fuels, chemicals, and the conservation of energy (1-3). Substantial progress has been made in understanding the basal recombination machinery, much of which is conserved in organisms as diverse as yeast, plants and mammals (4, 5). Significantly less is known about the factors that regulate how often and where that basal machinery acts on higher eukaryotic chromosomes. One important mechanism for regulating the frequency and distribution of meiotic recombination is crossover interference - or the ability of one recombination event to influence nearby events. The MUS81 gene is thought to play an important role in regulating the influence of interference on crossing over. The immediate goals of this project are to use reverse genetics to identify mutants in two putative MUS81 homologs in the model plant Arabidopsis thaliana, characterize those mutants and initiate a novel forward genetic screen for additional regulators of meiotic recombination. The long-term goal of the project is to understand how meiotic recombination is regulated in higher eukaryotes with an emphasis on the molecular basis of crossover interference. The ability to monitor recombination in all four meiotic products (tetrad analysis) has been a powerful tool in the arsenal of yeast geneticists. Previously, the qrt mutant of Arabidopsis, which causes the four pollen products of male meiosis to remain attached, was developed as a facile system

  14. Recombinational repair: workshop summary

    International Nuclear Information System (INIS)

    Howard-Flanders, P.

    1983-01-01

    Recombinational repair may or may not be synonymous with postreplication repair. Considerable progress has been made in the study of the relevant enzymes, particularly those from bacteria. In this workshop we focus on the recombination enzyme RecA protein. What structural changes take place in the protein and in DNA during repair. How does homologous pairing take place. How is ATP hydrolysis coupled to the stand exchange reaction and the formation of heteroduplx DNA. Turning to another enzyme needed for certain kinds of bacterial recombination, we will ask whether the purified recB protein and recC protein complement each other and are sufficient for exonuclease V activity. In higher cells, we would like to know whether sister exchanges, which occur in bacteria after uv irradiation, are also seen in animal cells

  15. Parton recombination model

    International Nuclear Information System (INIS)

    Hwa, R.C.

    1978-08-01

    Low P/sub T/ meson production in hadronic collisions is described in the framework of the parton model. The recombination of quark and antiquark is suggested as the dominant mechanism in the large x region. Phenomenological evidences for the mechanism are given. The application to meson initiated reactions yields the quark distribution in mesons. 21 references

  16. Sex recombination, and reproductive fitness: an experimental study using Paramecium

    Energy Technology Data Exchange (ETDEWEB)

    Nyberg, D.

    1982-08-01

    The effect of sex and recombination on reproductive fitness are measured using five wild stocks of Paramecium primaurelia. Among the wild stocks there were highly significant differences in growth rates. No hybrid had as low a fitness as the least fit parental stock. Recombination produced genotypes of higher fitness than those of either parent only in the cross between the two stocks of lowest fitness. The increase in variance of fitness as a result of recombination was almost exclusively attributable to the generation lines with low fitness. The fitness consequences of sexuality and mate choice were stock specific; some individuals leaving the most descendants by inbreeding, others by outcrossing. For most crosses the short-term advantage of sex, if any, accrue from the fusion of different gametes (hybrid vigor) and not from recombination. Since the homozygous genotype with the highest fitnes left the most progeny by inbreeding (no recombination), the persistence of conjugation in P. primaurelia is paradoxical. (JMT)

  17. Cultivating Insect Cells To Produce Recombinant Proteins

    Science.gov (United States)

    Spaulding, Glenn; Goodwin, Thomas; Prewett, Tacey; Andrews, Angela; Francis, Karen; O'Connor, Kim

    1996-01-01

    Method of producing recombinant proteins involves growth of insect cells in nutrient solution in cylindrical bioreactor rotating about cylindrical axis, oriented horizontally and infecting cells with viruses into which genes of selected type cloned. Genes in question those encoding production of desired proteins. Horizontal rotating bioreactor preferred for use in method, denoted by acronym "HARV", described in "High-Aspect-Ratio Rotating Cell-Culture Vessel" (MSC-21662).

  18. Limiting factors in Escherichia colifed-batch production of recombinant proteins

    DEFF Research Database (Denmark)

    Sanden, A.M.; Prytz, I.; Tubelekas, I.

    2003-01-01

    recombinant protein production, fed-batch, specific growth rate, feed profile, induction, mRNA, transcription, translation, acetic acid formation......recombinant protein production, fed-batch, specific growth rate, feed profile, induction, mRNA, transcription, translation, acetic acid formation...

  19. Construction and characterization of a recombinant invertebrate iridovirus.

    Science.gov (United States)

    Ozgen, Arzu; Muratoglu, Hacer; Demirbag, Zihni; Vlak, Just M; van Oers, Monique M; Nalcacioglu, Remziye

    2014-08-30

    Chilo iridescent virus (CIV), officially named Insect iridescent virus 6 (IIV6), is the type species of the genus Iridovirus (family Iridoviridae). In this paper we constructed a recombinant CIV, encoding the green fluorescent protein (GFP). This recombinant can be used to investigate viral replication dynamics. We showed that homologous recombination is a valid method to make CIV gene knockouts and to insert foreign genes. The CIV 157L gene, putatively encoding a non-functional inhibitor of apoptosis (IAP), was chosen as target for foreign gene insertion. The gfp open reading frame preceded by the viral mcp promoter was inserted into the 157L locus by homologous recombination in Anthonomus grandis BRL-AG-3A cells. Recombinant virus (rCIV-Δ157L-gfp) was purified by successive rounds of plaque purification. All plaques produced by the purified recombinant virus emitted green fluorescence due to the presence of GFP. One-step growth curves for recombinant and wild-type CIV were similar and the recombinant was fully infectious in vivo. Hence, CIV157L can be inactivated without altering the replication kinetics of the virus. Consequently, the CIV 157L locus can be used as a site for insertion of foreign DNA, e.g. to modify viral properties for insect biocontrol. Copyright © 2014 Elsevier B.V. All rights reserved.

  20. Site directed recombination

    Science.gov (United States)

    Jurka, Jerzy W.

    1997-01-01

    Enhanced homologous recombination is obtained by employing a consensus sequence which has been found to be associated with integration of repeat sequences, such as Alu and ID. The consensus sequence or sequence having a single transition mutation determines one site of a double break which allows for high efficiency of integration at the site. By introducing single or double stranded DNA having the consensus sequence flanking region joined to a sequence of interest, one can reproducibly direct integration of the sequence of interest at one or a limited number of sites. In this way, specific sites can be identified and homologous recombination achieved at the site by employing a second flanking sequence associated with a sequence proximal to the 3'-nick.

  1. Algae-based oral recombinant vaccines

    Science.gov (United States)

    Specht, Elizabeth A.; Mayfield, Stephen P.

    2014-01-01

    Recombinant subunit vaccines are some of the safest and most effective vaccines available, but their high cost and the requirement of advanced medical infrastructure for administration make them impractical for many developing world diseases. Plant-based vaccines have shifted that paradigm by paving the way for recombinant vaccine production at agricultural scale using an edible host. However, enthusiasm for “molecular pharming” in food crops has waned in the last decade due to difficulty in developing transgenic crop plants and concerns of contaminating the food supply. Microalgae could be poised to become the next candidate in recombinant subunit vaccine production, as they present several advantages over terrestrial crop plant-based platforms including scalable and contained growth, rapid transformation, easily obtained stable cell lines, and consistent transgene expression levels. Algae have been shown to accumulate and properly fold several vaccine antigens, and efforts are underway to create recombinant algal fusion proteins that can enhance antigenicity for effective orally delivered vaccines. These approaches have the potential to revolutionize the way subunit vaccines are made and delivered – from costly parenteral administration of purified protein, to an inexpensive oral algae tablet with effective mucosal and systemic immune reactivity. PMID:24596570

  2. Algae-based oral recombinant vaccines

    Directory of Open Access Journals (Sweden)

    Elizabeth A Specht

    2014-02-01

    Full Text Available Recombinant subunit vaccines are some of the safest and most effective vaccines available, but their high cost and the requirement of advanced medical infrastructure for administration make them impractical for many developing world diseases. Plant-based vaccines have shifted that paradigm by paving the way for recombinant vaccine production at agricultural scale using an edible host. However, enthusiasm for molecular pharming in food crops has waned in the last decade due to difficulty in developing transgenic crop plants and concerns of contaminating the food supply. Microalgae are poised to become the next candidate in recombinant subunit vaccine production, and they present several advantages over terrestrial crop plant-based platforms including scalable and contained growth, rapid transformation, easily obtained stable cell lines, and consistent transgene expression levels. Algae have been shown to accumulate and properly fold several vaccine antigens, and efforts are underway to create recombinant algal fusion proteins that can enhance antigenicity for effective orally-delivered vaccines. These approaches have the potential to revolutionize the way subunit vaccines are made and delivered – from costly parenteral administration of purified protein, to an inexpensive oral algae tablet with effective mucosal and system immune reactivity.

  3. Nonradiative recombination in semiconductors

    CERN Document Server

    Abakumov, VN; Yassievich, IN

    1991-01-01

    In recent years, great progress has been made in the understandingof recombination processes controlling the number of excessfree carriers in semiconductors under nonequilibrium conditions. As a result, it is now possible to give a comprehensivetheoretical description of these processes. The authors haveselected a number of experimental results which elucidate theunderlying physical problems and enable a test of theoreticalmodels. The following topics are dealt with: phenomenological theory ofrecombination, theoretical models of shallow and deep localizedstates, cascade model of carrier captu

  4. Recombination epoch revisited

    International Nuclear Information System (INIS)

    Krolik, J.H.

    1989-01-01

    Previous studies of cosmological recombination have shown that this process produces as a by-product a highly superthermal population of Ly-alpha photons which retard completion of recombination. Cosmological redshifting was thought to determine the frequency distribution of the photons, while two-photon decay of hydrogen's 2s state was thought to control their numbers. It is shown here that frequency diffusion due to photon scattering dominate the cosmological redshift in the frequency range near line center which fixes the ratio of ground state to excited state population, while incoherent scattering into the far-red damping wing effectively destroys Ly-alpha photons as a rate which is competitive with two-photon decay. The former effect tends to hold back recombination, while the latter tends to accelerate it; the net results depends on cosmological parameters, particularly the combination Omega(b) h/sq rt (2q0), where Omega(b) is the fraction of the critical density provided by baryons. 18 references

  5. Dielectronic recombination theory

    International Nuclear Information System (INIS)

    LaGattuta, K.J.

    1991-01-01

    A theory now in wide use for the calculation of dielectronic recombination cross sections (σ DR ) and rate coefficients (α DR ) was one introduced originally by Feshbach for nuclear physics applications, and then later adapted for atomic scattering problems by Hahn. In the following, we briefly review this theory in a very general form, which allows one to account for the effects of overlapping and interacting resonances, as well as continuum-continuum coupling. An extension of our notation will then also allow for the inclusion of the effects of direct radiative recombination, along with a treatment of the interference between radiative and dielectronic recombination. Other approaches to the calculation of σ DR have been described by Fano and by Seaton. We will not consider those theories here. Calculations of α DR have progressed considerably over the last 25 years, since the early work of Burgess. Advances in the reliability of theoretical predictions have also been promoted recently b a variety of direct laboratory measurements of σ DR . While the measurements of σ DR for δn ≠ 0 excitations have tended to agree very well with calculations, the case of δn = 0 has been much problematic. However, by invoking a mechanism originally proposed by Jacobs, which takes into account the effect of stray electric fields on high Rydberg states (HRS) participating in the DR process, new calculations have improved the agreement between theory and experiment for these cases. Nevertheless, certain discrepancies still remain

  6. Novel Drosophila receptor that binds multiple growth factors

    International Nuclear Information System (INIS)

    Rosner, M.R.; Thompson, K.L.; Garcia, V.; Decker, S.J.

    1986-01-01

    The authors have recently reported the identification of a novel growth factor receptor from Drosophila cell cultures that has dual binding specificity for both insulin and epidermal growth factor (EGF). This 100 kDa protein is also antigenically related to the cytoplasmic region of the mammalian EGF receptor-tyrosine kinase. They now report that this protein binds to mammalian nerve growth factor and human transforming growth factor alpha as well as insulin and EGF with apparent dissociation constants ranging from 10 -6 to 10 -8 M. The 100 kDa protein can be affinity-labeled with these 125 I-labeled growth factors after immunoprecipitation with anti-EGF receptor antiserum. These four growth factors appear to share a common binding site, as evidenced by their ability to block affinity labelling by 125 I-insulin. No significant binding to the 100 kDa protein was observed with platelet-derived growth factor, transforming growth factor beta, or glucagon. The 100 kDa Drosophila protein has a unique ligand-binding spectrum with no direct counterpart in mammalian cells and may represent an evolutionary precursor of the mammalian receptors for these growth factors

  7. Continuous 24-hour intravenous infusion of recombinant human growth hormone (GH)-releasing hormone-(1-44)-amide augments pulsatile, entropic, and daily rhythmic GH secretion in postmenopausal women equally in the estrogen-withdrawn and estrogen-supplemented states.

    Science.gov (United States)

    Evans, W S; Anderson, S M; Hull, L T; Azimi, P P; Bowers, C Y; Veldhuis, J D

    2001-02-01

    How estrogen amplifies GH secretion in the human is not known. The present study tests the clinical hypothesis that estradiol modulates the stimulatory actions of a primary GH feedforward signal, GHRH. To this end, we investigated the ability of short-term (7- to 12-day) supplementation with oral estradiol vs. placebo to modulate basal, pulsatile, entropic, and 24-h rhythmic GH secretion driven by a continuous iv infusion of recombinant human GHRH-(1--44)-amide vs. saline in nine healthy postmenopausal women. Volunteers underwent concurrent blood sampling every 10 min for 24 h on four occasions in a prospectively randomized, single blind, within-subject cross-over design (placebo/saline, placebo/GHRH, estradiol/saline, estradiol/GHRH). Intensively sampled serum GH concentrations were quantitated by ultrasensitive chemiluminescence assay. Basal, pulsatile, entropic (feedback-sensitive), and 24-h rhythmic modes of GH secretion were appraised by deconvolution analysis, the approximate entropy (ApEn) statistic, and cosine regression, respectively. ANOVA revealed that continuous iv infusion of GHRH in the estrogen-withdrawn (control) milieu 1) amplified individual basal (P = 0.00011) and pulsatile (P < 10(-13)) GH secretion rates by 12- and 11-fold, respectively; 2) augmented GH secretory burst mass and amplitude each by 10-fold (P < 10(-11)), without altering GH secretory burst frequency, duration, or half-life; 3) increased the disorderliness (ApEn) of GH release patterns (P = 0.0000002); 4) elevated the mesor (cosine mean) and amplitude of the 24-h rhythm in serum GH concentrations by nearly 30-fold (both P < 10(-12)); 5) induced a phase advance in the clocktime of the GH zenith (P = 0.021); and 6) evoked a new 24-h rhythm in GH secretory burst mass with a maximum at 0018 h GH (P < 10(-3)), while damping the mesor of the 24-h rhythm in GH interpulse intervals (P < 0.025). Estradiol supplementation alone 1) increased the 24-h mean and integrated serum GH concentration

  8. Ultrastructure and growth factor content of equine platelet-rich fibrin gels.

    Science.gov (United States)

    Textor, Jamie A; Murphy, Kaitlin C; Leach, J Kent; Tablin, Fern

    2014-04-01

    To compare fiber diameter, pore area, compressive stiffness, gelation properties, and selected growth factor content of platelet-rich fibrin gels (PRFGs) and conventional fibrin gels (FGs). PRFGs and conventional FGs prepared from the blood of 10 healthy horses. Autologous fibrinogen was used to form conventional FGs. The PRFGs were formed from autologous platelet-rich plasma of various platelet concentrations (100 × 10³ platelets/μL, 250 × 10³ platelets/μL, 500 × 10³ platelets/μL, and 1,000 × 10³ platelets/μL). All gels contained an identical fibrinogen concentration (20 mg/mL). Fiber diameter and pore area were evaluated with scanning electron microscopy. Maximum gelation rate was assessed with spectrophotometry, and gel stiffness was determined by measuring the compressive modulus. Gel weights were measured serially over 14 days as an index of contraction (volume loss). Platelet-derived growth factor-BB and transforming growth factor-β1 concentrations were quantified with ELISAs. Fiber diameters were significantly larger and mean pore areas were significantly smaller in PRFGs than in conventional FGs. Gel weight decreased significantly over time, differed significantly between PRFGs and conventional FGs, and was significantly correlated with platelet concentration. Platelet-derived growth factor-BB and transforming growth factor-β1 concentrations were highest in gels and releasates derived from 1,000 × 10³ platelets/μL. The inclusion of platelets in FGs altered the architecture and increased the growth factor content of the resulting scaffold. Platelets may represent a useful means of modifying these gels for applications in veterinary and human regenerative medicine.

  9. Recombination clumping factor during cosmic reionization

    International Nuclear Information System (INIS)

    Kaurov, Alexander A.; Gnedin, Nickolay Y.

    2014-01-01

    We discuss the role of recombinations in the intergalactic medium, and the related concept of the clumping factor, during cosmic reionization. The clumping factor is, in general, a local quantity that depends on both the local overdensity and the scale below which the baryon density field can be assumed smooth. That scale, called the filtering scale, depends on over-density and local thermal history. We present a method for building a self-consistent analytical model of inhomogeneous reionization, assuming the linear growth rate of the density fluctuation, which simultaneously accounts for these effects. We show that taking into account the local clumping factor introduces significant corrections to the total recombination rate, compared to the model with a globally uniform clumping factor.

  10. Clinical Application of Growth Factors and Cytokines in Wound Healing

    Science.gov (United States)

    Barrientos, Stephan; Brem, Harold; Stojadinovic, Olivera; Tomic-Canic, Marjana

    2016-01-01

    Wound healing is a complex and dynamic biological process that involves the coordinated efforts of multiple cell types and is executed and regulated by numerous growth factors and cytokines. There has been a drive in the past two decades to study the therapeutic effects of various growth factors in the clinical management of non-healing wounds (e.g. pressure ulcers, chronic venous ulcers, diabetic foot ulcers). For this review, we conducted a nonline search of Medline and Pub Medical and critically analyzed the literature regarding the role of growth factors and cytokines in the management of these wounds. We focused on currently approved therapies, emerging therapies and future research possibilities. In this review we discuss four growth factors and cytokines currently being used on and off label for the healing of wounds. These include: granulocyte-macrophage colony stimulating factor (GM-CSF), platelet derived growth factor (PDGF), vascular endothelial growth factor (VEGF), and basic fibroblast growth factor (bFGF). While the clinical results of using growth factors and cytokines are encouraging, many studies involved a small sample size and are disparate in measured endpoints. Therefore, further research is required to provide definitive evidence of efficacy. PMID:24942811

  11. Recombination properties of dislocations in GaN

    Science.gov (United States)

    Yakimov, Eugene B.; Polyakov, Alexander Y.; Lee, In-Hwan; Pearton, Stephen J.

    2018-04-01

    The recombination activity of threading dislocations in n-GaN with different dislocation densities and different doping levels was studied using electron beam induced current (EBIC). The recombination velocity on a dislocation, also known as the dislocation recombination strength, was calculated. The results suggest that dislocations in n-GaN giving contrast in EBIC are charged and surrounded by a space charge region, as evidenced by the observed dependence of dislocation recombination strength on dopant concentration. For moderate (below ˜108 cm-2) dislocation densities, these defects do not primarily determine the average diffusion length of nonequilibrium charge carriers, although locally, dislocations are efficient recombination sites. In general, it is observed that the effect of the growth method [standard metalorganic chemical vapor deposition (MOCVD), epitaxial lateral overgrowth versions of MOCVD, and hydride vapor phase epitaxy] on the recombination activity of dislocations is not very pronounced, although the average diffusion lengths can widely differ for various samples. The glide of basal plane dislocations at room temperature promoted by low energy electron irradiation does not significantly change the recombination properties of dislocations.

  12. Recombinant Innovation and Endogenous Transitions

    OpenAIRE

    Koen Frenken; Luis R. Izquierdo; Paolo Zeppini

    2012-01-01

    We propose a model of technological transitions based on two different types of innovations. Branching innovations refer to technological improvements along a particular path, while recombinant innovations represent fusions of multiple paths. Recombinant innovations create “short-cuts” which reduce switching costs allowing agents to escape a technological lock-in. As a result, recombinant innovations speed up technological progress allowing transitions that are impossible with only branching ...

  13. High-throughput measurement of recombination rates and genetic interference in Saccharomyces cerevisiae.

    Science.gov (United States)

    Raffoux, Xavier; Bourge, Mickael; Dumas, Fabrice; Martin, Olivier C; Falque, Matthieu

    2018-06-01

    Allelic recombination owing to meiotic crossovers is a major driver of genome evolution, as well as a key player for the selection of high-performing genotypes in economically important species. Therefore, we developed a high-throughput and low-cost method to measure recombination rates and crossover patterning (including interference) in large populations of the budding yeast Saccharomyces cerevisiae. Recombination and interference were analysed by flow cytometry, which allows time-consuming steps such as tetrad microdissection or spore growth to be avoided. Moreover, our method can also be used to compare recombination in wild-type vs. mutant individuals or in different environmental conditions, even if the changes in recombination rates are small. Furthermore, meiotic mutants often present recombination and/or pairing defects affecting spore viability but our method does not involve growth steps and thus avoids filtering out non-viable spores. Copyright © 2018 John Wiley & Sons, Ltd.

  14. Expression of recombinant Antibodies

    Directory of Open Access Journals (Sweden)

    André eFrenzel

    2013-07-01

    Full Text Available Recombinant antibodies are highly specific detection probes in research, diagnostics and have emerged over the last two decades as the fastest growing class of therapeutic proteins. Antibody generation has been dramatically accelerated by in vitro selection systems, particularly phage display. An increasing variety of recombinant production systems have been developed, ranging from Gram-negative and positive bacteria, yeasts and filamentous fungi, insect cell lines, mammalian cells to transgenic plants and animals. Currently, almost all therapeutic antibodies are still produced in mammalian cell lines in order to reduce the risk of immunogenicity due to altered, non-human glycosylation patterns. However, recent developments of glycosylation-engineered yeast, insect cell lines and transgenic plants are promising to obtain antibodies with human-like post-translational modifications. Furthermore, smaller antibody fragments including bispecific antibodies without any glycosylation are successfully produced in bacteria and have advanced to clinical testing. The first therapeutic antibody products from a non-mammalian source can be expected in coming next years. In this review, we focus on current antibody production systems including their usability for different applications.

  15. On the relict recombination lines

    International Nuclear Information System (INIS)

    Bershtejn, I.N.; Bernshtejn, D.N.; Dubrovich, V.K.

    1977-01-01

    Accurate numerical calculation of intensities and profiles of hydrogen recombination lines of cosmological origin is made. Relie radiation distortions stipulated by recombination quantum release at the irrevocable recombination are investigated. Mean number calculation is given for guantums educing for one irrevocably-lost electron. The account is taken of the educed quantums interraction with matter. The main quantum-matter interrraction mechanisms are considered: electronic blow broadening; free-free, free-bound, bound-bound absorptions Recombination dynamics is investigated depending on hydrogen density and total density of all the matter kinds in the Universe

  16. Advances in recombinant antibody manufacturing.

    Science.gov (United States)

    Kunert, Renate; Reinhart, David

    2016-04-01

    Since the first use of Chinese hamster ovary (CHO) cells for recombinant protein expression, production processes have steadily improved through numerous advances. In this review, we have highlighted several key milestones that have contributed to the success of CHO cells from the beginning of their use for monoclonal antibody (mAb) expression until today. The main factors influencing the yield of a production process are the time to accumulate a desired amount of biomass, the process duration, and the specific productivity. By comparing maximum cell densities and specific growth rates of various expression systems, we have emphasized the limiting parameters of different cellular systems and comprehensively described scientific approaches and techniques to improve host cell lines. Besides the quantitative evaluation of current systems, the quality-determining properties of a host cell line, namely post-translational modifications, were analyzed and compared to naturally occurring polyclonal immunoglobulin fractions from human plasma. In summary, numerous different expression systems for mAbs are available and also under scientific investigation. However, CHO cells are the most frequently investigated cell lines and remain the workhorse for mAb production until today.

