WorldWideScience

Sample records for cell aggregation induces

  1. p53 Aggregates penetrate cells and induce the co-aggregation of intracellular p53.

    Directory of Open Access Journals (Sweden)

    Karolyn J Forget

    Full Text Available Prion diseases are unique pathologies in which the infectious particles are prions, a protein aggregate. The prion protein has many particular features, such as spontaneous aggregation, conformation transmission to other native PrP proteins and transmission from an individual to another. Protein aggregation is now frequently associated to many human diseases, for example Alzheimer's disease, Parkinson's disease or type 2 diabetes. A few proteins associated to these conformational diseases are part of a new category of proteins, called prionoids: proteins that share some, but not all, of the characteristics associated with prions. The p53 protein, a transcription factor that plays a major role in cancer, has recently been suggested to be a possible prionoid. The protein has been shown to accumulate in multiple cancer cell types, and its aggregation has also been reproduced in vitro by many independent groups. These observations suggest a role for p53 aggregates in cancer development. This study aims to test the «prion-like» features of p53. Our results show in vitro aggregation of the full length and N-terminally truncated protein (p53C, and penetration of these aggregates into cells. According to our findings, the aggregates enter cells using macropinocytosis, a non-specific pathway of entry. Lastly, we also show that once internalized by the cell, p53C aggregates can co-aggregate with endogenous p53 protein. Together, these findings suggest prion-like characteristics for p53 protein, based on the fact that p53 can spontaneously aggregate, these aggregates can penetrate cells and co-aggregate with cellular p53.

  2. Ralstonia insidiosa induces cell aggregation by Listeria monocytogenes

    Science.gov (United States)

    Biofilm formation is an important strategy for foodborne bacterial pathogens to survive in stressful environments such as fresh produce processing facilities. Bacterial cell aggregation strongly promotes the initiation of microcolonies and the formation of biofilms on abiological surfaces. We previ...

  3. The cell aggregating propensity of probiotic actinobacterial isolates: isolation and characterization of the aggregation inducing peptide pheromone.

    Science.gov (United States)

    Muthu Selvam, Ramu; Vinothini, Gopal; Palliyarai Thaiyammal, Sethuramalingam; Latha, Selvanathan; Chinnathambi, Arunachalam; Dhanasekaran, Dharumadurai; Padmanabhan, Parasuraman; Ali Alharbi, Sulaiman; Archunan, Govindaraju

    2016-01-01

    The auto-aggregating ability of a probiotic is a prerequisite for colonization and protection of the gastrointestinal tract, whereas co-aggregation provides a close interaction with pathogenic bacteria. Peptide pheromone mediated signaling has been studied in several systems. However, it has not yet been explored in prokaryotes, especially actinobacteria. Hence, in the present study, the diffusible aggregation promoting factor was purified from the culture supernatant of a potent actinobacterial probiont and characterized using 20 different actinobacterial cultures isolated from the gut region of chicken and goat. The results showed that the pheromone-like compound induces the aggregation propensity of treated isolates. The factor was found to be a heat stable, acidic pH resistant, low molecular weight peptide which enhances the biofilm forming ability of other actinobacterial isolates. The aggregation promoting factor represents a bacterial sex factor (pheromone) and its characterization confirms its usage in the probiotic formulation.

  4. Cathepsin G Induces Cell Aggregation of Human Breast Cancer MCF-7 Cells via a 2-Step Mechanism: Catalytic Site-Independent Binding to the Cell Surface and Enzymatic Activity-Dependent Induction of the Cell Aggregation

    Directory of Open Access Journals (Sweden)

    Riyo Morimoto-Kamata

    2012-01-01

    Full Text Available Neutrophils often invade various tumor tissues and affect tumor progression and metastasis. Cathepsin G (CG is a serine protease secreted from activated neutrophils. Previously, we have shown that CG induces the formation of E-cadherin-mediated multicellular spheroids of human breast cancer MCF-7 cells; however, the molecular mechanisms involved in this process are unknown. In this study, we investigated whether CG required its enzymatic activity to induce MCF-7 cell aggregation. The cell aggregation-inducing activity of CG was inhibited by pretreatment of CG with the serine protease inhibitors chymostatin and phenylmethylsulfonyl fluoride. In addition, an enzymatically inactive S195G (chymotrypsinogen numbering CG did not induce cell aggregation. Furthermore, CG specifically bound to the cell surface of MCF-7 cells via a catalytic site-independent mechanism because the binding was not affected by pretreatment of CG with serine protease inhibitors, and cell surface binding was also detected with S195G CG. Therefore, we propose that the CG-induced aggregation of MCF-7 cells occurs via a 2-step process, in which CG binds to the cell surface, independently of its catalytic site, and then induces cell aggregation, which is dependent on its enzymatic activity.

  5. Effect of polymer aggregation on the open circuit voltage in organic photovoltaic cells: aggregation-induced conjugated polymer gel and its application for preventing open circuit voltage drop.

    Science.gov (United States)

    Kim, Bong-Gi; Jeong, Eun Jeong; Park, Hui Joon; Bilby, David; Guo, L Jay; Kim, Jinsang

    2011-03-01

    To investigate the structure-dependent aggregation behavior of conjugated polymers and the effect of aggregation on the device performance of conjugated polymer photovoltaic cells, new conjugated polymers (PVTT and CN-PVTT) having the same regioregularity but different intermolecular packing were prepared and characterized by means of UV-vis spectroscopy and atomic force microscopy (AFM). Photovoltaic devices were prepared with these polymers under different polymer-aggregate conditions. Polymer aggregation induced by thermal annealing increases the short circuit current but provides no advantage in the overall power conversion efficiency because of a decrease in the open circuit voltage. The device fabricated from a pre-aggregated polymer suspension, acquired from ultrasonic agitation of a conjugated polymer gel, showed enhanced performance because of better phase separation and reduced recombination between polymer/PCBM.

  6. Cell aggregation and sedimentation.

    Science.gov (United States)

    Davis, R H

    1995-01-01

    The aggregation of cells into clumps or flocs has been exploited for decades in such applications as biological wastewater treatment, beer brewing, antibiotic fermentation, and enhanced sedimentation to aid in cell recovery or retention. More recent research has included the use of cell aggregation and sedimentation to selectively separate subpopulations of cells. Potential biotechnological applications include overcoming contamination, maintaining plasmid-bearing cells in continuous fermentors, and selectively removing nonviable hybridoma cells from perfusion cultures.

  7. Neutrophil Cathepsin G, but Not Elastase, Induces Aggregation of MCF-7 Mammary Carcinoma Cells by a Protease Activity-Dependent Cell-Oriented Mechanism

    Directory of Open Access Journals (Sweden)

    Satoru Yui

    2014-01-01

    Full Text Available We previously found that a neutrophil serine protease, cathepsin G, weakens adherence to culture substrates and induces E-cadherin-dependent aggregation of MCF-7 human breast cancer cells through its protease activity. In this study, we examined whether aggregation is caused by degradation of adhesion molecules on the culture substrates or through an unidentified mechanism. We compared the effect of treatment with cathepsin G and other proteases, including neutrophil elastase against fibronectin- (FN- coated substrates. Cathepsin G and elastase potently degraded FN on the substrates and induced aggregation of MCF-7 cells that had been subsequently seeded onto the substrate. However, substrate-bound cathepsin G and elastase may have caused cell aggregation. After inhibiting the proteases on the culture substrates using the irreversible inhibitor phenylmethylsulfonyl fluoride (PMSF, we examined whether aggregation of MCF-7 cells was suppressed. PMSF attenuated cell aggregation on cathepsin G-treated substrates, but the effect was weak in cells pretreated with high concentrations of cathepsin G. In contrast, PMSF did not suppress cell aggregation on elastase-treated FN. Moreover, cathepsin G, but not elastase, induced aggregation on poly-L-lysine substrates which are not decomposed by these enzymes, and the action of cathepsin G was nearly completely attenuated by PMSF. These results suggest that cathepsin G induces MCF-7 aggregation through a cell-oriented mechanism.

  8. Nestin+cells forming spheroids aggregates resembling tumorspheres in experimental ENU-induced gliomas.

    Science.gov (United States)

    García-Blanco, Alvaro; Bulnes, Susana; Pomposo, Iñigo; Carrasco, Alex; Lafuente, José Vicente

    2016-12-01

    Nestin+cells from spheroid aggregates display typical histopathological features compatible with cell stemness. Nestin and CD133+cells found in glioblastomas, distributed frequently around aberrant vessels, are considered as potential cancer stem cells. They are possible targets for antitumoral therapy because they lead the tumorigenesis, invasiveness and angiogenesis. However, little is known about their role and presence in low-grade gliomas. The aim of this work is to localize and characterize the distribution of these cells inside tumors during the development of experimental endogenous glioma. For this study, a single dose of Ethyl-nitrosourea was injected into pregnant rats. Double immunofluorescences were performed in order to identify stem-like and differentiated cells. Low-grade gliomas display Nestin+cells distributed throughout the tumor. More malignant gliomas show, in addition to that, a perivascular location with some Nestin+cells co-expressing CD133 or VEGF, and the intratumoral spheroid aggregates of Nestin/CD133+cells. These structures are encapsulated by well-differentiated VEGF/GFAP+cells. Spheroid aggregates increase in size in the most malignant stages. Spheroid aggregates have morphological and phenotypic similarities to in vitro neurospheres and could be an in vivo analogue of them. These arrangements could be a reservoir of undifferentiated cells formed to escape adverse microenvironments.

  9. Differential inhibition of tumour cell-induced platelet aggregation by the nicotinate aspirin prodrug (ST0702) and aspirin

    Science.gov (United States)

    Medina, Carlos; Harmon, Shona; Inkielewicz, Iwona; Santos-Martinez, Maria Jose; Jones, Michael; Cantwell, Paula; Bazou, Despina; Ledwidge, Mark; Radomski, Marek W; Gilmer, John F

    2012-01-01

    BACKGROUND AND PURPOSE Tumour cell-induced platelet aggregation (TCIPA) facilitates cancer cell invasion, angiogenesis and the formation of metastatic foci. TCIPA can be modulated by pharmacological inhibitors of MMP-2 and ADP; however, the COX inhibitor aspirin did not prevent TCIPA. In this study, we have tested the pharmacological effects of a new group of isosorbide-based aspirin prodrugs on TCIPA. EXPERIMENTAL APPROACH TCIPA was induced in human platelets by mixing with human adenocarcinoma or fibrosarcoma cells under no flow and flow conditions. The release of gelatinases and P-selectin expression during TCIPA were studied by zymography and flow cytometry respectively. KEY RESULTS Tumour cells caused platelet aggregation. This aggregation resulted in the release of MMP-2 and a significant up-regulation of P-selectin on platelets, indicative of platelet activation. Pharmacological modulation of TCIPA revealed that ST0702, one of the aspirin prodrugs, down-regulated TCIPA while aspirin was ineffective. The deacetylated metabolite of ST0702, 5-nicotinate salicylate (ST0702 salicylate), down-regulated both ADP-stimulated platelet aggregation and TCIPA. CONCLUSIONS AND IMPLICATIONS Our results show that ST0702 was an effective inhibitor of TCIPA in vitro. Its deacetylated metabolite may contribute to the effects of ST0702 by inhibiting ADP-mediated TCIPA. PMID:22122360

  10. Solvents induced ZnO nanoparticles aggregation associated with their interfacial effect on organic solar cells.

    Science.gov (United States)

    Li, Pandeng; Jiu, Tonggang; Tang, Gang; Wang, Guojie; Li, Jun; Li, Xiaofang; Fang, Junfeng

    2014-10-22

    ZnO nanofilm as a cathode buffer layer has surface defects due to the aggregations of ZnO nanoparticles, leading to poor device performance of organic solar cells. In this paper, we report the ZnO nanoparticles aggregations in solution can be controlled by adjusting the solvents ratios (chloroform vs methanol). These aggregations could influence the morphology of ZnO film. Therefore, compact and homogeneous ZnO film can be obtained to help achieve a preferable power conversion efficiency of 8.54% in inverted organic solar cells. This improvement is attributed to the decreased leakage current and the increased electron-collecting efficiency as well as the improved interface contact with the active layer. In addition, we find the enhanced maximum exciton generation rate and exciton dissociation probability lead to the improvement of device performance due to the preferable ZnO dispersion. Compared to other methods of ZnO nanofilm fabrication, it is the more convenient, moderate, and effective to get a preferable ZnO buffer layer for high-efficiency organic solar cells.

  11. CD147 and CD98 complex-mediated homotypic aggregation attenuates the CypA-induced chemotactic effect on Jurkat T cells.

    Science.gov (United States)

    Guo, Na; Zhang, Kui; Lv, Minghua; Miao, Jinlin; Chen, Zhinan; Zhu, Ping

    2015-02-01

    Homotypic cell aggregation plays important roles in physiological and pathological processes, including embryogenesis, immune responses, angiogenesis, tumor cell invasion and metastasis. CD147 has been implicated in most of these phenomena, and it was identified as a T cell activation-associated antigen due to its obvious up-regulation in activated T cells. However, the explicit function and mechanism of CD147 in T cells have not been fully elucidated. In this study, large and compact aggregates were observed in Jurkat T cells after treatment with the specific CD147 monoclonal antibody HAb18 or after the expression of CD147 was silenced by RNA interference, which indicated an inhibitory effect of CD147 in T cell homotypic aggregation. Knocking down CD147 expression resulted in a significant decrease in CD98, along with prominent cell aggregation, similar to that treated by CD98 and CD147 monoclonal antibodies. Furthermore, decreased cell chemotactic activity was observed following CD147- and CD98-mediated cell aggregation, and increased aggregation was correlated with a decrease in the chemotactic ability of the Jurkat T cells, suggesting that CD147- and CD98-mediated homotypic cell aggregation plays a negative role in T cell chemotaxis. Our data also showed that p-ERK, p-ZAP70, p-CD3ζ and p-LCK were significantly decreased in the CD147- and CD98-knocked down Jurkat T cells, which suggested that decreased CD147- and/or CD98-induced homotypic T cell aggregation and aggregation-inhibited chemotaxis might be associated with these signaling pathways. A role for CD147 in cell aggregation and chemotaxis was further indicated in primary CD4(+) T cells. Similarly, low expression of CD147 in primary T cells induced prominent cell aggregation and this aggregation attenuated primary T cell chemotactic ability in response to CypA. Our results have demonstrated the correlation between homotypic cell aggregation and the chemotactic response of T cells to CypA, and these data

  12. In vitro cytotoxicity of fluorescent silica nanoparticles hybridized with aggregation-induced emission luminogens for living cell imaging.

    Science.gov (United States)

    Xia, Yun; Li, Min; Peng, Tao; Zhang, Weijie; Xiong, Jun; Hu, Qinggang; Song, Zifang; Zheng, Qichang

    2013-01-07

    Fluorescent silica nanoparticles (FSNPs) can provide high-intensity and photostable fluorescent signals as a probe for biomedical analysis. In this study, FSNPs hybridized with aggregation-induced emission (AIE) luminogens (namely FSNP-SD) were successfully fabricated by a surfactant-free sol-gel method. The FSNP-SD were spherical, monodisperse and uniform in size, with an average diameter of approximately 100 nm, and emitted strong fluorescence at the peak of 490 nm. The FSNP-SD selectively stained the cytoplasmic regions and were distributed in the cytoplasm. Moreover, they can stay inside cells, enabling the tacking of cells over a long period of time. The intracellular vesicles and multinucleated cells were increase gradually with the rise of FSNP-SD concentration. Both cell viability and survival only lost less than 20% when the cells were exposed to the high concentration of 100 μg/mL FSNP-SD. Additionally, the cell apoptosis and intracellular ROS assay indicated that FSNP-SD had no significant toxic effects at the maximum working concentration of 80 μg/mL. This study demonstrated that the FSNP-SD are promising biocompatible fluorescent probes for living cell imaging.

  13. In Vitro Cytotoxicity of Fluorescent Silica Nanoparticles Hybridized with Aggregation-Induced Emission Luminogens for Living Cell Imaging

    Directory of Open Access Journals (Sweden)

    Yun Xia

    2013-01-01

    Full Text Available Fluorescent silica nanoparticles (FSNPs can provide high-intensity and photostable fluorescent signals as a probe for biomedical analysis. In this study, FSNPs hybridized with aggregation-induced emission (AIE luminogens (namely FSNP-SD were successfully fabricated by a surfactant-free sol-gel method. The FSNP-SD were spherical, monodisperse and uniform in size, with an average diameter of approximately 100 nm, and emitted strong fluorescence at the peak of 490 nm. The FSNP-SD selectively stained the cytoplasmic regions and were distributed in the cytoplasm. Moreover, they can stay inside cells, enabling the tacking of cells over a long period of time. The intracellular vesicles and multinucleated cells were increase gradually with the rise of FSNP-SD concentration. Both cell viability and survival only lost less than 20% when the cells were exposed to the high concentration of 100 μg/mL FSNP-SD. Additionally, the cell apoptosis and intracellular ROS assay indicated that FSNP-SD had no significant toxic effects at the maximum working concentration of 80 μg/mL. This study demonstrated that the FSNP-SD are promising biocompatible fluorescent probes for living cell imaging.

  14. Inflammation Induces TDP-43 Mislocalization and Aggregation.

    Directory of Open Access Journals (Sweden)

    Ana Sofia Correia

    Full Text Available TAR DNA-binding protein 43 (TDP-43 is a major component in aggregates of ubiquitinated proteins in amyotrophic lateral sclerosis (ALS and frontotemporal lobar degeneration (FTLD. Here we report that lipopolysaccharide (LPS-induced inflammation can promote TDP-43 mislocalization and aggregation. In culture, microglia and astrocytes exhibited TDP-43 mislocalization after exposure to LPS. Likewise, treatment of the motoneuron-like NSC-34 cells with TNF-alpha (TNF-α increased the cytoplasmic levels of TDP-43. In addition, the chronic intraperitoneal injection of LPS at a dose of 1mg/kg in TDP-43(A315T transgenic mice exacerbated the pathological TDP-43 accumulation in the cytoplasm of spinal motor neurons and it enhanced the levels of TDP-43 aggregation. These results suggest that inflammation may contribute to development or exacerbation of TDP-43 proteinopathies in neurodegenerative disorders.

  15. Enhanced aggregation of androgen receptor in induced pluripotent stem cell-derived neurons from spinal and bulbar muscular atrophy.

    Science.gov (United States)

    Nihei, Yoshihiro; Ito, Daisuke; Okada, Yohei; Akamatsu, Wado; Yagi, Takuya; Yoshizaki, Takahito; Okano, Hideyuki; Suzuki, Norihiro

    2013-03-22

    Spinal and bulbar muscular atrophy (SBMA) is an X-linked motor neuron disease caused by a CAG repeat expansion in the androgen receptor (AR) gene. Ligand-dependent nuclear accumulation of mutant AR protein is a critical characteristic of the pathogenesis of SBMA. SBMA has been modeled in AR-overexpressing animals, but precisely how the polyglutamine (polyQ) expansion leads to neurodegeneration is unclear. Induced pluripotent stem cells (iPSCs) are a new technology that can be used to model human diseases, study pathogenic mechanisms, and develop novel drugs. We established SBMA patient-derived iPSCs, investigated their cellular biochemical characteristics, and found that SBMA-iPSCs can differentiate into motor neurons. The CAG repeat numbers in the AR gene of SBMA-iPSCs and also in the atrophin-1 gene of iPSCs derived from another polyQ disease, dentato-rubro-pallido-luysian atrophy (DRPLA), remain unchanged during reprogramming, long term passage, and differentiation, indicating that polyQ disease-associated CAG repeats are stable during maintenance of iPSCs. The level of AR expression is up-regulated by neuronal differentiation and treatment with the AR ligand dihydrotestosterone. Filter retardation assays indicated that aggregation of ARs following dihydrotestosterone treatment in neurons derived from SBMA-iPSCs increases significantly compared with neurological control iPSCs, easily recapitulating the pathological feature of mutant ARs in SBMA-iPSCs. This phenomenon was not observed in iPSCs and fibroblasts, thereby showing the neuron-dominant phenotype of this disease. Furthermore, the HSP90 inhibitor 17-allylaminogeldanamycin sharply decreased the level of aggregated AR in neurons derived from SBMA-iPSCs, indicating a potential for discovery and validation of candidate drugs. We found that SBMA-iPSCs possess disease-specific biochemical features and could thus open new avenues of research into not only SBMA, but also other polyglutamine diseases.

  16. A multifunctional probe with aggregation-induced emission characteristics for selective fluorescence imaging and photodynamic killing of bacteria over mammalian cells.

    Science.gov (United States)

    Gao, Meng; Hu, Qinglian; Feng, Guangxue; Tomczak, Nikodem; Liu, Rongrong; Xing, Bengang; Tang, Ben Zhong; Liu, Bin

    2015-04-02

    A multifunctional probe aggregation-induced emission-Zinc(II)-dipicolylamine (AIE-ZnDPA) is developed for selective targeting, fluorescence imaging, and photodynamic killing of both Gram-positive and Gram-negative bacteria over mammalian cells. The probe has significant advantages in simple probe design, enhanced fluorescence upon bacteria binding, excellent photostability, and broad-spectrum antibacterial activity with almost no harm to mammalian cells.

  17. Antigen 43 from Escherichia coli induces inter- and intraspecies cell aggregation and changes in colony morphology of Pseudomonas fluorescens

    DEFF Research Database (Denmark)

    Kjærgaard, Kristian; Schembri, Mark; Hasman, Henrik;

    2000-01-01

    Antigen 43 (Ag43) is a surface-displayed autotransporter protein of Escherichia coli. By virtue of its self-association characteristics, this protein is able to mediate autoaggregation and flocculation off. coli cells in static cultures. Additionally, surface display of Ag43 is associated...... with a distinct frizzy colony morphology in E. coli. Here we show that Ag43 can be expressed in a functional form on the surface of the environmentally important Pseudomonas fluorescens strain SBW25 with ensuing cell aggregation and frizzy colony types. Using green fluorescence protein-tagged cells, we...... demonstrate that Ag43 can be used as a tool to provide interspecies cell aggregation between E. coli and P. fluorescens. Furthermore, Ag43 expression enhances biofilm formation in P. fluorescens to glass surfaces. The versatility of this protein was also reflected in Ag43 surface display in a variety of other...

  18. Investigating the feasibility of scale up and automation of human induced pluripotent stem cells cultured in aggregates in feeder free conditions.

    Science.gov (United States)

    Soares, Filipa A C; Chandra, Amit; Thomas, Robert J; Pedersen, Roger A; Vallier, Ludovic; Williams, David J

    2014-03-10

    The transfer of a laboratory process into a manufacturing facility is one of the most critical steps required for the large scale production of cell-based therapy products. This study describes the first published protocol for scalable automated expansion of human induced pluripotent stem cell lines growing in aggregates in feeder-free and chemically defined medium. Cells were successfully transferred between different sites representative of research and manufacturing settings; and passaged manually and using the CompacT SelecT automation platform. Modified protocols were developed for the automated system and the management of cells aggregates (clumps) was identified as the critical step. Cellular morphology, pluripotency gene expression and differentiation into the three germ layers have been used compare the outcomes of manual and automated processes.

  19. Neuroprotection of inositol hexaphosphate and changes of mitochondrion mediated apoptotic pathway and α-synuclein aggregation in 6-OHDA induced parkinson's disease cell model.

    Science.gov (United States)

    Zhang, Zheng; Hou, Lin; Li, Xianghong; Ju, Chuanxia; Zhang, Jinyu; Li, Xin; Wang, Xiuli; Liu, Cun; Lv, Yuqiang; Wang, Yuehua

    2016-02-15

    Animal and cell experiments showed that inositol hexaphosphate (IP6) was protective on neurons in parkinson's disease (PD) model, but the underlying mechanism of this action was not extensively elucidated. To address this question, we established 6-hydroxydopamine (6-OHDA) induced human dopaminergic cell line SH-SY5Y as PD cell model and testified the neuroprotection of IP6. Through hoechst nuclear stain method and flow cytometric analysis, apoptosis induced by 6-OHDA was blocked by IP6 pretreatment. Significant protection against reactive oxygen species (ROS) and lipid peroxidation product malondialdehyde (MDA) was observed in 6-OHDA induced cells pretreated with IP6. To further investigate the mechanism of anti-apoptotic effect of IP6, expression of mediators in mitochondrion dependent apoptotic pathway was detected. Results indicated that loss of mitochondrial membrane potential, cytochrome c releasing, upregulation of Bcl-2-associated X protein (Bax), downregulation of B-cell CLL/lymphoma 2 (Bcl-2) and caspases activation were reversed by IP6. In addition, using flow cytometric method and western blot approach, our data showed that IP6 attenuated the rise of calcium and α-synuclein aggregation in cytosol. Collectively, IP6 exerted its neuroprotection on dopaminergic cells in PD cell model and the mechanism may be associated with changes of mitochondrion mediated apoptotic pathway and α-synuclein aggregation.

  20. Inhibition of β-amyloid Aggregation By Albiflorin, Aloeemodin And Neohesperidin And Their Neuroprotective Effect On Primary Hippocampal Cells Against β-amyloid Induced Toxicity.