  17. Dissociative recombination of dications

    International Nuclear Information System (INIS)

    Seiersen, K.; Heber, O.; Jensen, M.J.; Safvan, C.P.; Andersen, L. H.

    2003-01-01

    Dissociative recombination (DR) of doubly-charged positive ions has been studied at the heavy ion storage ring ASTRID. Low-energy electrons were scattered on the dication of the N 2 molecule, and the absolute cross section was measured in the energy range of 10 -4 -50 eV. From the measured cross section, a thermal rate coefficient of 5.8x10 -7 cm 3 s -1 at 300 K was extracted. Furthermore, we present new results on the CO 2+ DR rate, and a summary and comparison of measured DR rate coefficients for both the singly and doubly-charged ions of CO, CO 2 , and N 2 is presented

  18. Cell biology of mitotic recombination

    DEFF Research Database (Denmark)

    Lisby, Michael; Rothstein, Rodney

    2015-01-01

    Homologous recombination provides high-fidelity DNA repair throughout all domains of life. Live cell fluorescence microscopy offers the opportunity to image individual recombination events in real time providing insight into the in vivo biochemistry of the involved proteins and DNA molecules as w...

  19. Hadron Correlations and Parton Recombination

    Energy Technology Data Exchange (ETDEWEB)

    Fries, R.J. [School of Physics and Astronomy, University of Minnesota, Minneapolis, MN 55455 (United States)]. E-mail: rjfries@comp.tamu.edu

    2007-02-15

    Parton recombination has been found to be an extremely useful model to understand hadron production at the Relativistic Heavy Ion Collider. It is particularly important to explore its connections with hard processes. This article reviews some of the aspects of the quark recombination model and places particular emphasis on hadron correlations.

  20. Auger recombination in sodium iodide

    Science.gov (United States)

    McAllister, Andrew; Kioupakis, Emmanouil; Åberg, Daniel; Schleife, André

    2014-03-01

    Scintillators are an important tool used to detect high energy radiation - both in the interest of national security and in medicine. However, scintillator detectors currently suffer from lower energy resolutions than expected from basic counting statistics. This has been attributed to non-proportional light yield compared to incoming radiation, but the specific mechanism for this non-proportionality has not been identified. Auger recombination is a non-radiative process that could be contributing to the non-proportionality of scintillating materials. Auger recombination comes in two types - direct and phonon-assisted. We have used first-principles calculations to study Auger recombination in sodium iodide, a well characterized scintillating material. Our findings indicate that phonon-assisted Auger recombination is stronger in sodium iodide than direct Auger recombination. Computational resources provided by LLNL and NERSC. Funding provided by NA-22.

  1. Similarity of recombinant human perlecan domain 1 by alternative expression systems bioactive heterogenous recombinant human perlecan D1

    DEFF Research Database (Denmark)

    Ellis, April L; Pan, Wensheng; Yang, Guang

    2010-01-01

    BACKGROUND: Heparan sulfate glycosaminoglycans are diverse components of certain proteoglycans and are known to interact with growth factors as a co-receptor necessary to induce signalling and growth factor activity. In this report we characterize heterogeneously glycosylated recombinant human...... perlecan domain 1 (HSPG2 abbreviated as rhPln.D1) synthesized in either HEK 293 cells or HUVECs by transient gene delivery using either adenoviral or expression plasmid technology. RESULTS: By SDS-PAGE analysis following anion exchange chromatography, the recombinant proteoglycans appeared to possess...... glycosaminoglycan chains ranging, in total, from 6 kDa to >90 kDa per recombinant. Immunoblot analysis of enzyme-digested high Mr rhPln.D1 demonstrated that the rhPln.D1 was synthesized as either a chondroitin sulfate or heparan sulfate proteoglycan, in an approximately 2:1 ratio, with negligible hybrids. Secondary...

  2. c-erbA and v-erbA modulate growth and gene expression of a mouse glial precursor cell line.

    Science.gov (United States)

    Iglesias, T; Llanos, S; López-Barahona, M; Pérez-Aranda, A; Rodríguez-Peña, A; Bernal, J; Höhne, A; Seliger, B; Muñoz, A

    1994-07-01

    The c-erbA alpha protooncogene coding for the thyroid hormone (T3) receptor (TR alpha 1) and the viral, mutated v-erbA oncogene were expressed in an immortal mouse glial cell line (B3.1) using retroviral vectors. c-erbA alpha expression led to a decrease in cell proliferation in high and low serum conditions, both in the presence and in the absence of T3. In serum-free medium, c-erbA-expressing cells (B3.1 + TR alpha 1) were completely arrested, whereas cells expressing v-erbA (B3.1 + v-erbA) showed a higher DNA synthesis rate than normal B3.1 cells. Although proliferation of all three cell types was stimulated by platelet-derived growth factor and basic fibroblast growth factor, differences were also observed in the response to these agents. B3.1 + TR alpha 1 cells were more sensitive to platelet-derived growth factor than B3.1 and B3.1 + v-erbA cells. In contrast, B3.1 cells responded to basic fibroblast growth factor better than B3.1 + TR alpha 1 or B3.1 + v-erbA cells. Insulin-like growth factor I potentiated the action of platelet-derived growth factor and basic fibroblast growth factor. Again, different responses to treatment with insulin-like growth factor I alone were observed; B3.1 + TR alpha 1 cells did not respond to it, whereas B3.1 + v-erbA cells showed a dramatic stimulation by this agent. Interestingly, in the presence of T3, the blockade in B3.1 + TR alpha 1 cell proliferation was accompanied by the down-regulation of the typical astrocytic genes, glial fibrillary acidic protein and vimentin. These hormone effects were not found in v-erbA-expressing cells. In addition, v-erbA inhibited the basal expression of the cyclic nucleotide phosphodiesterase gene, an oligodendrocytic marker.(ABSTRACT TRUNCATED AT 250 WORDS)

  3. Optimizing the feeding operation of recombinant Escherichia coli ...

    African Journals Online (AJOL)

    Recombinant Escherichia coli BL21 was used to produce human-like collagen in fed-batch culture. After building and analyzing the kinetic models of fed-batch cultures, the maximum specific growth rate, Yx/s and Yp/s were 0.411 h-1 , 0.428 g·g-1 and 0.0716 g/g, respectively. The square error of cell growth models, glucose ...

  4. The effect of 30 months of low-dose replacement therapy with recombinant human growth hormone (rhGH) on insulin and C-peptide kinetics, insulin secretion, insulin sensitivity, glucose effectiveness, and body composition in GH-deficient adults

    DEFF Research Database (Denmark)

    Rosenfalck, A M; Maghsoudi, S; Fisker, S

    2000-01-01

    The aim of the present study was to evaluate the long-term (30 months) metabolic effects of recombinant human GH (rhGH) given in a mean dose of 6.7 microg/kg x day (= 1.6 IU/day), in 11 patients with adult GH deficiency. Glucose metabolism was evaluated by an oral glucose tolerance test and an iv...... (frequently sampled iv glucose tolerance test) glucose tolerance test, and body composition was estimated by dual-energy x-ray absorptiometry. Treatment with rhGH induced persistent favorable changes in body composition, with a 10% increase in lean body mass (P ... in glucose tolerance, beta-cell response was still inappropriate. Our conclusion is that long-term rhGH-replacement therapy in GH deficiency adults induced a significant deterioration in glucose tolerance, profound changes in kinetics of C-peptide, and insulin and prehepatic insulin secretion, despite...

  5. Platelet-rich plasma stimulated by pulse electric fields: Platelet activation, procoagulant markers, growth factor release and cell proliferation.

    Science.gov (United States)

    Frelinger, A L; Torres, A S; Caiafa, A; Morton, C A; Berny-Lang, M A; Gerrits, A J; Carmichael, S L; Neculaes, V B; Michelson, A D

    2016-01-01

    Therapeutic use of activated platelet-rich plasma (PRP) has been explored for wound healing, hemostasis and antimicrobial wound applications. Pulse electric field (PEF) stimulation may provide more consistent platelet activation and avoid complications associated with the addition of bovine thrombin, the current state of the art ex vivo activator of therapeutic PRP. The aim of this study was to compare the ability of PEF, bovine thrombin and thrombin receptor activating peptide (TRAP) to activate human PRP, release growth factors and induce cell proliferation in vitro. Human PRP was prepared in the Harvest SmartPreP2 System and treated with vehicle, PEF, bovine thrombin, TRAP or Triton X-100. Platelet activation and procoagulant markers and microparticle generation were measured by flow cytometry. Released growth factors were measured by ELISA. The releasates were tested for their ability to stimulate proliferation of human epithelial cells in culture. PEF produced more platelet-derived microparticles, P-selectin-positive particles and procoagulant annexin V-positive particles than bovine thrombin or TRAP. These differences were associated with higher levels of released epidermal growth factor after PEF than after bovine thrombin or TRAP but similar levels of platelet-derived, vascular-endothelial, and basic fibroblast growth factors, and platelet factor 4. Supernatant from PEF-treated platelets significantly increased cell proliferation compared to plasma. In conclusion, PEF treatment of fresh PRP results in generation of microparticles, exposure of prothrombotic platelet surfaces, differential release of growth factors compared to bovine thrombin and TRAP and significant cell proliferation. These results, together with PEF's inherent advantages, suggest that PEF may be a superior alternative to bovine thrombin activation of PRP for therapeutic applications.

  6. Oxygen-hydrogen recombination system

    International Nuclear Information System (INIS)

    Sato, Shuichiro; Takejima, Masaki.

    1981-01-01

    Purpose: To avoid reduction in the performance of catalyst used for an oxygen-hydrogen recombiner in the off gas processing system of a nuclear reactor. Constitution: A thermometer is provided for the detection of temperature in an oxygen-hydrogen recombiner. A cooling pipe is provided in the recombiner and cooling medium is introduced externally. The cooling medium may be water or air. In accordance with the detection value from the thermometer, ON-OFF control is carried out for a valve to control the flow rate of the cooling medium thereby rendering the temperature in the recombiner to a predetermined value. This can prevent the catalyst from being exposed to high temperature and avoid the reduction in the performance of the catalyst. (Ikeda, J.)

  7. Controlled Release from Recombinant Polymers

    Science.gov (United States)

    Price, Robert; Poursaid, Azadeh; Ghandehari, Hamidreza

    2014-01-01

    Recombinant polymers provide a high degree of molecular definition for correlating structure with function in controlled release. The wide array of amino acids available as building blocks for these materials lend many advantages including biorecognition, biodegradability, potential biocompatibility, and control over mechanical properties among other attributes. Genetic engineering and DNA manipulation techniques enable the optimization of structure for precise control over spatial and temporal release. Unlike the majority of chemical synthetic strategies used, recombinant DNA technology has allowed for the production of monodisperse polymers with specifically defined sequences. Several classes of recombinant polymers have been used for controlled drug delivery. These include, but are not limited to, elastin-like, silk-like, and silk-elastinlike proteins, as well as emerging cationic polymers for gene delivery. In this article, progress and prospects of recombinant polymers used in controlled release will be reviewed. PMID:24956486

  8. Hydrogen recombiner development at AECL

    International Nuclear Information System (INIS)

    Dewit, W.A.; Koroll, G.W.; Loesel Sitar, J.; Graham, W.R.C.

    1997-01-01

    Catalytic recombiners have been developed at AECL for the purpose of hydrogen removal in post-accident nuclear containment buildings. The recombiners are based on a particular catalyst designed by AECL which has extraordinary resistance to fouling from water and water vapour and a large thermodynamic range of operation. The catalysts were developed, originally, for the purpose of heavy water manufacturing by way of a catalytic exchange process. Application of these catalyst materials in recombiners for containment applications began in the late 1980's. The first application was a passive recombiner, qualified for use in control of radiolytic hydrogen in the headspace of a pool-type experimental reactor of AECL design in 1988. The passive, or natural convection recombiner concept has continued development to commercial stage for application in power reactor containments. This paper reviews the AECL recombiner development, describes the current model and shows results from tests of full-scale recombiners in the Large Scale Vented Combustion Test Facility at AECL-WL. The AECL recombiner is designed for compactness and ease of engineering into containment. The design is a simple, open-ended rectangular enclosure with catalyst elements arranged inside to promote optimum convective flow driven by heat of recombination at the catalyst surface. Self start, as evidenced by catalyst heating and initiation of flow, is achieved in less than 1% hydrogen, with available oxygen, at room temperature and 100% relative humidity. This low temperature start-up in condensing atmospheres is viewed as the most challenging condition for wet-proofing effectiveness. Cold start-up is a vital performance requirement in containments, such as CANDU, where engineered air-cooling systems are operating and where long-term hydrogen control is required, after containment atmospheres have cooled. Once started, the removal capacity scales linearly with the inlet cross-section area and the partial

  9. Review of Parton Recombination Models

    International Nuclear Information System (INIS)

    Bass, Steffen A

    2006-01-01

    Parton recombination models have been very successful in explaining data taken at RHIC on hadron spectra and emission patterns in Au+Au collisions at transverse momenta above 2 GeV/c, which have exhibited features which could not be understood in the framework of basic perturbative QCD. In this article I will review the current status on recombination models and outline which future challenges need to be addressed by this class of models

  10. Recombinant snake venom prothrombin activators

    OpenAIRE

    L?vgren, Ann

    2012-01-01

    Three prothrombin activators; ecarin, which was originally isolated from the venom of the saw-scaled viper Echis carinatus, trocarin from the rough-scaled snake Tropidechis carinatus, and oscutarin from the Taipan snake Oxyuranus scutellatus, were expressed in mammalian cells with the purpose to obtain recombinant prothrombin activators that could be used to convert prothrombin to thrombin. We have previously reported that recombinant ecarin can efficiently generate thrombin without the need ...

  11. Delayed recombination and cosmic parameters

    International Nuclear Information System (INIS)

    Galli, Silvia; Melchiorri, Alessandro; Bean, Rachel; Silk, Joseph

    2008-01-01

    Current cosmological constraints from cosmic microwave background anisotropies are typically derived assuming a standard recombination scheme, however additional resonance and ionizing radiation sources can delay recombination, altering the cosmic ionization history and the cosmological inferences drawn from the cosmic microwave background data. We show that for recent observations of the cosmic microwave background anisotropy, from the Wilkinson microwave anisotropy probe satellite mission (WMAP) 5-year survey and from the arcminute cosmology bolometer array receiver experiment, additional resonance radiation is nearly degenerate with variations in the spectral index, n s , and has a marked effect on uncertainties in constraints on the Hubble constant, age of the universe, curvature and the upper bound on the neutrino mass. When a modified recombination scheme is considered, the redshift of recombination is constrained to z * =1078±11, with uncertainties in the measurement weaker by 1 order of magnitude than those obtained under the assumption of standard recombination while constraints on the shift parameter are shifted by 1σ to R=1.734±0.028. From the WMAP5 data we obtain the following constraints on the resonance and ionization sources parameters: ε α i <0.058 at 95% c.l.. Although delayed recombination limits the precision of parameter estimation from the WMAP satellite, we demonstrate that this should not be the case for future, smaller angular scales measurements, such as those by the Planck satellite mission.

  12. Doping in the recombinant era: strategies and counterstrategies.

    Science.gov (United States)

    Azzazy, Hassan M E; Mansour, Mai M H; Christenson, Robert H

    2005-11-01

    Advances in recombinant DNA technology have created one of the most powerful weapons in the current doping arsenal: recombinant proteins [Sweeney HL. Gene doping. Sci Am 2004;291:62-9; Unal M, Ozer Unal D. Gene doping in sports. Sports Med 2004;34:357-62]. Recombinant erythropoietin (EPO) and human growth hormone (hGH) are currently being abused but are fortunately detectable either directly by employing isoelectric focusing and immunoassays or indirectly by assessing changes in selected hematopoietic parameters. The detection is technically demanding due to the extent of similarity between the recombinant proteins and their endogenous counterparts. Another issue facing detection efforts is the speed and conditions at which blood samples are collected and analyzed in a sports setting. Recently, gene doping, which stemmed out of legitimate gene therapy trials, has emerged as the next level of doping. Erythropoietin (EPO), human growth hormone (hGH), insulin-like growth factor-1 (IGF-1), peroxisome proliferator-activated receptor-delta (PPAR delta), and myostatin inhibitor genes have been identified as primary targets for doping. Sports clinical scientists today are racing against the clock because assuring the continued integrity of sports competition depends on their ability to outpace the efforts of dopers by developing new detection strategies.

  13. Human platelet lysate as a promising growth-stimulating additive for culturing of stem cells and other cell types.

    Science.gov (United States)

    Shanskii, Ya D; Sergeeva, N S; Sviridova, I K; Kirakozov, M S; Kirsanova, V A; Akhmedova, S A; Antokhin, A I; Chissov, V I

    2013-11-01

    We compared the composition and biological activity of fetal calf serum and platelet lysate from donor platelet concentrate. In platelet lysate, the concentrations of alkaline phosphatase, lactate dehydrogenase, creatinine, and mineral metabolism parameters were lower, while parameters of lipid and protein metabolism were higher than in fetal calf serum. The concentrations of growth factors (platelet-derived (AA, AB, BB), vascular endothelial, insulin-like, and transforming growth factor β) in platelet lysate 1.7-148.7-fold surpassed the corresponding parameters in fetal calf serum. After replacement of fetal calf serum with platelet lysate in the culture medium (0, 25, 50, 75, and 100%), the count of multipotent mesenchymal stromal cells on day 7 (in comparison with day 1) increased by 154.8, 206.6, 228.2, 367.7, and 396.5%, respectively. Thus, platelet lysate can be an adequate non-xenogenic alternative for fetal calf serum.

  14. Distinct phosphotyrosines on a growth factor receptor bind to specific molecules that mediate different signaling pathways.

    Science.gov (United States)

    Fantl, W J; Escobedo, J A; Martin, G A; Turck, C W; del Rosario, M; McCormick, F; Williams, L T

    1992-05-01

    The receptor for platelet-derived growth factor (PDGF) binds two proteins containing SH2 domains, GTPase activating protein (GAP) and phosphatidylinositol 3-kinase (PI3-kinase). The sites on the receptor that mediate this interaction were identified by using phosphotyrosine-containing peptides representing receptor sequences to block specifically binding of either PI3-kinase or GAP. These results suggested that PI3-kinase binds two phosphotyrosine residues, each located in a 5 aa motif with an essential methionine at the fourth position C-terminal to the tyrosine. Point mutations at these sites caused a selective elimination of PI3-kinase binding and loss of PDGF-stimulated DNA synthesis. Mutation of the binding site for GAP prevented the receptor from associating with or phosphorylating GAP, but had no effect on PI3-kinase binding and little effect on DNA synthesis. Therefore, GAP and PI3-kinase interact with the receptor by binding to different phosphotyrosine-containing sequence motifs.

  15. Manufacturing of recombinant therapeutic proteins in microbial systems.

    Science.gov (United States)

    Graumann, Klaus; Premstaller, Andreas

    2006-02-01

    Recombinant therapeutic proteins have gained enormous importance for clinical applications. The first recombinant products have been produced in E. coli more than 20 years ago. Although with the advent of antibody-based therapeutics mammalian expression systems have experienced a major boost, microbial expression systems continue to be widely used in industry. Their intrinsic advantages, such as rapid growth, high yields and ease of manipulation, make them the premier choice for expression of non-glycosylated peptides and proteins. Innovative product classes such as antibody fragments or alternative binding molecules will further expand the use of microbial systems. Even more, novel, engineered production hosts and integrated technology platforms hold enormous potential for future applications. This review summarizes current applications and trends for development, production and analytical characterization of recombinant therapeutic proteins in microbial systems.

  16. PROGENITORS OF RECOMBINING SUPERNOVA REMNANTS

    Energy Technology Data Exchange (ETDEWEB)

    Moriya, Takashi J., E-mail: takashi.moriya@ipmu.jp [Kavli Institute for the Physics and Mathematics of the Universe, Todai Institutes for Advanced Study, University of Tokyo, Kashiwanoha 5-1-5, Kashiwa, Chiba 277-8583 (Japan)

    2012-05-01

    Usual supernova remnants have either ionizing plasma or plasma in collisional ionization equilibrium, i.e., the ionization temperature is lower than or equal to the electron temperature. However, the existence of recombining supernova remnants, i.e., supernova remnants with ionization temperature higher than the electron temperature, has been recently confirmed. One suggested way to have recombining plasma in a supernova remnant is to have a dense circumstellar medium at the time of the supernova explosion. If the circumstellar medium is dense enough, collisional ionization equilibrium can be established in the early stage of the evolution of the supernova remnant and subsequent adiabatic cooling, which occurs after the shock wave gets out of the dense circumstellar medium, makes the electron temperature lower than the ionization temperature. We study the circumstellar medium around several supernova progenitors and show which supernova progenitors can have a circumstellar medium dense enough to establish collisional ionization equilibrium soon after the explosion. We find that the circumstellar medium around red supergiants (especially massive ones) and the circumstellar medium dense enough to make Type IIn supernovae can establish collisional ionization equilibrium soon after the explosion and can evolve to become recombining supernova remnants. Wolf-Rayet stars and white dwarfs have the possibility to be recombining supernova remnants but the fraction is expected to be very small. As the occurrence rate of the explosions of red supergiants is much higher than that of Type IIn supernovae, the major progenitors of recombining supernova remnants are likely to be red supergiants.

  17. Meiotic recombination in human oocytes.

    Directory of Open Access Journals (Sweden)

    Edith Y Cheng

    2009-09-01

    Full Text Available Studies of human trisomies indicate a remarkable relationship between abnormal meiotic recombination and subsequent nondisjunction at maternal meiosis I or II. Specifically, failure to recombine or recombination events located either too near to or too far from the centromere have been linked to the origin of human trisomies. It should be possible to identify these abnormal crossover configurations by using immunofluorescence methodology to directly examine the meiotic recombination process in the human female. Accordingly, we initiated studies of crossover-associated proteins (e.g., MLH1 in human fetal oocytes to analyze their number and distribution on nondisjunction-prone human chromosomes and, more generally, to characterize genome-wide levels of recombination in the human female. Our analyses indicate that the number of MLH1 foci is lower than predicted from genetic linkage analysis, but its localization pattern conforms to that expected for a crossover-associated protein. In studies of individual chromosomes, our observations provide evidence for the presence of "vulnerable" crossover configurations in the fetal oocyte, consistent with the idea that these are subsequently translated into nondisjunctional events in the adult oocyte.

  18. Recombination Phenotypes of Escherichia coli greA Mutants

    Directory of Open Access Journals (Sweden)

    Poteete Anthony R

    2011-03-01

    Full Text Available Abstract Background The elongation factor GreA binds to RNA polymerase and modulates transcriptional pausing. Some recent research suggests that the primary role of GreA may not be to regulate gene expression, but rather, to promote the progression of replication forks which collide with RNA polymerase, and which might otherwise collapse. Replication fork collapse is known to generate dsDNA breaks, which can be recombinogenic. It follows that GreA malfunction could have consequences affecting homologous recombination. Results Escherichia coli mutants bearing substitutions of the active site acidic residues of the transcription elongation factor GreA, D41N and E44K, were isolated as suppressors of growth inhibition by a toxic variant of the bacteriophage lambda Red-beta recombination protein. These mutants, as well as a D41A greA mutant and a greA deletion, were tested for proficiency in recombination events. The mutations were found to increase the efficiency of RecA-RecBCD-mediated and RecA-Red-mediated recombination, which are replication-independent, and to decrease the efficiency of replication-dependent Red-mediated recombination. Conclusion These observations provide new evidence for a role of GreA in resolving conflicts between replication and transcription.