    Science.gov (United States)

    Ho, See-Lok; Poon, Chung-Yan; Lin, Chengyuan; Yan, Ting; Kwong, Daniel Wai-Jing; Yung, Ken Kin-Lam; Wong, Man S; Bian, Zhaoxiang; Li, Hung-Wing

    2015-01-01

    Being one of the hallmarks of Alzheimer's disease, β-amyloid (Aβ) aggregates induce complicated neurotoxicity. Evidences show that the underlying mechanism of neurotoxicity involves a glutamate receptor subtype, N-methyl-D-aspartate (NMDA) receptor, an increase in intracellular calcium(II) ion loading as well as an elevation in oxidation stress. In this work, among the 35 chemical components of Chinese herbal medicines (CHMs) being screened for inhibitors of Aβ aggregation, four of them, namely albiflorin, aloeemodin, neohesperidin and physcion, were found for the first time to exhibit a potent inhibitory effect on Aβ(1-40) and Aβ(1-42) aggregation. Their neuroprotective capability on primary hippocampal neuronal cells was also investigated by MTT assay, ROS assay and intracellular calcium(II) ion concentration measurement. It was interesting to find that physcion was rather toxic to neuronal cells while albiflorin, aloeemodin and neohesperidin reduced the toxicity and ROS induced by both monomeric and oligomeric Aβ species. In addition, albiflorin was particularly powerful in maintaining the intracellular Ca(2+) concentration.

  1. A pyrene-benzthiazolium conjugate portraying aggregation induced emission, a ratiometric detection and live cell visualization of HSO{sub 3}{sup −}

    Energy Technology Data Exchange (ETDEWEB)

    Diwan, Uzra; Kumar, Virendra [Department of Chemistry (Centre of Advanced Study), Faculty of Science, Banaras Hindu University, Varanasi, Uttar Pradesh 221005 (India); Mishra, Rakesh K. [Photosciences and Photonics, Chemical Sciences and Technology Division, CSIR–National Institute for Interdisciplinary Science and Technology, Thiruvananthapuram 695019 (India); Rana, Nishant Kumar; Koch, Biplob; Singh, Manish Kumar [Department of Zoology (Centre of Advanced Study), Faculty of Science, Banaras Hindu University, Varanasi 221005 (India); Upadhyay, K.K., E-mail: drkaushalbhu@yahoo.co.in [Department of Chemistry (Centre of Advanced Study), Faculty of Science, Banaras Hindu University, Varanasi, Uttar Pradesh 221005 (India)

    2016-07-27

    The present study deals with the photophysical property of a pyrene-benzthiazolium conjugate R1, as a strong intramolecular charge transfer (ICT) probe exhibiting long wavelength emission in the red region. Unlike traditional planar polyaromatic hydrocarbons whose aggregation generally quenches the light emission, the pyrene based R1 was found to display aggregation-induced emission (AIE) property along with simultaneous increase in its quantum yield upon increasing the water content of the medium. The R1 exhibits high specificity towards HSO{sub 3}{sup −}/SO{sub 3}{sup 2−} by interrupting its own ICT producing there upon a large ratiometric blue shift of ∼220 nm in its emission spectrum. The lowest detection limit for the above measurement was found to be 8.90 × 10{sup −8} M. The fluorescent detection of HSO{sub 3}{sup −} was also demonstrated excellently by test paper strip and silica coated TLC plate incorporating R1. The live cell imaging of HSO{sub 3}{sup −} through R1 in HeLa cells was studied using fluorescence microscopic studies. The particle size and morphological features of R1 and R1-HSO{sub 3}{sup −} aggregates in aqueous solution were characterized by DLS along with SEM analysis.- Highlights: • A pyrene-benzthiazolium conjugate probe (R1) itself showed interesting phenomenon of an aggregation-induced emission (AIE). • R1 emits in the red channel and effectively utilized as a colorimetric and ratiometric fluorescent sensor for HSO{sub 3}{sup −}. • The nano-dimensional spherical particles of R1 got enlarged upon its interaction with the HSO{sub 3}{sup −}. • R1 can efficiently stain HSO{sub 3}{sup −} in live cells and can be used for the on-spot detection of the same.

  2. Control of aggregation-induced emission by DNA hybridization

    OpenAIRE

    Li, Shaoguang; Langenegger, Simon Matthias; Häner, Robert

    2013-01-01

    Aggregation-induced emission (AIE) was studied by hybridization of dialkynyl-tetraphenylethylene (DATPE) modified DNA strands. Molecular aggregation and fluorescence of DATPEs are controlled by duplex formation.

  3. Cucurbitacin B induces rapid depletion of the G-actin pool through reactive oxygen species-dependent actin aggregation in melanoma cells

    Institute of Scientific and Technical Information of China (English)

    Yanting Zhang; Dongyun Ouyang; Lihui Xu; Yuhua Ji; Qingbing Zha; Jiye Cai; Xianhui He

    2011-01-01

    Cucurbitacin B (CuB), a triterpenoid compound isolated from Cucurbitaceae plants, has been reported as a promising anti-cancer agent, yet its action mechanism is still controversial. In this study, we explored the potential mechanism of CuB in murine B16F10 melanoma cells.Anti-proliferation and anti-invasion effects were assessed in cultured cells, and in vivo anti-tumor activity was evaluated in a murine subcutaneous melanoma model. Flow cytometry was adopted to analyze cell cycle distribution and reactive oxygen species (ROS) levels. Actin levels were determined by western blot analysis, and the profiles of differential expressed proteins were identified by a quantitative proteomic approach. The results showed that CuB exerted inhibitory effects on cell proliferation, colony formation, as well as migration and invasion potential of the melanoma cells. The growth of subcutaneous melanoma was significantly inhibited in mice treated with CuB when compared with control group. Furthermore,CuB treatment caused rapid cell membrane blebbing and deformation, and induced G2/M-phase arrest and formation of multiploid cells. Notably, the G-actin pool was rapidly depleted and actin aggregates were formed quickly after CuB treatment. A number of cytoskeleton-regulatory proteins were differentially regulated. Blockage of ROS production significantly reduced the G-actin depletion ability and the anti-tumor activity of CuB. These findings indicate that CuB induces rapid depletion of the G-actin pool through ROS-dependent actin aggregation in melanoma cells, which may at least partly account for its anti-tumor activity.

  4. Distinguishing aggregate formation and aggregate clearance using cell based assays

    NARCIS (Netherlands)

    E. Eenjes, E.; J.M. Dragich; H. Kampinga (Harm); A. Yamamoto, A.

    2016-01-01

    textabstractThe accumulation of ubiquitinated proteinaceous inclusions represents a complex process, reflecting the disequilibrium between aggregate formation and aggregate clearance. Although decreasing aggregate formation or augmenting aggregate clearance will ultimately lead to diminished aggrega

  5. CD40-induced aggregation of MHC class II and CD80 on the cell surface leads to an early enhancement in antigen presentation.

    Science.gov (United States)

    Clatza, Abigail; Bonifaz, Laura C; Vignali, Dario A A; Moreno, José

    2003-12-15

    Ligation of CD40 on B cells increases their ability to present Ag and to activate MHC class II (MHC-II)-restricted T cells. How this occurs is not entirely clear. In this study we demonstrate that CD40 ligation on Ag-presenting B cells (APC) for a short period between 30 min and 3 h has a rapid, augmenting effect on the ability of a B cell line and normal B cells to activate T cells. This is not due to alterations in Ag processing or to an increase in surface expression of CD80, CD86, ICAM-1, or MHC-II. This effect is particularly evident with naive, resting T lymphocytes and appears to be more pronounced under limiting Ag concentrations. Shortly after CD40 ligation on a B cell line, MHC-II and CD80 progressively accumulated in cholesterol-enriched microdomains on the cell surface, which correlated with an initial enhancement in their Ag presentation ability. Moreover, CD40 ligation induced a second, late, more sustained enhancement of Ag presentation, which correlates with a significant increase in CD80 expression by APC. Thus, CD40 signaling enhances the efficiency with which APC activate T cells by at least two related, but distinct, mechanisms: an early stage characterized by aggregation of MHC-II and CD80 clusters, and a late stage in which a significant increase in CD80 expression is observed. These results raise the possibility that one important role of CD40 is to contribute to the formation of the immunological synapse on the APC side.

  6. Tracking Hypoxic Signaling within Encapsulated Cell Aggregates

    Science.gov (United States)

    Skiles, Matthew L.; Sahai, Suchit; Blanchette, James O.

    2011-01-01

    nutrients, notably oxygen, is therefore reduced and limited by diffusion. This reduced oxygen availability may especially impact β-cells whose insulin secretory function is highly dependent on oxygen11-13. Capsule composition and geometry will also impact diffusion rates and lengths for oxygen. Therefore, we also describe a technique for identifying hypoxic cells within our PEG capsules. Infection of the cells with a recombinant adenovirus allows for a fluorescent signal to be produced when intracellular hypoxia-inducible factor (HIF) pathways are activated14. As HIFs are the primary regulators of the transcriptional response to hypoxia, they represent an ideal target marker for detection of hypoxic signaling15. This approach allows for easy and rapid detection of hypoxic cells. Briefly, the adenovirus has the sequence for a red fluorescent protein (Ds Red DR from Clontech) under the control of a hypoxia-responsive element (HRE) trimer. Stabilization of HIF-1 by low oxygen conditions will drive transcription of the fluorescent protein (Figure 1). Additional details on the construction of this virus have been published previously15. The virus is stored in 10% glycerol at -80° C as many 150 μL aliquots in 1.5 mL centrifuge tubes at a concentration of 3.4 x 1010 pfu/mL. Previous studies in our lab have shown that MIN6 cells encapsulated as aggregates maintain their viability throughout 4 weeks of culture in 20% oxygen. MIN6 aggregates cultured at 2 or 1% oxygen showed both signs of necrotic cells (still about 85-90% viable) by staining with ethidium bromide as well as morphological changes relative to cells in 20% oxygen. The smooth spherical shape of the aggregates displayed at 20% was lost and aggregates appeared more like disorganized groups of cells. While the low oxygen stress does not cause a pronounced drop in viability, it is clearly impacting MIN6 aggregation and function as measured by glucose-stimulated insulin secretion15. Western blot analysis of encapsulated

  7. Tracking hypoxic signaling within encapsulated cell aggregates.

    Science.gov (United States)

    Skiles, Matthew L; Sahai, Suchit; Blanchette, James O

    2011-12-16

    , is therefore reduced and limited by diffusion. This reduced oxygen availability may especially impact β-cells whose insulin secretory function is highly dependent on oxygen. Capsule composition and geometry will also impact diffusion rates and lengths for oxygen. Therefore, we also describe a technique for identifying hypoxic cells within our PEG capsules. Infection of the cells with a recombinant adenovirus allows for a fluorescent signal to be produced when intracellular hypoxia-inducible factor (HIF) pathways are activated. As HIFs are the primary regulators of the transcriptional response to hypoxia, they represent an ideal target marker for detection of hypoxic signaling. This approach allows for easy and rapid detection of hypoxic cells. Briefly, the adenovirus has the sequence for a red fluorescent protein (Ds Red DR from Clontech) under the control of a hypoxia-responsive element (HRE) trimer. Stabilization of HIF-1 by low oxygen conditions will drive transcription of the fluorescent protein (Figure 1). Additional details on the construction of this virus have been published previously. The virus is stored in 10% glycerol at -80° C as many 150 μL aliquots in 1.5 mL centrifuge tubes at a concentration of 3.4 x 10(10) pfu/mL. Previous studies in our lab have shown that MIN6 cells encapsulated as aggregates maintain their viability throughout 4 weeks of culture in 20% oxygen. MIN6 aggregates cultured at 2 or 1% oxygen showed both signs of necrotic cells (still about 85-90% viable) by staining with ethidium bromide as well as morphological changes relative to cells in 20% oxygen. The smooth spherical shape of the aggregates displayed at 20% was lost and aggregates appeared more like disorganized groups of cells. While the low oxygen stress does not cause a pronounced drop in viability, it is clearly impacting MIN6 aggregation and function as measured by glucose-stimulated insulin secretion. Western blot analysis of encapsulated cells in 20% and 1% oxygen also

  8. Aggregation of Red Blood Cells: From Rouleaux to Clot Formation

    CERN Document Server

    Wagner, C; Svetina, S

    2013-01-01

    Red blood cells are known to form aggregates in the form of rouleaux. This aggregation process is believed to be reversible, but there is still no full understanding on the binding mechanism. There are at least two competing models, based either on bridging or on depletion. We review recent experimental results on the single cell level and theoretical analyses of the depletion model and of the influence of the cell shape on the binding strength. Another important aggregation mechanism is caused by activation of platelets. This leads to clot formation which is life saving in the case of wound healing but also a major cause of death in the case of a thrombus induced stroke. We review historical and recent results on the participation of red blood cells in clot formation.

  9. Serum replacement with albumin-associated lipids prevents excess aggregation and enhances growth of induced pluripotent stem cells in suspension culture.

    Science.gov (United States)

    Horiguchi, Ikki; Sakai, Yasuyuki

    2016-07-08

    Suspension culture systems are currently under investigation for the mass production of pluripotent stem (PS) cells for tissue engineering; however, the control of cell aggregation in suspension culture remains challenging. Existing methods to control aggregation such as microwell culture are difficult to scale up. To address this issue, in this study a novel method that incorporates the addition of KnockOut Serum Replacement (KSR) to the PS cell culture medium was described. The method regulated cellular aggregation and significantly improved cell growth (a 2- to 10-fold increase) without any influence on pluripotency. In addition, albumin-associated lipids as the major working ingredient of KSR responsible for this inhibition of aggregation were identified. This is one of the simplest methods described to date to control aggregation and requires only chemically synthesizable reagents. Thus, this method has the potential to simplify the mass production process of PS cells and thus lower their cost. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:1009-1016, 2016.

  10. Aluminum induces tau aggregation in vitro but not in vivo.

    Science.gov (United States)

    Mizoroki, Tatsuya; Meshitsuka, Shunsuke; Maeda, Sumihiro; Murayama, Miyuki; Sahara, Naruhiko; Takashima, Akihiko

    2007-07-01

    Etiological studies suggest that aluminum (Al) intake might increase an individual's risk of developing Alzheimer's disease (AD). Biochemical analysis data on the effects of Al, however, are inconsistent. Hence, the pathological involvement of Al in AD remains unclear. If Al is involved in AD, then it is reasonable to hypothesize that Al might be involved in the formation of either amyloid plaques or neurofibrillary tangles (NFTs). Here, we investigated whether Al might be involved in NFT formation by using an in vitro tau aggregation paradigm, a tau-overexpressing neuronal cell line (N2a), and a tau-overexpressing mouse model. Although Al induced tau aggregation in a heparin-induced tau assembly assay, these aggregates were neither thioflavin T positive nor did they resemble tau fibrils seen in human AD brains. With cell lysates from stable cell lines overexpressing tau, the accumulation of SDS-insoluble tau increased when the lysates were treated with at least 100 muM Al-maltolate. Yet Al-maltolate caused illness or death in transgenic mice overexpressing human tau and in non-transgenic littermates well before the Al concentration in the brain reached 100 muM. These results indicate that Al has no direct link to AD pathology.

  11. Orthogonal cross-seeding: an approach to explore protein aggregates in living cells.

    Science.gov (United States)

    Hinz, Justyna; Gierasch, Lila M; Ignatova, Zoya

    2008-04-08

    Protein aggregation is associated with the pathology of many diseases, especially neurodegenerative diseases. A variety of structurally polymorphic aggregates or preaggregates including amyloid fibrils is accessible to any aggregating protein. Preaggregates are now believed to be the toxic culprits in pathologies rather than mature aggregates. Although clearly valuable, understanding the mechanism of formation and the structural characteristics of these prefibrillar species is currently lacking. We report here a simple new approach to map the nature of the aggregate core of transient aggregated species directly in the cell. The method is conceptually based on the highly discriminating ability of aggregates to recruit new monomeric species with equivalent molecular structure. Different soluble segments comprising parts of an amyloidogenic protein were transiently pulse-expressed in a tightly controlled, time-dependent manner along with the parent aggregating full-length protein, and their recruitment into the insoluble aggregate was monitored immunochemically. We used this approach to determine the nature of the aggregate core of the metastable aggregate species formed during the course of aggregation of a chimera containing a long polyglutamine repeat tract in a bacterial host. Strikingly, we found that different segments of the full-length protein dominated the aggregate core at different times during the course of aggregation. In its simplicity, the approach is also potentially amenable to screen also for compounds that can reshape the aggregate core and induce the formation of alternative nonamyloidogenic species.

  12. Thrombolytic therapy reduces red blood cell aggregation in plasma without affecting intrinsic aggregability.

    Science.gov (United States)

    Ben-Ami, R; Sheinman, G; Yedgar, S; Eldor, A; Roth, A; Berliner, A S; Barshtein, G

    2002-03-15

    Red blood cell (RBC) aggregation may contribute to occlusion of the coronary microcirculation during myocardial infarction. We studied the effect of thrombolytic therapy on RBC aggregation in patients with acute myocardial infarction (AMI). Compared with patients with myocardial infarction who did not receive thrombolytic therapy, those treated with systemic thrombolysis exhibited significantly reduced RBC aggregation, reduced plasma fibrinogen levels and increased plasma D-dimer levels. Using measurement of RBC aggregation in a standardized dextran-500 solution, reduction in RBC aggregation after thrombolysis was shown to be plasma dependent. Thrombolytic therapy had no direct effect on intrinsic RBC aggregability in patients with AMI. We conclude that thrombolytic therapy has rheologic consequences that may contribute to its overall efficacy. Inhibition of RBC aggregation by thrombolytic therapy may result from the degradation of fibrinogen, a key factor in the formation of RBC aggregates, and from the generation of fibrinogen degradation products capable of disaggregating RBCs.

  13. Phenomenological understanding of aggregation and dispersion of chemotactic cells

    CERN Document Server

    Iwasa, Masatomo

    2011-01-01

    We present a simple model that describes the motion of a single chemotactic cell exposed to a traveling wave of the chemoattractant. The model incorporates two types of responses to stimulation by the chemoattractant, i.e., change in polarity and change in motility of the cell. The periodic change in motility is assumed to be induced by the periodic stimulation by the chemoattractant on the basis of previous observations. Consequently, net migration of the cell occurs in a particular direction with respect to wave propagation, which explains the migration of Dictyostelium cells in aggregation processes. The difference between two time delays from the stimulation to the two responses and the wave frequency determined by the frequency of the secretion of the chemoattractant are important parameters that determine the direction of migration and the effective interaction between cells in a population. This result explains the dispersed state of a population of vegetative cells and cells in preaggregation without ...

  14. Mathematical modelling of cell aggregation in liver tissue engineering

    OpenAIRE

    Green, John Edward E.

    2006-01-01

    A promising method for growing functional liver tissue in vitro involves culturing hepatocytes as spheroidal cell aggregates. In this thesis, we develop mathematical models of cell aggregation, and use them to determine how hepatocytes' interactions with the extracellular matrix (ECM) on which they are seeded, and with stellate cells, affect the process. Chapters 2-4 focus on the effect that cell-ECM coupling has on the aggregation process. We use a novel formulation that couples a mechani...

  15. Mineral trioxide aggregate induces osteoblastogenesis via Atf6

    Directory of Open Access Journals (Sweden)

    Toyonobu Maeda

    2015-06-01

    Full Text Available Mineral trioxide aggregate (MTA has been recommended for various uses in endodontics. To understand the effects of MTA on alveolar bone, we examined whether MTA induces osteoblastic differentiation using MC3T3-E1 cells. MTA enhanced mineralization concomitant with alkaline phosphatase activity in a dose- and time-dependent manner. MTA increased production of collagens (Type I and Type III and matrix metalloproteinases (MMP-9 and MMP-13, suggesting that MTA affects bone matrix remodeling. MTA also induced Bglap (osteocalcin but not Bmp2 (bone morphogenetic protein-2 mRNA expression. We observed induction of Atf6 (activating transcription factor 6, an endoplasmic reticulum (ER stress response transcription factor mRNA expression and activation of Atf6 by MTA treatment. Forced expression of p50Atf6 (active form of Atf6 markedly enhanced Bglap mRNA expression. Chromatin immunoprecipitation assay was performed to investigate the increase in p50Atf6 binding to the Bglap promoter region by MTA treatment. Furthermore, knockdown of Atf6 gene expression by introduction of Tet-on Atf6 shRNA expression vector abrogated MTA-induced mineralization. These results suggest that MTA induces in vitro osteoblastogenesis through the Atf6–osteocalcin axis as ER stress signaling. Therefore, MTA in endodontic treatment may affect alveolar bone healing in the resorbed region caused by pulpal infection.

  16. Nutrient deprivation induces α-synuclein aggregation through endoplasmic reticulum stress response and SREBP2 pathway

    OpenAIRE

    Jiang, Peizhou; Gan, Ming; Lin, Wen-Lang; Yen, Shu-Hui C.

    2014-01-01

    Abnormal accumulation of filamentous α-synuclein (α-syn) in neurons, regarded as Lewy bodies (LBs), are a hallmark of Parkinson disease (PD). Although the exact mechanism(s) underlying LBs formation remains unknown, autophagy and ER stress response have emerged as two important pathways affecting α-syn aggregation. In present study we tested whether cells with the tetracycline-off inducible overexpression of α-syn and accumulating α-syn aggregates can benefit from autophagy activation elicite...

  17. Nutrient deprivation induces α-synuclein aggregation through endoplasmic reticulum stress response and SREBP2 pathway

    OpenAIRE

    Peizhou eJiang; Ming eGan; Wen-Lang eLin; Yen, Shu-Hui C.

    2014-01-01

    Abnormal accumulation of filamentous α-synuclein (α-syn) in neurons, regarded as Lewy bodies(LBs), are a hallmark of Parkinson disease (PD). Although the exact mechanism(s) underlying LBs formation remains unknown, autophagy and ER stress response have emerged as two important pathways affecting α-syn aggregation. In present study we tested whether cells with the tetracycline-off inducible overexpression of α-syn and accumulating α-syn aggregates can benefit from autophagy activation elicited...

  18. Observation of fine particle aggregating behavior induced by high intensity conditioning using high speed CCD

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    The aggregating behavior between bubbles and particles induced by high intensity conditioning (HIC) was studied using high speed CCD technique. Bubble size measurement was conducted, and the attachment behavior between bubbles and particles in HIC cell and flotation cell were observed. The results show that in HIC cell, high intensity conditioning creates an advantage environment for the formation of small size bubble due to hydrodynamic cavitations, and these fine bubbles have high probability of bubble-particle collision,which will enhance fine particle flotation. The bubble-particle attachment experiments indicate that in high intensity conditioning cell, a lot of fine bubbles are produced in situ on the surface of fine particles, and most of fine particles are aggregated under the bridging action of fine bubbles. The observation of bubble-particle interaction in flotation cell illustrates that aggregates created by HIC can be loaded more easily by big air bubble in flotation cell than those created by normal conditioning.

  19. Effects of Suilysin on Streptococcus suis-induced platelet aggregation

    Directory of Open Access Journals (Sweden)

    Shengwei Zhang

    2016-10-01

    Full Text Available Blood platelets play important roles during pathological thrombocytopenia in streptococcal toxic shock syndrome (STSS. Streptococcus suis (S. suis an emerging human pathogen, can cause STSS similarly to S. pyogenes. However, S. suis interactions with platelets are poorly understood. Here, we found that suilysin (SLY, different from other bacterial cholesterol-dependent cytolysins (CDCs, was the sole stimulus that induced platelet aggregation. Furthermore, the inside-out activation of GPIIb/IIIa of platelets mediated SLY-induced platelet aggregation. This process was triggered by Ca2+ influx that depend on the pore forming on platelets by SLY. Additionally, although SLY induced α-granule release occurred via the MLCK-dependent pathway, PLC-β-IP3/DAG-MLCK and Rho-ROCK-MLCK signaling were not involved in SLY-induced platelet aggregation. Interestingly, the pore dependent Ca2+ influx was also found to participate in the induction of platelet aggregation with pneumolysin (PLY and streptolysin O (SLO, two other CDCs. It is possible that the CDC-mediated platelet aggregation we observed in S. suis is a similar response mechanism to that used by a wide range of bacteria. These findings might lead to the discovery of potential therapeutic targets for S. suis-associated STSS.

  20. Effects of Suilysin on Streptococcus suis-Induced Platelet Aggregation

    Science.gov (United States)

    Zhang, Shengwei; Wang, Junping; Chen, Shaolong; Yin, Jiye; Pan, Zhiyuan; Liu, Keke; Li, Lin; Zheng, Yuling; Yuan, Yuan; Jiang, Yongqiang

    2016-01-01

    Blood platelets play important roles during pathological thrombocytopenia in streptococcal toxic shock syndrome (STSS). Streptococcus suis (S. suis) an emerging human pathogen, can cause STSS similarly to S. pyogenes. However, S. suis interactions with platelets are poorly understood. Here, we found that suilysin (SLY), different from other bacterial cholesterol-dependent cytolysins (CDCs), was the sole stimulus that induced platelet aggregation. Furthermore, the inside-out activation of GPIIb/IIIa of platelets mediated SLY-induced platelet aggregation. This process was triggered by Ca2+ influx that depend on the pore forming on platelets by SLY. Additionally, although SLY induced α-granule release occurred via the MLCK-dependent pathway, PLC-β-IP3/DAG-MLCK and Rho-ROCK-MLCK signaling were not involved in SLY-induced platelet aggregation. Interestingly, the pore dependent Ca2+ influx was also found to participate in the induction of platelet aggregation with pneumolysin (PLY) and streptolysin O (SLO), two other CDCs. It is possible that the CDC-mediated platelet aggregation we observed in S. suis is a similar response mechanism to that used by a wide range of bacteria. These findings might lead to the discovery of potential therapeutic targets for S. suis-associated STSS. PMID:27800304

  1. Photodynamic treatment of red blood cell concentrates for virus inactivation enhances red blood cell aggregation: protection with antioxidants.