  19. Comparison of poliovirus recombinants: accumulation of point mutations provides further advantages.

    Science.gov (United States)

    Savolainen-Kopra, Carita; Samoilovich, Elena; Kahelin, Heidi; Hiekka, Anna-Kaisa; Hovi, Tapani; Roivainen, Merja

    2009-08-01

    The roles of recombination and accumulation of point mutations in the origin of new poliovirus (PV) characteristics have been hypothesized, but it is not known which are essential to evolution. We studied phenotypic differences between recombinant PV strains isolated from successive stool specimens of an oral PV vaccine recipient. The studied strains included three PV2/PV1 recombinants with increasing numbers of mutations in the VP1 gene, two of the three with an amino acid change I-->T in the DE-loop of VP1, their putative PV1 parent and strains Sabin 1 and 2. Growth of these viruses was examined in three cell lines: colorectal adenocarcinoma, neuroblastoma and HeLa. The main observation was a higher growth rate between 4 and 6 h post-infection of the two recombinants with the I-->T substitution. All recombinants grew at a higher rate than parental strains in the exponential phase of the replication cycle. In a temperature sensitivity test, the I-->T-substituted recombinants replicated equally well at an elevated temperature. Complete genome sequencing of the three recombinants revealed 12 (3), 19 (3) and 27 (3) nucleotide (amino acid) differences from Sabin. Mutations were located in regions defining attenuation, temperature sensitivity, antigenicity and the cis-acting replicating element. The recombination site was in the 5' end of 3D. In a competition assay, the most mutated recombinant beat parental Sabin in all three cell lines, strongly suggesting that this virus has an advantage. Two independent intertypic recombinants, PV3/PV1 and PV3/PV2, also showed similar growth advantages, but they also contained several point mutations. Thus, our data defend the hypothesis that accumulation of certain advantageous mutations plays a key role in gaining increased fitness.

  20. Electric hydrogen recombiner special tests

    International Nuclear Information System (INIS)

    Wilson, J.F.

    1975-12-01

    Westinghouse has produced an electric hydrogen recombiner to control hydrogen levels in reactor containments following a postulated loss-of-coolant accident. The recombiner underwent extensive testing for NRC qualification (see WCAP 7709-L and Supplements 1, 2, 3, 4). As a result, WCAP 7709-L and Supplements 1, 2, 3, and 4 have been accepted by the NRC for reference in applications not committed to IEEE-323-1974. Supplement 5 and the next supplement will demonstrate conformance to IEEE-323-1974. This supplement describes additional tests, beyond those necessary to qualify the system, which will be referenced in supplement 6. Each test has demonstrated a considerable margin of safety over required performance. Concurrently, the test results increased the fund of technical information on the electric hydrogen recombiner

  1. Concentration of platelets and growth factors in platelet-rich plasma from Goettingen minipigs.

    Science.gov (United States)

    Jungbluth, Pascal; Grassmann, Jan-Peter; Thelen, Simon; Wild, Michael; Sager, Martin; Windolf, Joachim; Hakimi, Mohssen

    2014-01-01

    In minipigs little is known about the concentration of growth factors in plasma, despite their major role in several patho-physiological processes such as healing of fractures. This prompted us to study the concentration of platelets and selected growth factors in plasma and platelet-rich plasma (PRP) preparation of sixteen Goettingen minipigs. Platelet concentrations increased significantly in PRP in comparison to native blood plasma. Generally, significant increase in the concentration of all growth factors tested was observed in the PRP in comparison to the corresponding plasma or serum. Five of the plasma samples examined contained detectable levels of bone morphogenic protein 2 (BMP-2) whereas eleven of the plasma or serum samples contained minimal amounts of vascular endothelial growth factor (VEGF) and platelet-derived growth factor (PDGF-bb) respectively. On the other hand variable concentrations of bone morphogenic protein 7 (BMP-7) and transforming growth factor β1 (TGF-β1) were measured in all plasma samples. In contrast, all PRP samples contained significantly increased amounts of growth factors. The level of BMP-2, BMP-7, TGF-β1, VEGF and PDGF-bb increased by 17.6, 1.5, 7.1, 7.2 and 103.3 fold, in comparison to the corresponding non-enriched preparations. Moreover significant positive correlations were found between platelet count and the concentrations of BMP-2 (r=0.62, pplatelet-rich plasma of minipigs which might thus serve as a source of autologous growth factors.

  2. Organotypic Cultures of Intervertebral Disc Cells: Responses to Growth Factors and Signaling Pathways Involved

    Directory of Open Access Journals (Sweden)

    Harris Pratsinis

    2015-01-01

    Full Text Available Intervertebral disc (IVD degeneration is strongly associated with low back pain, a major cause of disability worldwide. An in-depth understanding of IVD cell physiology is required for the design of novel regenerative therapies. Accordingly, aim of this work was the study of IVD cell responses to mitogenic growth factors in a three-dimensional (3D organotypic milieu, comprising characteristic molecules of IVD’s extracellular matrix. In particular, annulus fibrosus (AF cells were cultured inside collagen type-I gels, while nucleus pulposus (NP cells in chondroitin sulfate A (CSA supplemented collagen gels, and the effects of Platelet-Derived Growth Factor (PDGF, basic Fibroblast Growth Factor (bFGF, and Insulin-Like Growth Factor-I (IGF-I were assessed. All three growth factors stimulated DNA synthesis in both AF and NP 3D cell cultures, with potencies similar to those observed previously in monolayers. CSA supplementation inhibited basal DNA synthesis rates, without affecting the response to growth factors. ERK and Akt were found to be phosphorylated following growth factor stimulation. Blockade of these two signaling pathways using pharmacologic inhibitors significantly, though not completely, inhibited growth factor-induced DNA synthesis. The proposed culture systems may prove useful for further in vitro studies aiming at future interventions for IVD regeneration.

  3. Genome engineering for improved recombinant protein expression in Escherichia coli.

    Science.gov (United States)

    Mahalik, Shubhashree; Sharma, Ashish K; Mukherjee, Krishna J

    2014-12-19

    A metabolic engineering perspective which views recombinant protein expression as a multistep pathway allows us to move beyond vector design and identify the downstream rate limiting steps in expression. In E.coli these are typically at the translational level and the supply of precursors in the form of energy, amino acids and nucleotides. Further recombinant protein production triggers a global cellular stress response which feedback inhibits both growth and product formation. Countering this requires a system level analysis followed by a rational host cell engineering to sustain expression for longer time periods. Another strategy to increase protein yields could be to divert the metabolic flux away from biomass formation and towards recombinant protein production. This would require a growth stoppage mechanism which does not affect the metabolic activity of the cell or the transcriptional or translational efficiencies. Finally cells have to be designed for efficient export to prevent buildup of proteins inside the cytoplasm and also simplify downstream processing. The rational and the high throughput strategies that can be used for the construction of such improved host cell platforms for recombinant protein expression is the focus of this review.

  4. Recombinant human endostatin improves tumor vasculature and alleviates hypoxia in Lewis lung carcinoma

    International Nuclear Information System (INIS)

    Peng Fang; Wang Jin; Zou Yi; Bao Yong; Huang Wenlin; Chen Guangming; Luo Xianrong; Chen Ming

    2011-01-01

    Objective: To investigate whether recombinant human endostatin can create a time window of vascular normalization prior to vascular pruning to alleviate hypoxia in Lewis lung carcinoma in mice. Methods: Kinetic changes in morphology of tumor vasculature in response to recombinant human endostatin were detected under a confocal microscope with immunofluorescent staining in Lewis lung carcinomas in mice. The hypoxic cell fraction of different time was assessed with immunohistochemical staining . Effects on tumor growth were monitored as indicated in the growth curve of tumors . Results: Compared with the control group vascularity of the tumors was reduced over time by recombinant human endostatin treatment and significantly regressed for 9 days. During the treatment, pericyte coverage increased at day 3, increased markedly at day 5, and fell again at day 7. The vascular basement membrane was thin and closely associated with endothelial cells after recombinant human endostatin treatment, but appeared thickened, loosely associated with endothelial cells in control tumors. The decrease in hypoxic cell fraction at day 5 after treatment was also found. Tumor growth was not accelerated 5 days after recombinant human endostatin treatment. Conclusions: Recombinant human endostatin can normalize tumor vasculature within day 3 to 7, leading to improved tumor oxygenation. The results provide important experimental basis for combining recombinant human endostatin with radiation therapy in human tumors. (authors)

  5. Secretion of human epidermal growth factor (EGF) in autotrophic culture by a recombinant hydrogen-utilizing bacterium, Pseudomonas pseudoflava, carrying broad-host-range EGF secretion vector pKSEGF2.

    OpenAIRE

    Hayase, N; Ishiyama, A; Niwano, M

    1994-01-01

    We constructed the broad-host-range human epidermal growth factor (EGF) secretion plasmid pKSEGF2 by inserting the Escherichia coli tac promoter, the signal sequence of Pseudomonas stutzeri amylase, and the synthesized EGF gene into the broad-host-range vector pKT230. E. coli JM109 carrying pKSEGF2 secreted EGF into the periplasm and the culture medium under the control of the tac promoter. Pseudomonas aeruginosa PAO1161 carrying pKSEGF2 and Pseudomonas putida AC10 carrying pKSEGF2 secreted E...

  6. Platelet-rich plasma preparation for regenerative medicine: optimization and quantification of cytokines and growth factors

    Science.gov (United States)

    2013-01-01

    Introduction Platelet-rich plasma (PRP) is nowadays widely applied in different clinical scenarios, such as orthopedics, ophthalmology and healing therapies, as a growth factor pool for improving tissue regeneration. Studies into its clinical efficiency are not conclusive and one of the main reasons for this is that different PRP preparations are used, eliciting different responses that cannot be compared. Platelet quantification and the growth factor content definition must be defined in order to understand molecular mechanisms behind PRP regenerative strength. Standardization of PRP preparations is thus urgently needed. Methods PRP was prepared by centrifugation varying the relative centrifugal force, temperature, and time. Having quantified platelet recovery and yield, the two-step procedure that rendered the highest output was chosen and further analyzed. Cytokine content was determined in different fractions obtained throughout the whole centrifugation procedure. Results Our method showed reproducibility when applied to different blood donors. We recovered 46.9 to 69.5% of total initial platelets and the procedure resulted in a 5.4-fold to 7.3-fold increase in platelet concentration (1.4 × 106 to 1.9 × 106 platelets/μl). Platelets were highly purified, because only blood cells and leukocytes was present in the final PRP preparation. We also quantified growth factors, cytokines and chemokines secreted by the concentrated platelets after activation with calcium and calcium/thrombin. High concentrations of platelet-derived growth factor, endothelial growth factor and transforming growth factor (TGF) were secreted, together with the anti-inflammatory and proinflammatory cytokines interleukin (IL)-4, IL-8, IL-13, IL-17, tumor necrosis factor (TNF)-α and interferon (IFN)-α. No cytokines were secreted before platelet activation. TGF-β3 and IFNγ were not detected in any studied fraction. Clots obtained after platelet coagulation retained a high concentration of

  7. A Naturally Occurring Recombinant Enterovirus Expresses a Torovirus Deubiquitinase.

    Science.gov (United States)

    Shang, Pengcheng; Misra, Saurav; Hause, Ben; Fang, Ying

    2017-07-15

    Enteroviruses (EVs) are implicated in a wide range of diseases in humans and animals. In this study, a novel enterovirus (enterovirus species G [EVG]) (EVG 08/NC_USA/2015) was isolated from a diagnostic sample from a neonatal pig diarrhea case and identified by using metagenomics and complete genome sequencing. The viral genome shares 75.4% nucleotide identity with a prototypic EVG strain (PEV9 UKG/410/73). Remarkably, a 582-nucleotide insertion, flanked by 3C pro cleavage sites at the 5' and 3' ends, was found in the 2C/3A junction region of the viral genome. This insertion encodes a predicted protease with 54 to 68% amino acid identity to torovirus (ToV) papain-like protease (PLP) (ToV-PLP). Structural homology modeling predicts that this protease adopts a fold and a catalytic site characteristic of minimal PLP catalytic domains. This structure is similar to those of core catalytic domains of the foot-and-mouth disease virus leader protease and coronavirus PLPs, which act as deubiquitinating and deISGylating (interferon [IFN]-stimulated gene 15 [ISG15]-removing) enzymes on host cell substrates. Importantly, the recombinant ToV-PLP protein derived from this novel enterovirus also showed strong deubiquitination and deISGylation activities and demonstrated the ability to suppress IFN-β expression. Using reverse genetics, we generated a ToV-PLP knockout recombinant virus. Compared to the wild-type virus, the ToV-PLP knockout mutant virus showed impaired growth and induced higher expression levels of innate immune genes in infected cells. These results suggest that ToV-PLP functions as an innate immune antagonist; enterovirus G may therefore gain fitness through the acquisition of ToV-PLP from a recombination event. IMPORTANCE Enteroviruses comprise a highly diversified group of viruses. Genetic recombination has been considered a driving force for viral evolution; however, recombination between viruses from two different orders is a rare event. In this study, we

  8. Production and recombination of gluons

    International Nuclear Information System (INIS)

    Temiraliev, A.T.

    2006-01-01

    Full text: Nonlinear Markov process of parton production has been considered. The Kolmogorov equation is applied for the evolution equation based on the approximation of independent gluons production in every decay act. We introduced a 'crossing' parameter and used the combination relations to obtain nonlinear recombination equation for the evolution of gluon structure function. (author)

  9. Recombinator of hydrogen and oxygen

    International Nuclear Information System (INIS)

    Stejskal, J.; Klein, O.; Scholtz, G.; Schmidt, P.; Olaussson, A.

    1976-01-01

    Improvements are proposed for the well known reactors for the catalytic recombination of hydrogen and oxygen, which should permit this being used in contiuous operation in nuclear reactors (BWRs). The improvements concern the geometric arrangement of gas-inlet and -outlet pipes, the inclination of the axis of the catalyst container and the introduction of remote operation. (UWI) [de

  10. Improving recombinant protein purification yield

    Science.gov (United States)

    Production of adequate amounts of recombinant proteins is essential for antibody production, biochemical activity study, and structural determination during the post-genomic era. It’s technologically challenging and a limiting factor for tung oil research because analytical reagents such as high qua...

  11. Recombination in hepatitis C virus.

    Science.gov (United States)

    González-Candelas, Fernando; López-Labrador, F Xavier; Bracho, María Alma

    2011-10-01

    Hepatitis C virus (HCV) is a Flavivirus with a positive-sense, single-stranded RNA genome of about 9,600 nucleotides. It is a major cause of liver disease, infecting almost 200 million people all over the world. Similarly to most RNA viruses, HCV displays very high levels of genetic diversity which have been used to differentiate six major genotypes and about 80 subtypes. Although the different genotypes and subtypes share basic biological and pathogenic features they differ in clinical outcomes, response to treatment and epidemiology. The first HCV recombinant strain, in which different genome segments derived from parentals of different genotypes, was described in St. Petersburg (Russia) in 2002. Since then, there have been only a few more than a dozen reports including descriptions of HCV recombinants at all levels: between genotypes, between subtypes of the same genotype and even between strains of the same subtype. Here, we review the literature considering the reasons underlying the difficulties for unequivocally establishing recombination in this virus along with the analytical methods necessary to do it. Finally, we analyze the potential consequences, especially in clinical practice, of HCV recombination in light of the coming new therapeutic approaches against this virus.

  12. Cloning an artificial gene encoding angiostatic anginex: From designed peptide to functional recombinant protein

    International Nuclear Information System (INIS)

    Brandwijk, Ricardo J.M.G.E.; Nesmelova, Irina; Dings, Ruud P.M.; Mayo, Kevin H.; Thijssen, Victor L.J.L.; Griffioen, Arjan W.

    2005-01-01

    Anginex, a designed peptide 33-mer, is a potent angiogenesis inhibitor and anti-tumor agent in vivo. Anginex functions by inhibiting endothelial cell (EC) proliferation and migration leading to detachment and apoptosis of activated EC's. To better understand tumor endothelium targeting properties of anginex and enable its use in gene therapy, we constructed an artificial gene encoding the biologically exogenous peptide and produced the protein recombinantly in Pichia pastoris. Mass spectrometry shows recombinant anginex to be a dimer and circular dichroism shows the recombinant protein folds with β-strand structure like the synthetic peptide. Moreover, like parent anginex, the recombinant protein is active at inhibiting EC growth and migration, as well as inhibiting angiogenesis in vivo in the chorioallantoic membrane of the chick embryo. This study demonstrated that it is possible to produce a functionally active protein version of a rationally designed peptide, using an artificial gene and the recombinant protein approach

  13. Acceptance of a reusable self-injection device for recombinant human growth hormone: final data from a questionnaire-based, cross-sectional, international, multicenter, observational study in pediatric patients

    Directory of Open Access Journals (Sweden)

    Schnabel D

    2016-09-01

    Full Text Available Dirk Schnabel,1 Carl-Joachim Partsch,2 Muriel Houang,3 Sarah Ehtisham,4 Helen Johnstone,5 Markus Zabransky,6 Wieland Kiess7 1Pediatric Endocrinology, Center for Chronic Sick Children, Otto-Heubner-Centrum für Kinder- und Jugendmedizin, Charite, University Medicine, Berlin, Germany; 2Endokrinologikum Hamburg, Hamburg, Germany; 3Explorations Fonctionnelles Endocriniennes, Hôpital Armand Trousseau, Paris, France; 4Mediclinic City Hospital, Dubai Healthcare City, Dubai, United Arab Emirates; 5The Newcastle upon Tyne Hospitals NHS Foundation Trust, Newcastle, UK; 6Sandoz International GmbH, Holzkirchen, 7Department of Women and Child Health, Hospital for Children and Adolescents, University Hospitals, University of Leipzig, Leipzig, Germany Background: A questionnaire-based survey was conducted to assess attitudes toward a reusable self-injection system (SurePal™ among pediatric patients with growth disturbances who were prescribed treatment with Omnitrope® within routine clinical practice.Methods: This was a multicenter, observational study, incorporated into the noninterventional PAtients TReated with Omnitrope® (PATRO Children study. Included subjects, or their caregivers, completed a questionnaire on the following five main areas: attractiveness of SurePal™, training received, using the device, the low drug wastage system, and experience versus other devices used previously (pretreated patients. Responses were based on a 5-point scale, with 2 being the best possible outcome and −2 the worst possible outcome.Results: In total, 550 patients were included in this study (338 from France, 169 from Germany, and 43 from the UK. The mean age ± standard deviation of participants was 10.8±3.5 years; the majority (57% were male and growth hormone treatment naïve (88%. Almost half (49.8% of children prepared their SurePal™ for injection themselves and 45.5% performed injections themselves. As patients progressed into their teens, the majority

  14. Potency of full-length MGF to induce maximal activation of the IGF-I R Is similar to recombinant human IGF-I at high equimolar concentrations

    NARCIS (Netherlands)

    J.A.M.J.L. Janssen (Joseph); L.J. Hofland (Leo); C.J. Strasburger; E.S.R.D. Van Dungen (Elisabeth S.R. Den); M. Thevis (Mario)

    2016-01-01

    textabstractAims To compare full-length mechano growth factor (full-length MGF) with human recombinant insulin-like growth factor-I (IGF-I) and human recombinant insulin (HI) in their ability to activate the human IGF-I receptor (IGF-IR), the human insulin receptor (IR-A) and the human insulin

  15. Live recombinant BHV/BRSV vaccine

    NARCIS (Netherlands)

    Keil, G.M.; Rijsewijk, F.A.M.

    1998-01-01

    The present invention refers to synthetic Bovine Respiratory Syncytium virus genes. Also the invention relates to live attenuated Bovine Herpesvirus recombinants carrying such synthetic genes. Furthermore, the invention relates to vaccines based on these live attenuated recombinants, for the

  16. Hadron production at RHIC: recombination of quarks

    Energy Technology Data Exchange (ETDEWEB)

    Fries, Rainer J [School of Physics and Astronomy, University of Minnesota, Minneapolis, MN 55455 (United States)

    2005-01-01

    We discuss quark recombination applied to the hadronization of a quark gluon plasma. It has been shown that the quark recombination model can explain essential features of hadron production measured in high energy heavy ion collisions.

  17. Affinity purification of recombinant human plasminogen activator ...

    African Journals Online (AJOL)

    Affinity purification of recombinant human plasminogen activator from ... Screening antibody was performed using rhPA milk in an ELISA-elution assay. ... useful for purifying other tPA mutants or other novel recombinant milkderived proteins.

  18. Graded Recombination Layers for Multijunction Photovoltaics

    KAUST Repository

    Koleilat, Ghada I.; Wang, Xihua; Sargent, Edward H.

    2012-01-01

    it to achieve multicolor and spectrally tunable behavior. In series-connected current-matched multijunction devices, the recombination layers must allow the hole current from one cell to recombine, with high efficiency and low voltage loss, with the electron

  19. Recombinant innovation and endogenous technological transitions

    NARCIS (Netherlands)

    Frenken, K.; Izquierdo, L.R.; Zeppini, P.

    2012-01-01

    We propose a model of technological transitions based on two different types of innovations. Branching innovations refer to technological improvements along a particular path, while recombinant innovations represent fusions of multiple paths. Recombinant innovations create "short-cuts" which reduce

  20. Platelet-rich plasma preparation for regenerative medicine: optimization and quantification of cytokines and growth factors.

    Science.gov (United States)

    Amable, Paola Romina; Carias, Rosana Bizon Vieira; Teixeira, Marcus Vinicius Telles; da Cruz Pacheco, Italo; Corrêa do Amaral, Ronaldo José Farias; Granjeiro, José Mauro; Borojevic, Radovan

    2013-06-07

    Platelet-rich plasma (PRP) is nowadays widely applied in different clinical scenarios, such as orthopedics, ophthalmology and healing therapies, as a growth factor pool for improving tissue regeneration. Studies into its clinical efficiency are not conclusive and one of the main reasons for this is that different PRP preparations are used, eliciting different responses that cannot be compared. Platelet quantification and the growth factor content definition must be defined in order to understand molecular mechanisms behind PRP regenerative strength. Standardization of PRP preparations is thus urgently needed. PRP was prepared by centrifugation varying the relative centrifugal force, temperature, and time. Having quantified platelet recovery and yield, the two-step procedure that rendered the highest output was chosen and further analyzed. Cytokine content was determined in different fractions obtained throughout the whole centrifugation procedure. Our method showed reproducibility when applied to different blood donors. We recovered 46.9 to 69.5% of total initial platelets and the procedure resulted in a 5.4-fold to 7.3-fold increase in platelet concentration (1.4 × 10(6) to 1.9 × 10(6) platelets/μl). Platelets were highly purified, because only platelets after activation with calcium and calcium/thrombin. High concentrations of platelet-derived growth factor, endothelial growth factor and transforming growth factor (TGF) were secreted, together with the anti-inflammatory and proinflammatory cytokines interleukin (IL)-4, IL-8, IL-13, IL-17, tumor necrosis factor (TNF)-α and interferon (IFN)-α. No cytokines were secreted before platelet activation. TGF-β3 and IFNγ were not detected in any studied fraction. Clots obtained after platelet coagulation retained a high concentration of several growth factors, including platelet-derived growth factor and TGF. Our study resulted in a consistent PRP preparation method that yielded a cytokine and growth factor pool

  1. Population inversion in recombining hydrogen plasma

    International Nuclear Information System (INIS)

    Furukane, Utaro; Yokota, Toshiaki; Oda, Toshiatsu.

    1978-11-01

    The collisional-radiative model is applied to a recombining hydrogen plasma in order to investigate the plasma condition in which the population inversion between the energy levels of hydrogen can be generated. The population inversion is expected in a plasma where the three body recombination has a large contribution to the recombining processes and the effective recombination rate is beyond a certain value for a given electron density and temperature. Calculated results are presented in figures and tables. (author)

  2. Regulation of homologous recombination in eukaryotes

    OpenAIRE

    Heyer, Wolf-Dietrich; Ehmsen, Kirk T.; Liu, Jie

    2010-01-01

    Homologous recombination is required for accurate chromosome segregation during the first meiotic division and constitutes a key repair and tolerance pathway for complex DNA damage including DNA double-stranded breaks, interstrand crosslinks, and DNA gaps. In addition, recombination and replication are inextricably linked, as recombination recovers stalled and broken replication forks enabling the evolution of larger genomes/replicons. Defects in recombination lead to genomic instability and ...