    Science.gov (United States)

    Ben-Hur, E; Barshtein, G; Chen, S; Yedgar, S

    1997-10-01

    Photodynamic treatment (PDT) using phthalocyanines and red light appears to be a promising procedure for decontamination of red blood cell (RBC) concentrates for transfusion. A possible complication of this treatment may be induced aggregation of RBC. The production of RBC aggregates was measured with a novel computerized cell flow properties analyzer (CFA). The PDT of RBC concentrates with sulfonated aluminum phthalocyanine (AIPcS4) and the silicon phthalocyanine Pc 4 under virucidal conditions markedly enhanced RBC aggregation and higher shear stress was required to disperse these aggregates. The clusters of cells were huge and abnormally shaped, unlike the rouleaux formed by untreated RBC. This aggregation was prevented when a mixture of antioxidants was included during PDT. Addition of the antioxidants after PDT reduced aggregation only partially. It is concluded that inclusion of antioxidants during PDT of RBC concentrates prior to transfusion may reduce or eliminate the hemodynamic risk that the virucidal treatment may present to the recipient.

  2. Multivalent scaffolds induce galectin-3 aggregation into nanoparticles

    Directory of Open Access Journals (Sweden)

    Candace K. Goodman

    2014-07-01

    Full Text Available Galectin-3 meditates cell surface glycoprotein clustering, cross linking, and lattice formation. In cancer biology, galectin-3 has been reported to play a role in aggregation processes that lead to tumor embolization and survival. Here, we show that lactose-functionalized dendrimers interact with galectin-3 in a multivalent fashion to form aggregates. The glycodendrimer–galectin aggregates were characterized by dynamic light scattering and fluorescence microscopy methodologies and were found to be discrete particles that increased in size as the dendrimer generation was increased. These results show that nucleated aggregation of galectin-3 can be regulated by the nucleating polymer and provide insights that improve the general understanding of the binding and function of sugar-binding proteins.

  3. Multivalent scaffolds induce galectin-3 aggregation into nanoparticles

    Science.gov (United States)

    Goodman, Candace K; Wolfenden, Mark L; Nangia-Makker, Pratima; Michel, Anna K; Raz, Avraham

    2014-01-01

    Summary Galectin-3 meditates cell surface glycoprotein clustering, cross linking, and lattice formation. In cancer biology, galectin-3 has been reported to play a role in aggregation processes that lead to tumor embolization and survival. Here, we show that lactose-functionalized dendrimers interact with galectin-3 in a multivalent fashion to form aggregates. The glycodendrimer–galectin aggregates were characterized by dynamic light scattering and fluorescence microscopy methodologies and were found to be discrete particles that increased in size as the dendrimer generation was increased. These results show that nucleated aggregation of galectin-3 can be regulated by the nucleating polymer and provide insights that improve the general understanding of the binding and function of sugar-binding proteins. PMID:25161713

  4. Rubrene analogues with the aggregation-induced emission enhancement behaviour

    DEFF Research Database (Denmark)

    Zhang, Xiaoxu; Sørensen, Jakob Kryger; Fu, Xiaowei;

    2014-01-01

    In the light of the principle of aggregation-induced emission enhancement (AIEE), the rubrene analogue with orange light-emitting properties is designed and synthesized by substituting the phenyl side groups of rubrene with thienyl groups. To the best of our knowledge, this is the first report...

  5. Linear aggregation theory in cell biology

    Energy Technology Data Exchange (ETDEWEB)

    Hill, T.L.

    1987-01-01

    This is an account of the theory, probability, and thermodynamics of linear aggregation. The emphasis is on basic principles illustrated by simple models, not on particular applications or polymers. The general physical aggregate systems - attached single-stranded polymers, single-stranded polymers modified by a second component, long multistranded polymers, attached multistranded polymers - are given extensive treatment. Also included are a discussion of the GTPase and ATPase activity accompanying the aggregation of microtubules and action, and the properties of the kinetic two-phase model of the end of a microtubule.

  6. Formaldehyde at low concentration induces protein tau into globular amyloid-like aggregates in vitro and in vivo.

    Directory of Open Access Journals (Sweden)

    Chun Lai Nie

    Full Text Available Recent studies have shown that neurodegeneration is closely related to misfolding and aggregation of neuronal tau. Our previous results show that neuronal tau aggregates in formaldehyde solution and that aggregated tau induces apoptosis of SH-SY5Y and hippocampal cells. In the present study, based on atomic force microscopy (AFM observation, we have found that formaldehyde at low concentrations induces tau polymerization whilst acetaldehyde does not. Neuronal tau misfolds and aggregates into globular-like polymers in 0.01-0.1% formaldehyde solutions. Apart from globular-like aggregation, no fibril-like polymerization was observed when the protein was incubated with formaldehyde for 15 days. SDS-PAGE results also exhibit tau polymerizing in the presence of formaldehyde. Under the same experimental conditions, polymerization of bovine serum albumin (BSA or alpha-synuclein was not markedly detected. Kinetic study shows that tau significantly misfolds and polymerizes in 60 minutes in 0.1% formaldehyde solution. However, presence of 10% methanol prevents protein tau from polymerization. This suggests that formaldehyde polymerization is involved in tau aggregation. Such aggregation process is probably linked to the tau's special "worm-like" structure, which leaves the epsilon-amino groups of Lys and thiol groups of Cys exposed to the exterior. Such a structure can easily bond to formaldehyde molecules in vitro and in vivo. Polymerizing of formaldehyde itself results in aggregation of protein tau. Immunocytochemistry and thioflavin S staining of both endogenous and exogenous tau in the presence of formaldehyde at low concentrations in the cell culture have shown that formaldehyde can induce tau into amyloid-like aggregates in vivo during apoptosis. The significant protein tau aggregation induced by formaldehyde and the severe toxicity of the aggregated tau to neural cells may suggest that toxicity of methanol and formaldehyde ingestion is related to

  7. Improvement and enhancement of antibladder carcinoma cell effects of heteronemin by the nanosized hyaluronan aggregation

    Directory of Open Access Journals (Sweden)

    Huang HH

    2016-03-01

    Full Text Available Han Hsiang Huang,1 Shyh Ming Kuo,2 Yi-Jhen Wu,2 Jui-Hsin Su3 1Department of Veterinary Medicine, National Chiayi University, Chiayi City, 2Department of Biomedical Engineering, I-Shou University, Kaohsiung City, 3National Museum of Marine Biology and Aquarium, Pingtung, Taiwan Abstract: The effects against tumors exerted by marine active compounds have been highlighted and investigated. Polymeric nanoparticles made from biodegradable and biocompatible molecules such as hyaluronan (HA and chitosan (CHI are able to aggregate the compounds to enhance their activities against tumor cells and reduce the toxicity on normal cells. Here, we extensively examined the antitumor activities and the mechanisms of HA/CHI nanoparticles-aggregated heteronemin (HET extracted from the sponge Hippospongia sp. The half-maximal inhibitory concentration (IC50 of pure HET toward T24 bladder carcinoma cells is ~0.28 µg/mL. Pure HET from 0.2 to 0.8 µg/mL and HA nanoparticles-aggregated HET at 0.1 and 0.2 µg/mL significantly reduced T24 cell viability. Compared to pure HET, HA nanoparticles/HET aggregates showed much weaker viability-inhibitory effects on L929 normal fibroblasts. HET dose-dependently suppressed cancer cell migration as HA/CHI nanoparticles-aggregated HET displayed stronger migration-inhibitory effects than pure HET. Flow cytometric analysis showed that pure HET increased early/total apoptosis and JC-1 monomer fluorescence, while HA/CHI nanoparticles-aggregated HET induced higher apoptosis and JC-1 monomer rates than pure HET, suggesting that aggregation of HA nanoparticles offers HET stronger apoptosis-inducing capacity through mitochondrial depolarization. Western blot analysis showed that HA nanoparticles-aggregated HET further increased mitochondrial-associated, caspase-dependent and caspase-independent, as well as endoplasmic reticulum stress-related factors in comparison with pure HET. These data indicated that pure HET possesses cytotoxic

  8. Aggregation of red blood cells in patients with Gaucher disease.

    Science.gov (United States)

    Adar, Tomer; Ben-Ami, Ronen; Elstein, Deborah; Zimran, Ari; Berliner, Shlomo; Yedgar, Saul; Barshtein, Gershon

    2006-08-01

    Gaucher disease is associated with increased red blood cell (RBC) aggregation, but the pathophysiological significance of this phenomenon and its correlation with disease manifestations are unclear. RBC aggregation was evaluated in 43 patients with Gaucher disease and 53 healthy controls. Dynamic RBC aggregation was examined in a narrow-gap flow chamber at varying shear stress. Compared with the controls, RBC aggregation in Gaucher disease was increased by 25%. Comparison of RBC aggregation in autologous plasma and in dextran (500 kDa) showed an increase both in plasma-dependent (extrinsic) and -independent (intrinsic) RBC aggregation. Subgroup analysis revealed that increased RBC aggregation was limited to patients with an intact spleen. RBC aggregation in patients did not correlate with plasma fibrinogen concentration, disease severity, enzyme replacement therapy or genotype. We conclude that RBC aggregation is increased in patients with Gaucher disease and an intact spleen, possibly reflecting the accumulation of glucocerebroside and other substances in the plasma and RBC membranes of these patients. Our results do not support a role for RBC aggregation in the pathogenesis of vascular complications of Gaucher disease.

  9. Simulation of red blood cell aggregation in shear flow.

    Science.gov (United States)

    Lim, B; Bascom, P A; Cobbold, R S

    1997-01-01

    A simulation model has been developed for red blood cell (RBC) aggregation in shear flow. It is based on a description of the collision rates of RBC, the probability of particles sticking together, and the breakage of aggregates by shear forces. The influence of shear rate, hematocrit, aggregate fractal dimension, and binding strength on aggregation kinetics were investigated and compared to other theoretical and experimental results. The model was used to simulate blood flow in a long large diameter tube under steady flow conditions at low Reynolds numbers. The time and spatial distribution of the state of aggregation are shown to be in qualitative agreement with previous B-mode ultrasound studies in which a central region of low echogenicity was noted. It is suggested that the model can provide a basis for interpreting prior measurements of ultrasound echogenicity and may help relate them to the local state of aggregation.

  10. Transmethylation inhibitors decrease chemotactic sensitivity and delay cell aggregation in Dictyostelium discoideum

    NARCIS (Netherlands)

    van Waarde, A; van Haastert, P J

    1984-01-01

    In Dictyostelium discoideum, extracellular cyclic AMP (cAMP) induces chemotaxis and cell aggregation. Suspensions of cAMP-sensitive cells respond to a cAMP pulse with a rapid, transient increase of protein carboxyl methylation. The transmethylation inhibitors cycloleucine, L-homocysteine thiolactone

  11. Detection and characterization of red blood cell (RBC) aggregation with photoacoustics

    Science.gov (United States)

    Hysi, Eno; Saha, Ratan K.; Rui, Min; Kolios, Michael C.

    2012-02-01

    Red blood cells (RBCs) aggregate in the presence of increased plasma fibrinogen and low shear forces during blood flow. RBC aggregation has been observed in deep vein thrombosis, sepsis and diabetes. We propose using photoacoustics (PA) as a non-invasive imaging modality to detect RBC aggregation. The theoretical and experimental feasibility of PA for detecting and characterizing aggregation was assessed. A simulation study was performed to generate PA signals from non-aggregated and aggregated RBCs using a frequency domain approach and to study the PA signals' dependence on hematocrit and aggregate size. The effect of the finite bandwidth nature of transducers on the PA power spectra was also investigated. Experimental confirmation of theoretical results was conducted using porcine RBC samples exposed to 1064 nm optical wavelength using the Imagio Small Animal PA imaging system (Seno Medical Instruments, Inc., San Antonio, TX). Aggregation was induced with Dextran-70 (Sigma-Aldrich, St. Louis, MO) and the effect of hematocrit and aggregation level was investigated. The theoretical and experimental PA signal amplitude increased linearly with increasing hematocrit. The theoretical dominant frequency content of PA signals shifted towards lower frequencies (<30 MHz) and 9 dB enhancements in spectral power were observed as the size of aggregates increased compared to non-aggregating RBCs. Calibration of the PA spectra with the transducer response obtained from a 200 nm gold film was performed to remove system dependencies. Analysis of the spectral parameters from the calibrated spectra suggested that PA can assess the degree of aggregation at multiple hematocrit and aggregation levels.

  12. Complement-induced equine neutrophil adhesiveness and aggregation.

    Science.gov (United States)

    Slauson, D O; Skrabalak, D S; Neilsen, N R; Zwahlen, R D

    1987-05-01

    Equine neutrophils (PMN) were isolated from citrated normal blood by density gradient separation on Ficoll-Hypaque to greater than 96% purity and 98% viability and an average of 3.78 x 10(7) PMN/ml. The agonist C5a des Arg was used in serial dilutions of whole zymosan-activated equine plasma (ZAP) or was partially purified from ZAP by column chromatography. Purified equine PMN exhibited rapid aggregation following incubation with C5a des Arg which was further dependent on the availability of divalent cations, especially Mg++. The microfilament disruptive agent cytochalasin B (5 micrograms/50 microliters) greatly augmented aggregation responses to C5a des Arg. Subaggregating doses of C5a des Arg promoted PMN adhesiveness as assayed on 0.5 x 10 cm borosilicate glass columns containing a 2.0 cm bed of Sephadex G-25. This C5a des Arg-induced increased adhesiveness was inhibitable by prior incubation of the PMN with either non-steroidal (0.065 M phenylbutazone) or steroidal (0.005 M dexamethasone) anti-inflammatory agents. Ultrastructural studies correlated well with functional assays and revealed marked organelle-free lamellipodia formation without PMN-PMN contact at subaggregating doses of the agonist and progressive PMN-PMN contact at aggregating doses. Equine PMN are responsive to C5a des Arg, and induced adhesiveness responses can be manipulated by anti-inflammatory agents.

  13. Aggregation-induced emission—fluorophores and applications

    Science.gov (United States)

    Hong, Yuning

    2016-06-01

    Aggregation-induced emission (AIE) is a novel photophysical phenomenon found in a group of luminogens that are not fluorescent in solution but are highly emissive in the aggregate or solid state. Since the first publication of AIE luminogens in 2001, AIE has become a hot research area in which the number of research papers regarding new AIE molecules and their applications has been increasing in an exponential manner. Thomson Reuters Essential Science Indicators ranked AIE no.3 among the Top 100 Research Frontiers in the field of Chemistry and Materials Science in 2013. In this review, I will give a general introduction of the AIE phenomenon, discuss the structure-property relationship of the AIE lumingens and summarize the recent progress in the applications including as light-emitting materials in optoelectronics, as chemosensors and bioprobes, and for bioimaging (total 69 references cited).

  14. Chronotropic Biosensing Via Stem-Cell Derived Myocyte Aggregates

    Science.gov (United States)

    2007-11-02

    the example of Figure 2, the myocyte aggregates showed a high degree of ab - solute tracking; for the 6 sustained-activity aggregates, the cor- relation...chronotropy. Indeed, excitable-cell biosensor systems could lead to the development of long-term, physiologically- tuned, functionally integrated bioprocessing ...H. Ko- dama, J. Pan, M. Sano, T. Takahashi, S. Hori, H. Abe , J. Hata, A. Umezawa, and S. Ogawa, “Cardiomyocytes can be generated from marrow stromal

  15. Mechanisms of xenogeneic baboon platelet aggregation and phagocytosis by porcine liver sinusoidal endothelial cells.

    Directory of Open Access Journals (Sweden)

    Qiang Peng

    Full Text Available BACKGROUND: Baboons receiving xenogeneic livers from wild type and transgenic pigs survive less than 10 days. One of the major issues is the early development of profound thrombocytopenia that results in fatal hemorrhage. Histological examination of xenotransplanted livers has shown baboon platelet activation, phagocytosis and sequestration within the sinusoids. In order to study the mechanisms of platelet consumption in liver xenotransplantation, we have developed an in vitro system to examine the interaction between pig endothelial cells with baboon platelets and to thereby identify molecular mechanisms and therapies. METHODS: Fresh pig hepatocytes, liver sinusoidal and aortic endothelial cells were isolated by collagenase digestion of livers and processing of aortae from GTKO and Gal+ MGH-miniature swine. These primary cell cultures were then tested for the differential ability to induce baboon or pig platelet aggregation. Phagocytosis was evaluated by direct observation of CFSE labeled-platelets, which are incubated with endothelial cells under confocal light microscopy. Aurintricarboxylic acid (GpIb antagonist blocking interactions with von Willebrand factor/vWF, eptifibatide (Gp IIb/IIIa antagonist, and anti-Mac-1 Ab (anti-α(Mβ(2 integrin Ab were tested for the ability to inhibit phagocytosis. RESULTS: None of the pig cells induced aggregation or phagocytosis of porcine platelets. However, pig hepatocytes, liver sinusoidal and aortic endothelial cells (GTKO and Gal+ all induced moderate aggregation of baboon platelets. Importantly, pig liver sinusoidal endothelial cells efficiently phagocytosed baboon platelets, while pig aortic endothelial cells and hepatocytes had minimal effects on platelet numbers. Anti-MAC-1 Ab, aurintricarboxylic acid or eptifibatide, significantly decreased baboon platelet phagocytosis by pig liver endothelial cells (P<0.01. CONCLUSIONS: Although pig hepatocytes and aortic endothelial cells directly caused

  16. Structural analysis of red blood cell aggregates under shear flow.

    Science.gov (United States)

    Chesnutt, J K W; Marshall, J S

    2010-03-01

    A set of measures of red blood cell (RBC) aggregates are developed and applied to examine the aggregate structure under plane shear and channel flows. Some of these measures are based on averages over the set of red blood cells which are in contact with each other at a given time. Other measures are developed by first fitting an ellipse to the planar projection of the aggregate, and then examining the area and aspect ratio of the fit ellipse as well as the orientations of constituent RBCs with respect to the fit ellipse axes. The aggregate structural measures are illustrated using a new mesoscale computational model for blood cell transport, collision and adhesion. The sensitivity of this model to change in adhesive surface energy density and shear rate on the aggregate structure is examined. It is found that the mesoscale model predictions exhibit reasonable agreement with experimental and theoretical data for blood flow in plane shear and channel flows. The new structural measures are used to examine the differences between predictions of two- and three-dimensional computations of the aggregate formation, showing that two-dimensional computations retain some of the important aspects of three-dimensional computations.

  17. N-acetyl-L-cysteine prevents stress-induced desmin aggregation in cellular models of desminopathy.

    Directory of Open Access Journals (Sweden)

    Bertrand-David Segard

    Full Text Available Mutations within the human desmin gene are responsible for a subcategory of myofibrillar myopathies called desminopathies. However, a single inherited mutation can produce different phenotypes within a family, suggesting that environmental factors influence disease states. Although several mouse models have been used to investigate organ-specific desminopathies, a more general mechanistic perspective is required to advance our knowledge toward patient treatment. To improve our understanding of disease pathology, we have developed cellular models to observe desmin behaviour in early stages of disease pathology, e.g., upon formation of cytoplasmic desmin aggregates, within an isogenic background. We cloned the wildtype and three mutant desmin cDNAs using a Tet-On Advanced® expression system in C2C12 cells. Mutations were selected based on positioning within desmin and capacity to form aggregates in transient experiments, as follows: DesS46Y (head domain; low aggregation, DesD399Y (central rod domain; high aggregation, and DesS460I (tail domain; moderate aggregation. Introduction of these proteins into a C2C12 background permitted us to compare between desmin variants as well as to determine the role of external stress on aggregation. Three different types of stress, likely encountered during muscle activity, were introduced to the cell models-thermal (heat shock, redox-associated (H2O2 and cadmium chloride, and mechanical (stretching stresses-after which aggregation was measured. Cells containing variant DesD399Y were more sensitive to stress, leading to marked cytoplasmic perinuclear aggregations. We then evaluated the capacity of biochemical compounds to prevent this aggregation, applying dexamethasone (an inducer of heat shock proteins, fisetin or N-acetyl-L-cysteine (antioxidants before stress induction. Interestingly, N-acetyl-L-cysteine pre-treatment prevented DesD399Y aggregation during most stress. N-acetyl-L-cysteine has recently been

  18. Manganese exposure induces α-synuclein aggregation in the frontal cortex of non-human primates.

    Science.gov (United States)

    Verina, Tatyana; Schneider, Jay S; Guilarte, Tomás R

    2013-03-13

    Aggregation of α-synuclein (α-syn) in the brain is a defining pathological feature of neurodegenerative disorders classified as synucleinopathies. They include Parkinson's disease (PD), dementia with Lewy bodies (DLB), and multiple system atrophy (MSA). Occupational and environmental exposure to manganese (Mn) is associated with a neurological syndrome consisting of psychiatric symptoms, cognitive impairment and parkinsonism. In this study, we examined α-syn immunoreactivity in the frontal cortex of Cynomolgus macaques as part of a multidisciplinary assessment of the neurological effects produced by exposure to moderate levels of Mn. We found increased α-syn-positive cells in the gray matter of Mn-exposed animals, typically observed in pyramidal and medium-sized neurons in deep cortical layers. Some of these neurons displayed loss of Nissl staining with α-syn-positive spherical aggregates. In the white matter we also observed α-syn-positive glial cells and in some cases α-syn-positive neurites. These findings suggest that Mn exposure promotes α-syn aggregation in neuronal and glial cells that may ultimately lead to degeneration in the frontal cortex gray and white matter. To our knowledge, this is the first report of Mn-induced neuronal and glial cell α-syn accumulation and aggregation in the frontal cortex of non-human primates.

  19. Quantification of the aggregation of magnetic nanoparticles with different polymeric coatings in cell culture medium

    Energy Technology Data Exchange (ETDEWEB)

    Eberbeck, D; Zirpel, P; Trahms, L [Physikalisch-Technische Bundesanstalt, Abbestrasse 2-12, 10587 Berlin (Germany); Kettering, M; Hilger, I [Institute of Diagnostic and Interventional Radiology, University Hospital Jena, Erlanger Allee 101, 07747 Jena (Germany); Bergemann, C, E-mail: dietmar.eberbeck@ptb.d [Chemicell GmbH, Eresburgstrasse 22-23, 12103 Berlin (Germany)

    2010-10-13

    The knowledge of the physico-chemical characteristics of magnetic nanoparticles (MNPs) is essential to enhance the efficacy of MNP-based therapeutic treatments (e.g. magnetic heating, magnetic drug targeting). According to the literature, the MNP uptake by cells may depend on the coating of MNPs, the surrounding medium as well as on the aggregation behaviour of the MNPs. Therefore, in this study, the aggregation behaviour of MNPs in various media was investigated. MNPs with different coatings were suspended in cell culture medium (CCM) containing fetal calf serum (FCS) and the distribution of the hydrodynamic sizes was measured by magnetorelaxometry (MRX). FCS as well as bovine serum albumin (BSA) buffer (phosphate buffered saline with 0.1% bovine serum albumin) may induce MNP aggregation. Its strength depends crucially on the type of coating. The degree of aggregation in CCM depends on its FCS content showing a clear, local maximum at FCS concentrations, where the IgG concentration (part of FCS) is of the order of the MNP number concentration. Thus, we attribute the observed aggregation behaviour to the mechanism of agglutination of MNPs by serum compartments as for example IgG. No aggregation was induced for MNPs coated with dextran, polyarabic acid or sodium phosphate, respectively, which were colloidally stable in CCM.

  20. Suppression of Aggrus/podoplanin-induced platelet aggregation and pulmonary metastasis by a single-chain antibody variable region fragment.

    Science.gov (United States)

    Miyata, Kenichi; Takagi, Satoshi; Sato, Shigeo; Morioka, Hiroshi; Shiba, Kiyotaka; Minamisawa, Tamiko; Takami, Miho; Fujita, Naoya

    2014-12-01

    Almost all highly metastatic tumor cells possess high platelet aggregating abilities, thereby form large tumor cell-platelet aggregates in the microvasculature. Embolization of tumor cells in the microvasculature is considered to be the first step in metastasis to distant organs. We previously identified the platelet aggregation-inducing factor expressed on the surfaces of highly metastatic tumor cells and named as Aggrus. Aggrus was observed to be identical to the marker protein podoplanin (alternative names, T1α, OTS-8, and others). Aggrus is frequently overexpressed in several types of tumors and enhances platelet aggregation by interacting with the platelet receptor C-type lectin-like receptor 2 (CLEC-2). Here, we generated a novel single-chain antibody variable region fragment (scFv) by linking the variable regions of heavy and light chains of the neutralizing anti-human Aggrus monoclonal antibody MS-1 with a flexible peptide linker. Unfortunately, the generated KM10 scFv failed to suppress Aggrus-induced platelet aggregation in vitro. Therefore, we performed phage display screening and finally obtained a high-affinity scFv, K-11. K-11 scFv was able to suppress Aggrus-induced platelet aggregation in vitro. Moreover, K-11 scFv prevented the formation of pulmonary metastasis in vivo. These results suggest that K-11 scFv may be useful as metastasis inhibitory scFv and is expected to aid in the development of preclinical and clinical examinations of Aggrus-targeted cancer therapies.

  1. Methylglyoxal-induced modification causes aggregation of myoglobin

    Science.gov (United States)

    Banerjee, Sauradipta; Maity, Subhajit; Chakraborti, Abhay Sankar

    2016-02-01

    Post-translational modification of proteins by Maillard reaction, known as glycation, is thought to be the root cause of different complications, particularly in diabetes mellitus and age-related disorders. Methylglyoxal (MG), a reactive α-oxoaldehyde, increases in diabetic condition and reacts with proteins to form advanced glycation end products (AGEs) following Maillard-like reaction. We have investigated the in vitro effect of MG (200 μM) on the monomeric heme protein myoglobin (Mb) (100 μM) in a time-dependent manner (7 to 18 days incubation at 25 °C). MG induces significant structural alterations of the heme protein, including heme loss, changes in tryptophan fluorescence, decrease of α-helicity with increased β-sheet content etc. These changes occur gradually with increased period of incubation. Incubation of Mb with MG for 7 days results in formation of the AGE adducts: carboxyethyllysine at Lys-16, carboxymethyllysine at Lys-87 and carboxyethyllysine or pyrraline-carboxymethyllysine at Lys-133. On increasing the period of incubation up to 14 days, additional AGEs namely, carboxyethyllysine at Lys-42 and hydroimidazolone or argpyrimidine at Arg-31 and Arg-139 have been detected. MG also induces aggregation of Mb, which is clearly evident with longer period of incubation (18 days), and appears to have amyloid nature. MG-derived AGEs may thus have an important role as the precursors of protein aggregation, which, in turn, may be associated with physiological complications.

  2. Temperature Induced Aggregation and Clouding in Humic Acid Solutions

    Directory of Open Access Journals (Sweden)

    Leah Shaffer

    2015-01-01

    Full Text Available Humic acids in aqueous solution demonstrate inverse temperature-solubility relationships when solution conditions are manipulated to reduce coulombic repulsion among the humic polyanions. These effects were followed by dynamic light scattering (DLS measurements of the resulting aggregates, as well as the addition of a polarity sensitive fluorescent probe (pyrene. The humic solutions could be primed for temperature induced clouding by carefully lowering the pH to a point where hydration effects became dominant. The exact value of the cloud point (CP was a function of both pH and humate concentration. The CPs mostly lay in the range 50–90°C, but DLS showed that temperature induced aggregation proceeded from approximately 30°C onward. Similar effects could be achieved by adding multivalent cations at concentrations below those which cause spontaneous precipitation. The declouding of clouded humate solutions could be affected by lowering the temperature combined with mechanical agitation to disentangle the humic polymers.

  3. Hemoglobin Aggregation in Single Red Blood Cells of Sickle Cell Anemia

    Science.gov (United States)

    Nishio, Izumi; Tanaka, Toyoichi; Sun, Shao-Tang; Imanishi, Yuri; Tsuyoshi Ohnishi, S.

    1983-06-01

    A laser light scattering technique was used to observe the extent of hemoglobin aggregation in solitary red blood cells of sickle cell anemia. Hemoglobin aggregation was confirmed in deoxygenated cells. The light scattering technique can also be applied to cytoplasmic studies of any biological cell.

  4. Salt-induced aggregation of gold nanoparticles for photoacoustic imaging and photothermal therapy of cancer

    Science.gov (United States)

    Sun, Mengmeng; Liu, Fei; Zhu, Yukun; Wang, Wansheng; Hu, Jin; Liu, Jing; Dai, Zhifei; Wang, Kun; Wei, Yen; Bai, Jing; Gao, Weiping

    2016-02-01

    The challenge in photothermal therapy (PTT) is to develop biocompatible photothermal transducers that can absorb and convert near-infrared (NIR) light into heat with high efficiency. Herein, we report salt-induced aggregation of gold nanoparticles (GNPs) in biological media to form highly efficient and biocompatible NIR photothermal transducers for PTT and photothermal/photoacoustic (PT/PA) imaging of cancer. The GNP depots in situ formed by salt-induced aggregation of GNPs show strong NIR absorption induced by plasmonic coupling between adjacent GNPs and very high photothermal conversion efficiency (52%), enabling photothermal destruction of tumor cells. More interestingly, GNPs in situ aggregate in tumors to form GNP depots, enabling simultaneous PT/PA imaging and PTT of the tumors. These findings may provide a simple and effective way to develop a new class of intelligent and biocompatible NIR photothermal transducers with high efficiency for PT/PA imaging and PTT.The challenge in photothermal therapy (PTT) is to develop biocompatible photothermal transducers that can absorb and convert near-infrared (NIR) light into heat with high efficiency. Herein, we report salt-induced aggregation of gold nanoparticles (GNPs) in biological media to form highly efficient and biocompatible NIR photothermal transducers for PTT and photothermal/photoacoustic (PT/PA) imaging of cancer. The GNP depots in situ formed by salt-induced aggregation of GNPs show strong NIR absorption induced by plasmonic coupling between adjacent GNPs and very high photothermal conversion efficiency (52%), enabling photothermal destruction of tumor cells. More interestingly, GNPs in situ aggregate in tumors to form GNP depots, enabling simultaneous PT/PA imaging and PTT of the tumors. These findings may provide a simple and effective way to develop a new class of intelligent and biocompatible NIR photothermal transducers with high efficiency for PT/PA imaging and PTT. Electronic supplementary

  5. Prefoldin Plays a Role as a Clearance Factor in Preventing Proteasome Inhibitor-induced Protein Aggregation*

    Science.gov (United States)

    Abe, Akira; Takahashi-Niki, Kazuko; Takekoshi, Yuka; Shimizu, Takashi; Kitaura, Hirotake; Maita, Hiroshi; Iguchi-Ariga, Sanae M. M.; Ariga, Hiroyoshi

    2013-01-01

    Prefoldin is a molecular chaperone composed of six subunits, PFD1–6, and prevents misfolding of newly synthesized nascent polypeptides. Although it is predicted that prefoldin, like other chaperones, modulates protein aggregation, the precise function of prefoldin against protein aggregation under physiological conditions has never been elucidated. In this study, we first established an anti-prefoldin monoclonal antibody that recognizes the prefoldin complex but not its subunits. Using this antibody, it was found that prefoldin was localized in the cytoplasm with dots in co-localization with polyubiquitinated proteins and that the number and strength of dots were increased in cells that had been treated with lactacystin, a proteasome inhibitor, and thapsigargin, an inducer of endoplasmic reticulum stress. Knockdown of prefoldin increased the level of SDS-insoluble ubiquitinated protein and reduced cell viability in lactacystin and thapsigargin-treated cells. Opposite results were obtained in prefoldin-overexpressed cells. It has been reported that mice harboring a missense mutation L110R of MM-1α/PFD5 exhibit neurodegeneration in the cerebellum. Although the prefoldin complex containing L110R MM-1α was properly formed in vitro and in cells derived from L110R MM-1α mice, the levels of ubiquitinated proteins and cytotoxicity were higher in L110R MM-1α cells than in wild-type cells under normal conditions and were increased by lactacystin and thapsigargin treatment, and growth of L110R MM-1α cells was attenuated. Furthermore, the polyubiquitinated protein aggregation level was increased in the brains of L110R MM-1α mice. These results suggest that prefoldin plays a role in quality control against protein aggregation and that dysfunction of prefoldin is one of the causes of neurodegenerative diseases. PMID:23946485

  6. Familial aggregation of urothelial cell carcinoma.

    NARCIS (Netherlands)

    Aben, K.K.H.; Witjes, J.A.; Schoenberg, M.P.; Hulsbergen-van de Kaa, C.A.; Verbeek, A.L.M.; Kiemeney, L.A.L.M.

    2002-01-01

    Urothelial cell carcinoma (UCC) is not considered to be a familial disease. Familial clustering of UCC was described in several case reports, however, some with an extremely early age at onset suggesting a genetic component. Epidemiological studies yielded inconsistent evidence of familial UCC, poss

  7. Tracking Hypoxic Signaling within Encapsulated Cell Aggregates

    OpenAIRE

    Skiles, Matthew L.; Sahai, Suchit; Blanchette, James O.

    2011-01-01

    In Diabetes mellitus type 1, autoimmune destruction of the pancreatic β-cells results in loss of insulin production and potentially lethal hyperglycemia. As an alternative treatment option to exogenous insulin injection, transplantation of functional pancreatic tissue has been explored1,2. This approach offers the promise of a more natural, long-term restoration of normoglycemia. Protection of the donor tissue from the host's immune system is required to prevent rejection and encapsulation is...

  8. Gelation process visualized by aggregation-induced emission fluorogens

    Science.gov (United States)

    Wang, Zhengke; Nie, Jingyi; Qin, Wei; Hu, Qiaoling; Tang, Ben Zhong

    2016-06-01

    Alkaline-urea aqueous solvent system provides a novel and important approach for the utilization of polysaccharide. As one of the most important polysaccharide, chitosan can be well dissolved in this solvent system, and the resultant hydrogel material possesses unique and excellent properties. Thus the sound understanding of the gelation process is fundamentally important. However, current study of the gelation process is still limited due to the absence of direct observation and the lack of attention on the entire process. Here we show the entire gelation process of chitosan LiOH-urea aqueous system by aggregation-induced emission fluorescent imaging. Accompanied by other pseudo in situ investigations, we propose the mechanism of gelation process, focusing on the formation of junction points including hydrogen bonds and crystalline.

  9. Numerical simulations of deformation and aggregation of red blood cells in shear flow.

    Science.gov (United States)

    Low, Hong-Tong; Ju, M; Sui, Y; Nazir, T; Namgung, B; Kim, Sangho

    2013-01-01

    This article reviews numerical simulations of red blood cells (RBCs) mainly using the lattice Boltzmann method (LBM), focusing on the 2-dimensional deformation and aggregation of the cells in simple shear flow. We outline the incorporation of the immersed boundary method into the LBM, in which the membrane forces are obtained from the membrane model. The RBCs are simulated as a single biconcave capsule and as a doublet of biconcave capsules. The transition from swinging to tumbling motions of the RBCs, as induced by reducing the shear rate or increasing the membrane bending stiffness, is discussed. Also discussed is the aggregation tendency of the doublet of RBCs, for which homogenous deformability maintained RBC aggregation, whereas an increased deformability difference resulted in RBC dissociation.

  10. Fractal Aggregation of Copper Particles using Electroless Cell

    Directory of Open Access Journals (Sweden)

    S.Q.Chishty

    2013-12-01

    Full Text Available A phenomenon In which the particles performing Brownian motion when hit the aggregates become the part of it is known as Diffusion limited aggregation (DLA, which produces a fractal shape. Experimental efforts are discussed through which some DLA shapes arc obtained. For this purpose different electrolytic solutions are used. Electro less cells are also designed and constructed using standard methods. The cells have to be flexible in the sense that changing of plates and solutions should be easier for photography. We used compounds of copper,. for growth of fractals. Within a very short time metallic dendrites appeared in the cell at different operating conditions. These images were photographed, while desired branching structures in copper sulphate solution were seen. Results thus obtained are compared with the growth of DLA.

  11. A role for amyloid in cell aggregation and biofilm formation.

    Directory of Open Access Journals (Sweden)

    Melissa C Garcia

    Full Text Available Cell adhesion molecules in Saccharomyces cerevisiae and Candida albicans contain amyloid-forming sequences that are highly conserved. We have now used site-specific mutagenesis and specific peptide perturbants to explore amyloid-dependent activity in the Candida albicans adhesin Als5p. A V326N substitution in the amyloid-forming region conserved secondary structure and ligand binding, but abrogated formation of amyloid fibrils in soluble Als5p and reduced cell surface thioflavin T fluorescence. When displayed on the cell surface, Als5p with this substitution prevented formation of adhesion nanodomains and formation of large cellular aggregates and model biofilms. In addition, amyloid nanodomains were regulated by exogenous peptides. An amyloid-forming homologous peptide rescued aggregation and biofilm activity of Als5p(V326N cells, and V326N substitution peptide inhibited aggregation and biofilm activity in Als5p(WT cells. Therefore, specific site mutation, inhibition by anti-amyloid peturbants, and sequence-specificity of pro-amyloid and anti-amyloid peptides showed that amyloid formation is essential for nanodomain formation and activation.

  12. Reversible NaCl-induced aggregation of a monoclonal antibody at low pH: Characterization of aggregates and factors affecting aggregation.

    Science.gov (United States)

    Bickel, Fabian; Herold, Eva Maria; Signes, Alba; Romeijn, Stefan; Jiskoot, Wim; Kiefer, Hans

    2016-10-01

    We investigated the influence of pH and sodium chloride concentration on aggregation kinetics of a monoclonal antibody. Aggregation was induced by sodium chloride addition at low pH. Protein conformation before and after salt addition was determined as well as the reversibility of aggregation. Aggregation was monitored at pH values between 2 and 7 with NaCl up to 1.5M by turbidity measurement and size-exclusion chromatography. Particle size distribution was assessed by using size-exclusion chromatography as well as nanoparticle tracking analysis and flow imaging microscopy. Structural changes were monitored by circular dichroism, Fourier transform infrared and fluorescence spectroscopy. Thermal stability was measured by differential scanning fluorimetry. Aggregation propensity was maximal at low pH and high ionic strength. While thermal stability decreased with pH, the secondary structure remained unchanged down to pH 3.5 and up to 1.5M NaCl. Precipitated protein could be largely reverted to monomers by dilution into salt-free buffer. The re-solubilized antibody was indistinguishable in structure, solubility and monodispersity from the unstressed protein. Also, binding to Protein A was steady. Aggregation could be reduced in the presence of trehalose. The results suggest a reversible aggregation mechanism characterized by a limited change in tertiary structure at low pH and a subsequent loss of colloidal stability resulting from electrostatic repulsion once salt is added to the sample. The experimental setup is robust and allows high-throughput quantification of the effect of additives on aggregation kinetics.

  13. Effects of osmolytes on protein folding and aggregation in cells.

    Science.gov (United States)

    Ignatova, Zoya; Gierasch, Lila M

    2007-01-01

    Nature has developed many strategies to ensure that the complex and challenging protein folding reaction occurs in vivo with adequate efficiency and fidelity for the success of the organism. Among the strategies widely employed in a huge range of species and cell types is the elaboration of small organic molecules called osmolytes that offset the potentially damaging effects of osmotic stress. While considerable knowledge has been gained in vitro regarding the influence of osmolytes on protein structure and folding, it is of great interest to probe the effects of osmolytes in cells. We have developed an in-cell fluorescent-labeling method that enables the study of protein stability and also protein aggregation in vivo. We utilize a genetically encoded tag called a tetra-Cys motif that binds specifically to a bis-arsenical fluorescein-based dye "FlAsH"; we inserted the tetra-Cys motif into a protein of interest in such a way that the FlAsH signal reported on the state of folding or aggregation of the protein. Then, we designed protocols to assess how various osmolytes influence the stability and propensity to aggregate of our protein of interest. These are described here. Not only are there potential biotechnological applications of osmolytes in the quest to produce greater quantities of well-folded proteins, but also osmolytes may serve as tools and points of departure for therapeutic intervention in protein folding and aggregation diseases. Having in vivo methods to analyze how osmolytes affect folding and aggregation enhances our ability to further these goals greatly.

  14. Patterns of gravity induced aggregate migration during casting of fluid concretes

    DEFF Research Database (Denmark)

    Spangenberg, Jon; Roussel, N.; Hattel, Jesper Henri

    2012-01-01

    In this paper, aggregate migration patterns during fluid concrete castings are studied through experiments, dimensionless approach and numerical modeling. The experimental results obtained on two beams show that gravity induced migration is primarily affecting the coarsest aggregates resulting in...... that it finds its origin in the non Newtonian nature of fresh concrete and that increasing casting rate shall decrease the magnitude of gravity induced particle migration.......In this paper, aggregate migration patterns during fluid concrete castings are studied through experiments, dimensionless approach and numerical modeling. The experimental results obtained on two beams show that gravity induced migration is primarily affecting the coarsest aggregates resulting...

  15. Protective effects of Nitraria retusa extract and its constituent isorhamnetin against amyloid β-induced cytotoxicity and amyloid β aggregation.

    Science.gov (United States)

    Iida, Akihisa; Usui, Takeo; Zar Kalai, Feten; Han, Junkyu; Isoda, Hiroko; Nagumo, Yoko

    2015-01-01

    Nitraria retusa is a halophyte species that is distributed in North Africa and used as a traditional medicinal plant. In this study, N. retusa ethanol extract and its constituent isorhamnetin (IRA) protected against amyloid β (Aβ)-induced cytotoxicity in human neuroblastoma SH-SY5Y cells. An in vitro Aβ aggregation assay suggested that IRA destabilizes Aβ fibrils.

  16. Differential inhibition of lipopolysaccharide-induced granulocyte aggregation and prostanoid production by emoxypin.

    Science.gov (United States)

    Kubatiev, A; Turgiev, A; Smirnov, L; Pomoynetsky, V; Dumaev, K

    1990-01-01

    Emoxypin is known to be an effective membrane-stabilizing 3-oxy-pyridine derivative. We attempted to evaluate its influence on lipopolysaccharide (LPS)-induced granulocyte aggregation and prostanoid production. Granulocytes isolated from rabbit venous blood by dextran sedimentation and Pezcoll gradient centrifugation were stirred in the aggregometer cuvette with emoxypin (5mM), indomethacin (50 microM) or their solvents at 37 degrees C for 2 min. Then S. typhimurium LPS (200 micrograms/ml) was added and the aggregation was traced for 5 min. Thromboxane B2 (TxB2), prostaglandins (PG) E, F2 alpha and 13,14-dihydro-15-keto-PGF2 alpha were determined in supernatants radioimmunochemically. Indomethacin did not affect the pattern of aggregation, whereas emoxypin virtually precluded the response. Granulocytes incubated with LPS produced by the 15th sec and 5th min 1.3 and 2.5 times as much TxB2 respectively as did the intact cells (p less than 0.01). LPS had no effect on PGE production. Fifteen-sec contact of granulocytes with LPS had no significant influence on the formation of PGF2 alpha and its 13,14-dihydro-15-keto metabolite. The amount of PGF2 alpha released into the medium by the end of the 5th min of incubation with LPS was 1.5 times higher than in the control (p less than 0.05); the level of 13,14-dihydro-15-keto-PGF2 alpha was decreased 1.6 times (p less than 0.01). Emoxypin abolished totally LPS-induced TxB2 and PGF2 alpha production. We conclude that aggregation and eicosanoid production are independent manifestations of LPS-induced rabbit granulocyte activation.

  17. Phytochelatins inhibit the metal-induced aggregation of alpha-crystallin.

    Science.gov (United States)

    Hori, Yasuhisa; Yoshikawa, Tomoaki; Tsuji, Naoki; Bamba, Takeshi; Aso, Yoshikazu; Kudou, Motonori; Uchida, Yoshiki; Takagi, Masahiro; Harada, Kazuo; Hirata, Kazumasa

    2009-02-01

    Phytochelatins (PCs) are heavy-metal-binding peptides found in some eukaryotes. This study investigates the use of plant-derived PCs for the inhibition of metal-induced protein aggregation. The results of this study show that PCs inhibit zinc-induced alpha-crystallin aggregation, and suggest that PCs might be useful as anti-cataract agents.

  18. Enhanced Shear-induced Platelet Aggregation Due to Low-temperature Storage

    Science.gov (United States)

    2013-07-01

    pathogen inactivation technologies.4,5 In principle, storage of PLTs under refrigeration (4°C), which is standard practice for red blood cells (RBCs), can...by more than 100% (i.e., twofold) compared to freshly isolated PLTs at high shear rates. Effect of cellcell collisions and fluid shear stress on...in aggregating stored PLTs. PLT aggregation under shear is controlled by cellcell collision frequency and the force applied to the cells .26 These

  19. Laminopathy-inducing lamin A mutants can induce redistribution of lamin binding proteins into nuclear aggregates.

    Science.gov (United States)

    Hübner, S; Eam, J E; Hübner, A; Jans, D A

    2006-01-15

    Lamins, members of the family of intermediate filaments, form a supportive nucleoskeletal structure underlying the nuclear envelope and can also form intranuclear structures. Mutations within the A-type lamin gene cause a variety of degenerative diseases which are collectively referred to as laminopathies. At the molecular level, laminopathies have been shown to be linked to a discontinuous localization pattern of A-type lamins, with some laminopathies containing nuclear lamin A aggregates. Since nuclear aggregate formation could lead to the mislocalization of proteins interacting with A-type lamins, we set out to examine the effects of FLAG-lamin A N195K and R386K protein aggregate formation on the subnuclear distribution of the retinoblastoma protein (pRb) and the sterol responsive element binding protein 1a (SREBP1a) after coexpression as GFP-fusion proteins in HeLa cells. We observed strong recruitment of both proteins into nuclear aggregates. Nuclear aggregate recruitment of the NPC component nucleoporin NUP153 was also observed and found to be dependent on the N-terminus. That these effects were specific was implied by the fact that a number of other coexpressed karyophilic GFP-fusion proteins, such as the nucleoporin NUP98 and kanadaptin, did not coaggregate with FLAG-lamin A N195K or R386K. Immunofluorescence analysis further indicated that the precursor form of lamin A, pre-lamin A, could be found in intranuclear aggregates. Our results imply that redistribution into lamin A-/pre-lamin A-containing aggregates of proteins such as pRb and SREBP1a could represent a key aspect underlying the molecular pathogenesis of certain laminopathies.

  20. Induced Interval-Valued Intuitionistic Fuzzy Hybrid Aggregation Operators with TOPSIS Order-Inducing Variables

    Directory of Open Access Journals (Sweden)

    Jun-Ling Zhang

    2012-01-01

    Full Text Available Two induced aggregation operators with novelly designed TOPSIS order-inducing variables are proposed: Induced Interval-valued Intuitionistic Fuzzy Hybrid Averaging (I-IIFHA operator and Induced Interval-valued Intuitionistic Fuzzy Hybrid Geometric (I-IIFHG operator. The merit of two aggregation operators is that they can consider additional preference information of decision maker’s attitudinal characteristics besides argument-dependent information and argument-independent information. Some desirable properties of I-IIFHA and I-IIFHG are studied and theoretical analysis also shows that they can include a wide range of aggregation operators as special cases. Further, we extend these operators to form a novel group decision-making method for selecting the most desirable alternative in multiple attribute multi-interest group decision-making problems with attribute values and decision maker’s interest values taking the form of interval-valued intuitionistic fuzzy numbers, and application research to real estate purchase selection shows its practicality.

  1. Active protein aggregates induced by terminally attached self-assembling peptide ELK16 in Escherichia coli

    Directory of Open Access Journals (Sweden)

    Zhou Bihong

    2011-02-01

    Full Text Available Abstract Background In recent years, it has been gradually realized that bacterial inclusion bodies (IBs could be biologically active. In particular, several proteins including green fluorescent protein, β-galactosidase, β-lactamase, alkaline phosphatase, D-amino acid oxidase, polyphosphate kinase 3, maltodextrin phosphorylase, and sialic acid aldolase have been successfully produced as active IBs when fused to an appropriate partner such as the foot-and-mouth disease virus capsid protein VP1, or the human β-amyloid peptide Aβ42(F19D. As active IBs may have many attractive advantages in enzyme production and industrial applications, it is of considerable interest to explore them further. Results In this paper, we report that an ionic self-assembling peptide ELK16 (LELELKLK2 was able to effectively induce the formation of cytoplasmic inclusion bodies in Escherichia coli (E. coli when attached to the carboxyl termini of four model proteins including lipase A, amadoriase II, β-xylosidase, and green fluorescent protein. These aggregates had a general appearance similar to the usually reported cytoplasmic inclusion bodies (IBs under transmission electron microscopy or fluorescence confocal microscopy. Except for lipase A-ELK16 fusion, the three other fusion protein aggregates retained comparable specific activities with the native counterparts. Conformational analyses by Fourier transform infrared spectroscopy revealed the existence of newly formed antiparallel beta-sheet structures in these ELK16 peptide-induced inclusion bodies, which is consistent with the reported assembly of the ELK16 peptide. Conclusions This has been the first report where a terminally attached self-assembling β peptide ELK16 can promote the formation of active inclusion bodies or active protein aggregates in E. coli. It has the potential to render E. coli and other recombinant hosts more efficient as microbial cell factories for protein production. Our observation might

  2. UV Light–Induced Aggregation of Titania Submicron Particles

    Directory of Open Access Journals (Sweden)

    Can Zhou

    2016-11-01

    Full Text Available In this study, aggregation of TiO2 (rutile and anatase submicron particles in deionized (DI water under ultra-violet (UV light irradiation was investigated. While no aggregation was observed in the dark, rutile and anatase submicron particles started aggregating upon application of UV light and ceased aggregation in about 2 and 8.4 h, respectively. It has been demonstrated that UV light directly mitigated the particle mobility of TiO2, resulting in a neutralization effect of the Zeta potential. It was also observed that rutile particles aggregated much faster than anatase particles under UV radiation, indicating that the Zeta potential of as-prepared rutile is less than that of anatase in deionized (DI water. In addition, the interaction energy of rutile and anatase particles was simulated using the Derjaguin–Landau–Verwey–Overbeek (DLVO model. The results showed a significant reduction of barrier energy from 118.2 kBT to 33.6 kBT for rutile and from 333.5 kBT to 46.1 kBT for anatase, respectively, which further validated the remarkable influence of UV irradiation on the aggregation kinetics of rutile and anatase submicron particles. This work presents a further understanding of the aggregation mechanism of light-controlled submicron particles and has a promising potential application in environmental remediation.

  3. Removal of sialic acid from the surface of human MCF-7 mammary cancer cells abolishes E-cadherin-dependent cell-cell adhesion in an aggregation assay.