  3. The effect of a single recombination event

    DEFF Research Database (Denmark)

    Schierup, Mikkel Heide; Jensen, Thomas Mailund; Wiuf, Carsten

    We investigate the variance in how visible a single recombination event is in a SNP data set as a function of the type of recombination event and its age. Data is simulated under the coalescent with recombination and inference is by the popular composite likelihood methods. The major determinant...

  4. A trans-Complementing Recombination Trap Demonstrates a Low Propensity of Flaviviruses for Intermolecular Recombination▿

    Science.gov (United States)

    Taucher, Christian; Berger, Angelika; Mandl, Christian W.

    2010-01-01

    Intermolecular recombination between the genomes of closely related RNA viruses can result in the emergence of novel strains with altered pathogenic potential and antigenicity. Although recombination between flavivirus genomes has never been demonstrated experimentally, the potential risk of generating undesirable recombinants has nevertheless been a matter of concern and controversy with respect to the development of live flavivirus vaccines. As an experimental system for investigating the ability of flavivirus genomes to recombine, we developed a “recombination trap,” which was designed to allow the products of rare recombination events to be selected and amplified. To do this, we established reciprocal packaging systems consisting of pairs of self-replicating subgenomic RNAs (replicons) derived from tick-borne encephalitis virus (TBEV), West Nile virus (WNV), and Japanese encephalitis virus (JEV) that could complement each other in trans and thus be propagated together in cell culture over multiple passages. Any infectious viruses with intact, full-length genomes that were generated by recombination of the two replicons would be selected and enriched by end point dilution passage, as was demonstrated in a spiking experiment in which a small amount of wild-type virus was mixed with the packaged replicons. Using the recombination trap and the JEV system, we detected two aberrant recombination events, both of which yielded unnatural genomes containing duplications. Infectious clones of both of these genomes yielded viruses with impaired growth properties. Despite the fact that the replicon pairs shared approximately 600 nucleotides of identical sequence where a precise homologous crossover event would have yielded a wild-type genome, this was not observed in any of these systems, and the TBEV and WNV systems did not yield any viable recombinant genomes at all. Our results show that intergenomic recombination can occur in the structural region of flaviviruses

  5. Epidermal growth factor and insulin-like growth factor I upregulate the expression of the epidermal growth factor system in rat liver

    DEFF Research Database (Denmark)

    Bor, M V; Sørensen, B S; Vinter-Jensen, L

    2000-01-01

    BACKGROUND/AIM: Both epidermal growth factor and insulin-like growth factor I play a role in connection with the liver. In the present study, the possible interaction of these two growth factor systems was studied by investigating the effect of epidermal growth factor or insulin-like growth factor...... I treatment on the expression of the epidermal growth factor receptor, and its activating ligands, transforming growth factor-alpha and epidermal growth factor. METHODS: Fifty-five male rats received no treatment, human recombinant epidermal growth factor or human recombinant insulin-like growth.......8+/-1.6 fmol/mg protein epidermal growth factor and 144+/-22 fmol/mg protein transforming growth factor-alpha. Both epidermal growth factor and insulin-like growth factor I treatment increased the expression of mRNA for transforming growth factor-alpha and epidermal growth factor receptor, as well...

  6. Recombination Catalysts for Hypersonic Fuels

    Science.gov (United States)

    Chinitz, W.

    1998-01-01

    The goal of commercially-viable access to space will require technologies that reduce propulsion system weight and complexity, while extracting maximum energy from the products of combustion. This work is directed toward developing effective nozzle recombination catalysts for the supersonic and hypersonic aeropropulsion engines used to provide such access to space. Effective nozzle recombination will significantly reduce rk=le length (hence, propulsion system weight) and reduce fuel requirements, further decreasing the vehicle's gross lift-off weight. Two such catalysts have been identified in this work, barium and antimony compounds, by developing chemical kinetic reaction mechanisms for these materials and determining the engine performance enhancement for a typical flight trajectory. Significant performance improvements are indicated, using only 2% (mole or mass) of these compounds in the combustor product gas.

  7. Mechanisms of sister chromatid recombination

    International Nuclear Information System (INIS)

    Nakai, Sayaka; Machida, Isamu; Tsuji, Satsuki

    1985-01-01

    Studies using T948 as a model system have been carried out aimed at elucidating the mechanism of sister chromatid recombination (SCR). Characterization of U.V. light- and x-ray-induced SCR, the relationiship between SCR induction and DNA repair using rad mutations, and the relationship between SCR induction and the time of cell division using cdc mutations are presented. It has been supposed that SCR is induced at the phase of S-G 2 following DNA replication, that postreplication break of DNA strands is strongly involved in the induction of SCR, and that induction type of SCR, i.e., conversion type or recombination type, is dependent upon the type of molecular damage of DNA. (Namekawa, K.)

  8. Interface recombination influence on carrier transport

    International Nuclear Information System (INIS)

    Konin, A

    2013-01-01

    A theory of interface recombination in the semiconductor–semiconductor junction is developed. The interface recombination rate dependence on the nonequilibrium carrier densities is derived on the basis of a model in which the interface recombination occurs through the mechanism of trapping. The general relation between the interface recombination parameters at small carrier density deviation from the equilibrium ones is obtained. The validity of this relation is proved considering the generation of the Hall electric field in the extrinsic semiconductor sample. The anomalous Hall electromotive force in a weak magnetic field was investigated and interpreted by means of a new interface recombination model. The experimental data corroborate the developed theory. (paper)

  9. Yeast synthetic biology for the production of recombinant therapeutic proteins.

    Science.gov (United States)

    Kim, Hyunah; Yoo, Su Jin; Kang, Hyun Ah

    2015-02-01

    The production of recombinant therapeutic proteins is one of the fast-growing areas of molecular medicine and currently plays an important role in treatment of several diseases. Yeasts are unicellular eukaryotic microbial host cells that offer unique advantages in producing biopharmaceutical proteins. Yeasts are capable of robust growth on simple media, readily accommodate genetic modifications, and incorporate typical eukaryotic post-translational modifications. Saccharomyces cerevisiae is a traditional baker's yeast that has been used as a major host for the production of biopharmaceuticals; however, several nonconventional yeast species including Hansenula polymorpha, Pichia pastoris, and Yarrowia lipolytica have gained increasing attention as alternative hosts for the industrial production of recombinant proteins. In this review, we address the established and emerging genetic tools and host strains suitable for recombinant protein production in various yeast expression systems, particularly focusing on current efforts toward synthetic biology approaches in developing yeast cell factories for the production of therapeutic recombinant proteins. © FEMS 2015. All rights reserved. For permissions, please e-mail: journals.permission@oup.com.

  10. Recombinant Cyclophilins Lack Nuclease Activity

    OpenAIRE

    Manteca, Angel; Sanchez, Jesus

    2004-01-01

    Several single-domain prokaryotic and eukaryotic cyclophilins have been identified as also being unspecific nucleases with a role in DNA degradation during the lytic processes that accompany bacterial cell death and eukaryotic apoptosis. Evidence is provided here that the supposed nuclease activity of human and bacterial recombinant cyclophilins is due to contamination of the proteins by the host Escherichia coli endonuclease and is not an intrinsic property of these proteins.

  11. Workshop on Radio Recombination Lines

    CERN Document Server

    1980-01-01

    Since their first detection 15 years ago, radio recombination lines from several elements have been observed in a wide variety of objects including HII regions, planetary nebulae, molecular clouds, the diffuse interstellar medium, and recently, other galaxies. The observations span almost the entire range from 0.1 to 100 GHz, and employ both single­ djsh and aperture synthesis techniques. The theory of radio recombination lines has also advanced strongly, to the point where it is perhaps one of the best-understood in astro­ physics. In a parallel development, it has become possible over the last decade to study these same highly-excited atoms in the laboratory; this work provides further confirmation of the theoretical framework. However there has been continuing controversy over the astrophysical interpre­ tation of radio recombination line observations, especially regarding the role of stimulated emission. A workshop was held in Ottawa on 24-25 August, 1979, bringing together many of the active scientist...

  12. Consequences of recombination on traditional phylogenetic analysis

    DEFF Research Database (Denmark)

    Schierup, M H; Hein, J

    2000-01-01

    We investigate the shape of a phylogenetic tree reconstructed from sequences evolving under the coalescent with recombination. The motivation is that evolutionary inferences are often made from phylogenetic trees reconstructed from population data even though recombination may well occur (mt......DNA or viral sequences) or does occur (nuclear sequences). We investigate the size and direction of biases when a single tree is reconstructed ignoring recombination. Standard software (PHYLIP) was used to construct the best phylogenetic tree from sequences simulated under the coalescent with recombination....... With recombination present, the length of terminal branches and the total branch length are larger, and the time to the most recent common ancestor smaller, than for a tree reconstructed from sequences evolving with no recombination. The effects are pronounced even for small levels of recombination that may...

  13. Physiological role of growth factors and bone morphogenetic proteins in osteogenesis and bone fracture healing: а review

    Directory of Open Access Journals (Sweden)

    S. Sagalovsky

    2015-01-01

    Full Text Available The repair of large bone defects remains a major clinical orthopedic challenge. Bone regeneration and fracture healing is a complex physiological mechanisms regulated by a large number of biologically active molecules. Multiple factors regulate this cascade of molecular events, which affects different stages in the osteoblast and chondroblast lineage during such processes as migration, proliferation, chemotaxis, differentiation, inhibition, and extracellular protein synthesis. A recent review has focused on the mechanisms by which growth and differentiation factors regulate the fracture healing process. Rapid progress in skeletal cellular and molecular biology has led to identification of many signaling molecules associated with formation of skeletal tissues, including a large family of growth factors (transforming growth factor-β and bone morphogenetic proteins, fibroblast growth factor, insulin-like growth factor, vascular endothelial growth factor, platelet-derived growth factor, cytokines and interleukins. There is increasing evidence indicating that they are critical regulators of cellular proliferation, differentiation, extracellular matrix biosynthesis and bone mineralization. A clear understanding of cellular and molecular pathways involved in fracture healing is not only critical for improvement of fracture treatments, but it may also enhance further our knowledge of mechanisms involved in skeletal growth and repair, as well as mechanisms of aging. This suggests that, in the future, they may play a major role in the treatment of bone disease and fracture repair.

  14. Fine-Scale Recombination Maps of Fungal Plant Pathogens Reveal Dynamic Recombination Landscapes and Intragenic Hotspots.

    Science.gov (United States)

    Stukenbrock, Eva H; Dutheil, Julien Y

    2018-03-01

    Meiotic recombination is an important driver of evolution. Variability in the intensity of recombination across chromosomes can affect sequence composition, nucleotide variation, and rates of adaptation. In many organisms, recombination events are concentrated within short segments termed recombination hotspots. The variation in recombination rate and positions of recombination hotspot can be studied using population genomics data and statistical methods. In this study, we conducted population genomics analyses to address the evolution of recombination in two closely related fungal plant pathogens: the prominent wheat pathogen Zymoseptoria tritici and a sister species infecting wild grasses Z. ardabiliae We specifically addressed whether recombination landscapes, including hotspot positions, are conserved in the two recently diverged species and if recombination contributes to rapid evolution of pathogenicity traits. We conducted a detailed simulation analysis to assess the performance of methods of recombination rate estimation based on patterns of linkage disequilibrium, in particular in the context of high nucleotide diversity. Our analyses reveal overall high recombination rates, a lack of suppressed recombination in centromeres, and significantly lower recombination rates on chromosomes that are known to be accessory. The comparison of the recombination landscapes of the two species reveals a strong correlation of recombination rate at the megabase scale, but little correlation at smaller scales. The recombination landscapes in both pathogen species are dominated by frequent recombination hotspots across the genome including coding regions, suggesting a strong impact of recombination on gene evolution. A significant but small fraction of these hotspots colocalize between the two species, suggesting that hotspot dynamics contribute to the overall pattern of fast evolving recombination in these species. Copyright © 2018 Stukenbrock and Dutheil.

  15. Phosphoproteomic fingerprinting of epidermal growth factor signaling and anticancer drug action in human tumor cells.

    Science.gov (United States)

    Lim, Yoon-Pin; Diong, Lang-Shi; Qi, Robert; Druker, Brian J; Epstein, Richard J

    2003-12-01

    Many proteins regulating cancer cell growth are tyrosine phosphorylated. Using antiphosphotyrosine affinity chromatography, thiourea protein solubilization, two-dimensional PAGE, and mass spectrometry, we report here the characterization of the epidermal growth factor (EGF)-induced phosphoproteome in A431 human epidermoid carcinoma cells. Using this approach, more than 50 distinct tyrosine phosphoproteins are identifiable within five main clusters-cytoskeletal proteins, signaling enzymes, SH2-containing adaptors, chaperones, and focal adhesion proteins. Comparison of the phosphoproteomes induced in vitro by transforming growth factor-alpha and platelet-derived growth factor demonstrates the pathway- and cell-specific nature of the phosphoproteomes induced. Elimination of both basal and ligand-dependent phosphoproteins by cell exposure to the EGF receptor catalytic inhibitor gefitinib (Iressa, ZD1839) suggests either an autocrine growth loop or the presence of a second inhibited kinase in A431 cells. By identifying distinct patterns of phosphorylation involving novel signaling substrates, and by clarifying the mechanism of action of anticancer drugs, these findings illustrate the potential of immunoaffinity-based phosphoproteomics for guiding the discovery of new drug targets and the rational utilization of pathway-specific chemotherapies.

  16. Effects of nuclear mutations for recombination and repair functions and of caffeine on mitochondrial recombination

    International Nuclear Information System (INIS)

    Fraenkel, A.H.M.

    1974-01-01

    Studies of both prokaryotic and eukaryotic organisms indicate that pathways governing repair of damage to nuclear DNA caused by x-ray or ultraviolet irradiation overlap with those controlling recombination. Fourteen nuclear mutants of Saccharomyces cerevisiae were tested in order to determine whether these mutant genes affected mitochondrial recombination. None of the mutations studied significantly affected mitochondrial recombination. The nuclear recombination and repair pathways studied do not overlap with the nuclear pathway which controls recombination of mitochondrial DNA. A second set of experiments was designed to test the effect of caffeine on both nuclear and mitochondrial recombination in Saccharomyces cerevisiae. (U.S.)

  17. Vaccine platform recombinant measles virus.

    Science.gov (United States)

    Mühlebach, Michael D

    2017-10-01

    The classic development of vaccines is lengthy, tedious, and may not necessarily be successful as demonstrated by the case of HIV. This is especially a problem for emerging pathogens that are newly introduced into the human population and carry the inherent risk of pandemic spread in a naïve population. For such situations, a considerable number of different platform technologies are under development. These are also under development for pathogens, where directly derived vaccines are regarded as too complicated or even dangerous due to the induction of inefficient or unwanted immune responses causing considerable side-effects as for dengue virus. Among platform technologies are plasmid-based DNA vaccines, RNA replicons, single-round infectious vector particles, or replicating vaccine-based vectors encoding (a) critical antigen(s) of the target pathogens. Among the latter, recombinant measles viruses derived from vaccine strains have been tested. Measles vaccines are among the most effective and safest life-attenuated vaccines known. Therefore, the development of Schwarz-, Moraten-, or AIK-C-strain derived recombinant vaccines against a wide range of mostly viral, but also bacterial pathogens was quite straightforward. These vaccines generally induce powerful humoral and cellular immune responses in appropriate animal models, i.e., transgenic mice or non-human primates. Also in the recent first clinical phase I trial, the results have been quite encouraging. The trial indicated the expected safety and efficacy also in human patients, interestingly independent from the level of prevalent anti-measles immunity before the trial. Thereby, recombinant measles vaccines expressing additional antigens are a promising platform for future vaccines.

  18. Characterization of the growth of murine fibroblasts that express human insulin receptors. II. Interaction of insulin with other growth factors

    International Nuclear Information System (INIS)

    Randazzo, P.A.; Jarett, L.

    1990-01-01

    The effects of insulin-like growth factor-1 (IGF-1), epidermal growth factor (EGF), platelet-derived growth factor (PDGF), and insulin on DNA synthesis were studied in murine fibroblasts transfected with an expression vector containing human insulin receptor cDNA (NIH 3T3/HIR) and the parental NIH 3T3 cells. In NIH 3T3/HIR cells, individual growth factors in serum-free medium stimulated DNA synthesis with the following relative efficacies: insulin greater than or equal to 10% fetal calf serum greater than PDGF greater than IGF-1 much greater than EGF. In comparison, the relative efficacies of these factors in stimulating DNA synthesis by NIH 3T3 cells were 10% fetal calf serum greater than PDGF greater than EGF much greater than IGF-1 = insulin. In NIH 3T3/HIR cells, EGF was synergistic with 1-10 ng/ml insulin but not with 100 ng/ml insulin or more. Synergy of PDGF or IGF-1 with insulin was not detected. In the parental NIH 3T3 cells, insulin and IGF-1 were found to be synergistic with EGF (1 ng/ml), PDGF (100 ng/ml), and PDGF plus EGF. In NIH 3T3/HIR cells, the lack of interaction of insulin with other growth factors was also observed when the percentage of cells synthesizing DNA was examined. Despite insulin's inducing only 60% of NIH 3T3/HIR cells to incorporate thymidine, addition of PDGF, EGF, or PDGF plus EGF had no further effect. In contrast, combinations of growth factors resulted in 95% of the parental NIH 3T3 cells synthesizing DNA. The independence of insulin-stimulated DNA synthesis from other mitogens in the NIH 3T3/HIR cells is atypical for progression factor-stimulated DNA synthesis and is thought to be partly the result of insulin receptor expression in an inappropriate context or quantity

  19. CRMAGE: CRISPR Optimized MAGE Recombineering

    DEFF Research Database (Denmark)

    Ronda, Carlotta; Pedersen, Lasse Ebdrup; Sommer, Morten Otto Alexander

    2016-01-01

    A bottleneck in metabolic engineering and systems biology approaches is the lack of efficient genome engineering technologies. Here, we combine CRISPR/Cas9 and λ Red recombineering based MAGE technology (CRMAGE) to create a highly efficient and fast method for genome engineering of Escherichia coli...... that are assembled by a USER-cloning approach enabling quick and cost efficient gRNA replacement. CRMAGE furthermore utilizes CRISPR/Cas9 for efficient plasmid curing, thereby enabling multiple engineering rounds per day. To facilitate the design process, a web-based tool was developed to predict both the λ Red...

  20. Atomic excitation and recombination in external fields

    International Nuclear Information System (INIS)

    Nayfeh, M.H.; Clark, C.W.

    1985-01-01

    This volume offers a timely look at Rydberg states of atoms in external fields and dielectronic recombination. Each topic provides authoritative coverage, presents a fresh account of a flourishing field of current atomic physics and introduces new opportunities for discovery and development. Topics considered include electron-atom scattering in external fields; observations of regular and irregular motion as exemplified by the quadratic zeeman effect and other systems; Rydberg atoms in external fields and the Coulomb geometry; crossed-field effects in the absorption spectrum of lithium in a magnetic field; precise studies of static electric field ionization; widths and shapes of stark resonances in sodium above the saddle point; studies of electric field effects and barium autoionizing resonances; autoionization and dielectronic recombination in plasma electric microfields; dielectronic recombination measurements on multicharged ions; merged beam studies of dielectronic recombination; Rydberg atoms and dielectronic recombination in astrophysics; and observations on dielectronic recombination

  1. Biological characterization of bovine herpesvirus 1 recombinants possessing the vaccine glycoprotein E negative phenotype.

    Science.gov (United States)

    Muylkens, Benoît; Meurens, François; Schynts, Frédéric; de Fays, Katalin; Pourchet, Aldo; Thiry, Julien; Vanderplasschen, Alain; Antoine, Nadine; Thiry, Etienne

    2006-03-31

    Intramolecular recombination is a frequent event during the replication cycle of bovine herpesvirus 1 (BoHV-1). Recombinant viruses frequently arise and survive in cattle after concomitant nasal infections with two BoHV-1 mutants. The consequences of this process, related to herpesvirus evolution, have to be assessed in the context of large use of live marker vaccines based on glycoprotein E (gE) gene deletion. In natural conditions, double nasal infections by vaccine and wild-type strains are likely to occur. This situation might generate virulent recombinant viruses inducing a serological response indistinguishable from the vaccine one. This question was addressed by generating in vitro BoHV-1 recombinants deleted in the gE gene from seven wild-type BoHV-1 strains and one mutant strain deleted in the genes encoding gC and gE. In vitro growth properties were assessed by virus production, one step growth kinetics and plaque size assay. Heterogeneity in the biological properties was shown among the investigated recombinant viruses. The results demonstrated that some recombinants, in spite of their gE minus phenotype, have biological characteristics close to wild-type BoHV-1.

  2. Overexpressed of RAD51 suppresses recombination defects: a possible mechanism to reverse genomic instability

    Energy Technology Data Exchange (ETDEWEB)

    Schild, David; Wiese, Claudia

    2009-10-15

    RAD51, a key protein in the homologous recombinational DNA repair (HRR) pathway, is the major strand-transferase required for mitotic recombination. An important early step in HRR is the formation of single-stranded DNA (ss-DNA) coated by RPA (a ss-DNA binding protein). Displacement of RPA by RAD51 is highly regulated and facilitated by a number of different proteins known as the 'recombination mediators'. To assist these recombination mediators, a second group of proteins also is required and we are defining these proteins here as 'recombination co-mediators'. Defects in either recombination mediators or comediators, including BRCA1 and BRCA2, lead to impaired HRR that can genetically be complemented for (i.e. suppressed) by overexpression of RAD51. Defects in HRR have long been known to contribute to genomic instability leading to tumor development. Since genomic instability also slows cell growth, precancerous cells presumably require genomic restabilization to gain a growth advantage. RAD51 is overexpressed in many tumors, and therefore, we hypothesize that the complementing ability of elevated levels of RAD51 in tumors with initial HRR defects limits genomic instability during carcinogenic progression. Of particular interest, this model may also help explain the high frequency of TP53 mutations in human cancers, since wild-type p53 represses RAD51.

  3. Density dependence of dielectronic recombination in selenium

    International Nuclear Information System (INIS)

    Hagelstein, P.L.; Rosen, M.D.; Jacobs, V.L.

    1986-01-01

    Dielectronic recombination has been found to be the dominant recombination process in the determination of the ionization balance of selenium near the Ne-like sequence under conditions relevant to the exploding-foil EUV laser plasmas. The dielectronic recombination process tends to populate excited levels, and these levels in turn are more susceptible to subsequent excitation and ionization than are the ground-state ions. If one defines an effective recombination rate which includes, in addition to the primary recombination, the subsequent excitation and ionization of the additional excited-state population due to the primary recombination, then this effective recombination rate can be density-sensitive at relatively low electron density. We present results for this effective dielectronic recombination rate at an electron density of 3 x 10/sup 20/ electrons/cm 3 for recombination from Ne-like to Na-like selenium and from F-like to Ne-like selenium. In the former case, the effective recombination rate coefficient is found to be 1.8 x 10/sup -11/ cm 3 /sec at 1.0 keV, which is to be compared with the zero-density value of 2.8 x 10/sup -11/ cm 3 /sec. In the latter case (F-like to Ne-like), the effective recombination rate coefficient is found to be 1.3 x 10/sup -11/ cm 3 /sec, which is substantially reduced from the zero-density result of 3.3 x 10/sup -11/ cm 3 /sec. We have examined the effects of dielectronic recombination on the laser gain of the dominant Ne-like 3p-3s transitions and have compared our results with those presented by Whitten et al. [Phys. Rev. A 33, 2171 (1986)

  4. Rapid purification of recombinant histones.

    Science.gov (United States)

    Klinker, Henrike; Haas, Caroline; Harrer, Nadine; Becker, Peter B; Mueller-Planitz, Felix

    2014-01-01

    The development of methods to assemble nucleosomes from recombinant histones decades ago has transformed chromatin research. Nevertheless, nucleosome reconstitution remains time consuming to this day, not least because the four individual histones must be purified first. Here, we present a streamlined purification protocol of recombinant histones from bacteria. We termed this method "rapid histone purification" (RHP) as it circumvents isolation of inclusion bodies and thereby cuts out the most time-consuming step of traditional purification protocols. Instead of inclusion body isolation, whole cell extracts are prepared under strongly denaturing conditions that directly solubilize inclusion bodies. By ion exchange chromatography, the histones are purified from the extracts. The protocol has been successfully applied to all four canonical Drosophila and human histones. RHP histones and histones that were purified from isolated inclusion bodies had similar purities. The different purification strategies also did not impact the quality of octamers reconstituted from these histones. We expect that the RHP protocol can be readily applied to the purification of canonical histones from other species as well as the numerous histone variants.