    Science.gov (United States)

    Deman, J J; Van Larebeke, N A; Bruyneel, E A; Bracke, M E; Vermeulen, S J; Vennekens, K M; Mareel, M M

    1995-09-01

    MCF-7 human breast cancer cells express E-cadherin and show, at least in some circumstances, E-cadherin-dependent cell-cell adhesion (Bracke et al., 1993). The MCF-7/AZ variant spontaneously displays E-cadherin-dependent fast aggregation; in the MCF-7/6 variant, E-cadherin appeared not to be spontaneously functional in the conditions of the fast aggregation assay, but function could be induced by incubation of the suspended cells in the presence of insulinlike growth factor I (IGF-I) (Bracke et al., 1993). E-cadherin from MCF-7 cells was shown to contain sialic acid. Treatment with neuraminidase was shown to remove this sialic acid, as well as most of the sialic acid present at the cell surface. Applied to MCF-7/AZ, and MCF-7/6 cells, pretreatment with neuraminidase abolished spontaneous as well as IGF-I induced, E-cadherin-dependent fast cell-cell adhesion of cells in suspension, as measured in the fast aggregation assay. Treatment with neuraminidase did not, however, inhibit the possibly different, but equally E-cadherin-mediated, process of cell-cell adhesion of MCF-7 cells on a flat plastic substrate as assessed by determining the percentage of cells remaining isolated (without contact with other cells) 24 h after plating.

  4. Heparin promotes suspension adaptation process of CHO-TS28 cells by eliminating cell aggregation.

    Science.gov (United States)

    Li, Ling; Qin, Jun; Feng, Qiang; Tang, Hao; Liu, Rong; Xu, Liqing; Chen, Zhinan

    2011-01-01

    While heparin has been shown to eliminate cell aggregation in suspension adaptations of insect and HEK293 cells for virus-based cell cultures, the role of heparin in long period serum-free suspension adaptation of the anchorage-dependent Chinese hamster ovary (CHO) cell lines remains inconclusive. In this paper, we explore the potential application of heparin in suspension adaptation of CHO cell line which produces an anti-human chimeric antibody cHAb18. Heparin showed a concentration-dependent inhibition of CHO-TS28 cell-to-cell adhesion, with a significant inhibitory effect occurring when the concentration exceeded 250 μg/ml (P cell aggregation elimination role at all concentrations (P cell growth and antibody secretion, with the highest cell density ((99.83 ± 12.21) × 10(4) cells/ml, P = 0.034) and maximum antibody yield ((9.46 ± 0.94) mg/l, P cell aggregates were effectively dispersed by 250 μg/ml heparin and a single-cell suspension culture process was promoted. In suspension adapted CHO-TS28 cells, cell growth rates and specific antibody productivity were maintained; while antigen-binding activity improved slightly. Together, our results show that heparin may promote suspension adaptation of anchorage-depended CHO cells by resisting cell aggregation without reducing cell growth, antibody secretion, and antigen-binding activity.

  5. Spontaneous and Induced Platelet Aggregation during Pregnancy and Labor

    Directory of Open Access Journals (Sweden)

    T. P. Bondar

    2016-01-01

    Full Text Available Objective: to evaluate changes in characteristics of spontaneous platelet (Pt aggregation in patients with obstetric complications associated with hereditary thrombophilia.Materials and methods. Blood samples were taken from 52 recently confined women on the first day after labor; at that, ethic regulations for the preanalytical phase were followed. Determination of PlA1/ PlA2 polymorphism enotype was performed by means of amplificationrestriction analysis. Geometrical characteristics of patients' peripheral blood Pt aggregation were studied by means of AFM Integra Prima. The degree of confidence of the parameters under test was determined using the ttest, and the significance level was considered valid at P<0.05.Results. A statistical analysis of the findings demonstrated that the length of Pt aggregates in healthy pregnant women was significantly higher than that in healthy nonpregnant women at all study phases. Patients with the P1A1/P1A2 polymorphism in the GP IIb/IIIa Pt receptor gene demonstrated increased widthm height, and density of Pt aggregates. The changes were most significant during the incubation phase lasting for 15 and 30 minutes. The study of geometric parameters of different exposures demonstrated the following: the longer the incubation period, the greater the difference between geometric parameters of the aggregates (e.g. height, length, and width. Conclusion. The analysis of obtained data demonstrated that the presence of P1A1/P1A2 polymorphism in GP IIb/IIIa Pt gene receptor contributes to the decrease in the platelet response threshold and enhances the spontaneous Pt aggregation. The imaging of aggregates provides strong evidence for the accelerated growth of the aggregates in thrombotic complications of pregnancy.

  6. Epicuticular lipids induce aggregation in Chagas disease vectors

    Directory of Open Access Journals (Sweden)

    Juárez M Patricia

    2009-01-01

    Full Text Available Abstract Background The triatomine bugs are vectors of the protozoan parasite Trypanosoma cruzi, the causative agent of Chagas disease. Aggregation behavior plays an important role in their survival by facilitating the location of refuges and cohesion of aggregates, helping to keep them safely assembled into shelters during daylight time, when they are vulnerable to predators. There are evidences that aggregation is mediated by thigmotaxis, by volatile cues from their faeces, and by hexane-extractable contact chemoreceptive signals from their cuticle surface. The epicuticular lipids of Triatoma infestans include a complex mixture of hydrocarbons, free and esterified fatty acids, alcohols, and sterols. Results We analyzed the response of T. infestans fifth instar nymphs after exposure to different amounts either of total epicuticular lipid extracts or individual lipid fractions. Assays were performed in a circular arena, employing a binary choice test with filter papers acting as aggregation attractive sites; papers were either impregnated with a hexane-extract of the total lipids, or lipid fraction; or with the solvent. Insects were significantly aggregated around papers impregnated with the epicuticular lipid extracts. Among the lipid fractions separately tested, only the free fatty acid fraction promoted significant bug aggregation. We also investigated the response to different amounts of selected fatty acid components of this fraction; receptiveness varied with the fatty acid chain length. No response was elicited by hexadecanoic acid (C16:0, the major fatty acid component. Octadecanoic acid (C18:0 showed a significant assembling effect in the concentration range tested (0.1 to 2 insect equivalents. The very long chain hexacosanoic acid (C26:0 was significantly attractant at low doses (≤ 1 equivalent, although a repellent effect was observed at higher doses. Conclusion The detection of contact aggregation pheromones has practical

  7. Antimicrobial preservatives induce aggregation of interferon alpha-2a: the order in which preservatives induce protein aggregation is independent of the protein.

    Science.gov (United States)

    Bis, Regina L; Mallela, Krishna M G

    2014-09-10

    Antimicrobial preservatives (APs) are included in liquid multi-dose protein formulations to combat the growth of microbes and bacteria. These compounds have been shown to cause protein aggregation, which leads to serious immunogenic and toxic side-effects in patients. Our earlier work on a model protein cytochrome c (Cyt c) demonstrated that APs cause protein aggregation in a specific manner. The aim of this study is to validate the conclusions obtained from our model protein studies on a pharmaceutical protein. Interferon α-2a (IFNA2) is available as a therapeutic treatment for numerous immune-compromised disorders including leukemia and hepatitis C, and APs have been used in its multi-dose formulation. Similar to Cyt c, APs induced IFNA2 aggregation, demonstrated by the loss of soluble monomer and increase in solution turbidity. The extent of IFNA2 aggregation increased with the increase in AP concentration. IFNA2 aggregation also depended on the nature of AP, and followed the order m-cresol>phenol>benzyl alcohol>phenoxyethanol. This specific order exactly matched with that observed for the model protein Cyt c. These and previously published results on antibodies and other recombinant proteins suggest that the general mechanism by which APs induce protein aggregation may be independent of the protein.

  8. Duration of exposure to high fluid shear stress is critical in shear-induced platelet activation-aggregation.

    Science.gov (United States)

    Zhang, Jian-ning; Bergeron, Angela L; Yu, Qinghua; Sun, Carol; McBride, Latresha; Bray, Paul F; Dong, Jing-fei

    2003-10-01

    Platelet functions are increasingly measured under flow conditions to account for blood hydrodynamic effects. Typically, these studies involve exposing platelets to high shear stress for periods significantly longer than would occur in vivo. In the current study, we demonstrate that the platelet response to high shear depends on the duration of shear exposure. In response to a 100 dyn/cm2 shear stress for periods less than 10-20 sec, platelets in PRP or washed platelets were aggregated, but minimally activated as demonstrated by P-selectin expression and binding of the activation-dependent alphaIIbbeta3 antibody PAC-1 to sheared platelets. Furthermore, platelet aggregation under such short pulses of high shear was subjected to rapid disaggregation. The disaggregated platelets could be re-aggregated by ADP in a pattern similar to unsheared platelets. In comparison, platelets that are exposed to high shear for longer than 20 sec are activated and aggregated irreversibly. In contrast, platelet activation and aggregation were significantly greater in whole blood with significantly less disaggregation. The enhancement is likely via increased collision frequency of platelet-platelet interaction and duration of platelet-platelet association due to high cell density. It may also be attributed to the ADP release from other cells such as red blood cells because increased platelet aggregation in whole blood was partially inhibited by ADP blockage. These studies demonstrate that platelets have a higher threshold for shear stress than previously believed. In a pathologically relevant timeframe, high shear alone is likely to be insufficient in inducing platelet activation and aggregation, but acts synergistically with other stimuli.

  9. Protein carbonylation, protein aggregation and neuronal cell death in a murine model of multiple sclerosis

    Science.gov (United States)

    Dasgupta, Anushka

    Many studies have suggested that oxidative stress plays an important role in the pathophysiology of both multiple sclerosis (MS) and its animal model experimental autoimmune encephalomyelitis (EAE). Yet, the mechanism by which oxidative stress leads to tissue damage in these disorders is unclear. Recent work from our laboratory has revealed that protein carbonylation, a major oxidative modification caused by severe and/or chronic oxidative stress conditions, is elevated in MS and EAE. Furthermore, protein carbonylation has been shown to alter protein structure leading to misfolding/aggregation. These findings prompted me to hypothesize that carbonylated proteins, formed as a consequence of oxidative stress and/or decreased proteasomal activity, promote protein aggregation to mediate neuronal apoptosis in vitro and in EAE. To test this novel hypothesis, I first characterized protein carbonylation, protein aggregation and apoptosis along the spinal cord during the course of myelin-oligodendrocyte glycoprotein (MOG)35-55 peptide-induced EAE in C57BL/6 mice [Chapter 2]. The results show that carbonylated proteins accumulate throughout the course of the disease, albeit by different mechanisms: increased oxidative stress in acute EAE and decreased proteasomal activity in chronic EAE. I discovered not only that there is a temporal correlation between protein carbonylation and apoptosis but also that carbonyl levels are significantly higher in apoptotic cells. A high number of juxta-nuclear and cytoplasmic protein aggregates containing the majority of the oxidized proteins are also present during the course of EAE, which seems to be due to reduced autophagy. In chapter 3, I show that when gluthathione levels are reduced to those in EAE spinal cord, both neuron-like PC12 (nPC12) cells and primary neuronal cultures accumulate carbonylated proteins and undergo cell death (both by necrosis and apoptosis). Immunocytochemical and biochemical studies also revealed a temporal

  10. Aggregate Size Optimization in Microwells for Suspension-based Cardiac Differentiation of Human Pluripotent Stem Cells

    OpenAIRE

    Bauwens, Celine L.; Toms, Derek; Ungrin, Mark

    2016-01-01

    Cardiac differentiation of human pluripotent stems cells (hPSCs) is typically carried out in suspension cell aggregates. Conventional aggregate formation of hPSCs involves dissociating cell colonies into smaller clumps, with size control of the clumps crudely controlled by pipetting the cell suspension until the desired clump size is achieved. One of the main challenges of conventional aggregate-based cardiac differentiation of hPSCs is that culture heterogeneity and spatial disorganization l...

  11. Does ocean acidification induce an upward flux of marine aggregates?

    Directory of Open Access Journals (Sweden)

    X. Mari

    2008-07-01

    Full Text Available The absorption of anthropogenic atmospheric carbon dioxide (CO2 by the ocean provokes its acidification. This acidification may alter several oceanic processes, including the export of biogenic carbon from the upper layer of the ocean, hence providing a feedback on rising atmospheric carbon concentrations. The effect of seawater acidification on transparent exopolymeric particles (TEP driven aggregation and sedimentation processes were investigated by studying the interactions between latex beads and TEP precursors collected in the lagoon of New Caledonia. A suspension of TEP and beads was prepared and the formation of mixed aggregates was monitored as a function of pH under increasing turbulence intensities. The pH was controlled by addition of sulfuric acid. Aggregation and sedimentation processes driven by TEP were drastically reduced when the pH of seawater decreases within the expected limits imposed by increased anthropogenic CO2 emissions. In addition to the diminution of TEP sticking properties, the diminution of seawater pH led to a significant increase of the TEP pool, most likely due to swollen structures. A diminution of seawater pH by 0.2 units or more led to a stop or a reversal of the downward flux of particles. If applicable to oceanic conditions, the sedimentation of marine aggregates may slow down or even stop as the pH decreases, and the vertical flux of organic carbon may reverse. This would enhance both rising atmospheric carbon and ocean acidification.

  12. Does ocean acidification induce an upward flux of marine aggregates?

    Directory of Open Access Journals (Sweden)

    X. Mari

    2008-04-01

    Full Text Available The adsorption of anthropogenic atmospheric carbon dioxide (CO2 by the ocean provokes its acidification. This acidification may alter several oceanic processes, including the export of biogenic carbon from the upper layer of the ocean, hence providing a feedback on rising atmospheric carbon concentrations. The effect of seawater acidification on transparent exopolymeric particles (TEP driven aggregation and sedimentation processes were investigated by studying the interactions between latex beads and TEP precursors collected in the lagoon of New Caledonia. A suspension of TEP and beads was prepared and the formation of mixed aggregates was monitored as a function of pH under increasing turbulence intensities. The pH was controlled by addition of sulfuric acid. Aggregation and sedimentation processes driven by TEP were drastically reduced when the pH of seawater decreases within the expected limits imposed by increased anthropogenic CO2 emissions. In addition to the diminution of TEP sticking properties, the diminution of seawater pH led to a significant increase of the TEP pool, most likely due to swollen structures. A diminution of seawater pH by 0.2 units or more led to a stop or a reversal of the downward flux of particles. If applicable to oceanic conditions, the sedimentation of marine aggregates may slow down or even stop as the pH decreases, and the vertical flux of organic carbon may reverse. This would enhance both rising atmospheric carbon and ocean acidification.

  13. Towards meso -Ester BODIPYs with Aggregation-Induced Emission Properties: The Effect of Substitution Positions

    KAUST Repository

    Chua, Ming Hui

    2015-06-17

    Three meso-ester boron dipyrromethene (BODIPY) dyes have been synthesized and functionalized with aggregation-induced emission (AIE)-active tetraphenylethene or triphenylethene moieties. It was found that functionalizing at the different positions of the BODIPY core resulted in the final dye having different emission properties in response to aggregation: from aggregation-induced quenching (ACQ) to being AIE active. X-ray crystallographic analysis was thus performed to provide an explanation for these differences. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  14. Effect of the LHCII pigment-protein complex aggregation on photovoltaic properties of sensitized TiO2 solar cells.

    Science.gov (United States)

    Yang, Yiqun; Jankowiak, Ryszard; Lin, Chen; Pawlak, Krzysztof; Reus, Michael; Holzwarth, Alfred R; Li, Jun

    2014-10-14

    A modified dye-sensitized solar cell consisting of a thin TiO2 barrier layer sensitized with natural trimeric light-harvesting complex II (LHCII) from spinach was used as a biomimetic model to study the effects of LHCII aggregation on the photovoltaic properties. The aggregation of individual trimers induced molecular reorganization, which dramatically increased the photocurrent. The morphology of small- and large-size LHCII aggregates deposited on a surface was confirmed by atomic force microscopy. Enhanced LHCII immobilization was accomplished via electrostatic interaction with amine-functionalized photoanodes. The photocurrent responses of the assembled solar cells under illumination at three characteristic wavelength bands in the UV-Vis absorption spectra of LHCII solutions confirmed that a significant photocurrent was generated by LHCII photosensitizers. The enhanced photocurrent by large aggregated LHCII is shown to correlate with the quenching in the far-red fluorescence deriving from chlorophyll-chlorophyll charge transfer states that are effectively coupled with the TiO2 surface and thus inject electrons into the TiO2 conduction band. The large aggregated LHCII with more chlorophyll-chlorophyll charge transfer states is a much better sensitizer since it injects electrons more efficiently into the conduction band of TiO2 than the small aggregated LHCII mostly consisting of unquenched chlorophyll excited state. The assembled solar cells demonstrated remarkable stability in both aqueous buffer and acetonitrile electrolytes over 30 days.

  15. Cell and organ printing 2: fusion of cell aggregates in three-dimensional gels.

    Science.gov (United States)

    Boland, Thomas; Mironov, Vladimir; Gutowska, Anna; Roth, Elisabeth A; Markwald, Roger R

    2003-06-01

    We recently developed a cell printer (Wilson and Boland, 2003) that enables us to place cells in positions that mimic their respective positions in organs. However, this technology was limited to the printing of two-dimensional (2D) tissue constructs. Here we describe the use of thermosensitive gels to generate sequential layers for cell printing. The ability to drop cells on previously printed successive layers provides a real opportunity for the realization of three-dimensional (3D) organ printing. Organ printing will allow us to print complex 3D organs with computer-controlled, exact placing of different cell types, by a process that can be completed in several minutes. To demonstrate the feasibility of this novel technology, we showed that cell aggregates can be placed in the sequential layers of 3D gels close enough for fusion to occur. We estimated the optimum minimal thickness of the gel that can be reproducibly generated by dropping the liquid at room temperature onto a heated substrate. Then we generated cell aggregates with the corresponding (to the minimal thickness of the gel) size to ensure a direct contact between printed cell aggregates during sequential printing cycles. Finally, we demonstrated that these closely-placed cell aggregates could fuse in two types of thermosensitive 3D gels. Taken together, these data strongly support the feasibility of the proposed novel organ-printing technology.

  16. Spherical TiO2 aggregates with different building units for dye-sensitized solar cells.

    Science.gov (United States)

    Liu, Zhaohui; Su, Xunjia; Hou, Genliang; Bi, Song; Xiao, Zhou; Jia, Haipeng

    2013-09-07

    Tailoring the architectures of spherical TiO2 aggregates is crucial to obtain superior photovoltaic properties and promote their application in dye-sensitized solar cells (DSSCs). Herein, we synthesized spherical TiO2 aggregates using different building units, including nanocrystallites, nanorods, nanosheets, and nanotubes, via a hydrothermal method, and studied the effect of the building units on the performances of DSSCs. The aggregates assembled by uniform nanosheet and nanotube building units were synthesized with the use of spherical TiO2 nanorod aggregates as titanium sources in an alkaline hydrothermal reaction. Compared with TiO2 nanoparticles, the spherical TiO2 aggregates possess higher surface area, more efficient light scattering ability, and better electron transport properties. Among the four types of spherical TiO2 aggregates; the nanorod, nanotube, and nanosheet aggregates demonstrate better electron transport properties than the nanocrystallite aggregates; the nanotube and nanosheet aggregates exhibit more efficient light scattering than the nanocrystallite and nanorod aggregates; and the nanotube aggregates show the highest surface area. Thus the DSSC based on nanotube aggregates exhibited the highest energy conversion efficiency of 7.48%, which is 16.0%, 9.7%, and 19.5% higher than those of the DSSCs based on the nanosheet, nanorod, and nanocrystallite aggregates, respectively.

  17. Amyloid-β peptide (1-42) aggregation induced by copper ions under acidic conditions

    Institute of Scientific and Technical Information of China (English)

    Yannan Bin; Xia Li; Yonghui He; Shu Chen; Juan Xiang

    2013-01-01

    It is well known that the aggregation of amyloid-β peptide (Aβ) induced by Cu2+ is related to incubation time,solution pH,and temperature.In this work,the aggregation of Aβ1-42 in the presence of Cu2+ under acidic conditions was studied at different incubation time and temperature (e.g.25 and 37℃).Incubation temperature,pH,and the presence of Cu2+ in Aβ solution were confirmed to alter the morphology of aggregation (fibrils or amorphous aggregates),and the morphology is pivotal for Aβ neurotoxicity and Alzheimer disease (AD) development.The results of atomic force microscopy (AFM) indicated that the formation of Aβ fibrous morphology is preferred at lower pH,but Cu2+ induced the formation of amorphous aggregates.The aggregation rate of Aβ was increased with the elevation of temperature.These results were further confirmed by fluorescence spectroscopy and circular dichroism spectroscopy and it was found that the formation of β-sheet structure was inhibited by Cu2+ binding to Aβ.The result was consistent with AFM observation and the fibrillation process was restrained.We believe that the local charge state in hydrophilic domain of Aβ may play a dominant role in the aggregate morphology due to the strong steric hindrance.This research will be valuable for understanding of Aβ toxicity in AD.

  18. Amyloid-β peptide (1-42) aggregation induced by copper ions under acidic conditions.

    Science.gov (United States)

    Bin, Yannan; Li, Xia; He, Yonghui; Chen, Shu; Xiang, Juan

    2013-07-01

    It is well known that the aggregation of amyloid-β peptide (Aβ) induced by Cu²⁺ is related to incubation time, solution pH, and temperature. In this work, the aggregation of Aβ₁₋₄₂ in the presence of Cu²⁺ under acidic conditions was studied at different incubation time and temperature (e.g. 25 and 37°C). Incubation temperature, pH, and the presence of Cu²⁺ in Aβ solution were confirmed to alter the morphology of aggregation (fibrils or amorphous aggregates), and the morphology is pivotal for Aβ neurotoxicity and Alzheimer disease (AD) development. The results of atomic force microscopy (AFM) indicated that the formation of Aβ fibrous morphology is preferred at lower pH, but Cu²⁺ induced the formation of amorphous aggregates. The aggregation rate of Aβ was increased with the elevation of temperature. These results were further confirmed by fluorescence spectroscopy and circular dichroism spectroscopy and it was found that the formation of β-sheet structure was inhibited by Cu²⁺ binding to Aβ. The result was consistent with AFM observation and the fibrillation process was restrained. We believe that the local charge state in hydrophilic domain of Aβ may play a dominant role in the aggregate morphology due to the strong steric hindrance. This research will be valuable for understanding of Aβ toxicity in AD.

  19. Thiosemicarbazone modification of 3-acetyl coumarin inhibits Aβ peptide aggregation and protect against Aβ-induced cytotoxicity.

    Science.gov (United States)

    Ranade, Dnyanesh S; Bapat, Archika M; Ramteke, Shefali N; Joshi, Bimba N; Roussel, Pascal; Tomas, Alain; Deschamps, Patrick; Kulkarni, Prasad P

    2016-10-01

    Aggregation of amyloid β peptide (Aβ) is an important event in the progression of Alzheimer's disease. Therefore, among the available therapeutic approaches to fight with disease, inhibition of Aβ aggregation is widely studied and one of the promising approach for the development of treatments for Alzheimer's disease. Thiosemicarbazone compounds are known for their variety of biological activities. However, the potential of thiosemicarbazone compounds towards inhibition of Aβ peptide aggregation and the subsequent toxicity is little explored. Herein, we report synthesis and x-ray crystal structure of novel compound 3-acetyl coumarin thiosemicarbazone and its efficacy toward inhibition of Aβ(1-42) peptide aggregation. Our results indicate that 3-acetyl coumarin thiosemicarbazone inhibits Aβ(1-42) peptide aggregation up to 80% compared to the parent 3-acetyl coumarin which inhibits 52%. Further, 3-acetyl coumarin thiosemicarbazone provides neuroprotection against Aβ-induced cytotoxicity in SH-SY5Y cell line. These findings indicate that thiosemicarbazone modification renders 3-acetyl coumarin neuroprotective properties.

  20. Non-cell autonomous cell death caused by transmission of Huntingtin aggregates in Drosophila.

    Science.gov (United States)

    Babcock, Daniel T; Ganetzky, Barry

    2015-01-01

    Recent evidence indicates that protein aggregates can spread between neurons in several neurodegenerative diseases but much remains unknown regarding the underlying mechanisms responsible for this spreading and its role in disease progression. We recently demonstrated that mutant Huntingtin aggregates spread between cells within the Drosophila brain resulting in non-cell autonomous loss of a pair of large neurons in the posterior protocerebrum. However, the full extent of neuronal loss throughout the brain was not determined. Here we examine the effects of driving expression of mutant Huntingtin in Olfactory Receptor Neurons (ORNs) by using a marker for cleaved caspase activity to monitor neuronal apoptosis as a function of age. We find widespread caspase activity in various brain regions over time, demonstrating that non-cell autonomous damage is widespread. Improved understanding of which neurons are most vulnerable and why should be useful in developing treatment strategies for neurodegenerative diseases that involve transcellular spreading of aggregates.

  1. Mild exposure of RIN-5F β-cells to human islet amyloid polypeptide aggregates upregulates antioxidant enzymes via NADPH oxidase-RAGE: An hormetic stimulus

    Directory of Open Access Journals (Sweden)

    Elisabetta Borchi

    2014-01-01

    Full Text Available The presence of amyloid aggregates of human islet amyloid polypeptide (hIAPP, a hallmark of type 2 diabetes, contributes to pancreatic β-cell impairment, where oxidative stress plays a key role. A contribution of NADPH oxidase to reactive oxygen species (ROS generation after cell exposure to micromolar concentrations of hIAPP aggregates has been suggested. However, little is known about β-cells exposure to lower amounts of hIAPP aggregates, similar to those found in human pancreas. Thus, we aimed to investigate the events resulting from RIN-5F cells exposure to nanomolar concentrations of toxic hIAPP aggregates. We found an early and transient rise of NADPH oxidase activity resulting from increased Nox1 expression following the engagement of receptor for advanced glycation end-products (RAGE by hIAPP aggregates. Unexpectedly, NADPH oxidase activation was not accompanied by a significant ROS increase and the lipoperoxidation level was significantly reduced. Indeed, cell exposure to hIAPP aggregates affected the antioxidant defences, inducing a significant increase of the expression and activity of catalase and glutathione peroxidase. We conclude that exposure of pancreatic β-cells to nanomolar concentrations of hIAPP aggregates for a short time induces an hormetic response via the RAGE-Nox1 axis; the latter stimulates the enzymatic antioxidant defences that preserve the cells against oxidative stress damage.

  2. Bacillus pasteurii urease shares with plant ureases the ability to induce aggregation of blood platelets.

    Science.gov (United States)

    Olivera-Severo, D; Wassermann, G E; Carlini, C R

    2006-08-15

    Ureases (EC 3.5.1.5) are highly homologous enzymes found in plants, bacteria and fungi. Canatoxin, an isoform Canavalia ensiformis urease, has several biological properties unrelated to its ureolytic activity, like platelet-aggregating and pro-inflammatory effects. Here, we describe that Bacillus pasteurii urease (BPU) also induces aggregation of rabbit platelets, similar to the canatoxin-induced effect (ED(50) 0.4 and 0.015 mg/mL, respectively). BPU induced-aggregation was blocked in platelets pretreated with dexamethasone and esculetin, a phospholipase A(2) and a lipoxygenase inhibitor, respectively, while platelets treated with indomethacin, a cyclooxygenase inhibitor, showed increased response to BPU. Methoxyverapamil (Ca(2+) channel blocker) and AMP (ADP antagonist) abrogated urease-induced aggregation, whereas the PAF-acether antagonist Web2170 had no effect. We concluded that platelet aggregation induced by BPU is mediated by lipoxygenase-derived eicosanoids and secretion of ADP from the platelets through a calcium-dependent mechanism. Potential relevance of these findings for bacterium-plant interactions and pathogenesis of bacterial infections are discussed.

  3. Platelet-collagen adhesion enhances platelet aggregation induced by binding of VWF to platelets

    Energy Technology Data Exchange (ETDEWEB)

    Laduca, F.M.; Bell, W.R.; Bettigole, R.E. (Johns Hopkins Univ. School of Medicine, Baltimore, MD (USA) State Univ. of New York, Buffalo (USA))

    1987-11-01

    Ristocetin-induced platelet aggregation (RIPA) was evaluated in the presence of platelet-collagen adhesion. RIPA of normal donor platelet-rich plasma (PRP) demonstrated a primary wave of aggregation mediated by the binding of von Willebrand factor (VWF) to platelets and a secondary aggregation wave, due to a platelet-release reaction, initiated by VWF-platelet binding and inhibitable by acetylsalicylic acid (ASA). An enhanced RIPA was observed in PRP samples to which collagen had been previously added. These subthreshold concentrations of collagen, which by themselves were insufficient to induce aggregation, caused measurable platelet-collagen adhesion. Subthreshold collagen did not cause microplatelet aggregation, platelet release of ({sup 3}H)serotonin, or alter the dose-responsive binding of {sup 125}I-labeled VWF to platelets, which occurred with increasing ristocetin concentrations. However, ASA inhibition of the platelet release reaction prevented collagen-enhanced RIPA. These results demonstrate that platelet-collagen adhesion altered the platelet-release reaction induced by the binding of VWF to platelets causing a platelet-release reaction at a level of VWF-platelet binding not normally initiating a secondary aggregation. These findings suggest that platelet-collagen adhesion enhances platelet function mediated by VWF.

  4. Screening and analysis of differentially expressed genes of dermal papillae cells with aggregative behavior

    Institute of Scientific and Technical Information of China (English)

    宋志强; 郝飞; 杨卫兵; 王继文; 邹锋

    2004-01-01

    Objective: To screen and clone differentially expressed genes of dermal papillae cells (DPC) with aggregative behavior, and to explore the molecular mechanism of their aggregation. Methods: Total RNAs were extracted from DPC with and without aggregative behavior and double strand cDNAs were synthesized by using SMART cDNA synthesis, respectively. The cDNA fragments of differentially expressed genes in DPCs with aggregative behavior were isolated by suppression subtractive hybridization. Positive clones were screened by PCR method and verified by cDNA dot blot, Northern blot and then analyzed through homologous retrieving. Results: A subtractive cDNA library of DPC with aggregative behavior has been successfully constructed. The result of screening and cloning of the library showed that, DPC with aggregative behavior could expresse genes related to homologous aggregation, proliferation and cycle control, including known genes (capping protein, paladin, vascular endothelial growth factor), hematopoietic stem/progenitor cells (HSPC) related clone (HSPC011 and HSPC016) and a new gene. Conclusion: The construction of subtracted library of DPC lays solid foundation for screening and cloning new and specific genes related to aggregative behavior of DPC. Several genes might be cooperatively involved in the homologous aggregation, proliferation and cycle control of DPC. Among these genes, capping protein and palladin might be closely related to the aggregative behavior of dermal papilla cells, and VEGF and HSPC related clone would be responsible for the status of higher proliferation of dermal papilla cells.

  5. Geometrical Aspects During Formation of Compact Aggregates of Red Blood Cells

    Directory of Open Access Journals (Sweden)

    Cardoso A.V.

    2002-01-01

    Full Text Available In the past forty years considerable progress has been achieved on the knowledge of human blood as a non-Newtonian shear-thinning suspension, whose initial state, that is at rest (stasis or at very low shear rates, has a gel-like internal structure which is destroyed as shear stress increases. The main goal of this communication is to describe the role of geometrical aspects during RBC (red blood cell aggregate formation, growth and compaction on naturally aggregate (porcine blood and non-aggregate (bovine blood samples. We consider how these aspects coupled with tension equilibrium are decisive to transform red cell linear roleaux to three-dimensional aggregates or clusters. Geometrical aspects are also crucial on the compaction of red blood cell aggregates. These densely packed aggregates could precipitate out of blood- either as dangerous deposits on arterial walls, or as clots which travel in suspension until they block some crucial capillary.

  6. Entamoeba Clone-Recognition Experiments: Morphometrics, Aggregative Behavior, and Cell-Signaling Characterization.

    Science.gov (United States)

    Espinosa, Avelina; Paz-Y-Miño-C, Guillermo; Hackey, Meagan; Rutherford, Scott

    2016-05-01

    Studies on clone- and kin-discrimination in protists have proliferated during the past decade. We report clone-recognition experiments in seven Entamoeba lineages (E. invadens IP-1, E. invadens VK-1:NS, E. terrapinae, E. moshkovskii Laredo, E. moshkovskii Snake, E. histolytica HM-1:IMSS and E. dispar). First, we characterized morphometrically each clone (length, width, and cell-surface area) and documented how they differed statistically from one another (as per single-variable or canonical-discriminant analyses). Second, we demonstrated that amebas themselves could discriminate self (clone) from different (themselves vs. other clones). In mix-cell-line cultures between closely-related (E. invadens IP-1 vs. E. invadens VK-1:NS) or distant-phylogenetic clones (E. terrapinae vs. E. moshkovskii Laredo), amebas consistently aggregated with same-clone members. Third, we identified six putative cell-signals secreted by the amebas (RasGap/Ankyrin, coronin-WD40, actin, protein kinases, heat shock 70, and ubiquitin) and which known functions in Entamoeba spp. included: cell proliferation, cell adhesion, cell movement, and stress-induced encystation. To our knowledge, this is the first multi-clone characterization of Entamoeba spp. morphometrics, aggregative behavior, and cell-signaling secretion in the context of clone-recognition. Protists allow us to study cell-cell recognition from ecological and evolutionary perspectives. Modern protistan lineages can be central to studies about the origins and evolution of multicellularity.

  7. SDF-1α/CXCR4 Signaling in Lipid Rafts Induces Platelet Aggregation via PI3 Kinase-Dependent Akt Phosphorylation

    Science.gov (United States)

    Hayashi, Moyuru; Kaneda, Mizuho; Iida, Kazuko; Shimonaka, Motoyuki; Hara, Takahiko; Arai, Morio; Koike, Yuichi; Yamamoto, Naomasa; Kasahara, Kohji

    2017-01-01

    Stromal cell-derived factor-1α (SDF-1α)-induced platelet aggregation is mediated through its G protein-coupled receptor CXCR4 and phosphatidylinositol 3 kinase (PI3K). Here, we demonstrate that SDF-1α induces phosphorylation of Akt at Thr308 and Ser473 in human platelets. SDF-1α-induced platelet aggregation and Akt phosphorylation are inhibited by pretreatment with the CXCR4 antagonist AMD3100 or the PI3K inhibitor LY294002. SDF-1α also induces the phosphorylation of PDK1 at Ser241 (an upstream activator of Akt), GSK3β at Ser9 (a downstream substrate of Akt), and myosin light chain at Ser19 (a downstream element of the Akt signaling pathway). SDF-1α-induced platelet aggregation is inhibited by pretreatment with the Akt inhibitor MK-2206 in a dose-dependent manner. Furthermore, SDF-1α-induced platelet aggregation and Akt phosphorylation are inhibited by pretreatment with the raft-disrupting agent methyl-β-cyclodextrin. Sucrose density gradient analysis shows that 35% of CXCR4, 93% of the heterotrimeric G proteins Gαi-1, 91% of Gαi-2, 50% of Gβ and 4.0% of PI3Kβ, and 4.5% of Akt2 are localized in the detergent-resistant membrane raft fraction. These findings suggest that SDF-1α/CXCR4 signaling in lipid rafts induces platelet aggregation via PI3K-dependent Akt phosphorylation. PMID:28072855

  8. SDF-1α/CXCR4 Signaling in Lipid Rafts Induces Platelet Aggregation via PI3 Kinase-Dependent Akt Phosphorylation.

    Science.gov (United States)

    Ohtsuka, Hiroko; Iguchi, Tomohiro; Hayashi, Moyuru; Kaneda, Mizuho; Iida, Kazuko; Shimonaka, Motoyuki; Hara, Takahiko; Arai, Morio; Koike, Yuichi; Yamamoto, Naomasa; Kasahara, Kohji

    2017-01-01

    Stromal cell-derived factor-1α (SDF-1α)-induced platelet aggregation is mediated through its G protein-coupled receptor CXCR4 and phosphatidylinositol 3 kinase (PI3K). Here, we demonstrate that SDF-1α induces phosphorylation of Akt at Thr308 and Ser473 in human platelets. SDF-1α-induced platelet aggregation and Akt phosphorylation are inhibited by pretreatment with the CXCR4 antagonist AMD3100 or the PI3K inhibitor LY294002. SDF-1α also induces the phosphorylation of PDK1 at Ser241 (an upstream activator of Akt), GSK3β at Ser9 (a downstream substrate of Akt), and myosin light chain at Ser19 (a downstream element of the Akt signaling pathway). SDF-1α-induced platelet aggregation is inhibited by pretreatment with the Akt inhibitor MK-2206 in a dose-dependent manner. Furthermore, SDF-1α-induced platelet aggregation and Akt phosphorylation are inhibited by pretreatment with the raft-disrupting agent methyl-β-cyclodextrin. Sucrose density gradient analysis shows that 35% of CXCR4, 93% of the heterotrimeric G proteins Gαi-1, 91% of Gαi-2, 50% of Gβ and 4.0% of PI3Kβ, and 4.5% of Akt2 are localized in the detergent-resistant membrane raft fraction. These findings suggest that SDF-1α/CXCR4 signaling in lipid rafts induces platelet aggregation via PI3K-dependent Akt phosphorylation.

  9. Inhibitory effect of copper nanoparticles on rosin modified surfactant induced aggregation of lysozyme.

    Science.gov (United States)

    Ishtikhar, Mohd; Usmani, Salman Sadullah; Gull, Nuzhat; Badr, Gamal; Mahmoud, Mohamed H; Khan, Rizwan Hasan

    2015-01-01

    Protein aggregation is associated with many serious diseases including Parkinson's and Alzheimer's. Protein aggregation is a primary problem related with the health of industrial workers who work with the surfactants, metal ions, and cosolvents. We have synthesized rosin-based surfactants, i.e., quaternary amines of rosin diethylaminoethyl esters (QRMAE), which is an ester of rosin acid with polyethylene glycol monomethyl ether. Here, we report the thermal aggregation of lysozyme induced by QRMAE at 65 °C and pH 7.4 for a given time period in which amorphous aggregates are formed and confirm that copper-nanoparticles have the ability to inhibit QRMAE-induced aggregation compared with zinc and silver-nanoparticles. Aggregation experiments was evaluated using several spectroscopic methods and dye binding assay, such as turbidity, Rayleigh light scattering, 1-anilino-8-naphthalene sulfonate (ANS), Thioflavin T (Th T), congo red (CR) and circular dichroism (CD), that was further supported by scanning electron microscopy (SEM) and SEM with EDX. The therapeutic use of nanoparticles and the fact that rosin possesses excellent film-forming properties, and that its derivatives have pharmaceuticals application such as micro encapsulation, coating and film forming, it's matrix materials are used for sustained and controlled release tablets, renders importance and application to the present study.

  10. Aggregates of nisin with various bactoprenol-containing cell wall precursors differ in size and membrane permeation capacity.

    Science.gov (United States)

    Scherer, Katharina; Wiedemann, Imke; Ciobanasu, Corina; Sahl, Hans-Georg; Kubitscheck, Ulrich

    2013-11-01

    Many lantibiotics use the membrane bound cell wall precursor Lipid II as a specific target for killing Gram-positive bacteria. Binding of Lipid II usually impedes cell wall biosynthesis, however, some elongated lantibiotics such as nisin, use Lipid II also as a docking molecule for pore formation in bacterial membranes. Although the unique nisin pore formation can be analyzed in Lipid II-doped vesicles, mechanistic details remain elusive. We used optical sectioning microscopy to directly visualize the interaction of fluorescently labeled nisin with membranes of giant unilamellar vesicles containing Lipid II and its various bactoprenol precursors. We quantitatively analyzed the binding and permeation capacity of nisin when applied at nanomolar concentrations. Specific interactions with Lipid I, Lipid II and bactoprenol-diphosphate (C55-PP), but not bactoprenol-phosphate (C55-P), resulted in the formation of large molecular aggregates. For Lipid II, we demonstrated the presence of both nisin and Lipid II in these aggregates. Membrane permeation induced by nisin was observed in the presence of Lipid I and Lipid II, but not in the presence of C55-PP. Notably, the size of the C55-PP-nisin aggregates was significantly smaller than that of the aggregates formed with Lipid I and Lipid II. We conclude that the membrane permeation capacity of nisin is determined by the size of the bactoprenol-containing aggregates in the membrane. Notably, transmitted light images indicated that the formation of large aggregates led to a pinch-off of small vesicles, a mechanism, which probably limits the growth of aggregates and induces membrane leakage.

  11. Dye-sensitized solar cell employing zinc oxide aggregates grown in the presence of lithium

    Science.gov (United States)

    Zhang, Qifeng; Cao, Guozhong

    2013-10-15

    Provided are a novel ZnO dye-sensitized solar cell and method of fabricating the same. In one embodiment, deliberately added lithium ions are used to mediate the growth of ZnO aggregates. The use of lithium provides ZnO aggregates that have advantageous microstructure, morphology, crystallinity, and operational characteristics. Employing lithium during aggregate synthesis results in a polydisperse collection of ZnO aggregates favorable for porosity and light scattering. The resulting nanocrystallites forming the aggregates have improved crystallinity and more favorable facets for dye molecule absorption. The lithium synthesis improves the surface stability of ZnO in acidic dyes. The procedures developed and disclosed herein also help ensure the formation of an aggregate film that has a high homogeneity of thickness, a high packing density, a high specific surface area, and good electrical contact between the film and the fluorine-doped tin oxide electrode and among the aggregate particles.

  12. Patterning processes in aggregates of Hydra cells visualized with the monoclonal antibody, TS19.

    Science.gov (United States)

    Sato, M; Bode, H R; Sawada, Y

    1990-10-01

    The monoclonal antibody, TS19, (Heimfeld et al., 1985), labels the apical surface of ectodermal epithelial cells of tentacles and lower peduncles in Hydra. To investigate the patterning process in a tissue whose original pattern was completely destroyed, the TS19 staining pattern was examined in developing aggregates of Hydra cells. Two types of aggregates were prepared. G-aggregates were made from tissue of the gastric portion of animals and RG-aggregates from gastric tissue allowed to regenerate for 24 hr before making aggregates. G-aggregates were initially TS19-negative, and later dim and uniformly TS19-positive. Thereafter, TS19 staining broke up into brightly stained and unstained regions. The brightly staining regions developed into head or foot structures. The TS19 pattern in RG-aggregates developed differently. Since the initial aggregates contained cells of regenerating tips, they started with TS19-positive cells as well as TS19-negative cells. The numbers of brightly staining TS19-positive cells increased with time. Some patches of these cells developed into head or foot structures, while others did not. These results and a simulation using a reaction-diffusion model suggest that the changes in activation levels affected the temporal changes in the pattern of TS19 staining, and that the de novo pattern formation in hydra can be explained in terms of a process involving activation and inhibition properties.

  13. UV irradiation induced transformation of TiO2 nanoparticles in water: aggregation and photoreactivity.

    Science.gov (United States)

    Sun, Jing; Guo, Liang-Hong; Zhang, Hui; Zhao, Lixia

    2014-10-21

    Transformation of nanomaterials in aqueous environment has significant impact on their behavior in engineered application and natural system. In this paper, UV irradiation induced transformation of TiO2 nanoparticles in aqueous solutions was demonstrated, and its effect on the aggregation and photocatalytic reactivity of TiO2 was investigated. UV irradiation of a TiO2 nanoparticle suspension accelerated nanoparticle aggregation that was dependent on the irradiation duration. The aggregation rate increased from UV irradiation which might be responsible for the change of surface charge and aggregation rate. UV irradiation also changed the photocatalytic degradation rate of Rhodamine B by TiO2, which initially increased with irradiation time, then decreased. Based on the photoluminescence decay and photocurrent collection data, the change was attributed to the variation in interparticle charge transfer kinetics. These results highlight the importance of light irradiation on the transformation and reactivity of TiO2 nanomaterials.

  14. Islet-like cell aggregates generated from human adipose tissue derived stem cells ameliorate experimental diabetes in mice.

    Directory of Open Access Journals (Sweden)

    Vikash Chandra

    Full Text Available BACKGROUND: Type 1 Diabetes Mellitus is caused by auto immune destruction of insulin producing beta cells in the pancreas. Currently available treatments include transplantation of isolated islets from donor pancreas to the patient. However, this method is limited by inadequate means of immuno-suppression to prevent islet rejection and importantly, limited supply of islets for transplantation. Autologous adult stem cells are now considered for cell replacement therapy in diabetes as it has the potential to generate neo-islets which are genetically part of the treated individual. Adopting methods of islet encapsulation in immuno-isolatory devices would eliminate the need for immuno-suppressants. METHODOLOGY/PRINCIPAL FINDINGS: In the present study we explore the potential of human adipose tissue derived adult stem cells (h-ASCs to differentiate into functional islet like cell aggregates (ICAs. Our stage specific differentiation protocol permit the conversion of mesodermic h-ASCs to definitive endoderm (Hnf3β, TCF2 and Sox17 and to PDX1, Ngn3, NeuroD, Pax4 positive pancreatic endoderm which further matures in vitro to secrete insulin. These ICAs are shown to produce human C-peptide in a glucose dependent manner exhibiting in-vitro functionality. Transplantation of mature ICAs, packed in immuno-isolatory biocompatible capsules to STZ induced diabetic mice restored near normoglycemia within 3-4 weeks. The detection of human C-peptide, 1155±165 pM in blood serum of experimental mice demonstrate the efficacy of our differentiation approach. CONCLUSIONS: h-ASC is an ideal population of personal stem cells for cell replacement therapy, given that they are abundant, easily available and autologous in origin. Our findings present evidence that h-ASCs could be induced to differentiate into physiologically competent functional islet like cell aggregates, which may provide as a source of alternative islets for cell replacement therapy in type 1 diabetes.

  15. Raman studies of gluten proteins aggregation induced by dietary fibres.

    Science.gov (United States)

    Nawrocka, Agnieszka; Szymańska-Chargot, Monika; Miś, Antoni; Kowalski, Radosław; Gruszecki, Wiesław I

    2016-03-01

    Interactions between gluten proteins and dietary fibre preparations are crucial in the baking industry. The addition of dietary fibre to bread causes significant reduction in its quality which is influenced by changes in the structure of gluten proteins. Fourier transform Raman spectroscopy was applied to determine changes in the structure of gluten proteins modified by seven dietary fibres. The commercially available gluten proteins without starch were mixed with the fibres in three concentrations: 3%, 6% and 9%. The obtained results showed that all fibres, regardless of their origin, caused the same kind of changes i.e. decrease in the α-helix content with a simultaneous increase in the content of antiparallel-β-sheet. The results indicated that presence of cellulose was the probable cause of these changes, and lead to aggregation or abnormal folding of the gluten proteins. Other changes observed in the gluten structure concerning β-structures, conformation of disulphide bridges, and aromatic amino acid environment, depended on the fibres chemical composition.

  16. Enhancement of cell adhesion, retention, and survival of HUVEC/cbMSC aggregates that are transplanted in ischemic tissues by concurrent delivery of an antioxidant for therapeutic angiogenesis.

    Science.gov (United States)

    Huang, Chieh-Cheng; Pan, Wen-Yu; Tseng, Michael T; Lin, Kun-Ju; Yang, Yi-Pei; Tsai, Hung-Wen; Hwang, Shiaw-Min; Chang, Yen; Wei, Hao-Ji; Sung, Hsing-Wen

    2016-01-01

    A recurring obstacle in cell-base strategies for treating ischemic diseases is the significant loss of viable cells that is caused by the elevated levels of regional reactive oxygen species (ROS), which ultimately limits therapeutic capacity. In this study, aggregates of human umbilical vein endothelial cells (HUVECs) and cord-blood mesenchymal stem cells (cbMSCs), which are capable of inducing therapeutic angiogenesis, are prepared. We hypothesize that the concurrent delivery of an antioxidant N-acetylcysteine (NAC) may significantly increase cell retention following the transplantation of HUVEC/cbMSC aggregates in a mouse model with hindlimb ischemia. Our in vitro results demonstrate that the antioxidant NAC can restore ROS-impaired cell adhesion and recover the reduced angiogenic potential of HUVEC/cbMSC aggregates under oxidative stress. In the animal study, we found that by scavenging the ROS generated in ischemic tissues, NAC is likely to be able to establish a receptive cell environment in the early stage of cell transplantation, promoting the adhesion, retention, and survival of cells of engrafted aggregates. Therapeutic angiogenesis is therefore enhanced and blood flow recovery and limb salvage are ultimately achieved. The combinatory strategy that uses an antioxidant and HUVEC/cbMSC aggregates may provide a new means of boosting the therapeutic efficacy of cell aggregates for the treatment of ischemic diseases.

  17. Characterization of circulating tumor cell aggregates identified in patients with epithelial tumors

    Science.gov (United States)

    Cho, Edward H.; Wendel, Marco; Luttgen, Madelyn; Yoshioka, Craig; Marrinucci, Dena; Lazar, Daniel; Schram, Ethan; Nieva, Jorge; Bazhenova, Lyudmila; Morgan, Alison; Ko, Andrew H.; Korn, W. Michael; Kolatkar, Anand; Bethel, Kelly; Kuhn, Peter

    2012-02-01

    Circulating tumor cells (CTCs) have been implicated as a population of cells that may seed metastasis and venous thromboembolism (VTE), two major causes of mortality in cancer patients. Thus far, existing CTC detection technologies have been unable to reproducibly detect CTC aggregates in order to address what contribution CTC aggregates may make to metastasis or VTE. We report here an enrichment-free immunofluorescence detection method that can reproducibly detect and enumerate homotypic CTC aggregates in patient samples. We identified CTC aggregates in 43% of 86 patient samples. The fraction of CTC aggregation was investigated in blood draws from 24 breast, 14 non-small cell lung, 18 pancreatic, 15 prostate stage IV cancer patients and 15 normal blood donors. Both single CTCs and CTC aggregates were measured to determine whether differences exist in the physical characteristics of these two populations. Cells contained in CTC aggregates had less area and length, on average, than single CTCs. Nuclear to cytoplasmic ratios between single CTCs and CTC aggregates were similar. This detection method may assist future studies in determining which population of cells is more physically likely to contribute to metastasis and VTE.

  18. Trehalose suppresses antibody aggregation during the culture of Chinese hamster ovary cells.

    Science.gov (United States)

    Onitsuka, Masayoshi; Tatsuzawa, Miki; Asano, Ryutaro; Kumagai, Izumi; Shirai, Akihiro; Maseda, Hideaki; Omasa, Takeshi

    2014-05-01

    The aggregation of therapeutic antibodies during the manufacturing process is problematic because of the potential risks posed by the aggregates, such as an unexpected immune response. One of the hallmark effects of trehalose, a disaccharide consisting of two alpha-glucose units, is as a chemical chaperone with anti-aggregation activity. In this study, Chinese hamster ovary (CHO) cell line producing a diabody-type bispecific antibody were cultured in medium containing trehalose and the aggregation of the secreted proteins during the culture process was analyzed. An analysis of the various forms of the antibody (monomeric, dimeric, and large aggregates) showed that trehalose decreased the relative content of large aggregates by two thirds. The aggregation kinetics indicated that trehalose directly inhibited the polymerization and aggregation steps in a nucleation-dependent aggregation mechanism. Moreover, both specific and volumetric antibody production were increased in CHO cells cultured in trehalose-containing medium. Thus, the addition of trehalose to recombinant CHO cell cultures would offer a practical strategy for quality improvement in the production of therapeutic antibodies.