  5. The extent and importance of intragenic recombination

    Directory of Open Access Journals (Sweden)

    de Silva Eric

    2004-11-01

    Full Text Available Abstract We have studied the recombination rate behaviour of a set of 140 genes which were investigated for their potential importance in inflammatory disease. Each gene was extensively sequenced in 24 individuals of African descent and 23 individuals of European descent, and the recombination process was studied separately in the two population samples. The results obtained from the two populations were highly correlated, suggesting that demographic bias does not affect our population genetic estimation procedure. We found evidence that levels of recombination correlate with levels of nucleotide diversity. High marker density allowed us to study recombination rate variation on a very fine spatial scale. We found that about 40 per cent of genes showed evidence of uniform recombination, while approximately 12 per cent of genes carried distinct signatures of recombination hotspots. On studying the locations of these hotspots, we found that they are not always confined to introns but can also stretch across exons. An investigation of the protein products of these genes suggested that recombination hotspots can sometimes separate exons belonging to different protein domains; however, this occurs much less frequently than might be expected based on evolutionary studies into the origins of recombination. This suggests that evolutionary analysis of the recombination process is greatly aided by considering nucleotide sequences and protein products jointly.

  6. Effects of recombinant human nerve growth factor on cervical cancer

    African Journals Online (AJOL)

    Jane

    2011-07-25

    Jul 25, 2011 ... thought to be an important determinant of cell survival and death (Arrighi et al., ... whereas the action was related to the concentration of. rhNGF. ..... (1999). Fusion of ETV6 to neurotrophin-3 receptor TRKC in acute myeloid ...

  7. Meningioma growth and interferon beta-1b treated multiple sclerosis: coincidence or relationship?

    Energy Technology Data Exchange (ETDEWEB)

    Drevelegas, A.; Xinou, E. [AHEPA University Hospital, Aristotele University School of Medicine, Department of Radiology, Thessaloniki (Greece); Karacostas, D.; Parissis, D.; Milonas, I. [AHEPA University Hospital, Aristotele University School of Medicine, B' Department of Neurology, Thessaloniki (Greece); Karkavelas, G. [AHEPA University Hospital, Aristotele University School of Medicine, Laboratory of Pathology, Thessaloniki (Greece)

    2005-07-01

    Although the coincidence of multiple sclerosis (MS) and central nervous system (CNS) tumors has been reported in over 30 cases in English literature, meningioma growth was associated with interferon-beta (INF-b) treated MS only in two of them. We report the case of a 19-year-old woman with clinically possible, laboratory supported MS, and a concomitant right intraventricular tumor with magnetic resonance imaging (MRI) characteristics consistent with meningioma (similar signal with grey matter on T1 and T2-weighted images and homogenous, intense enhancement). Two years after initiation of INF-b treatment, follow-up brain MRI revealed enlargement of the intraventricular mass and relative increase in the number of white matter lesions without significant clinical deterioration. She underwent almost total resection of the mass and histology confirmed the diagnosis of papillary meningioma. Based on the immunohistochemistry results, we speculate that INF-b resulted in meningioma growth by enhancing platelet derived growth factor (PDGF) receptors or/and down-regulating transforming growth factor receptors on the tumor itself. (orig.)

  8. Meningioma growth and interferon beta-1b treated multiple sclerosis: coincidence or relationship?

    International Nuclear Information System (INIS)

    Drevelegas, A.; Xinou, E.; Karacostas, D.; Parissis, D.; Milonas, I.; Karkavelas, G.

    2005-01-01

    Although the coincidence of multiple sclerosis (MS) and central nervous system (CNS) tumors has been reported in over 30 cases in English literature, meningioma growth was associated with interferon-beta (INF-b) treated MS only in two of them. We report the case of a 19-year-old woman with clinically possible, laboratory supported MS, and a concomitant right intraventricular tumor with magnetic resonance imaging (MRI) characteristics consistent with meningioma (similar signal with grey matter on T1 and T2-weighted images and homogenous, intense enhancement). Two years after initiation of INF-b treatment, follow-up brain MRI revealed enlargement of the intraventricular mass and relative increase in the number of white matter lesions without significant clinical deterioration. She underwent almost total resection of the mass and histology confirmed the diagnosis of papillary meningioma. Based on the immunohistochemistry results, we speculate that INF-b resulted in meningioma growth by enhancing platelet derived growth factor (PDGF) receptors or/and down-regulating transforming growth factor receptors on the tumor itself. (orig.)

  9. Epstein-Barr virus (EBV) recombinants: use of positive selection markers to rescue mutants in EBV-negative B-lymphoma cells.

    OpenAIRE

    Wang, F; Marchini, A; Kieff, E

    1991-01-01

    The objective of these experiments was to develop strategies for creation and identification of recombinant mutant Epstein-Barr viruses (EBV). EBV recombinant molecular genetics has been limited to mutations within a short DNA segment deleted from a nontransforming EBV and an underlying strategy which relies on growth transformation of primary B lymphocytes for identification of recombinants. Thus, mutations outside the deletion or mutations which affect transformation cannot be easily recove...

  10. Recombining without Hotspots: A Comprehensive Evolutionary Portrait of Recombination in Two Closely Related Species of Drosophila

    Science.gov (United States)

    Smukowski Heil, Caiti S.; Ellison, Chris; Dubin, Matthew; Noor, Mohamed A.F.

    2015-01-01

    Meiotic recombination rate varies across the genome within and between individuals, populations, and species in virtually all taxa studied. In almost every species, this variation takes the form of discrete recombination hotspots, determined in some mammals by a protein called PRDM9. Hotspots and their determinants have a profound effect on the genomic landscape, and share certain features that extend across the tree of life. Drosophila, in contrast, are anomalous in their absence of hotspots, PRDM9, and other species-specific differences in the determination of recombination. To better understand the evolution of meiosis and general patterns of recombination across diverse taxa, we present a truly comprehensive portrait of recombination across time, combining recently published cross-based contemporary recombination estimates from each of two sister species with newly obtained linkage-disequilibrium-based historic estimates of recombination from both of these species. Using Drosophila pseudoobscura and Drosophila miranda as a model system, we compare recombination rate between species at multiple scales, and we suggest that Drosophila replicate the pattern seen in human–chimpanzee in which recombination rate is conserved at broad scales. We also find evidence of a species-wide recombination modifier(s), resulting in both a present and historic genome-wide elevation of recombination rates in D. miranda, and identify broad scale effects on recombination from the presence of an inversion. Finally, we reveal an unprecedented view of the distribution of recombination in D. pseudoobscura, illustrating patterns of linked selection and where recombination is taking place. Overall, by combining these estimation approaches, we highlight key similarities and differences in recombination between Drosophila and other organisms. PMID:26430062

  11. Recombining without Hotspots: A Comprehensive Evolutionary Portrait of Recombination in Two Closely Related Species of Drosophila.

    Science.gov (United States)

    Smukowski Heil, Caiti S; Ellison, Chris; Dubin, Matthew; Noor, Mohamed A F

    2015-10-01

    Meiotic recombination rate varies across the genome within and between individuals, populations, and species in virtually all taxa studied. In almost every species, this variation takes the form of discrete recombination hotspots, determined in some mammals by a protein called PRDM9. Hotspots and their determinants have a profound effect on the genomic landscape, and share certain features that extend across the tree of life. Drosophila, in contrast, are anomalous in their absence of hotspots, PRDM9, and other species-specific differences in the determination of recombination. To better understand the evolution of meiosis and general patterns of recombination across diverse taxa, we present a truly comprehensive portrait of recombination across time, combining recently published cross-based contemporary recombination estimates from each of two sister species with newly obtained linkage-disequilibrium-based historic estimates of recombination from both of these species. Using Drosophila pseudoobscura and Drosophila miranda as a model system, we compare recombination rate between species at multiple scales, and we suggest that Drosophila replicate the pattern seen in human-chimpanzee in which recombination rate is conserved at broad scales. We also find evidence of a species-wide recombination modifier(s), resulting in both a present and historic genome-wide elevation of recombination rates in D. miranda, and identify broad scale effects on recombination from the presence of an inversion. Finally, we reveal an unprecedented view of the distribution of recombination in D. pseudoobscura, illustrating patterns of linked selection and where recombination is taking place. Overall, by combining these estimation approaches, we highlight key similarities and differences in recombination between Drosophila and other organisms. © The Author(s) 2015. Published by Oxford University Press on behalf of the Society for Molecular Biology and Evolution.

  12. Fundamental Studies of Recombinant Hydrogenases

    Energy Technology Data Exchange (ETDEWEB)

    Adams, Michael W. [Univ. of Georgia, Athens, GA (United States)

    2014-01-25

    This research addressed the long term goals of understanding the assembly and organization of hydrogenase enzymes, of reducing them in size and complexity, of determining structure/function relationships, including energy conservation via charge separation across membranes, and in screening for novel H2 catalysts. A key overall goal of the proposed research was to define and characterize minimal hydrogenases that are produced in high yields and are oxygen-resistant. Remarkably, in spite of decades of research carried out on hydrogenases, it is not possible to readily manipulate or design the enzyme using molecular biology approaches since a recombinant form produced in a suitable host is not available. Such resources are essential if we are to understand what constitutes a “minimal” hydrogenase and design such catalysts with certain properties, such as resistance to oxygen, extreme stability and specificity for a given electron donor. The model system for our studies is Pyrococcus furiosus, a hyperthermophile that grows optimally at 100°C, which contains three different nickel-iron [NiFe-] containing hydrogenases. Hydrogenases I and II are cytoplasmic while the other, MBH, is an integral membrane protein that functions to both evolve H2 and pump protons. Three important breakthroughs were made during the funding period with P. furiosus soluble hydrogenase I (SHI). First, we produced an active recombinant form of SHI in E. coli by the co-expression of sixteen genes using anaerobically-induced promoters. Second, we genetically-engineered P. furiosus to overexpress SHI by an order of magnitude compared to the wild type strain. Third, we generated the first ‘minimal’ form of SHI, one that contained two rather than four subunits. This dimeric form was stable and active, and directly interacted with a pyruvate-oxidizing enzyme with any intermediate electron carrier. The research resulted in five peer-reviewed publications.

  13. Electron-ion recombination at low energy

    International Nuclear Information System (INIS)

    Andersen, L.H.

    1993-01-01

    The work is based on results obtained with a merged-beams experiment. A beam of electronics with a well characterized density and energy distribution was merged with a fast, monoenergetic ion beam. Results have been obtained for radiative recombination and dielectronic recombination at low relative energies (0 to ∼70eV). The obtained energy resolution was improved by about a factor of 30. High vacuum technology was used to suppress interactions with electrons from the environments. The velocity distribution of the electron beam was determined. State-selective dielectronic-recombination measurements were performable. Recombination processes were studied. The theoretical background for radiative recombination and Kramers' theory are reviewed. The quantum mechanical result and its relation to the semiclassical theory is discussed. Radiative recombination was also measured with several different non-bare ions, and the applicability of the semiclassical theory to non-bare ions was investigated. The use of an effective charge is discussed. For dielectronic recombination, the standard theoretical approach in the isolated resonance and independent-processes approximation is debated. The applicability of this method was tested. The theory was able to reproduce most of the experimental data except when the recombination process was sensitive to couplings between different electronic configurations. The influence of external perturbing electrostatic fields is discussed. (AB) (31 refs.)

  14. Recombination rate plasticity: revealing mechanisms by design

    Science.gov (United States)

    Sefick, Stephen; Rushton, Chase

    2017-01-01

    For over a century, scientists have known that meiotic recombination rates can vary considerably among individuals, and that environmental conditions can modify recombination rates relative to the background. A variety of external and intrinsic factors such as temperature, age, sex and starvation can elicit ‘plastic’ responses in recombination rate. The influence of recombination rate plasticity on genetic diversity of the next generation has interesting and important implications for how populations evolve. Further, many questions remain regarding the mechanisms and molecular processes that contribute to recombination rate plasticity. Here, we review 100 years of experimental work on recombination rate plasticity conducted in Drosophila melanogaster. We categorize this work into four major classes of experimental designs, which we describe via classic studies in D. melanogaster. Based on these studies, we highlight molecular mechanisms that are supported by experimental results and relate these findings to studies in other systems. We synthesize lessons learned from this model system into experimental guidelines for using recent advances in genotyping technologies, to study recombination rate plasticity in non-model organisms. Specifically, we recommend (1) using fine-scale genome-wide markers, (2) collecting time-course data, (3) including crossover distribution measurements, and (4) using mixed effects models to analyse results. To illustrate this approach, we present an application adhering to these guidelines from empirical work we conducted in Drosophila pseudoobscura. This article is part of the themed issue ‘Evolutionary causes and consequences of recombination rate variation in sexual organisms’. PMID:29109222

  15. Electron-ion recombination in merged beams

    International Nuclear Information System (INIS)

    Wolf, A.; Habs, D.; Lampert, A.; Neumann, R.; Schramm, U.; Schuessler, T.; Schwalm, D.

    1993-01-01

    Detailed studies of recombination processes between electrons and highly charged ions have become possible by recent improvements of merged-beams experiments. We discuss in particular measurements with stored cooled ion beams at the Test Storage Ring (TSR) in Heidelberg. The cross section of dielectronic recombination was measured with high energy resolution for few-electron systems up to the nuclear charge of Cu at a relative energy up to 2.6 keV. At low energy (∼0.1 eV) total recombination rates of several ions were measured and compared with calculated radiative recombination rates. Laser-stimulated recombination of protons and of C 6+ ions was investigated as a function of the photon energy using visible radiation. Both the total recombination rates and the stimulated recombination spectra indicate that in spite of the short interaction time in merged beams, also collisional capture of electrons into weakly bound levels (related to three-body recombination) could be important

  16. Electronic recombination in some physics problems

    International Nuclear Information System (INIS)

    Guzman, O.

    1988-01-01

    This work is related to calculations of electronic recombination rates, as a function of electronic density, electronic temperature, and ion nuclear charge. Recombination times can be calculated and compared to cooling time, in cooling processes of ion beans by electrons from storage rings. (A.C.A.S.) [pt

  17. Generation of Modified Pestiviruses by Targeted Recombination

    DEFF Research Database (Denmark)

    Rasmussen, Thomas Bruun; Friis, Martin Barfred; Risager, Peter Christian

    involves targeted modification of viral cDNA genomes, cloned within BACs, by Red/ET recombination-mediated mutagenesis in E.coli DH10B cells. Using recombination-mediated mutagenesis for the targeted design, the work can be expedited and focused in principal on any sequence within the viral genome...

  18. Cell biology of homologous recombination in yeast

    DEFF Research Database (Denmark)

    Eckert-Boulet, Nadine Valerie; Rothstein, Rodney; Lisby, Michael

    2011-01-01

    Homologous recombination is an important pathway for error-free repair of DNA lesions, such as single- and double-strand breaks, and for rescue of collapsed replication forks. Here, we describe protocols for live cell imaging of single-lesion recombination events in the yeast Saccharomyces...

  19. Recombinant Vaccinia Virus: Immunization against Multiple Pathogens

    Science.gov (United States)

    Perkus, Marion E.; Piccini, Antonia; Lipinskas, Bernard R.; Paoletti, Enzo

    1985-09-01

    The coding sequences for the hepatitis B virus surface antigen, the herpes simplex virus glycoprotein D, and the influenza virus hemagglutinin were inserted into a single vaccinia virus genome. Rabbits inoculated intravenously or intradermally with this polyvalent vaccinia virus recombinant produced antibodies reactive to all three authentic foreign antigens. In addition, the feasibility of multiple rounds of vaccination with recombinant vaccinia virus was demonstrated.

  20. Recombinant organisms for production of industrial products

    OpenAIRE

    Adrio, Jose-Luis; Demain, Arnold L

    2009-01-01

    A revolution in industrial microbiology was sparked by the discoveries of ther double-stranded structure of DNA and the development of recombinant DNA technology. Traditional industrial microbiology was merged with molecular biology to yield improved recombinant processes for the industrial production of primary and secondary metabolites, protein biopharmaceuticals and industrial enzymes. Novel genetic techniques such as metabolic engineering, combinatorial biosynthesis and molecular breeding...

  1. Radiative and interfacial recombination in CdTe heterostructures

    Energy Technology Data Exchange (ETDEWEB)

    Swartz, C. H., E-mail: craig.swartz@txstate.edu; Edirisooriya, M.; LeBlanc, E. G.; Noriega, O. C.; Jayathilaka, P. A. R. D.; Ogedengbe, O. S.; Hancock, B. L.; Holtz, M.; Myers, T. H. [Materials Science, Engineering, and Commercialization Program, Texas State University, 601 University Dr., San Marcos, Texas 78666 (United States); Zaunbrecher, K. N. [National Renewable Energy Laboratory, 15013 Denver West Parkway, Mississippi RSF200, Golden, Colorado 80401 (United States)

    2014-12-01

    Double heterostructures (DH) were produced consisting of a CdTe film between two wide band gap barriers of CdMgTe alloy. A combined method was developed to quantify radiative and non-radiative recombination rates by examining the dependence of photoluminescence (PL) on both excitation intensity and time. The measured PL characteristics, and the interface state density extracted by modeling, indicate that the radiative efficiency of CdMgTe/CdTe DHs is comparable to that of AlGaAs/GaAs DHs, with interface state densities in the low 10{sup 10 }cm{sup −2} and carrier lifetimes as long as 240 ns. The radiative recombination coefficient of CdTe is found to be near 10{sup −10} cm{sup 3}s{sup −1}. CdTe film growth on bulk CdTe substrates resulted in a homoepitaxial interface layer with a high non-radiative recombination rate.

  2. Biomimetic calcium phosphate coatings on recombinant spider silk fibres

    Energy Technology Data Exchange (ETDEWEB)

    Yang Liang; Habibovic, Pamela; Van Blitterswijk, Clemens A [Department of Tissue Regeneration, University of Twente, PO Box 217, 7500 AE Enschede (Netherlands); Hedhammar, My; Johansson, Jan [Department of Anatomy, Physiology and Biochemistry, Swedish University of Agricultural Sciences, the Biomedical Centre, Box 575, 751 23 Uppsala (Sweden); Blom, Tobias; Leifer, Klaus [Department of Engineering Sciences, Uppsala University, Box 534, S-751 21 Uppsala (Sweden)

    2010-08-01

    Calcium phosphate ceramic coatings, applied on surfaces of metallic and polymeric biomaterials, can improve their performance in bone repair and regeneration. Spider silk is biocompatible, strong and elastic, and hence an attractive biomaterial for applications in connective tissue repair. Recently, artificial spider silk, with mechanical and structural characteristics similar to those of native spider silk, has been produced from recombinant minispidroins. In the present study, supersaturated simulated body fluid was used to deposit calcium phosphate coatings on recombinant spider silk fibres. The mineralization process was followed in time using scanning electron microscopy equipped with an energy dispersive x-ray (EDX) detector and Raman spectroscope. Focused ion beam technology was used to produce a cross section of a coated fibre, which was further analysed by EDX. Preliminary in vitro experiments using a culture of bone marrow-derived human mesenchymal stem cells (hMSCs) on coated fibres were also performed. This study showed that recombinant spider silk fibres were successfully coated with a homogeneous and thick crystalline calcium phosphate layer. In the course of the mineralization process from modified simulated body fluid, sodium chloride crystals were first deposited on the silk surface, followed by the deposition of a calcium phosphate layer. The coated silk fibres supported the attachment and growth of hMSCs.

  3. Biomimetic calcium phosphate coatings on recombinant spider silk fibres

    International Nuclear Information System (INIS)

    Yang Liang; Habibovic, Pamela; Van Blitterswijk, Clemens A; Hedhammar, My; Johansson, Jan; Blom, Tobias; Leifer, Klaus

    2010-01-01

    Calcium phosphate ceramic coatings, applied on surfaces of metallic and polymeric biomaterials, can improve their performance in bone repair and regeneration. Spider silk is biocompatible, strong and elastic, and hence an attractive biomaterial for applications in connective tissue repair. Recently, artificial spider silk, with mechanical and structural characteristics similar to those of native spider silk, has been produced from recombinant minispidroins. In the present study, supersaturated simulated body fluid was used to deposit calcium phosphate coatings on recombinant spider silk fibres. The mineralization process was followed in time using scanning electron microscopy equipped with an energy dispersive x-ray (EDX) detector and Raman spectroscope. Focused ion beam technology was used to produce a cross section of a coated fibre, which was further analysed by EDX. Preliminary in vitro experiments using a culture of bone marrow-derived human mesenchymal stem cells (hMSCs) on coated fibres were also performed. This study showed that recombinant spider silk fibres were successfully coated with a homogeneous and thick crystalline calcium phosphate layer. In the course of the mineralization process from modified simulated body fluid, sodium chloride crystals were first deposited on the silk surface, followed by the deposition of a calcium phosphate layer. The coated silk fibres supported the attachment and growth of hMSCs.

  4. Molecular requirements for radiation-activated recombination

    International Nuclear Information System (INIS)

    Stevens, Craig W.; Zeng Ming; Stamato, Thomas; Cerniglia, George

    1997-01-01

    Purpose/Objective: The major stumbling block to successful gene therapy today is poor gene transfer. We hypothesized that ionizing radiation might activate cellular recombination, and so improve stable gene transfer. We further hypothesized that known DNA-damage-repair proteins might also be important in radiation-activated recombination. Materials and Methods: The effect of irradiation on stable gene transfer efficiency was determined in human (A549 and 39F) and rodent (NIH/3T3) cell lines. Continuous low dose rate and multiple radiation fractions were also tested. Nuclear extracts were made and the effect of irradiation on inter-plasmid recombination/ligation determined. Multiple DNA damage-repair deficient cell lines were tested for radiation-activated recombination. Results: A significant radiation dose-dependent improvement in stable plasmid transfection (by as much as 1300 fold) is demonstrated in neoplastic and primary cells. An improvement in transient plasmid transfection is also seen, with as much as 85% of cells transiently expressing b-galactosidase (20-50 fold improvement). Stable transfection is only improved for linearized or nicked plasmids. Cells have improved gene transfer for at least 96 hours after irradiation. Both fractionated and continuous low dose rate irradiation are effective at improving stable gene transfer in mammalian cells, thus making relatively high radiation dose delivery clinically feasible. Inter-plasmid recombination is radiation dose dependent in nuclear extract assays, and the type of overhang (3', 5' or blunt end) significantly affects recombination efficiency and the type of product. The most common end-joining activity involves filling-in of the overhang followed by blunt end ligation. Adenovirus is a linear, double stranded DNA virus. We demonstrate that adenoviral infection efficiency is increased by irradiation. The duration of transgene expression is lengthened because the virus integrates with high efficiency (∼10

  5. Effects of UV radiation on genetic recombination

    International Nuclear Information System (INIS)

    Vlahovic, K.; Zahradka, D.; Petranovic, M.; Petranovic, D.

    1996-01-01

    We have used the model consisting of Escherichia coli cells and l phage to study the effects of UV radiation on genetic recombination. We found two radiation induced processes that reduce or inhibit genetic recombination. One such process leads to the inability of prophage to excise itself from the irradiated bacterial chromosome by the site-specific recombination. The other process was shown to inhibit a type of general recombination by which the prophage transfers one of its genetic markers to the infecting homologous phage. Loss of the prophage ability to take part in both site-specific and general recombination was shown to develop in recB + but not in recB cells. From this we infer that the loss of prophage recombinogenicity in irradiated cells is a consequence of one process in which RecBCD enzyme (the product of recB, recC and recD genes) plays an essential role. (author)

  6. Containment air circulation for optimal hydrogen recombination

    International Nuclear Information System (INIS)

    Spinks, N.; Krause, M.

    1997-01-01

    An accepted first-line defense for hydrogen mitigation is to design for the hydrogen to be rapidly mixed with the containment atmosphere and diluted to below flammability concentrations. Then, as hydrogen continues to be produced in the longer term, recombiners can be used to remove hydrogen: recombiners can be located in forced-air ducts or passive recombiners can be distributed within containment and the heat of recombination used to promote local air circulation. However, this principle does not eliminate the possibility of high hydrogen concentrations at locations removed from the recombiners. An improvement on this strategy is to arrange for a specific, buoyancy-driven, overall circulation of the containment atmosphere such that the recombiners can be located within the recirculation flow, immediately downstream of the hydrogen source. This would make the mixing process more predictable and solve the mass-transfer problem associated with distributed recombiners. Ideally, the recombiners would be located just above the hydrogen source so that the heat of recombination would assist the overall circulation. In this way, the hydrogen would be removed as close as possible to the source, thereby minimizing the amount of hydrogen immediately downstream of the source and reducing the hydrogen concentration to acceptable levels at other locations. Such a strategy requires the containment volume to be divided into an upflow path, past the hydrogen source and the recombiner, and a downflow path to complete the circuit. The flow could be generated actively using fans or passively using buoyancy forces arising from the difference in density of gases in the upfiow and downflow paths; the gases in the downflow path being cooled at an elevated heat sink. (author)

  7. The unconventional xer recombination machinery of Streptococci/Lactococci

    NARCIS (Netherlands)

    Le Bourgeois, Pascal; Bugarel, Marie; Campo, Nathalie; Daveran-Mingot, Marie-Line; Labonte, Jessica; Lanfranchi, Daniel; Lautier, Thomas; Pages, Carine; Ritzenthaler, Paul

    Homologous recombination between circular sister chromosomes during DNA replication in bacteria can generate chromosome dimers that must be resolved into monomers prior to cell division. In Escherichia coli, dimer resolution is achieved by site-specific recombination, Xer recombination, involving

  8. Improved expression of recombinant plant-made hEGF.

    Science.gov (United States)

    Thomas, David Rhys; Walmsley, Amanda Maree

    2014-11-01

    The yield of recombinant hEGF was increased approximately tenfold through a range of optimisations. Further, the recombinant protein was found to have biological activity comparable to commercial hEGF. Human epidermal growth factor (hEGF) is a powerful mitogen that can enhance the healing of a wide range of injuries, including burns, cuts, diabetic ulcers and gastric ulcers. However, despite its clinical value, hEGF is only consistently used for the treatment of chronic diabetic ulcers due to its high cost. In this study, hEGF was transiently expressed in Nicotiana benthamiana plants and targeted to the apoplast, ER and vacuole. Several other approaches were also included in a stepwise fashion to identify the optimal conditions for the expression of recombinant hEGF. Expression was found to be highest in the vacuole, while targeting hEGF to the ER caused a decrease in total soluble protein (TSP). Using a codon optimised sequence was found to increase vacuolar targeted hEGF yield by ~34 %, while it was unable to increase the yield of ER targeted hEGF. The use of the P19 silencing inhibitor was able to further increase expression by over threefold, and using 5-week-old plants significantly increased expression compared to 4- or 6-week-old-plants. The combined effect of these optimisations increased expression tenfold over the initial apoplast targeted construct to an average yield of 6.24 % of TSP. The plant-made hEGF was then shown to be equivalent to commercial E. coli derived hEGF in its ability to promote the proliferation of mouse keratinocytes. This study supports the potential for plants to be used for the commercial production of hEGF, and identifies a potential limitation for the further improvement of recombinant protein yields.

  9. Growth factor and proteinase profile of Vivostat® platelet-rich fibrin linked to tissue repair.

    Science.gov (United States)

    Agren, M S; Rasmussen, K; Pakkenberg, B; Jørgensen, B

    2014-07-01

    Autologous platelet-rich fibrin (PRF(®)) is prepared by the automatic Vivostat(®) system. Conflicting results with Vivostat PRF in acute wound healing prompted us to examine its cellular and biomolecular composition. Specifically, platelets, selected growth factors and matrix metalloproteinase (MMP)-9 were quantified using novel analytical methods. Ten healthy non-thrombocytopenic volunteers donated blood for generation of intermediate fibrin-I and final PRF. Anticoagulated whole blood and serum procured in parallel served as baseline controls. Leucocyte, erythrocyte and platelet counts in whole blood and fibrin-I were determined by automated haematology analyser. Platelet concentration in PRF was quantified manually by stereologic analysis of Giemsa-stained tissue sections, and the total content of five growth factors and MMP-9 by enzyme-linked immunosorbent assays. The number of leucocytes and erythrocytes was reduced (P platelets increased (P fibrin-I versus whole blood. PRF contained 982 ± 206 × 10(9) platelets/l representing 3·9-fold (P platelet-derived growth factor (PDGF)-AB [2·5-fold, P PDGF-BB [1·6-fold, P vascular endothelial growth factor > basic fibroblast growth factor [75-fold, P platelet enrichment and biomolecular constituents may guide clinicians in their optimal use of Vivostat PRF for tissue regenerative applications. © 2013 International Society of Blood Transfusion.

  10. Transforming growth factor type β can act as a potent competence factor for AKR-2B cells

    International Nuclear Information System (INIS)

    Goustin, A.S.; Nuttall, G.A.; Leof, E.B.; Ranganathan, G.; Moses, H.L.

    1987-01-01

    Transforming growth factor type β (TGFβ) is a pleiotropic regulator of cell growth with specific high-affinity cell-surface receptors on a large number of cells; its mechanism of action, however, is poorly defined. In this report, the authors utilized the mouse fibroblast line AKR-2B to explore the question of the temporal requirements during the cell cycle in regard to both the growth inhibitory and the growth stimulatory action of TGFβ. The results indicate that AKR-2B cells are most sensitive to the inhibitory action of TGFβ during early to mid-G 1 . In addition, TGFβ need be present only briefly in order to exert its inhibitory effect on EGF-induced DNA synthesis. Likewise, the stimulatory effect of TGFβ in the absence of EGF requires only an equally brief exposure to TGFβ. Use of homogeneous 125 I-labeled TGFβ in a cell-binding assay demonstrates that TGFβ bound to cell-surface receptors can readily exchange into the culture medium, helping to rule out the possibility that persistent receptor-bound TGFβ is the source of a continuous stimulus. The data indicate that TGFβ exposure induces a stable state in the cell similar to but distinct from the state of competence induced by platelet-derived growth factor (PDGF)

  11. Regulation of insulin-like growth factor I receptors on vascular smooth muscle cells by growth factors and phorbol esters.

    Science.gov (United States)

    Ververis, J J; Ku, L; Delafontaine, P

    1993-06-01

    Insulin-like growth factor I (IGF I) is an important mitogen for vascular smooth muscle cells. To characterize regulation of vascular IGF I receptors, we performed radioligand displacement experiments using rat aortic smooth muscle cells (RASMs). Serum deprivation for 48 hours caused a 40% decrease in IGF I receptor number. Exposure of quiescent RASMs to platelet-derived growth factor (PDGF), fibroblast growth factor (FGF), or angiotensin II (Ang II) caused a 1.5-2.0-fold increase in IGF I receptors per cell. After FGF exposure, there was a marked increase in the mitogenic response to IGF I. IGF I downregulated its receptors in the presence of platelet-poor plasma. Stimulation of protein kinase C (PKC) by exposure of quiescent RASMs to phorbol 12-myristate 13-acetate caused a biphasic response in IGF I binding; there was a 42% decrease in receptor number at 45 minutes and a 238% increase at 24 hours. To determine the role of PKC in growth factor-induced regulation of IGF I receptors, we downregulated PKC by exposing RASMs to phorbol 12,13-dibutyrate (PDBu) for 48 hours. PDGF- and FGF- but not Ang II-mediated upregulation of IGF I receptors was completely inhibited in PDBu-treated cells. Thus, acute PKC activation by phorbol esters inhibits IGF I binding, whereas chronic PKC activation increases IGF I binding. PDGF and FGF but not Ang II regulate vascular IGF I receptors through a PKC-dependent pathway. These data provide new insights into the regulation of vascular smooth muscle cell IGF I receptors in vitro and are of potential importance in characterizing vascular proliferative responses in vivo.

  12. Biomaterials with persistent growth factor gradients in vivo accelerate vascularized tissue formation.

    Science.gov (United States)

    Akar, Banu; Jiang, Bin; Somo, Sami I; Appel, Alyssa A; Larson, Jeffery C; Tichauer, Kenneth M; Brey, Eric M

    2015-12-01

    Gradients of soluble factors play an important role in many biological processes, including blood vessel assembly. Gradients can be studied in detail in vitro, but methods that enable the study of spatially distributed soluble factors and multi-cellular processes in vivo are limited. Here, we report on a method for the generation of persistent in vivo gradients of growth factors in a three-dimensional (3D) biomaterial system. Fibrin loaded porous poly (ethylene glycol) (PEG) scaffolds were generated using a particulate leaching method. Platelet derived growth factor BB (PDGF-BB) was encapsulated into poly (lactic-co-glycolic acid) (PLGA) microspheres which were placed distal to the tissue-material interface. PLGA provides sustained release of PDGF-BB and its diffusion through the porous structure results in gradient formation. Gradients within the scaffold were confirmed in vivo using near-infrared fluorescence imaging and gradients were present for more than 3 weeks. The diffusion of PDGF-BB was modeled and verified with in vivo imaging findings. The depth of tissue invasion and density of blood vessels formed in response to the biomaterial increased with magnitude of the gradient. This biomaterial system allows for generation of sustained growth factor gradients for the study of tissue response to gradients in vivo. Published by Elsevier Ltd.

  13. Platelets promote tumor growth and metastasis via direct interaction between Aggrus/podoplanin and CLEC-2.

    Directory of Open Access Journals (Sweden)

    Satoshi Takagi

    Full Text Available The platelet aggregation-inducing factor Aggrus, also known as podoplanin, is frequently upregulated in several types of tumors and enhances hematogenous metastasis by interacting with and activating the platelet receptor CLEC-2. Thus, Aggrus-CLEC-2 binding could be a therapeutic molecular mechanism for cancer therapy. We generated a new anti-human Aggrus monoclonal antibody, MS-1, that suppressed Aggrus-CLEC-2 binding, Aggrus-induced platelet aggregation, and Aggrus-mediated tumor metastasis. Interestingly, the MS-1 monoclonal antibody attenuated the growth of Aggrus-positive tumors in vivo. Moreover, the humanized chimeric MS-1 antibody, ChMS-1, also exhibited strong antitumor activity against Aggrus-positive lung squamous cell carcinoma xenografted into NOD-SCID mice compromising antibody-dependent cellular cytotoxic and complement-dependent cytotoxic activities. Because Aggrus knockdown suppressed platelet-induced proliferation in vitro and tumor growth of the lung squamous cell carcinoma in vivo, Aggrus may be involved in not only tumor metastasis but also tumor growth by promoting platelet-tumor interaction, platelet activation, and secretion of platelet-derived factors in vivo. Our results indicate that molecular target drugs inhibiting specific platelet-tumor interactions can be developed as antitumor drugs that suppress both metastasis and proliferation of tumors such as lung squamous cell carcinoma.

  14. Polysome profiling of mAb producing CHO cell lines links translational control of cell proliferation and recombinant mRNA loading onto ribosomes with global and recombinant protein synthesis.

    Science.gov (United States)

    Godfrey, Charlotte L; Mead, Emma J; Daramola, Olalekan; Dunn, Sarah; Hatton, Diane; Field, Ray; Pettman, Gary; Smales, C Mark

    2017-08-01

    mRNA translation is a key process determining growth, proliferation and duration of a Chinese hamster ovary (CHO) cell culture and influences recombinant protein synthesis rate. During bioprocessing, CHO cells can experience stresses leading to reprogramming of translation and decreased global protein synthesis. Here we apply polysome profiling to determine reprogramming and translational capabilities in host and recombinant monoclonal antibody-producing (mAb) CHO cell lines during batch culture. Recombinant cell lines with the fastest cell specific growth rates were those with the highest global translational efficiency. However, total ribosomal capacity, determined from polysome profiles, did not relate to the fastest growing or highest producing mAb cell line, suggesting it is the ability to utilise available machinery that determines protein synthetic capacity. Cell lines with higher cell specific productivities tended to have elevated recombinant heavy chain transcript copy numbers, localised to the translationally active heavy polysomes. The highest titre cell line was that which sustained recombinant protein synthesis and maintained high recombinant transcript copy numbers in polysomes. Investigation of specific endogenous transcripts revealed a number that maintained or reprogrammed into heavy polysomes, identifying targets for potential cell engineering or those with 5' untranslated regions that might be utilised to enhance recombinant transcript translation. © 2017 The Authors. Biotechnology Journal published by WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  15. Efficacy and safety of recombinant activated factor VII for acute intracerebral hemorrhage

    DEFF Research Database (Denmark)

    Mayer, Stephan A; Brun, Nikolai C; Begtrup, Kamilla

    2008-01-01

    BACKGROUND: Intracerebral hemorrhage is the least treatable form of stroke. We performed this phase 3 trial to confirm a previous study in which recombinant activated factor VII (rFVIIa) reduced growth of the hematoma and improved survival and functional outcomes. METHODS: We randomly assigned 841...

  16. Induction of mitotic recombination by UV and diepoxybutane and its enhancement by hydroxyurea in Saccharomyces cerevisae

    Energy Technology Data Exchange (ETDEWEB)

    Zaborowska, D.; Swietlinska, Z.; Zuk, J. (Polska Akademia Nauk, Warsaw. Inst. Biochemii i Biofizyki)

    1983-04-01

    Mitotic inter- and intra-genic recombination was induced by UV-irradiation and treatment with diepoxybutane (DEB) in 2 heteroallelic diploid strains of Saccharomyces cerevisiae SBTD and D7. Induction of the events tested was strongly potentiated by plating of mutagen-treated cells on growth media containing 0.03 M hydroxyurea (HU).

  17. Dose of recombinant FSH and oestradiol concentration on day of HCG affect embryo development kinetics

    DEFF Research Database (Denmark)

    Muñoz, Manuel; Cruz, María; Humaidan, Peter

    2012-01-01

    During follicular growth, the follicle is exposed to an almost ever-changing composition of isoforms of FSH and LH, which causes a number of different and divergent biological effects. Through a time-lapse system, embryo kinetics were examined following the use of FSH only (recombinant FSH, rFSH)...

  18. Induction of mitotic recombination by UV and diepoxybutane and its enhancement by hydroxyurea in Saccharomyces cerevisae

    International Nuclear Information System (INIS)

    Zaborowska, D.; Swietlinska, Z.; Zuk, J.

    1983-01-01

    Mitotic inter- and intra-genic recombination was induced by UV-irradiation and treatment with diepoxybutane (DEB) in 2 heteroallelic diploid strains of Saccharomyces cerevisiae SBTD and D7. Induction of the events tested was strongly potentiated by plating of mutagen-treated cells on growth media containing 0.03 M hydroxyurea (HU). (orig.)

  19. Morphological characterization of recombinant strains of Aspergillus oryzae producing alpha-amylase during batch cultivations

    DEFF Research Database (Denmark)

    Spohr, Anders Bendsen; Carlsen, Morten; Nielsen, Jens Bredal

    1997-01-01

    Three alpha-amylase producing strains of Aspergillus oryzae used for recombinant protein production have been studied with respect to growth and protein production. By comparing the three strains with respect to morphology and protein production it is shown that a morphological mutant with a more...

  20. Growth factor and pro-inflammatory cytokine contents in platelet-rich plasma (PRP), plasma rich in growth factors (PRGF), advanced platelet-rich fibrin (A-PRF), and concentrated growth factors (CGF).

    Science.gov (United States)

    Masuki, Hideo; Okudera, Toshimitsu; Watanebe, Taisuke; Suzuki, Masashi; Nishiyama, Kazuhiko; Okudera, Hajime; Nakata, Koh; Uematsu, Kohya; Su, Chen-Yao; Kawase, Tomoyuki

    2016-12-01

    The development of platelet-rich fibrin (PRF) drastically simplified the preparation procedure of platelet-concentrated biomaterials, such as platelet-rich plasma (PRP), and facilitated their clinical application. PRF's clinical effectiveness has often been demonstrated in pre-clinical and clinical studies; however, it is still controversial whether growth factors are significantly concentrated in PRF preparations to facilitate wound healing and tissue regeneration. To address this matter, we performed a comparative study of growth factor contents in PRP and its derivatives, such as advanced PRF (A-PRF) and concentrated growth factors (CGF). PRP and its derivatives were prepared from the same peripheral blood samples collected from healthy donors. A-PRF and CGF preparations were homogenized and centrifuged to produce extracts. Platelet and white blood cell counts in A-PRF and CGF preparations were determined by subtracting those counts in red blood cell fractions, supernatant acellular serum fractions, and A-PRF/CGF exudate fractions from those counts of whole blood samples. Concentrations of growth factors (TGF-β1, PDGF-BB, VEGF) and pro-inflammatory cytokines (IL-1β, IL-6) were determined using ELISA kits. Compared to PRP preparations, both A-PRF and CGF extracts contained compatible or higher levels of platelets and platelet-derived growth factors. In a cell proliferation assay, both A-PRF and CGF extracts significantly stimulated the proliferation of human periosteal cells without significant reduction at higher doses. These data clearly demonstrate that both A-PRF and CGF preparations contain significant amounts of growth factors capable of stimulating periosteal cell proliferation, suggesting that A-PRF and CGF preparations function not only as a scaffolding material but also as a reservoir to deliver certain growth factors at the site of application.

  1. Immunohistochemical study of the growth factors, aFGF, bFGF, PDGF-AB, VEGF-A and its receptor (Flk-1) during arteriogenesis.

    Science.gov (United States)

    Wu, Song; Wu, Xiaoqiong; Zhu, Wu; Cai, Wei-Jun; Schaper, Jutta; Schaper, Wolfgang

    2010-10-01

    Growth factors are viewed as main arteriogenic stimulators for collateral vessel growth. However, the information about their native expression and distribution in collateral vessels is still limited. This study was designed to profile expression of acidic and basic FGF, platelet-derived growth factor (PDGF-AB) and vascular endothelial growth factor (VEGF-A) and its receptor, fetal liver kinase-1 (Flk-1) during arteriogenesis by confocal immunofluorescence in both dog ameroid constrictor model and rabbit arteriovenous shunt model of arteriogenesis. We found that: (1) in normal arteries (NA) in dog heart, aFGF, bFGF, and PDGF-AB all were mainly expressed in endothelial cells (EC) and media smooth muscle cells (SMC), but the expression of aFGF was very weak, with those of the other two being moderate; (2) in collateral arteries (CAs), aFGF, bFGF, and PDGF-AB all were significantly upregulated (P growth factors, aFGF, bFGF, and PDGF-AB are significantly upregulated in collateral vessels in dog heart, and enhanced VEGF-A and its receptor, Flk-1, are associated with rapid and lasting increased shear stress. These findings suggest that endogenous production of growth factors could be an important factor promoting collateral vessel growth.

  2. Electron-ion recombination rates for merged-beams experiments

    International Nuclear Information System (INIS)

    Pajek, M.

    1994-01-01

    Energy dependence of the electron-ion recombination rates are studied for different recombination processes (radiative recombination, three-body recombination, dissociative recombination) for Maxwellian relative velocity distribution of arbitrary asymmetry. The results are discussed in context of the electron-ion merged beams experiments in cooling ion storage rings. The question of indication of a possible contribution of the three-body recombination to the measured recombination rates versus relative energy is particularly addressed. Its influence on the electron beam temperature derived from the energy dependence of recombination rate is discussed

  3. First-principles study of Frenkel pair recombination in tungsten

    International Nuclear Information System (INIS)

    Qin, Shi-Yao; Jin, Shuo; Li, Yu-Hao; Zhou, Hong-Bo; Zhang, Ying; Lu, Guang-Hong

    2017-01-01

    The recombination of one Frenkel pair in tungsten has been investigated through first-principles simulation. Two different recombination types have been identified: instantaneous and thermally activated. The small recombination barriers for thermally activated recombination cases indicate that recombination can occur easily with a slightly increased temperature. For both of the two recombination types, recombination occurs through the self-interstitial atom moving towards the vacancy. The recombination process can be direct or through replacement sequences, depending on the vertical distance between the vacancy and the 〈1 1 1〉 line of self-interstitial atom pair.

  4. Conjugation of gold nanoparticles and recombinant human endostatin modulates vascular normalization via interruption of anterior gradient 2-mediated angiogenesis.

    Science.gov (United States)

    Pan, Fan; Yang, Wende; Li, Wei; Yang, Xiao-Yan; Liu, Shuhao; Li, Xin; Zhao, Xiaoxu; Ding, Hui; Qin, Li; Pan, Yunlong

    2017-07-01

    Several studies have revealed the potential of normalizing tumor vessels in anti-angiogenic treatment. Recombinant human endostatin is an anti-angiogenic agent which has been applied in clinical tumor treatment. Our previous research indicated that gold nanoparticles could be a nanoparticle carrier for recombinant human endostatin delivery. The recombinant human endostatin-gold nanoparticle conjugates normalized vessels, which improved chemotherapy. However, the mechanism of recombinant human endostatin-gold nanoparticle-induced vascular normalization has not been explored. Anterior gradient 2 has been reported to be over-expressed in many malignant tumors and involved in tumor angiogenesis. To date, the precise efficacy of recombinant human endostatin-gold nanoparticles on anterior gradient 2-mediated angiogenesis or anterior gradient 2-related signaling cohort remained unknown. In this study, we aimed to explore whether recombinant human endostatin-gold nanoparticles could normalize vessels in metastatic colorectal cancer xenografts, and we further elucidated whether recombinant human endostatin-gold nanoparticles could interrupt anterior gradient 2-induced angiogenesis. In vivo, it was indicated that recombinant human endostatin-gold nanoparticles increased pericyte expression while inhibit vascular endothelial growth factor receptor 2 and anterior gradient 2 expression in metastatic colorectal cancer xenografts. In vitro, we uncovered that recombinant human endostatin-gold nanoparticles reduced cell migration and tube formation induced by anterior gradient 2 in human umbilical vein endothelial cells. Treatment with recombinant human endostatin-gold nanoparticles attenuated anterior gradient 2-mediated activation of MMP2, cMyc, VE-cadherin, phosphorylation of p38, and extracellular signal-regulated protein kinases 1 and 2 (ERK1/2) in human umbilical vein endothelial cells. Our findings demonstrated recombinant human endostatin-gold nanoparticles might normalize

  5. Study of recombinant proteins derived from Ser-2 gene of Bombyx mori

    OpenAIRE

    STAŠKOVÁ, Tereza

    2012-01-01

    Four different variants of recombinant proteins derived from Bombyx mori Ser-2 gene were expressed in bacteria. The ability of these proteins to coat hydrofobic surfaces and to support growth of various types of adherent cells in vitro were examined. It was shown that these proteins support cell adhesion and proliferation, and could be used as coating agents to realize surfaces suitable for growth of vertebrate and insect cells.