  19. Photo-induced reduction of Noble metal ions to metal nanoparticles on tubular J-aggregates

    Directory of Open Access Journals (Sweden)

    Stefan Kirstein

    2006-01-01

    Full Text Available Palladium and silver nanoparticles are formed on the surface of tubular J-aggregates of an amphiphilic tetrachlorobenzimidacarbocyanine dye by reduction of the respective metal cations in aqueous solution. Upon addition of the palladium complex Na2PdCl4 to the aggregate solution, the absorption spectrum shows significant changes which is explained by partial destruction of the aggregates. Cryogenic transmission electron microscopy (cryo-TEM images show that the tubular J-aggregates are randomly covered by well-separated Pd nanoparticles of approximately 1–3 nm size. Larger particles and higher particle density along the aggregates are obtained when an auxiliary reducing agent is added to the solution. The presence of the metallic particles leads to efficient fluorescence quenching giving clear evidence for super quenching. In similar experiments using AgNO3, silver nanoparticles are grown which are larger in size but less dense distributed along the aggregates. At least in the case of the silver particles, the spontaneous formation of metal nanoparticles is assumed to be initiated by a photo-induced electron transfer process (PET.

  20. Fractal aggregates induced by liposome-liposome interaction in the presence of Ca2+.

    Science.gov (United States)

    Sabín, J; Prieto, G; Ruso, J M; Sarmiento, F

    2007-10-01

    We present a study of the fractal dimension of clusters of large unilamellar vesicles (LUVs) formed by egg yolk phosphatidylcholine (EYPC), dimyristoylphosphocholine (DMPC) and dipalmitoylphosphocholine (DPPC) induced by Ca2+ . Fractal dimensions were calculated by application of two methods, measuring the angular dependency of the light scattered by the clusters and following the evolution of the cluster size. In all cases, the fractal dimensions fell in the range from 2.1 to 1.8, corresponding to two regimes: diffusion-limited cluster aggregation (DLCA) and reaction-limited cluster aggregation (RLCA). Whereas DMPC clusters showed a typical transition from the RLCA to the DLCA aggregation, EYPC exhibited an unusual behaviour, since the aggregation was limited for a higher concentration than the critical aggregation concentration. The behaviour of DPPC was intermediate, with a transition from the RLCA to the DLCA regimes with cluster sizes depending on Ca2+ concentration. Studies on the reversibility of the aggregates show that EYPC and DPPC clusters can be re-dispersed by dilution with water. DMPC does not present reversibility. Reversibility is evidence of the existence of secondary minima in the DLVO potential between two liposomes. To predict these secondary minima, a correction of the DLVO model was necessary taking into account a repulsive force of hydration.

  1. Membrane aggregation and perturbation induced by antimicrobial peptide of S-thanatin

    Energy Technology Data Exchange (ETDEWEB)

    Wu, Guoqiu, E-mail: guoqiuwu@163.com [Center of Clinical Laboratory Medicine of Zhongda Hospital, Southeast University, Nanjing (China); Wu, Hongbin; Li, Linxian; Fan, Xiaobo; Ding, Jiaxuan; Li, Xiaofang; Xi, Tao [Biotechnology Center, Department of Life Science and Biotechnology, China Pharmaceutical University, Nanjing 210009 (China); Shen, Zilong, E-mail: Zilongshen@sina.com [Biotechnology Center, Department of Life Science and Biotechnology, China Pharmaceutical University, Nanjing 210009 (China)

    2010-04-23

    Thanatin, a 21-residue peptide, is an inducible insect peptide. In our previous study, we have identified a novel thanatin analog of S-thanatin, which exhibited a broad antimicrobial activity against bacteria and fungi with low hemolytic activity. This study was aimed to delineate the antimicrobial mechanism of S-thanatin and identify its interaction with bacterial membranes. In this study, membrane phospholipid was found to be the target for S-thanatin. In the presence of vesicles, S-thanatin interestingly led to the aggregation of anionic vesicles and sonicated bacteria. Adding S-thanatin to Escherichia coli suspension would result in the collapse of membrane and kill bacteria. The sensitivity assay of protoplast elucidated the importance of outer membrane (OM) for S-thanatin's antimicrobial activity. Compared with other antimicrobial peptide, S-thanatin produced chaotic membrane morphology and cell debris in electron microscopic appearance. These results supported our hypothesis that S-thanatin bound to negatively charged LPS and anionic lipid, impeded membrane respiration, exhausted the intracellular potential, and released periplasmic material, which led to cell death.

  2. In vitro aggregation behavior of a non-amyloidogenic λ light chain dimer deriving from U266 multiple myeloma cells.

    Directory of Open Access Journals (Sweden)

    Paolo Arosio

    Full Text Available Excessive production of monoclonal light chains due to multiple myeloma can induce aggregation-related disorders, such as light chain amyloidosis (AL and light chain deposition diseases (LCDD. In this work, we produce a non-amyloidogenic IgE λ light chain dimer from human mammalian cells U266, which originated from a patient suffering from multiple myeloma, and we investigate the effect of several physicochemical parameters on the in vitro stability of this protein. The dimer is stable in physiological conditions and aggregation is observed only when strong denaturating conditions are applied (acidic pH with salt at large concentration or heating at melting temperature T(m at pH 7.4. The produced aggregates are spherical, amorphous oligomers. Despite the larger β-sheet content of such oligomers with respect to the native state, they do not bind Congo Red or ThT. The impossibility to obtain fibrils from the light chain dimer suggests that the occurrence of amyloidosis in patients requires the presence of the light chain fragment in the monomer form, while dimer can form only amorphous oligomers or amorphous deposits. No aggregation is observed after denaturant addition at pH 7.4 or at pH 2.0 with low salt concentration, indicating that not a generic unfolding but specific conformational changes are necessary to trigger aggregation. A specific anion effect in increasing the aggregation rate at pH 2.0 is observed according to the following order: SO(4(-≫Cl(->H(2PO(4(-, confirming the peculiar role of sulfate in promoting protein aggregation. It is found that, at least for the investigated case, the mechanism of the sulfate effect is related to protein secondary structure changes induced by anion binding.

  3. In vitro aggregation behavior of a non-amyloidogenic λ light chain dimer deriving from U266 multiple myeloma cells.

    Science.gov (United States)

    Arosio, Paolo; Owczarz, Marta; Müller-Späth, Thomas; Rognoni, Paola; Beeg, Marten; Wu, Hua; Salmona, Mario; Morbidelli, Massimo

    2012-01-01

    Excessive production of monoclonal light chains due to multiple myeloma can induce aggregation-related disorders, such as light chain amyloidosis (AL) and light chain deposition diseases (LCDD). In this work, we produce a non-amyloidogenic IgE λ light chain dimer from human mammalian cells U266, which originated from a patient suffering from multiple myeloma, and we investigate the effect of several physicochemical parameters on the in vitro stability of this protein. The dimer is stable in physiological conditions and aggregation is observed only when strong denaturating conditions are applied (acidic pH with salt at large concentration or heating at melting temperature T(m) at pH 7.4). The produced aggregates are spherical, amorphous oligomers. Despite the larger β-sheet content of such oligomers with respect to the native state, they do not bind Congo Red or ThT. The impossibility to obtain fibrils from the light chain dimer suggests that the occurrence of amyloidosis in patients requires the presence of the light chain fragment in the monomer form, while dimer can form only amorphous oligomers or amorphous deposits. No aggregation is observed after denaturant addition at pH 7.4 or at pH 2.0 with low salt concentration, indicating that not a generic unfolding but specific conformational changes are necessary to trigger aggregation. A specific anion effect in increasing the aggregation rate at pH 2.0 is observed according to the following order: SO(4)(-)≫Cl(-)>H(2)PO(4)(-), confirming the peculiar role of sulfate in promoting protein aggregation. It is found that, at least for the investigated case, the mechanism of the sulfate effect is related to protein secondary structure changes induced by anion binding.

  4. Enzyme-induced aggregation of whey proteins with Bacillus licheniformis protease

    NARCIS (Netherlands)

    Creusot, N.P.

    2006-01-01

    Whey proteins are commonly used as ingredient in food. In relation with the gelation properties of whey proteins, this thesis deals with understanding the mechanism of peptide-induced aggregation of whey protein hydrolysates made with Bacillus licheniformis protease (BLP). The results show that BLP

  5. The clinical usefulness of the platelet aggregation test for the diagnosis of heparin-induced thrombocytopenia

    NARCIS (Netherlands)

    Chong, B H; Burgess, J; Ismail, F

    1993-01-01

    The platelet aggregation test is widely used for the diagnosis of heparin-induced thrombocytopenia (HIT), a potentially serious complication of heparin therapy. We have evaluated its sensitivity and specificity in comparison with those of the 14C-serotonin release test. The sensitivity of the platel

  6. The inhibitory activity of ginsenoside Rp4 in adenosine diphosphate-induced platelet aggregation

    Directory of Open Access Journals (Sweden)

    Young-Min Son

    2017-01-01

    Conclusion: G-Rp4 significantly inhibited ADP-induced platelet aggregation and this is mediated via modulating the intracellular signaling molecules. These results indicate that G-Rp4 could be a potential candidate as a therapeutic agent against platelet-related cardiovascular diseases.

  7. Aggregation of phospholipid vesicles induced by the ribosome inactivating protein saporin.

    Science.gov (United States)

    Hao, Q; Yan, L; Yang, H; Zhang, Y; Gao, G; Yao, Q; Li, Q

    1996-04-01

    Saporin-S6(SO-6) is a single chain ribosome inactivating protein, which can inhibit protein synthesis by inactivating eukaryotic ribosomes. The interaction of SO-6 with phospholipid model systems was described. SO-6 can specifically interact with negatively-charged phospholipid vesicles and it induces the aggregation of the lipid vesicles. The kinetics of the vesicle aggregation induced by SO-6 was studied. The saturating protein/lipid molar ratio was determined to be 1:100 based on titration experiments. The aggregation is dependent on the temperature in a range that was many times higher than the phase transition temperature of the phospholipid. The effect of pH on the aggregation of the vesicles can not be explained by simple deprotonation of side chain amino groups of the protein, and may be related to conformational changes of the protein. The maintenance of physiological ionic strength was required for the aggregation of SO-6 with vesicles. Finally, the interaction was prompted by Ca2+ ions, and was totally inhibited by EDTA, which suggests that SO-6 may interact with phospholipid vesicles in a Ca(2+)-dependent manner.

  8. Aggregation of gold nanoparticles followed by methotrexate release enables Raman imaging of drug delivery into cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Durgadas, C. V.; Sharma, C. P.; Paul, W.; Rekha, M. R. [Sree Chitra Tirunal Institute for Medical Sciences and Technology, Biosurface Technology Division (India); Sreenivasan, K., E-mail: sreeni@sctimst.ac.in [Sree Chitra Tirunal Institute for Medical Sciences and Technology, Laboratory for Polymer Analysis, Biomedical Technology Wing (India)

    2012-09-15

    This study refers an aqueous synthesis of methotrexate (MTX)-conjugated gold nanoparticles (GNPs), their interaction with HepG2 cells, and the use of Raman imaging to observe cellular internalization and drug delivery. GNPs of average size 3.5-5 nm were stabilized using the amine terminated bifunctional biocompatible copolymer and amended by conjugating MTX, an anticancer drug. The nanoparticles were released MTX at a faster rate in acidic pH and subsequently found to form aggregates. The Raman signals of cellular components were found to be enhanced by the aggregated particles enabling the mapping to visualize site-specific drug delivery. The methodology seems to have potential in optimizing the characteristics of nanodrug carriers for emptying the cargo precisely at specified sites.Graphical AbstractDrug release induced particle aggregation enhances Raman signals to aid in imaging.

  9. Jack bean (Canavalia ensiformis) urease induces eicosanoid-modulated hemocyte aggregation in the Chagas' disease vector Rhodnius prolixus.

    Science.gov (United States)

    Defferrari, M S; da Silva, R; Orchard, I; Carlini, C R

    2014-05-01

    Ureases are multifunctional proteins that display biological activities independently of their enzymatic function, such as induction of exocytosis and insecticidal effects. Rhodnius prolixus, a major vector of Chagas' disease, is a model for studies on the entomotoxicity of jack bean urease (JBU). We have previously shown that JBU induces the production of eicosanoids in isolated tissues of R. prolixus. In insects, the immune response comprises cellular and humoral reactions, and is centrally modulated by eicosanoids. Cyclooxygenase products signal immunity in insects, mainly cellular reactions, such as hemocyte aggregation. In searching for a link between JBU's toxic effects and immune reactions in insects, we have studied the effects of this toxin on R. prolixus hemocytes. JBU triggers aggregation of hemocytes after injection into the hemocoel and when applied to isolated cells. On in vitro assays, the eicosanoid synthesis inhibitors dexamethasone (phospholipase A2 indirect inhibitor) and indomethacin (cyclooxygenase inhibitor) counteracted JBU's effect, indicating that eicosanoids, more specifically cyclooxygenase products, are likely to mediate the aggregation response. Contrarily, the inhibitors esculetin and baicalein were inactive, suggesting that lipoxygenase products are not involved in JBU's effect. Extracellular calcium was also necessary for JBU's effect, in agreement to other cell models responsive to ureases. A progressive darkening of the medium of JBU-treated hemocytes was observed, suggestive of a humoral response. JBU was immunolocalized in the cultured cells upon treatment along with cytoskeleton damage. The highest concentration of JBU tested on cultured cells also led to nuclei aggregation of adherent hemocytes. This is the first time urease has been shown to affect insect hemocytes, contributing to our understanding of the entomotoxic mechanisms of action of this protein.

  10. Neo-lymphoid aggregates in the adult liver can initiate potent cell-mediated immunity.

    Directory of Open Access Journals (Sweden)

    Melanie Greter

    2009-05-01

    Full Text Available Subcutaneous immunization delivers antigen (Ag to local Ag-presenting cells that subsequently migrate into draining lymph nodes (LNs. There, they initiate the activation and expansion of lymphocytes specific for their cognate Ag. In mammals, the structural environment of secondary lymphoid tissues (SLTs is considered essential for the initiation of adaptive immunity. Nevertheless, cold-blooded vertebrates can initiate potent systemic immune responses even though they lack conventional SLTs. The emergence of lymph nodes provided mammals with drastically improved affinity maturation of B cells. Here, we combine the use of different strains of alymphoplastic mice and T cell migration mutants with an experimental paradigm in which the site of Ag delivery is distant from the site of priming and inflammation. We demonstrate that in mammals, SLTs serve primarily B cell priming and affinity maturation, whereas the induction of T cell-driven immune responses can occur outside of SLTs. We found that mice lacking conventional SLTs generate productive systemic CD4- as well as CD8-mediated responses, even under conditions in which draining LNs are considered compulsory for the initiation of adaptive immunity. We describe an alternative pathway for the induction of cell-mediated immunity (CMI, in which Ag-presenting cells sample Ag and migrate into the liver where they induce neo-lymphoid aggregates. These structures are insufficient to support antibody affinity maturation and class switching, but provide a novel surrogate environment for the initiation of CMI.

  11. Induced pluripotent stem cells

    Institute of Scientific and Technical Information of China (English)

    Siddhartha Bhowmik; LI Yong

    2011-01-01

    Induced pluripotent stem (iPS) cells are a recent development which has brought a promise of great therapeutic values. The previous technique of somatic cell nuclear transfer (SCNT) has been ineffective in humans. Recent discoveries show that human fibroblasts can be reprogrammed by a transient over expression of a small number of genes; they can undergo induced pluripotency. iPS were first produced in 2006. By 2008, work was underway to remove the potential oncogenes from their structure. In 2009, protein iPS (piPS) cells were discovered. Surface markers and reporter genes play an important role in stem cell research. Clinical applications include generation of self renewing stem cells, tissue replacement and many more. Stem cell therapy has the ability to dramatically change the treatment of human diseases.

  12. Theoretical study on electromagnetically induced transparency in molecular aggregate models using quantum Liouville equation method

    Energy Technology Data Exchange (ETDEWEB)

    Minami, Takuya; Nakano, Masayoshi [Department of Materials Engineering Science, Graduate School of Engineering Science, Osaka University, Toyonaka, Osaka 560-8531 (Japan)

    2015-01-22

    Electromagnetically induced transparency (EIT), which is known as an efficient control method of optical absorption property, is investigated using the polarizability spectra and population dynamics obtained by solving the quantum Liouville equation. In order to clarify the intermolecular interaction effect on EIT, we examine several molecular aggregate models composed of three-state monomers with the dipole-dipole coupling. On the basis of the present results, we discuss the applicability of EIT in molecular aggregate systems to a new type of optical switch.

  13. Extracellular polysaccharide composition of Azospirillum brasilense and its relation with cell aggregation.

    Science.gov (United States)

    Burdman, S; Jurkevitch, E; Soria-Díaz, M E; Serrano, A M; Okon, Y

    2000-08-15

    The exopolysaccharide (EPS) and capsular polysaccharide (CPS) composition of four Azospirillum brasilense strains differing in their aggregation capacity was analyzed by high performance anion exchange chromatography. When growing the different strains in an aggregation inducing medium containing a high carbon:nitrogen (C:N) ratio, both EPS and CPS showed a positive correlation between aggregation and the relative amount of arabinose. Arabinose was not detected in polysaccharides from Sp72002, a pleiotrophic Tn5 mutant strain impaired in aggregation. Arabinose was also not detected in extracellular polysaccharides of bacteria grown in a low C:N ratio, non-inducing aggregation medium, with exception for a relatively small amount found in the CPS of FAJ0204, a super-aggregating mutant strain. The only monosaccharides able to significantly inhibit aggregation at low sugar concentration when tested in a bioassay were arabinose (at a higher extent) and galactose. The possibility that residues of arabinose present in the extracellular polysaccharides are involved in the aggregation of A. brasilense is discussed.

  14. Vascular pentraxin 3 controls arterial thrombosis by targeting collagen and fibrinogen induced platelets aggregation

    Science.gov (United States)

    Bonacina, F.; Barbieri, S.S.; Cutuli, L.; Amadio, P.; Doni, A.; Sironi, M.; Tartari, S.; Mantovani, A.; Bottazzi, B.; Garlanda, C.; Tremoli, E.; Catapano, A.L.; Norata, G.D.

    2016-01-01

    Aim The long pentraxin PTX3 plays a non-redundant role during acute myocardial infarction, atherosclerosis and in the orchestration of tissue repair and remodeling during vascular injury, clotting and fibrin deposition. The aim of this work is to investigate the molecular mechanisms underlying the protective role of PTX3 during arterial thrombosis. Methods and results PTX3 KO mice transplanted with bone marrow from WT or PTX3 KO mice presented a significant reduction in carotid artery blood flow following FeCl3 induced arterial thrombosis (− 80.36 ± 11.5% and − 95.53 ± 4.46%), while in WT mice transplanted with bone marrow from either WT or PTX3 KO mice, the reduction was less dramatic (− 45.55 ± 1.37% and − 53.39 ± 9.8%), thus pointing to a protective effect independent of a hematopoietic cell's derived PTX3. By using P-selectin/PTX3 double KO mice, we further excluded a role for P-selectin, a target of PTX3 released by neutrophils, in vascular protection played by PTX3. In agreement with a minor role for hematopoietic cell-derived PTX3, platelet activation (assessed by flow cytometric expression of markers of platelet activation) was similar in PTX3 KO and WT mice as were haemostatic properties. Histological analysis indicated that PTX3 localizes within the thrombus and the vessel wall, and specific experiments with the N-terminal and the C-terminal PTX3 domain showed the ability of PTX3 to selectively dampen either fibrinogen or collagen induced platelet adhesion and aggregation. Conclusion PTX3 interacts with fibrinogen and collagen and, by dampening their pro-thrombotic effects, plays a protective role during arterial thrombosis. PMID:26976330

  15. PPARγ ligands decrease hydrostatic pressure-induced platelet aggregation and proinflammatory activity.

    Directory of Open Access Journals (Sweden)

    Fang Rao

    Full Text Available Hypertension is known to be associated with platelet overactivity, but the direct effects of hydrostatic pressure on platelet function remain unclear. The present study sought to investigate whether elevated hydrostatic pressure is responsible for platelet activation and to address the potential role of peroxisome proliferator-activated receptor-γ (PPARγ. We observed that hypertensive patients had significantly higher platelet volume and rate of ADP-induced platelets aggregation compared to the controls. In vitro, Primary human platelets were cultured under standard (0 mmHg or increased (120, 180, 240 mmHg hydrostatic pressure for 18 h. Exposure to elevated pressure was associated with morphological changes in platelets. Platelet aggregation and PAC-1 (the active confirmation of GPIIb/IIIa binding were increased, CD40L was translocated from cytoplasm to the surface of platelet and soluble CD40L (sCD40L was released into the medium in response to elevated hydrostatic pressure (180 and 240 mmHg. The PPARγ activity was up-regulated as the pressure was increased from 120 mmHg to 180 mmHg. Pressure-induced platelet aggregation, PAC-1 binding, and translocation and release of CD40L were all attenuated by the PPARγ agonist Thiazolidinediones (TZDs. These results demonstrate that platelet activation and aggregation are increased by exposure to elevated pressure and that PPARγ may modulate platelet activation induced by high hydrostatic pressure.

  16. Micelle depletion-induced vs. micelle-mediated aggregation in nanoparticles

    Energy Technology Data Exchange (ETDEWEB)

    Ray, D., E-mail: debes.phys@gmail.com; Aswal, V. K. [Solid State Physics Division, Bhabha Atomic Research Centre, Mumbai 400 085 (India)

    2015-06-24

    The phase behavior anionic silica nanoparticle (Ludox LS30) with non-ionic surfactants decaethylene glycol monododecylether (C12E10) and cationic dodecyltrimethyl ammonium bromide (DTAB) in aqueous electrolyte solution has been studied by small-angle neutron scattering (SANS). The measurements have been carried out for fixed concentrations of nanoparticle (1 wt%), surfactants (1 wt%) and electrolyte (0.1 M NaCl). Each of these nanoparticle–surfactant systems has been examined for different contrast conditions where individual components (nanoparticle or surfactant) are made visible. It is observed that the nanoparticle-micelle system in both the cases lead to the aggregation of nanoparticles. The aggregation is found to be micelle depletion-induced for C12E10 whereas micelle-mediated aggregation for DTAB. Interestingly, it is also found that phase behavior of mixed surfactant (C12E10 + DTAB) system is similar to that of C12E10 (unlike DTAB) micelles with nanoparticles.

  17. Homotypic aggregation of human cell lines by HLA class II-, class Ia- and HLA-G-specific monoclonal antibodies

    DEFF Research Database (Denmark)

    Odum, Niels; Ledbetter, J A; Martin, P

    1991-01-01

    adhesion between T and B cells by activating the CD18/CD11a (LFA-1) adhesion pathway. Here we report that monoclonal antibodies (mAb) against HLA-DR (L243, p4.1, HB10a, VI15) and certain broad class II reacting mAb (TU35, TU39), but not anti-DQ (TU22, Leu-10) mAb, induced homotypic aggregation of human......, but not the class I-negative parental line, 221, showed homotypic aggregation in response to an HLA-G specific mAb (87G) and a broad reacting class I-specific mAb (IOT2). Both cell lines responded with aggregation to anti-class II mAb (TU35). The anti-class I mAb, W6/32, had no effect on all cell lines tested...... and two anti-beta 2-microglobulin mAb had variable, weak effects. The aggregation response was an active, temperature-sensitive process which was almost totally abrogated by azide and by cytochalasins B and E, but unaffected by colchicine, EDTA, aphidicolin, actinomycin D and protein tyrosine kinase...

  18. Sorbitol crystallization-induced aggregation in frozen mAb formulations.

    Science.gov (United States)

    Piedmonte, Deirdre Murphy; Hair, Alison; Baker, Priti; Brych, Lejla; Nagapudi, Karthik; Lin, Hong; Cao, Wenjin; Hershenson, Susan; Ratnaswamy, Gayathri

    2015-02-01

    Sorbitol crystallization-induced aggregation of mAbs in the frozen state was evaluated. The effect of protein aggregation resulting from sorbitol crystallization was measured as a function of formulation variables such as protein concentration and pH. Long-term studies were performed on both IgG1 and IgG2 mAbs over the protein concentration range of 0.1-120 mg/mL. Protein aggregation was measured by size-exclusion HPLC (SE-HPLC) and further characterized by capillary-electrophoresis SDS. Sorbitol crystallization was monitored and characterized by subambient differential scanning calorimetry and X-ray diffraction. Aggregation due to sorbitol crystallization is inversely proportional to both protein concentration and formulation pH. At high protein concentrations, sorbitol crystallization was suppressed, and minimal aggregation by SE-HPLC resulted, presumably because of self-stabilization of the mAbs. The glass transition temperature (Tg ') and fragility index measurements were made to assess the influence of molecular mobility on the crystallization of sorbitol. Tg ' increased with increasing protein concentration for both mAbs. The fragility index decreased with increasing protein concentration, suggesting that it is increasingly difficult for sorbitol to crystallize at high protein concentrations.