  6. Induction of homologous recombination in Saccharomyces cerevisiae.

    Science.gov (United States)

    Simon, J R; Moore, P D

    1988-09-01

    We have investigated the effects of UV irradiation of Saccharomyces cerevisiae in order to distinguish whether UV-induced recombination results from the induction of enzymes required for homologous recombination, or the production of substrate sites for recombination containing regions of DNA damage. We utilized split-dose experiments to investigate the induction of proteins required for survival, gene conversion, and mutation in a diploid strain of S. cerevisiae. We demonstrate that inducing doses of UV irradiation followed by a 6 h period of incubation render the cells resistant to challenge doses of UV irradiation. The effects of inducing and challenge doses of UV irradiation upon interchromosomal gene conversion and mutation are strictly additive. Using the yeast URA3 gene cloned in non-replicating single- and double-stranded plasmid vectors that integrate into chromosomal genes upon transformation, we show that UV irradiation of haploid yeast cells and homologous plasmid DNA sequences each stimulate homologous recombination approximately two-fold, and that these effects are additive. Non-specific DNA damage has little effect on the stimulation of homologous recombination, as shown by studies in which UV-irradiated heterologous DNA was included in transformation/recombination experiments. We further demonstrate that the effect of competing single- and double-stranded heterologous DNA sequences differs in UV-irradiated and unirradiated cells, suggesting an induction of recombinational machinery in UV-irradiated S. cerevisiae cells.

  7. Hydroxyapatite coating on the titanium substrate modulated by a recombinant collagen-like protein

    International Nuclear Information System (INIS)

    Pan Mingli; Kong Xiangdong; Cai Yurong; Yao Juming

    2011-01-01

    Research highlights: → Hydroxyapatite was deposited on alkali-heat treated Ti substrate by immersing in 1.5 x SBF solution containing the recombinant collagen-like protein. → The recombinant collagen-like protein accelerated the preferential nucleation and growth of hydroxyapatite along c axis on the Ti substrate. → Hydroxyapatite-collagen composite on the Ti substrate promoted the attachment, subsequently proliferation and differentiation of MG-63 cells. - Abstract: Plenty of techniques have been developed to modify the surface character of titanium (Ti) and its alloys in order to realize their biological bond to natural bone. In this work, a biomimetic process was employed to form a hydroxyapatite (HAp) coating on the alkali-heat treated Ti substrate in 1.5 times simulated body fluid (1.5 x SBF) with the addition of a recombinant collagen-like protein. The coating was characterized using SEM-EDX, FESEM, and XRD. Results showed that the recombinant collagen-like protein could accelerate the preferential nucleation and directional growth along c axis of HAp on the pretreated Ti substrates. The investigation of in vitro cell cultivation showed that the existence of recombinant collagen-like protein in coating could improve the initial cell adhesion, proliferation and differentiation of MG-63 cells, which implied the materials possessed excellent biocompatibility and had a wide potential in biomedical application.

  8. Anti-angiogenesis and anti-tumor activity of recombinant anginex

    International Nuclear Information System (INIS)

    Brandwijk, Ricardo J.M.G.E.; Dings, Ruud P.M.; Linden, Edith van der; Mayo, Kevin H.; Thijssen, Victor L.J.L.; Griffioen, Arjan W.

    2006-01-01

    Anginex, a synthetic 33-mer angiostatic peptide, specifically inhibits vascular endothelial cell proliferation and migration along with induction of apoptosis in endothelial cells. Here we report on the in vivo characterization of recombinant anginex and use of the artificial anginex gene for gene therapy approaches. Tumor growth of human MA148 ovarian carcinoma in athymic mice was inhibited by 80% when treated with recombinant anginex. Histological analysis of the tumors showed an approximate 2.5-fold reduction of microvessel density, suggesting that angiogenesis inhibition is the cause of the anti-tumor effect. Furthermore, there was a significant correlation between the gene expression patterns of 16 angiogenesis-related factors after treatment with both recombinant and synthetic anginex. To validate the applicability of the anginex gene for gene therapy, stable transfectants of murine B16F10 melanoma cells expressing recombinant anginex were made. Supernatants of these cells inhibited endothelial cell proliferation in vitro. Furthermore, after subcutaneous injection of these cells in C57BL/6 mice, an extensive delay in tumor growth was observed. These data show that the artificial anginex gene can be used to produce a recombinant protein with similar activity as its synthetic counterpart and that the gene can be applied in gene therapy approaches for cancer treatment

  9. Mini review: Recombinant production of tailored bio-pharmaceuticals in different Bacillus strains and future perspectives.

    Science.gov (United States)

    Lakowitz, Antonia; Godard, Thibault; Biedendieck, Rebekka; Krull, Rainer

    2018-05-01

    Bio-pharmaceuticals like antibodies, hormones and growth factors represent about one-fifth of commercial pharmaceuticals. Host candidates of growing interest for recombinant production of these proteins are strains of the genus Bacillus, long being established for biotechnological production of homologous and heterologous proteins. Bacillus strains benefit from development of efficient expression systems in the last decades and emerge as major industrial workhorses for recombinant proteins due to easy cultivation, non-pathogenicity and their ability to secrete recombinant proteins directly into extracellular medium allowing cost-effective downstream processing. Their broad product portfolio of pharmaceutically relevant recombinant proteins described in research include antibody fragments, growth factors, interferons and interleukins, insulin, penicillin G acylase, streptavidin and different kinases produced in various cultivation systems like microtiter plates, shake flasks and bioreactor systems in batch, fed-batch and continuous mode. To further improve production and secretion performance of Bacillus, bottlenecks and limiting factors concerning proteases, chaperones, secretion machinery or feedback mechanisms can be identified on different cell levels from genomics and transcriptomics via proteomics to metabolomics and fluxomics. For systematical identification of recurring patterns characteristic of given regulatory systems and key genetic targets, systems biology and omics-technology provide suitable and promising approaches, pushing Bacillus further towards industrial application for recombinant pharmaceutical protein production. Copyright © 2017. Published by Elsevier B.V.

  10. Hydroxyapatite coating on the titanium substrate modulated by a recombinant collagen-like protein

    Energy Technology Data Exchange (ETDEWEB)

    Pan Mingli [Key Laboratory of Advanced Textile Materials and Manufacturing Technology of Ministry of Education, College of Materials and Textile, Zhejiang Sci-Tech University, Hangzhou 310018 (China); Kong Xiangdong [College of Life Sciences, Zhejiang Sci-Tech University, Hangzhou 310018 (China); Cai Yurong [Key Laboratory of Advanced Textile Materials and Manufacturing Technology of Ministry of Education, College of Materials and Textile, Zhejiang Sci-Tech University, Hangzhou 310018 (China); Yao Juming, E-mail: yaoj@zstu.edu.cn [Key Laboratory of Advanced Textile Materials and Manufacturing Technology of Ministry of Education, College of Materials and Textile, Zhejiang Sci-Tech University, Hangzhou 310018 (China)

    2011-04-15

    Research highlights: {yields} Hydroxyapatite was deposited on alkali-heat treated Ti substrate by immersing in 1.5 x SBF solution containing the recombinant collagen-like protein. {yields} The recombinant collagen-like protein accelerated the preferential nucleation and growth of hydroxyapatite along c axis on the Ti substrate. {yields} Hydroxyapatite-collagen composite on the Ti substrate promoted the attachment, subsequently proliferation and differentiation of MG-63 cells. - Abstract: Plenty of techniques have been developed to modify the surface character of titanium (Ti) and its alloys in order to realize their biological bond to natural bone. In this work, a biomimetic process was employed to form a hydroxyapatite (HAp) coating on the alkali-heat treated Ti substrate in 1.5 times simulated body fluid (1.5 x SBF) with the addition of a recombinant collagen-like protein. The coating was characterized using SEM-EDX, FESEM, and XRD. Results showed that the recombinant collagen-like protein could accelerate the preferential nucleation and directional growth along c axis of HAp on the pretreated Ti substrates. The investigation of in vitro cell cultivation showed that the existence of recombinant collagen-like protein in coating could improve the initial cell adhesion, proliferation and differentiation of MG-63 cells, which implied the materials possessed excellent biocompatibility and had a wide potential in biomedical application.

  11. Recombination every day: abundant recombination in a virus during a single multi-cellular host infection.

    Directory of Open Access Journals (Sweden)

    Remy Froissart

    2005-03-01

    Full Text Available Viral recombination can dramatically impact evolution and epidemiology. In viruses, the recombination rate depends on the frequency of genetic exchange between different viral genomes within an infected host cell and on the frequency at which such co-infections occur. While the recombination rate has been recently evaluated in experimentally co-infected cell cultures for several viruses, direct quantification at the most biologically significant level, that of a host infection, is still lacking. This study fills this gap using the cauliflower mosaic virus as a model. We distributed four neutral markers along the viral genome, and co-inoculated host plants with marker-containing and wild-type viruses. The frequency of recombinant genomes was evaluated 21 d post-inoculation. On average, over 50% of viral genomes recovered after a single host infection were recombinants, clearly indicating that recombination is very frequent in this virus. Estimates of the recombination rate show that all regions of the genome are equally affected by this process. Assuming that ten viral replication cycles occurred during our experiment-based on data on the timing of coat protein detection-the per base and replication cycle recombination rate was on the order of 2 x 10(-5 to 4 x 10(-5. This first determination of a virus recombination rate during a single multi-cellular host infection indicates that recombination is very frequent in the everyday life of this virus.

  12. Recombinant DNA production of spider silk proteins.

    Science.gov (United States)

    Tokareva, Olena; Michalczechen-Lacerda, Valquíria A; Rech, Elíbio L; Kaplan, David L

    2013-11-01

    Spider dragline silk is considered to be the toughest biopolymer on Earth due to an extraordinary combination of strength and elasticity. Moreover, silks are biocompatible and biodegradable protein-based materials. Recent advances in genetic engineering make it possible to produce recombinant silks in heterologous hosts, opening up opportunities for large-scale production of recombinant silks for various biomedical and material science applications. We review the current strategies to produce recombinant spider silks. © 2013 The Authors. Microbial Biotechnology published by John Wiley & Sons Ltd and Society for Applied Microbiology.

  13. Xylose utilization in recombinant zymomonas

    Science.gov (United States)

    Caimi, Perry G; McCole, Laura; Tao, Luan; Tomb, Jean-Francois; Viitanen, Paul V

    2014-03-25

    Xylose-utilizing Zymomonas strains studied were found to accumulate ribulose when grown in xylose-containing media. Engineering these strains to increase ribose-5-phosphate isomerase activity led to reduced ribulose accumulation, improved growth, improved xylose utilization, and increased ethanol production.

  14. Laser-induced electron--ion recombination used to study enhanced spontaneous recombination during electron cooling

    International Nuclear Information System (INIS)

    Schramm, U.; Wolf, A.; Schuess ler, T.; Habs, D.; Schwalm, D.; Uwira, O.; Linkemann, J.; Mueller, A.

    1997-01-01

    Spontaneous recombination of highly charged ions with free electrons in merged velocity matched electron and ion beams has been observed in earlier experiments to occur at rates significantly higher than predicted by theoretical estimates. To study this enhanced spontaneous recombination, laser induced recombination spectra were measured both in velocity matched beams and in beams with well defined relative velocities, corresponding to relative electron-ion detuning energies ranging from 1 meV up to 6.5 meV where the spontaneous recombination enhancement was found to be strongly reduced. Based on a comparison with simplified calculations, the development of the recombination spectra for decreasing detuning energies indicates additional contributions at matched velocities which could be related to the energy distribution of electrons causing the spontaneous recombination rate enhancement

  15. Recombinant Human Papillomavirus (HPV) Bivalent Vaccine

    Science.gov (United States)

    This page contains brief information about recombinant human papillomavirus (HPV) bivalent vaccine and a collection of links to more information about the use of this vaccine, research results, and ongoing clinical trials.

  16. Recombinant Human Papillomavirus (HPV) Nonavalent Vaccine

    Science.gov (United States)

    This page contains brief information about recombinant human papillomavirus (HPV) nonavalent vaccine and a collection of links to more information about the use of this vaccine, research results, and ongoing clinical trials.

  17. Recombinant Human Papillomavirus (HPV) Quadrivalent Vaccine

    Science.gov (United States)

    This page contains brief information about recombinant human papillomavirus (HPV) quadrivalent vaccine and a collection of links to more information about the use of this vaccine, research results, and ongoing clinical trials.

  18. Ultramicroscopic observation of recombinant adenoassociated virus ...

    African Journals Online (AJOL)

    Ultramicroscopic observation of recombinant adenoassociated virus type 2 on the surface of formvarcarbon coated copper grids under different relative humidity and incubation time using negative stain transmission electron microscopy.

  19. Recombinant vaccines: experimental and applied aspects

    DEFF Research Database (Denmark)

    Lorenzen, Niels

    1999-01-01

    Development of vaccines for aquaculture fish represent an important applied functional aspect of fish immunology research. Particularly in the case of recombinant vaccines, where a single antigen is usually expected to induce immunity to a specific pathogen, knowledge of mechanisms involved...... in induction of a protective immune response may become vital. The few recombinant vaccines licensd so far, despite much research during the last decade, illustrate that this is not a straightforward matter. However, as vaccine technology as well as our knowledge of the fish immune system is steadily improved......, these fields will open up a number of interesting research objectives of mutual benefit. Recent aspects of recombinant protein vaccines, live recombinant vaccines and DNA vaccines are discussed....

  20. New perspectives on recombinant human antibodies

    NARCIS (Netherlands)

    J. de Kruif (John); A.-R. van der Vuurst de Vries (Anne); L. Cilenti (L.); E. Boel (E.); W. van Ewijk (Willem); T. Logtenberg (Ton)

    1996-01-01

    textabstractThe limited potential of murine monoclonal antibodies for human immunotherapy has driven recent progress in recombinant antibody technology. Here, de Kruif and colleagues report on advances in the development and use of phage-antibody-display libraries.

  1. Construction of retroviral recombinant containing human tissue ...

    African Journals Online (AJOL)

    USER

    2010-03-29

    Mar 29, 2010 ... Recombinant retroviral vector containing human tissue inhibitor of matrix metalloproteinase-2 (TIMP-2) gene was ..... heavy metal ions, the protein could be express in an .... involves adhesion, degradation and movement. To.

  2. A Comparison Study of Growth Factor Expression following Treatment with Transcutaneous Electrical Nerve Stimulation, Saline Solution, Povidone-Iodine, and Lavender Oil in Wounds Healing

    Directory of Open Access Journals (Sweden)

    Adalet Koca Kutlu

    2013-01-01

    Full Text Available This study compared the effects of transcutaneous electrical nerve stimulation (TENS, saline solution (SS, povidone-iodine (PI, and lavender oil (Lavandula angustifolia through expression of growth factors in a rat model of wound healing. Six experimental groups were established, each containing 8 rats: a healthy group with no incision wounds, an incision-control group, an incision and TENS group, an incision and SS group, an incision and PI group, and an incision and lavender oil group. Experiments continued for 5 days, after which the skin in the excision area was removed. Tissue concentrations of epidermal growth factor (EGF and platelet-derived growth factor (PDGF-A were measured using enzyme-linked immunosorbent assay (ELISA. Tissue expressions of EGF, PDGF-A, and fibroblast growth factor (FGF-2 were determined using immunohistochemistry. Wound closure progressed more rapidly in the TENS and lavender oil groups than in the control and other study groups. In particular, PDGF-A expressions in the dermis and EGF expression in the epidermis were significantly intense in the TENS group (P<0.05. In addition, ELISA levels of growth factors such as PDGF-A and EGF were significantly higher in TENS group compared to the control group (P<0.05. These immunohistochemical and ELISA results suggest that TENS may improve wound healing through increasing growth factors in the dermis and epidermis more than other topical applications.

  3. Live recombinant BHV/BRSV vaccine

    OpenAIRE

    Keil, G.M.; Rijsewijk, F.A.M.

    1998-01-01

    The present invention refers to synthetic Bovine Respiratory Syncytium virus genes. Also the invention relates to live attenuated Bovine Herpesvirus recombinants carrying such synthetic genes. Furthermore, the invention relates to vaccines based on these live attenuated recombinants, for the protection of cattle against both Bovine herpesvirus infection and against Bovine Respiratory Syncytium virus infection. Also the invention relates to methods for the preparation of such live attenuated r...

  4. Co-factor activated recombinant adenovirus proteinases

    Science.gov (United States)

    Anderson, Carl W.; Mangel, Walter F.

    1996-08-06

    This application describes methods and expression constructs for producing activatable recombinant adenovirus proteinases. Purified activatable recombinant adenovirus proteinases and methods of purification are described. Activated adenovirus proteinases and methods for obtaining activated adenovirus proteinases are further included. Isolated peptide cofactors of adenovirus proteinase activity, methods of purifying and identifying said peptide cofactors are also described. Antibodies immunoreactive with adenovirus proteinases, immunospecific antibodies, and methods for preparing them are also described. Other related methods and materials are also described.

  5. Hadron correlations from recombination and fragmentation

    Energy Technology Data Exchange (ETDEWEB)

    Fries, Rainer J [School of Physics and Astronomy, University of Minnesota, Minneapolis, MN 55455 (United States)

    2005-04-01

    We review the formalism of quark recombination applied to the hadronization of a quark-gluon plasma. Evidence in favour of the quark recombination model is outlined. Recent work on parton correlations, leading to detectable correlations between hadrons, is discussed. Hot spots from completely quenched jets are a likely source of such correlations which appear to be jet like. It will be discussed how such a picture compares with measurement of associated hadron yields at RHIC.

  6. Recombination of a fast expanding plasma

    International Nuclear Information System (INIS)

    Salvat, M.

    1979-05-01

    The goal of the following calculations is to determine numerically the recombination of dense plasmas (for instance of laser-produced plasmas). The recombination is computed for plasmas with initial densities of 10 24 27 [m -3 ] and with initial temperatures >= 50 eV. The ionization of the plasma remains essentially constant during the early phase of expansion. The time for which the ionization is 'frozen-in' grows with decreasing initial density and with increasing initial temperature. (orig.) [de

  7. Dissociation of recombinant prion autocatalysis from infectivity.

    Science.gov (United States)

    Noble, Geoffrey P; Supattapone, Surachai

    2015-01-01

    Within the mammalian prion field, the existence of recombinant prion protein (PrP) conformers with self-replicating (ie. autocatalytic) activity in vitro but little to no infectious activity in vivo challenges a key prediction of the protein-only hypothesis of prion replication--that autocatalytic PrP conformers should be infectious. To understand this dissociation of autocatalysis from infectivity, we recently performed a structural and functional comparison between a highly infectious and non-infectious pair of autocatalytic recombinant PrP conformers derived from the same initial prion strain. (1) We identified restricted, C-terminal structural differences between these 2 conformers and provided evidence that these relatively subtle differences prevent the non-infectious conformer from templating the conversion of native PrP(C) substrates containing a glycosylphosphatidylinositol (GPI) anchor. (1) In this article we discuss a model, consistent with these findings, in which recombinant PrP, lacking post-translational modifications and associated folding constraints, is capable of adopting a wide variety of autocatalytic conformations. Only a subset of these recombinant conformers can be adopted by post-translationally modified native PrP(C), and this subset represents the recombinant conformers with high specific infectivity. We examine this model's implications for the generation of highly infectious recombinant prions and the protein-only hypothesis of prion replication.

  8. Mitigating Mitochondrial Genome Erosion Without Recombination.

    Science.gov (United States)

    Radzvilavicius, Arunas L; Kokko, Hanna; Christie, Joshua R

    2017-11-01

    Mitochondria are ATP-producing organelles of bacterial ancestry that played a key role in the origin and early evolution of complex eukaryotic cells. Most modern eukaryotes transmit mitochondrial genes uniparentally, often without recombination among genetically divergent organelles. While this asymmetric inheritance maintains the efficacy of purifying selection at the level of the cell, the absence of recombination could also make the genome susceptible to Muller's ratchet. How mitochondria escape this irreversible defect accumulation is a fundamental unsolved question. Occasional paternal leakage could in principle promote recombination, but it would also compromise the purifying selection benefits of uniparental inheritance. We assess this tradeoff using a stochastic population-genetic model. In the absence of recombination, uniparental inheritance of freely-segregating genomes mitigates mutational erosion, while paternal leakage exacerbates the ratchet effect. Mitochondrial fusion-fission cycles ensure independent genome segregation, improving purifying selection. Paternal leakage provides opportunity for recombination to slow down the mutation accumulation, but always at a cost of increased steady-state mutation load. Our findings indicate that random segregation of mitochondrial genomes under uniparental inheritance can effectively combat the mutational meltdown, and that homologous recombination under paternal leakage might not be needed. Copyright © 2017 by the Genetics Society of America.

  9. Electron - ion recombination processes - an overview

    International Nuclear Information System (INIS)

    Hahn, Yukap

    1997-01-01

    Extensive theoretical and experimental studies have been carried out for the past 20 years on electron - ion recombination processes, as they are applied to the analysis of astrophysical and laboratory plasmas. We review the basic understanding gained through these efforts, with emphasis on some of the more recent progress made in recombination theory as the recombining system is affected by time-dependent electric fields and plasma particles at low temperature. Together with collisional ionization and excitation processes, recombination is important in determining ionization balance and excited-state population in non-equilibrium plasmas. The radiation emitted by plasmas is usually the principal medium with which to study the plasma condition, as it is produced mainly during the recombination and decay of excited states of ions inside the plasma. This is especially true when the plasma under study is not readily accessible by direct probes, as in astrophysical plasmas. Moreover, external probes may sometimes cause undesirable disturbances of the plasma. Electron-ion recombination proceeds in several different modes. The direct modes include three-body recombination (TBR) and one-step radiative recombination (RR), all to the ground- and singly-excited states of the target ions. By contrast, the indirect resonant mode is a two-step dielectronic recombination (DR), which proceeds first with the formation of doubly-excited states by radiationless excitation/capture. The resonant states thus formed may relax by autoionization and/or radiative cascades. For more exotic modes of recombination, we consider off-shell dielectronic recombination (radiative DR = RDR), in which an electron capture is accompanied by simultaneous radiative emission and excitation of the target ion. Some discussion on attachment of electrons to neutral atoms, resulting in the formation of negative ions, is also given. When resonance states involve one or more electrons in high Rydberg states

  10. Influence of storage conditions on the release of growth factors in platelet-rich blood derivatives

    Directory of Open Access Journals (Sweden)

    Düregger Katharina

    2016-09-01

    Full Text Available Thrombocytes can be concentrated in blood derivatives and used as autologous transplants e.g. for wound treatment due to the release of growth factors such as platelet derived growth factor (PDGF. Conditions for processing and storage of these platelet-rich blood derivatives influence the release of PDGF from the platelet-bound α-granules into the plasma. In this study Platelet rich plasma (PRP and Platelet concentrate (PC were produced with a fully automated centrifugation system. Storage of PRP and PC for 1 h up to 4 months at temperatures between −20°C and +37°C was applied with the aim of evaluating the influence on the amount of released PDGF. Storage at −20°C resulted in the highest release of PDGF in PRP and a time dependency was determined: prolonged storage up to 1 month in PRP and 10 days in PC increased the release of PDGF. Regardless of the storage conditions, the release of PDGF per platelet was higher in PC than in PRP.

  11. Oligonucleotide recombination enabled site-specific mutagenesis in bacteria

    Science.gov (United States)

    Recombineering refers to a strategy for engineering DNA sequences using a specialized mode of homologous recombination. This technology can be used for rapidly constructing precise changes in bacterial genome sequences in vivo. Oligo recombination is one type of recombineering that uses ssDNA olig...

  12. Correlation between increasing tissue ischemia and circulating levels of angiogenic growth factors in peripheral artery disease.

    Science.gov (United States)

    Jalkanen, Juho; Hautero, Olli; Maksimow, Mikael; Jalkanen, Sirpa; Hakovirta, Harri

    2018-04-21

    The aim of the present study was to assess the circulating levels of vascular endothelial growth factor (VEGF) and other suggested therapeutic growth factors with the degree of ischemia in patients with different clinical manifestations of peripheral arterial disease (PAD) according to the Rutherford grades. The study cohort consists of 226 consecutive patients admitted to a Department of Vascular Surgery for elective invasive procedures. PAD patients were grouped according to the Rutherford grades after a clinical assessment. Ankle-brachial pressure indices (ABI) and absolute toe pressure (TP) values were measured. Serum levels of circulating VEGF, hepatocyte growth factor (HGF), basic fibroblast growth factor (bFGF), and platelet derived growth factor (PDGF) were measured from serum and analysed against Rutherford grades and peripheral hemodynamic measurements. The levels of VEGF (P = 0.009) and HGF (P correlations between Rutherford grades was detected as follows; VEGF (Pearson's correlation = 0.183, P = 0.004), HGF (Pearson's correlation = 0.253, P Pearson's correlation = 0.169, P = 0.008) and PDGF (Pearson's correlation = 0.296, P correlation with ABI (Pearson's correlation -0.19, P = 0.009) and TP (Pearson's correlation -0.20, P = 0.005) measurements. Our present observations show that the circulating levels of VEGF and other suggested therapeutic growth factors are significantly increased along with increasing ischemia. These findings present a new perspective to anticipated positive effects of gene therapies utilizing VEGF, HGF, and bFGF, because the levels of these growth factors are endogenously high in end-stage PAD. Copyright © 2018 Elsevier Ltd. All rights reserved.

  13. Recombination analysis based on the complete genome of bocavirus

    Directory of Open Access Journals (Sweden)

    Chen Shengxia

    2011-04-01

    Full Text Available Abstract Bocavirus include bovine parvovirus, minute virus of canine, porcine bocavirus, gorilla bocavirus, and Human bocaviruses 1-4 (HBoVs. Although recent reports showed that recombination happened in bocavirus, no systematical study investigated the recombination of bocavirus. The present study performed the phylogenetic and recombination analysis of bocavirus over the complete genomes available in GenBank. Results confirmed that recombination existed among bocavirus, including the likely inter-genotype recombination between HBoV1 and HBoV4, and intra-genotype recombination among HBoV2 variants. Moreover, it is the first report revealing the recombination that occurred between minute viruses of canine.

  14. Optimisation of culture conditions with respect to biotin requirement for the production of recombinant avidin in Pichia pastoris.

    Science.gov (United States)

    Jungo, Carmen; Urfer, Julien; Zocchi, Andrea; Marison, Ian; von Stockar, Urs

    2007-01-20

    Due to its very high affinity to biotin, avidin is one of the most widely exploited proteins in modern biotechnological and biomedical applications. Since biotin is an essential vitamin for the growth of many microorganisms, we examined the effect of biotin deficiency on growth for a recombinant Pichia pastoris strain expressing and secreting a recombinant glycosylated avidin. The results showed that biotin deficiency lowers growth rate and biomass yield for P. pastoris. Substitution of biotin in the medium by the two structurally unrelated compounds, aspartic acid and oleic acid, which do not bind to recombinant avidin was analyzed quantitatively. These two compounds had a growth promoting effect in biotin-deficient medium, but did not replace biotin completely. Indeed, in chemostat culture, wash-out occurred after about six liquid residence times and recombinant avidin productivity was lowered. However, addition of low amounts of biotin (20 microg L(-1) of biotin for a cell density of 8 g L(-1)) resulted in stable chemostat cultures on methanol with the production of recombinant biotin-free avidin. The specific avidin production rate was 22 microg g(-1) h(-1) at a dilution rate of 0.06 h(-1).

  15. Genetic recombination of the hepatitis C virus: clinical implications.

    Science.gov (United States)

    Morel, V; Fournier, C; François, C; Brochot, E; Helle, F; Duverlie, G; Castelain, S

    2011-02-01

    Genetic recombination is a well-known feature of RNA viruses that plays a significant role in their evolution. Although recombination is well documented for Flaviviridae family viruses, the first natural recombinant strain of hepatitis C virus (HCV) was identified as recently as 2002. Since then, a few other natural inter-genotypic, intra-genotypic and intra-subtype recombinant HCV strains have been described. However, the frequency of recombination may have been underestimated because not all known HCV recombinants are screened for in routine practice. Furthermore, the choice of treatment regimen and its predictive outcome remain problematic as the therapeutic strategy for HCV infection is genotype dependent. HCV recombination also raises many questions concerning its mechanisms and effects on the epidemiological and physiopathological features of the virus. This review provides an update on recombinant HCV strains, the process that gives rise to recombinants and clinical implications of recombination. © 2010 Blackwell Publishing Ltd.

  16. Improved Eco-Friendly Recombinant Anabaena sp. Strain PCC7120 with Enhanced Nitrogen Biofertilizer Potential▿

    Science.gov (United States)

    Chaurasia, Akhilesh Kumar; Apte, Shree Kumar

    2011-01-01

    Photosynthetic, nitrogen-fixing Anabaena strains are native to tropical paddy fields and contribute to the carbon and nitrogen economy of such soils. Genetic engineering was employed to improve the nitrogen biofertilizer potential of Anabaena sp. strain PCC7120. Constitutive enhanced expression of an additional integrated copy of the hetR gene from a light-inducible promoter elevated HetR protein expression and enhanced functional heterocyst frequency in the recombinant strain. The recombinant strain displayed consistently higher nitrogenase activity than the wild-type strain and appeared to be in homeostasis with compatible modulation of photosynthesis and respiration. The enhanced combined nitrogen availability from the recombinant strain positively catered to the nitrogen demand of rice seedlings in short-term hydroponic experiments and supported better growth. The engineered strain is stable, eco-friendly, and useful for environmental application as nitrogen biofertilizer in paddy fields. PMID:21057013

  17. Improved eco-friendly recombinant Anabaena sp. strain PCC7120 with enhanced nitrogen biofertilizer potential.

    Science.gov (United States)

    Chaurasia, Akhilesh Kumar; Apte, Shree Kumar

    2011-01-01

    Photosynthetic, nitrogen-fixing Anabaena strains are native to tropical paddy fields and contribute to the carbon and nitrogen economy of such soils. Genetic engineering was employed to improve the nitrogen biofertilizer potential of Anabaena sp. strain PCC7120. Constitutive enhanced expression of an additional integrated copy of the hetR gene from a light-inducible promoter elevated HetR protein expression and enhanced functional heterocyst frequency in the recombinant strain. The recombinant strain displayed consistently higher nitrogenase activity than the wild-type strain and appeared to be in homeostasis with compatible modulation of photosynthesis and respiration. The enhanced combined nitrogen availability from the recombinant strain positively catered to the nitrogen demand of rice seedlings in short-term hydroponic experiments and supported better growth. The engineered strain is stable, eco-friendly, and useful for environmental application as nitrogen biofertilizer in paddy fields.

  18. BRED: a simple and powerful tool for constructing mutant and recombinant bacteriophage genomes.

    Directory of Open Access Journals (Sweden)

    Laura J Marinelli

    Full Text Available Advances in DNA sequencing technology have facilitated the determination of hundreds of complete genome sequences both for bacteria and their bacteriophages. Some of these bacteria have well-developed and facile genetic systems for constructing mutants to determine gene function, and recombineering is a particularly effective tool. However, generally applicable methods for constructing defined mutants of bacteriophages are poorly developed, in part because of the inability to use selectable markers such as drug resistance genes during viral lytic growth. Here we describe a method for simple and effective directed mutagenesis of bacteriophage genomes using Bacteriophage Recombineering of Electroporated DNA (BRED, in which a highly efficient recombineering system is utilized directly on electroporated phage DNA; no selection is required and mutants can be readily detected by PCR. We describe the use of BRED to construct unmarked gene deletions, in-frame internal deletions, base substitutions, precise gene replacements, and the addition of gene tags.

  19. Heterogeneous recombination among Hepatitis B virus genotypes.

    Science.gov (United States)

    Castelhano, Nadine; Araujo, Natalia M; Arenas, Miguel

    2017-10-01

    The rapid evolution of Hepatitis B virus (HBV) through both evolutionary forces, mutation and recombination, allows this virus to generate a large variety of adapted variants at both intra and inter-host levels. It can, for instance, generate drug resistance or the diverse viral genotypes that currently exist in the HBV epidemics. Concerning the latter, it is known that recombination played a major role in the emergence and genetic diversification of novel genotypes. In this regard, the quantification of viral recombination in each genotype can provide relevant information to devise expectations about the evolutionary trends of the epidemic. Here we measured the amount of this evolutionary force by estimating global and local recombination rates in >4700 HBV complete genome sequences corresponding to nine (A to I) HBV genotypes. Counterintuitively, we found that genotype E presents extremely high levels of recombination, followed by genotypes B and C. On the other hand, genotype G presents the lowest level, where recombination is almost negligible. We discuss these findings in the light of known characteristics of these genotypes. Additionally, we present a phylogenetic network to depict the evolutionary history of the studied HBV genotypes. This network clearly classified all genotypes into specific groups and indicated that diverse pairs of genotypes are derived from a common ancestor (i.e., C-I, D-E and, F-H) although still the origin of this virus presented large uncertainty. Altogether we conclude that the amount of observed recombination is heterogeneous among HBV genotypes and that this heterogeneity can influence on the future expansion of the epidemic. Copyright © 2017 Elsevier B.V. All rights reserved.

  20. The new pLAI (lux regulon based auto-inducible expression system for recombinant protein production in Escherichia coli

    Directory of Open Access Journals (Sweden)

    Nocadello Salvatore

    2012-01-01

    Full Text Available Abstract Background After many years of intensive research, it is generally assumed that no universal expression system can exist for high-level production of a given recombinant protein. Among the different expression systems, the inducible systems are the most popular for their tight regulation. However, induction is in many cases less favorable due to the high cost and/or toxicity of inducers, incompatibilities with industrial scale-up or detrimental growth conditions. Expression systems using autoinduction (or self-induction prove to be extremely versatile allowing growth and induction of recombinant proteins without the need to monitor cell density or add inducer. Unfortunately, almost all the actual auto inducible expression systems need endogenous or induced metabolic changes during the growth to trigger induction, both frequently linked to detrimental condition to cell growth. In this context, we use a simple modular approach for a cell density-based genetic regulation in order to assemble an autoinducible recombinant protein expression system in E. coli. Result The newly designed pLAI expression system places the expression of recombinant proteins in Escherichia coli under control of the regulatory genes of the lux regulon of Vibrio fischeri's Quorum Sensing (QS system. The pLAI system allows a tight regulation of the recombinant gene allowing a negligible basal expression and expression only at high cell density. Sequence optimization of regulative genes of QS of V. fischeri for expression in E. coli upgraded the system to high level expression. Moreover, partition of regulative genes between the plasmid and the host genome and introduction of a molecular safety lock permitted tighter control of gene expression. Conclusion Coupling gene expression to cell density using cell-to-cell communication provides a promising approach for recombinant protein production. The system allows the control of expression of the target recombinant gene

  1. Suppression of SOX18 by siRNA inhibits cell growth and invasion of breast cancer cells.

    Science.gov (United States)

    Zhang, Jianxiang; Ma, Yanmei; Wang, Shoujun; Chen, Fu; Gu, Yuanting

    2016-06-01

    Breast cancer is the most common malignancy in women around the world, and its incidence and mortality rates are still rising. An increasing number of studies have reported that SOX18 plays an important role in various cancers. However, the role of SOX18 in breast cancer remains poorly understood. In this study, we aimed to investigate the biological role and potential molecular mechanism of SOX18 in breast cancer. We found that the mRNA and protein expression levels of SOX18 were prevalently and significantly overexpressed in human breast cancer cell lines. Next, we performed loss-of-function experiments by transfection of two breast cancer cell lines, BT-474 and MCF-7, with SOX18 small interfering RNAs (siRNA). Results showed that SOX18 siRNA transfection significantly suppressed mRNA and protein expression of SOX18 in breast cancer cells. Furthermore, knockdown of SOX18 significantly inhibited cell proliferation and invasion, but promoted apoptosis in breast cancer cells. Importantly, several oncogenic proteins, including the Ras homolog gene family member A (RhoA), platelet-derived growth factor B (PDGFB), Insulin-like growth factor 1 receptor (IGF-1R), and matrix metalloproteinase-7 (MMP-7), were markedly decreased by SOX18 siRNA. Taken together, the results of our study suggest that knockdown of SOX18 inhibits breast cancer cell growth and invasion, possibly by downregulating downstream oncogenic proteins, providing novel insights into the development of breast cancer therapy through targeting of SOX18.

  2. SH2 domains of the p85 alpha subunit of phosphatidylinositol 3-kinase regulate binding to growth factor receptors.

    Science.gov (United States)

    McGlade, C J; Ellis, C; Reedijk, M; Anderson, D; Mbamalu, G; Reith, A D; Panayotou, G; End, P; Bernstein, A; Kazlauskas, A

    1992-01-01

    The binding of cytoplasmic signaling proteins such as phospholipase C-gamma 1 and Ras GTPase-activating protein to autophosphorylated growth factor receptors is directed by their noncatalytic Src homology region 2 (SH2) domains. The p85 alpha regulatory subunit of phosphatidylinositol (PI) 3-kinase, which associates with several receptor protein-tyrosine kinases, also contains two SH2 domains. Both p85 alpha SH2 domains, when expressed individually as fusion proteins in bacteria, bound stably to the activated beta receptor for platelet-derived growth factor (PDGF). Complex formation required PDGF stimulation and was dependent on receptor tyrosine kinase activity. The bacterial p85 alpha SH2 domains recognized activated beta PDGF receptor which had been immobilized on a filter, indicating that SH2 domains contact autophosphorylated receptors directly. Several receptor tyrosine kinases within the PDGF receptor subfamily, including the colony-stimulating factor 1 receptor and the Steel factor receptor (Kit), also associate with PI 3-kinase in vivo. Bacterially expressed SH2 domains derived from the p85 alpha subunit of PI 3-kinase bound in vitro to the activated colony-stimulating factor 1 receptor and to Kit. We infer that the SH2 domains of p85 alpha bind to high-affinity sites on these receptors, whose creation is dependent on receptor autophosphorylation. The SH2 domains of p85 are therefore primarily responsible for the binding of PI 3-kinase to activated growth factor receptors. Images PMID:1372092

  3. Recombinant Vaccinia Viruses Coding Transgenes of Apoptosis-Inducing Proteins Enhance Apoptosis But Not Immunogenicity of Infected Tumor Cells

    Science.gov (United States)

    Tkachenko, Anastasiya; Richter, Vladimir

    2017-01-01

    Genetic modifications of the oncolytic vaccinia virus (VV) improve selective tumor cell infection and death, as well as activation of antitumor immunity. We have engineered a double recombinant VV, coding human GM-CSF, and apoptosis-inducing protein apoptin (VV-GMCSF-Apo) for comparing with the earlier constructed double recombinant VV-GMCSF-Lact, coding another apoptosis-inducing protein, lactaptin, which activated different cell death pathways than apoptin. We showed that both these recombinant VVs more considerably activated a set of critical apoptosis markers in infected cells than the recombinant VV coding GM-CSF alone (VV-GMCSF-dGF): these were phosphatidylserine externalization, caspase-3 and caspase-7 activation, DNA fragmentation, and upregulation of proapoptotic protein BAX. However, only VV-GMCSF-Lact efficiently decreased the mitochondrial membrane potential of infected cancer cells. Investigating immunogenic cell death markers in cancer cells infected with recombinant VVs, we demonstrated that all tested recombinant VVs were efficient in calreticulin and HSP70 externalization, decrease of cellular HMGB1, and ATP secretion. The comparison of antitumor activity against advanced MDA-MB-231 tumor revealed that both recombinants VV-GMCSF-Lact and VV-GMCSF-Apo efficiently delay tumor growth. Our results demonstrate that the composition of GM-CSF and apoptosis-inducing proteins in the VV genome is very efficient tool for specific killing of cancer cells and for activation of antitumor immunity. PMID:28951871

  4. [Improvement of thermal adaptability and fermentation of industrial ethanologenic yeast by genomic DNA mutagenesis-based genetic recombination].

    Science.gov (United States)

    Liu, Xiuying; He, Xiuping; Lu, Ying; Zhang, Borun

    2011-07-01

    Ethanol is an attractive alternative to fossil fuels. Saccharomyces cerevisiae is the most important ethanol producer. However, in the process of industrial production of ethanol, both cell growth and fermentation of ethanologenic S. cerevisiae are dramatically affected by environmental stresses, such as thermal stress. In this study, we improved both the thermotolerance and fermentation performance of industrial ethanologenic S. cerevisiae by combined usage of chemical mutagenesis and genomic DNA mutagenesis-based genetic recombination method. The recombinant S. cerevisiae strain T44-2 could grow at 44 degrees C, 3 degrees C higher than that of the original strain CE6. The survival rate of T44-2 was 1.84 and 1.87-fold of that of CE6 when heat shock at 48 degrees C and 52 degrees C for 1 h respectively. At temperature higher than 37 degrees C, recombinant strain T44-2 always gave higher cell growth and ethanol production than those of strain CE6. Meanwhile, from 30 degrees C to 40 degrees C, recombinant strain T44-2 produces 91.2-83.8 g/L of ethanol from 200 g/L of glucose, which indicated that the recombinant strain T44-2 had both thermotolerance and broad thermal adaptability. The work offers a novel method, called genomic DNA mutagenesis-based genetic recombination, to improve the physiological functions of S. cerevisiae.

  5. Graded Recombination Layers for Multijunction Photovoltaics

    KAUST Repository

    Koleilat, Ghada I.

    2012-06-13

    Multijunction devices consist of a stack of semiconductor junctions having bandgaps tuned across a broad spectrum. In solar cells this concept is used to increase the efficiency of photovoltaic harvesting, while light emitters and detectors use it to achieve multicolor and spectrally tunable behavior. In series-connected current-matched multijunction devices, the recombination layers must allow the hole current from one cell to recombine, with high efficiency and low voltage loss, with the electron current from the next cell. We recently reported a tandem solar cell in which the recombination layer was implemented using a progression of n-type oxides whose doping densities and work functions serve to connect, with negligible resistive loss at solar current densities, the constituent cells. Here we present the generalized conditions for design of efficient graded recombination layer solar devices. We report the number of interlayers and the requirements on work function and doping of each interlayer, to bridge an work function difference as high as 1.6 eV. We also find solutions that minimize the doping required of the interlayers in order to minimize optical absorption due to free carriers in the graded recombination layer (GRL). We demonstrate a family of new GRL designs experimentally and highlight the benefits of the progression of dopings and work functions in the interlayers. © 2012 American Chemical Society.

  6. Polyploidization increases meiotic recombination frequency in Arabidopsis

    Directory of Open Access Journals (Sweden)

    Rehmsmeier Marc

    2011-04-01

    Full Text Available Abstract Background Polyploidization is the multiplication of the whole chromosome complement and has occurred frequently in vascular plants. Maintenance of stable polyploid state over generations requires special mechanisms to control pairing and distribution of more than two homologous chromosomes during meiosis. Since a minimal number of crossover events is essential for correct chromosome segregation, we investigated whether polyploidy has an influence on the frequency of meiotic recombination. Results Using two genetically linked transgenes providing seed-specific fluorescence, we compared a high number of progeny from diploid and tetraploid Arabidopsis plants. We show that rates of meiotic recombination in reciprocal crosses of genetically identical diploid and autotetraploid Arabidopsis plants were significantly higher in tetraploids compared to diploids. Although male and female gametogenesis differ substantially in meiotic recombination frequency, both rates were equally increased in tetraploids. To investigate whether multivalent formation in autotetraploids was responsible for the increased recombination rates, we also performed corresponding experiments with allotetraploid plants showing strict bivalent pairing. We found similarly increased rates in auto- and allotetraploids, suggesting that the ploidy effect is independent of chromosome pairing configurations. Conclusions The evolutionary success of polyploid plants in nature and under domestication has been attributed to buffering of mutations and sub- and neo-functionalization of duplicated genes. Should the data described here be representative for polyploid plants, enhanced meiotic recombination, and the resulting rapid creation of genetic diversity, could have also contributed to their prevalence.

  7. SequenceLDhot: detecting recombination hotspots.

    Science.gov (United States)

    Fearnhead, Paul

    2006-12-15

    There is much local variation in recombination rates across the human genome--with the majority of recombination occurring in recombination hotspots--short regions of around approximately 2 kb in length that have much higher recombination rates than neighbouring regions. Knowledge of this local variation is important, e.g. in the design and analysis of association studies for disease genes. Population genetic data, such as that generated by the HapMap project, can be used to infer the location of these hotspots. We present a new, efficient and powerful method for detecting recombination hotspots from population data. We compare our method with four current methods for detecting hotspots. It is orders of magnitude quicker, and has greater power, than two related approaches. It appears to be more powerful than HotspotFisher, though less accurate at inferring the precise positions of the hotspot. It was also more powerful than LDhot in some situations: particularly for weaker hotspots (10-40 times the background rate) when SNP density is lower (maths.lancs.ac.uk/~fearnhea/Hotspot.

  8. Recombinant IκBα-loaded curcumin nanoparticles for improved cancer therapeutics

    Science.gov (United States)

    Banerjee, Subhamoy; Sahoo, Amaresh Kumar; Chattopadhyay, Arun; Sankar Ghosh, Siddhartha

    2014-08-01

    The field of recombinant protein therapeutics has been evolving rapidly, making significant impact on clinical applications for several diseases, including cancer. However, the functional aspects of proteins rely exclusively on their structural integrity, in which nanoparticle mediated delivery offers unique advantages over free proteins. In the present work, a novel strategy has been developed where the nanoparticles (NPs) used for the delivery of the recombinant protein could contribute to enhancing the therapeutic efficacy of the recombinant protein. The transcription factor, NFκB, involved in cell growth and its inhibitor, IκBα, regulates its proliferation. Another similar naturally available molecule, which inhibits the function of NFκB, is curcumin. Hence, we have developed a ‘green synthesis’ method for preparing water-soluble curcumin nanoparticles to stabilize recombinant IκBα protein. The NPs were characterized by UV-vis and fluorescence spectroscopy, transmission electron microscopy (TEM) and dynamic light scattering before administration into human cervical carcinoma (HeLa) and glioblastoma (U87MG) cells. Experimental results demonstrated that this combined module had enhanced therapeutic efficacy, causing apoptotic cell death, which was confirmed by cytotoxicity assay and flowcytometry analyses. The expression of apoptotic genes studied by semi-quantitative reverse transcription PCR delineated the molecular pathways involved in cell death. Thus, our study revealed that the functional delivery of recombinant IκBα-loaded curcumin NPs has promise as a natural-product-based protein therapeutics against cancer cells.

  9. Recombinant IκBα-loaded curcumin nanoparticles for improved cancer therapeutics

    International Nuclear Information System (INIS)

    Banerjee, Subhamoy; Ghosh, Siddhartha Sankar; Sahoo, Amaresh Kumar; Chattopadhyay, Arun

    2014-01-01

    The field of recombinant protein therapeutics has been evolving rapidly, making significant impact on clinical applications for several diseases, including cancer. However, the functional aspects of proteins rely exclusively on their structural integrity, in which nanoparticle mediated delivery offers unique advantages over free proteins. In the present work, a novel strategy has been developed where the nanoparticles (NPs) used for the delivery of the recombinant protein could contribute to enhancing the therapeutic efficacy of the recombinant protein. The transcription factor, NFκB, involved in cell growth and its inhibitor, IκBα, regulates its proliferation. Another similar naturally available molecule, which inhibits the function of NFκB, is curcumin. Hence, we have developed a ‘green synthesis’ method for preparing water-soluble curcumin nanoparticles to stabilize recombinant IκBα protein. The NPs were characterized by UV–vis and fluorescence spectroscopy, transmission electron microscopy (TEM) and dynamic light scattering before administration into human cervical carcinoma (HeLa) and glioblastoma (U87MG) cells. Experimental results demonstrated that this combined module had enhanced therapeutic efficacy, causing apoptotic cell death, which was confirmed by cytotoxicity assay and flowcytometry analyses. The expression of apoptotic genes studied by semi-quantitative reverse transcription PCR delineated the molecular pathways involved in cell death. Thus, our study revealed that the functional delivery of recombinant IκBα-loaded curcumin NPs has promise as a natural-product-based protein therapeutics against cancer cells. (paper)

  10. A new potential secretion pathway for recombinant proteins in Bacillus subtilis.