  19. Thermally-induced aggregation and fusion of protein-free lipid vesicles.

    Science.gov (United States)

    Ibarguren, Maitane; Bomans, Paul H H; Ruiz-Mirazo, Kepa; Frederik, Peter M; Alonso, Alicia; Goñi, Félix M

    2015-12-01

    Membrane fusion is an important phenomenon in cell biology and pathology. This phenomenon can be modeled using vesicles of defined size and lipid composition. Up to now fusion models typically required the use of chemical (polyethyleneglycol, cations) or enzymatic catalysts (phospholipases). We present here a model of lipid vesicle fusion induced by heat. Large unilamellar vesicles consisting of a phospholipid (dioleoylphosphatidylcholine), cholesterol and diacylglycerol in a 43:57:3 mol ratio were employed. In this simple system, fusion was the result of thermal fluctuations, above 60 °C. A similar system containing phospholipid and cholesterol but no diacylglycerol was observed to aggregate at and above 60 °C, in the absence of fusion. Vesicle fusion occurred under our experimental conditions only when (31)P NMR and cryo-transmission electron microscopy of the lipid mixtures used in vesicle preparation showed non-lamellar lipid phase formation (hexagonal and cubic). Non-lamellar structures are probably the result of lipid reassembly of the products of individual fusion events, or of fusion intermediates. A temperature-triggered mechanism of lipid reassembly might have occurred at various stages of protocellular evolution.

  20. Synthetic Quorum Sensing and Induced Aggregation in Model Microcapsule Colonies with Repressilator Feedback

    Science.gov (United States)

    Shum, Henry; Yashin, Victor; Balazs, Anna

    We model a system of synthetic microcapsules that communicate chemically by releasing nanoparticles or signaling molecules. These signaling species bind to receptors on the shells of capsules and modulate the target shell's permeability, thereby controlling nanoparticle release from the target capsule. Using the repressilator regulatory network motif, whereby three species suppress the production of the next in a cyclic fashion, we show that large amplitude oscillations in nanoparticle release can emerge when many capsules are close together. This exemplifies quorum sensing, which is the ability of cells to gauge their population density and collectively initiate a new behavior once a critical density is reached. We present a physically realizable model in which the oscillations exhibited in crowded populations induce aggregation of the microcapsules, mimicking complex biological behavior of the slime mold Dictyostelium discoideum with only simple, synthetic components. We also show that the clusters can be dispersed and reformed repeatedly and controllably by addition of chemical stimuli, demonstrating possible applications in creating reconfigurable or programmable materials.

  1. Inhibitory Effect of Waste Glass Powder on ASR Expansion Induced by Waste Glass Aggregate

    Directory of Open Access Journals (Sweden)

    Shuhua Liu

    2015-10-01

    Full Text Available Detailed research is carried out to ascertain the inhibitory effect of waste glass powder (WGP on alkali-silica reaction (ASR expansion induced by waste glass aggregate in this paper. The alkali reactivity of waste glass aggregate is examined by two methods in accordance with the China Test Code SL352-2006. The potential of WGP to control the ASR expansion is determined in terms of mean diameter, specific surface area, content of WGP and curing temperature. Two mathematical models are developed to estimate the inhibitory efficiency of WGP. These studies show that there is ASR risk with an ASR expansion rate over 0.2% when the sand contains more than 30% glass aggregate. However, WGP can effectively control the ASR expansion and inhibit the expansion rate induced by the glass aggregate to be under 0.1%. The two mathematical models have good simulation results, which can be used to evaluate the inhibitory effect of WGP on ASR risk.

  2. An effect of glycoprotein IIb/IIIa inhibitors on the kinetics of red blood cells aggregation.

    Science.gov (United States)

    Sokolova, Irina A; Muravyov, Alexei V; Khokhlova, Maria D; Rikova, Sofya Yu; Lyubin, Evgeny V; Gafarova, Marina A; Skryabina, Maria N; Fedyanin, Angrey A; Kryukova, Darya V; Shahnazarov, Alexander A

    2014-01-01

    The reversible aggregation of red blood cells (RBCs) continues to be of the basic science and clinical interest. Recently it has been reported about a specific binding between fibrinogen and unknown erythrocyte glycoprotein receptors. The aim of this study was to investigate whether the red blood cell aggregation (RBCA) include the cell-cell interaction using the membrane receptors that bind such ligands as fibrinogen or fibronectin. To test this hypothesis the RBCs were incubated with monafram - the drug of the monoclonal antibodies against glycoprotein (GP) IIb/IIIa, with the GPIIb-IIIa receptor antagonist tirofiban, epifibatide and with the fibrinogen inhibiting peptide. It has been found that the RBC incubation with monafram resulted in a marked RBCA decrease mainly in persons with high level of aggregation. Another research session has shown that RBC incubation with fibronectin was accompanied by a significant RBCA rise. The monafram addition to red cell incubation medium resulted in a significant RBCA lowering. The cell incubation with tirofiban and epifibatide issued in RBCA decrease. The similar results were obtained when RBCs were incubated with the fibrinogen inhibiting peptide. Although monafram, tirofiban, eptifibatide and the fibrinogen inhibiting peptide were related to fibrinogen function they didn't inhibit RBCA completely. Therefore, under moderate and low red blood cell aggregation the cell binding is probably related to nonspecific mode. It seems evident that the specific and nonspecific modes of red blood cell aggregate formation could co-exist. Additional theoretical and experimental investigations in this area are needed.

  3. Glycosylation analysis of an aggregated antibody produced by Chinese hamster ovary cells in bioreactor culture.

    Science.gov (United States)

    Onitsuka, Masayoshi; Kawaguchi, Akira; Asano, Ryutaro; Kumagai, Izumi; Honda, Kohsuke; Ohtake, Hisao; Omasa, Takeshi

    2014-05-01

    N-Glycosylation of therapeutic antibodies contributes not only to their biological function, but also to their stability and tendency to aggregate. Here, we investigated the impact of the glycosylation status of an aggregated antibody that accumulated during the bioreactor culture of Chinese hamster ovary cells. High-performance liquid chromatography analysis showed that there was no apparent difference in the glycosylation patterns of monomeric, dimeric, and large aggregated forms of the antibody. In contrast, lectin binding assays, which enable the total amounts of specific sugar residues to be detected, showed that both galactose and fucose residues in dimers and large aggregates were reduced to 70-80% of the amount in monomers. These results strongly suggest that the lack of N-linked oligosaccharides, a result of deglycosylation or aglycosylation, occurred in a proportion of the dimeric and large aggregated components. The present study demonstrates that glycosylation heterogeneities are a potential cause of antibody aggregation in cell culture of Chinese hamster ovary cells, and that the lack of N-glycosylation promotes the formation of dimers and finally results in large aggregates.

  4. Photometric measurements of red blood cell aggregation: light transmission versus light reflectance.

    Science.gov (United States)

    Baskurt, Oguz K; Uyuklu, Mehmet; Hardeman, Max R; Meiselman, Herbert J

    2009-01-01

    Red blood cell (RBC) aggregation is the reversible and regular clumping in the presence of certain macromolecules. This is a clinically important phenomenon, being significantly enhanced in the presence of acute phase reactants (e.g., fibrinogen). Both light reflection (LR) and light transmission (LT) from or through thin layers of RBC suspensions during the process of aggregation are accepted to reflect the time course of aggregation. It has been recognized that the time courses of LR and LT might be different from each other. We aim to compare the RBC aggregation measurements based on simultaneous recordings of LR and LT. The results indicate that LR during RBC aggregation is characterized by a faster time course compared to simultaneously recorded LT. This difference in time course of LR and LT is reflected in the calculated parameters reflecting the overall extent and kinetics of RBC aggregation. Additionally, the power of parameters calculated using LR and LT time courses in detecting a given difference in aggregation are significantly different from each other. These differences should be taken into account in selecting the appropriate calculated parameters for analyzing LR or LT time courses for the assessment of RBC aggregation.

  5. Oxygen transport and stem cell aggregation in stirred-suspension bioreactor cultures.

    Directory of Open Access Journals (Sweden)

    Jincheng Wu

    Full Text Available Stirred-suspension bioreactors are a promising modality for large-scale culture of 3D aggregates of pluripotent stem cells and their progeny. Yet, cells within these clusters experience limitations in the transfer of factors and particularly O2 which is characterized by low solubility in aqueous media. Cultured stem cells under different O2 levels may exhibit significantly different proliferation, viability and differentiation potential. Here, a transient diffusion-reaction model was built encompassing the size distribution and ultrastructural characteristics of embryonic stem cell (ESC aggregates. The model was coupled to experimental data from bioreactor and static cultures for extracting the effective diffusivity and kinetics of consumption of O2 within mouse (mESC and human ESC (hESC clusters. Under agitation, mESC aggregates exhibited a higher maximum consumption rate than hESC aggregates. Moreover, the reaction-diffusion model was integrated with a population balance equation (PBE for the temporal distribution of ESC clusters changing due to aggregation and cell proliferation. Hypoxia was found to be negligible for ESCs with a smaller radius than 100 µm but became appreciable for aggregates larger than 300 µm. The integrated model not only captured the O2 profile both in the bioreactor bulk and inside ESC aggregates but also led to the calculation of the duration that fractions of cells experience a certain range of O2 concentrations. The approach described in this study can be employed for gaining a deeper understanding of the effects of O2 on the physiology of stem cells organized in 3D structures. Such frameworks can be extended to encompass the spatial and temporal availability of nutrients and differentiation factors and facilitate the design and control of relevant bioprocesses for the production of stem cell therapeutics.

  6. Three cyanobacteriochromes work together to form a light color-sensitive input system for c-di-GMP signaling of cell aggregation.

    Science.gov (United States)

    Enomoto, Gen; Ni-Ni-Win; Narikawa, Rei; Ikeuchi, Masahiko

    2015-06-30

    Cyanobacteriochromes (CBCRs) are cyanobacterial photoreceptors that have diverse spectral properties and domain compositions. Although large numbers of CBCR genes exist in cyanobacterial genomes, no studies have assessed whether multiple CBCRs work together. We recently showed that the diguanylate cyclase (DGC) activity of the CBCR SesA from Thermosynechococcus elongatus is activated by blue-light irradiation and that, when irradiated, SesA, via its product cyclic dimeric GMP (c-di-GMP), induces aggregation of Thermosynechococcus vulcanus cells at a temperature that is suboptimum for single-cell viability. For this report, we first characterize the photobiochemical properties of two additional CBCRs, SesB and SesC. Blue/teal light-responsive SesB has only c-di-GMP phosphodiesterase (PDE) activity, which is up-regulated by teal light and GTP. Blue/green light-responsive SesC has DGC and PDE activities. Its DGC activity is enhanced by blue light, whereas its PDE activity is enhanced by green light. A ΔsesB mutant cannot suppress cell aggregation under teal-green light. A ΔsesC mutant shows a less sensitive cell-aggregation response to ambient light. ΔsesA/ΔsesB/ΔsesC shows partial cell aggregation, which is accompanied by the loss of color dependency, implying that a nonphotoresponsive DGC(s) producing c-di-GMP can also induce the aggregation. The results suggest that SesB enhances the light color dependency of cell aggregation by degrading c-di-GMP, is particularly effective under teal light, and, therefore, seems to counteract the induction of cell aggregation by SesA. In addition, SesC seems to improve signaling specificity as an auxiliary backup to SesA/SesB activities. The coordinated action of these three CBCRs highlights why so many different CBCRs exist.

  7. Mechanical agitation induces counterintuitive aggregation of pre-dispersed carbon nanotubes.

    Science.gov (United States)

    Fernandes, Ricardo M F; Buzaglo, Matat; Regev, Oren; Furó, István; Marques, Eduardo F

    2017-05-01

    Mechanical agitation is commonly used to fragment and disperse insoluble materials in liquids. However, here we show that when pristine single-walled carbon nanotubes pre-dispersed in water are subject to vortex-shaking for very short periods (typically 10-60s, power density ∼0.002WmL(-1)), re-aggregation counterintuitively occurs. The initial dispersions are produced using surfactants as dispersants and powerful tip sonication (∼1WmL(-1)) followed by centrifugation. Detailed imaging by light and electron microscopies shows that the vortex-induced aggregates consist of loose networks (1-10(2)μm in size) of intertwined tubes and thin bundles. The average aggregate size increases with vortexing time in an apparently logarithmic manner and depends on the dispersant used, initial concentration of nanotubes and size distribution of bundles. The aggregation is, nonetheless, reversible: if the vortex-shaken dispersions are mildly bath-sonicated (∼0.03WmL(-1)), the flocs break down and re-dispersal occurs. Molecular insight for the mechanism behind this surprising phenomenon is put forth.

  8. Useful multivariate kinetic analysis: Size determination based on cystein-induced aggregation of gold nanoparticles

    Science.gov (United States)

    Rabbani, Faride; Hormozi Nezhad, Mohammad Reza; Abdollahi, Hamid

    2013-11-01

    This study describes spectrometric monitored kinetic processes to determine the size of citrate-capped Au nanoparticles (Au NPs) based on aggregation induced by L-cysteine (L-Cys) as a molecular linker. The Au NPs association process is thoroughly dependent on pH, concentration and size of nanoparticles. Size dependency of aggregation inspirits to determine the average diameters of Au NPs. For this aim the procedure is achieved in aqueous medium at pH 7 (phosphate buffer), and multivariate data including kinetic spectra of Au NPs are collected during aggregation process. Subsequently partial least squares (PLS) modeling is carried out analyzing the obtained data. The model is built on the basis of relation between the kinetics behavior of aggregation and different Au NPs sizes. Training the model was performed using latent variables (LVs) of the original data. The analytical performance of the model was characterized by relative standard error. The proposed method was applied to determination of size in unknown samples. The predicted sizes of unknown samples that obtained by the introduced method are interestingly in agreement with the sizes measured by Transmission Electron Microscopy (TEM) images and Dynamic Light Scattering (DLS) measurement.

  9. Enzymatic hydrolysis of heat-induced aggregates of whey protein isolate.

    Science.gov (United States)

    O'Loughlin, I B; Murray, B A; Kelly, P M; FitzGerald, R J; Brodkorb, A

    2012-05-16

    The effects of heat-induced denaturation and subsequent aggregation of whey protein isolate (WPI) solutions on the rate of enzymatic hydrolysis was investigated. Both heated (60 °C, 15 min; 65 °C, 5 and 15 min; 70 °C, 5 and 15 min, 75 °C, 5 and 15 min; 80 °C, 10 min) and unheated WPI solutions (100 g L(-1) protein) were incubated with a commercial proteolytic enzyme preparation, Corolase PP, until they reached a target degree of hydrolysis (DH) of 5%. WPI solutions on heating were characterized by large aggregate formation, higher viscosity, and surface hydrophobicity and hydrolyzed more rapidly (P whey proteins exhibited differences in their susceptibility to hydrolysis. Both viscosity and surface hydrophobicity along with insolubility declined as hydrolysis progressed. However, microstructural changes observed by light and confocal laser scanning microscopy (CLSM) provided insights to suggest that aggregate size and porosity may be complementary to denaturation in promoting faster enzymatic hydrolysis. This could be clearly observed in the course of aggregate disintegration, gel network breakdown, and improved solution clarification.

  10. Quantification of alginate by aggregation induced by calcium ions and fluorescent polycations.

    Science.gov (United States)

    Zheng, Hewen; Korendovych, Ivan V; Luk, Yan-Yeung

    2016-01-01

    For quantification of polysaccharides, including heparins and alginates, the commonly used carbazole assay involves hydrolysis of the polysaccharide to form a mixture of UV-active dye conjugate products. Here, we describe two efficient detection and quantification methods that make use of the negative charges of the alginate polymer and do not involve degradation of the targeted polysaccharide. The first method utilizes calcium ions to induce formation of hydrogel-like aggregates with alginate polymer; the aggregates can be quantified readily by staining with a crystal violet dye. This method does not require purification of alginate from the culture medium and can measure the large amount of alginate that is produced by a mucoid Pseudomonas aeruginosa culture. The second method employs polycations tethering a fluorescent dye to form suspension aggregates with the alginate polyanion. Encasing the fluorescent dye in the aggregates provides an increased scattering intensity with a sensitivity comparable to that of the conventional carbazole assay. Both approaches provide efficient methods for monitoring alginate production by mucoid P. aeruginosa.

  11. Investigation into process-induced de-aggregation of cohesive micronised API particles.

    Science.gov (United States)

    Hoffmann, Magnus; Wray, Patrick S; Gamble, John F; Tobyn, Mike

    2015-09-30

    The aim of this study was to assess the impact of unit processes on the de-aggregation of a cohesive micronised API within a pharmaceutical formulation using near-infrared chemical imaging. The impact on the primary API particles was also investigated using an image-based particle characterization system with integrated Raman analysis. The blended material was shown to contain large, API rich domains which were distributed in-homogeneously across the sample, suggesting that the blending process was not aggressive enough to disperse aggregates of micronised drug particles. Cone milling, routinely used to improve the homogeneity of such cohesive formulations, was observed to substantially reduce the number and size of API rich domains; however, several smaller API domains survived the milling process. Conveyance of the cone milled formulation through the Alexanderwerk WP120 powder feed system completely dispersed all remaining aggregates. Importantly, powder feed transmission of the un-milled formulation was observed to produce an equally homogeneous API distribution. The size of the micronised primary drug particles remained unchanged during powder feed transmission. These findings provide further evidence that this powder feed system does induce shear, and is in fact better able to disperse aggregates of a cohesive micronised API within a blend than the blend-mill-blend step.

  12. A particle-based model to simulate the micromechanics of single-plant parenchyma cells and aggregates.

    Science.gov (United States)

    Van Liedekerke, P; Ghysels, P; Tijskens, E; Samaey, G; Smeedts, B; Roose, D; Ramon, H

    2010-05-26

    This paper is concerned with addressing how plant tissue mechanics is related to the micromechanics of cells. To this end, we propose a mesh-free particle method to simulate the mechanics of both individual plant cells (parenchyma) and cell aggregates in response to external stresses. The model considers two important features in the plant cell: (1) the cell protoplasm, the interior liquid phase inducing hydrodynamic phenomena, and (2) the cell wall material, a viscoelastic solid material that contains the protoplasm. In this particle framework, the cell fluid is modeled by smoothed particle hydrodynamics (SPH), a mesh-free method typically used to address problems with gas and fluid dynamics. In the solid phase (cell wall) on the other hand, the particles are connected by pairwise interactions holding them together and preventing the fluid to penetrate the cell wall. The cell wall hydraulic conductivity (permeability) is built in as well through the SPH formulation. Although this model is also meant to be able to deal with dynamic and even violent situations (leading to cell wall rupture or cell-cell debonding), we have concentrated on quasi-static conditions. The results of single-cell compression simulations show that the conclusions found by analytical models and experiments can be reproduced at least qualitatively. Relaxation tests revealed that plant cells have short relaxation times (1 micros-10 micros) compared to mammalian cells. Simulations performed on cell aggregates indicated an influence of the cellular organization to the tissue response, as was also observed in experiments done on tissues with a similar structure.

  13. Sponge cell reaggregation: Cellular structure and morphogenetic potencies of multicellular aggregates.

    Science.gov (United States)

    Lavrov, Andrey I; Kosevich, Igor A

    2016-02-01

    Sponges (phylum Porifera) are one of the most ancient extant multicellular animals and can provide valuable insights into origin and early evolution of Metazoa. High plasticity of cell differentiations and anatomical structure is characteristic feature of sponges. Present study deals with sponge cell reaggregation after dissociation as the most outstanding case of sponge plasticity. Dynamic of cell reaggregation and structure of multicellular aggregates of three demosponge species (Halichondria panicea (Pallas, 1766), Haliclona aquaeductus (Sсhmidt, 1862), and Halisarca dujardinii Johnston, 1842) were studied. Sponge tissue dissociation was performed mechanically. Resulting cell suspensions were cultured at 8-10°C for at least 5 days. Structure of multicellular aggregates was studied by light, transmission and scanning electron microscopy. Studied species share common stages of cell reaggregation-primary multicellular aggregates, early-stage primmorphs and primmorphs, but the rate of reaggregation varies considerably among species. Only cells of H. dujardinii are able to reconstruct functional and viable sponge after primmorphs formation. Sponge reconstruction in this species occurs due to active cell locomotion. Development of H. aquaeductus and H. panicea cells ceases at the stages of early primmorphs and primmorphs, respectively. Development of aggregates of these species is most likely arrested due to immobility of the majority of cells inside them. However, the inability of certain sponge species to reconstruct functional and viable individuals during cell reaggregation may be not a permanent species-specific characteristic, but depends on various factors, including the stage of the life cycle and experimental conditions.

  14. Bivalent Copper Ions Promote Fibrillar Aggregation of KCTD1 and Induce Cytotoxicity.

    Science.gov (United States)

    Liu, Zhepeng; Song, Feifei; Ma, Zhi-Li; Xiong, Qiushuang; Wang, Jingwen; Guo, Deyin; Sun, Guihong

    2016-01-01

    Potassium channel tetramerization domain containing 1 (KCTD1) family members have a BTB/POZ domain, which can facilitate protein-protein interactions involved in the regulation of different signaling pathways. KCTD proteins have potential Zn(2+)/Cu(2+) binding sites with currently unknown structural and functional roles. We investigated potential Cu(2+)-specific effects on KCTD1 using circular dichroism, turbidity measurement, fluorescent dye binding, proteinase K (PK) digestion, cell proliferation and apoptosis assays. These experiments indicate that the KCTD1 secondary structure assumes greater β-sheet content and the proteins aggregate into a PK-resistant form under 20 μM Cu(2+), and this β-sheet-rich aggregation with Cu(2+) promotes fibril formation, which results in increased cell toxicity by apoptosis. Our results reveal a novel role for Cu(2+) in determining the structure and function of KCTD1.

  15. The yeast peroxiredoxin Tsa1 protects against protein-aggregate-induced oxidative stress

    OpenAIRE

    2014-01-01

    ABSTRACT Peroxiredoxins are ubiquitous thiol-specific proteins that have multiple functions in stress protection, including protection against oxidative stress. Tsa1 is the major yeast peroxiredoxin and we show that it functions as a specific antioxidant to protect the cell against the oxidative stress caused by nascent-protein misfolding and aggregation. Yeast mutants lacking TSA1 are sensitive to misfolding caused by exposure to the proline analogue azetidine-2-carboxylic acid (AZC). AZC pr...

  16. Gingival tissue-produced inhibition of platelet aggregation and the loss of inhibition in streptozotocin-induced diabetic rats

    Energy Technology Data Exchange (ETDEWEB)

    Kawamura, Keiichiroh; Tamai, Kazuharu; Shirakawa, Masaharu; Okamoto, Hiroshi; Dohi, Toshihiro; Tsujimoto, Akira

    1988-01-01

    Addition of medium incubated with normal rat gingival tissue to platelet-rich plasma inhibited ADP-induced platelet aggregation. The ability of rat gingiva to produce activity inhibiting platelet aggregation was enhanced by the addition of arachidonic acid. Diabetic rat gingiva failed to inhibit platelet aggregation but did produce the anti-platelet aggregating activity in the presence of arachidonic acid. Indomethacin blocked the production of anti-platelet aggregating activity. There was no difference in conversion of (1-/sup 14/C)arachidonic acid to prostaglandins by normal and diabetic rat gingiva. These results suggest that an arachidonic acid metabolite released from gingiva during incubation inhibits platelet aggregation, and the synthesis of the metabolite is impaired in diabetic rat gingiva. A decrease in availability of arachidonic acid may be a causal factor of the defect in diabetic rat gingiva.

  17. Calcium-induced aggregation of archaeal bipolar tetraether liposomes derived from the thermoacidophilic archaeon Sulfolobus acidocaldarius

    Directory of Open Access Journals (Sweden)

    Roby Kanichay

    2003-01-01

    Full Text Available Previously, we showed that the proton permeability of small unilamellar vesicles (SUVs composed of polar lipid fraction E (PLFE from the thermoacidophilic archaeon Sulfolobus acidocaldarius was remarkably low and insensitive to temperature (Komatsu and Chong 1998. In this study, we used photon correlation spectroscopy to investigate the time dependence of PLFE SUV size as a function of Ca2+ concentration. In the absence of Ca2+, vesicle diameter changed little over 6 months. Addition of Ca2+, however, immediately induced formation of vesicle aggregates with an irregular shape, as revealed by confocal fluorescence microscopy. Aggregation was reversible upon addition of EDTA; however, the reversibility varied with temperature as well as incubation time with Ca2+. Freeze-fracture electron microscopy showed that, after a long period of incubation (2 weeks with Ca2+, the PLFE vesicles had not just aggregated, but had fused or coalesced. The initial rate of vesicle aggregation varied sigmoidally with Ca2+ concentration. At pH 6.6, the threshold calcium concentration (Cr for vesicle aggregation at 25 and 40 °C was 11 and 17 mM, respectively. At pH 3.0, the Cr at 25 °C increased to 25 mM. The temperature dependence of Cr may be attributable to changes in membrane surface potential, which was –22.0 and –13.2 mV at 25 and 40 °C, respectively, at pH 6.6, as determined by 2-(p-toluidinylnaphthalene-6-sulfonic acid fluorescence. The variation in surface potential with temperature is discussed in terms of changes in lipid conformation and membrane organization.

  18. Calcium-induced aggregation of archaeal bipolar tetraether liposomes derived from the thermoacidophilic archaeon Sulfolobus acidocaldarius.

    Science.gov (United States)

    Kanichay, Roby; Boni, Lawrence T; Cooke, Peter H; Khan, Tapan K; Chong, Parkson Lee-Gau

    2003-10-01

    Previously, we showed that the proton permeability of small unilamellar vesicles (SUVs) composed of polar lipid fraction E (PLFE) from the thermoacidophilic archaeon Sulfolobus acidocaldarius was remarkably low and insensitive to temperature (Komatsu and Chong 1998). In this study, we used photon correlation spectroscopy to investigate the time dependence of PLFE SUV size as a function of Ca2+ concentration. In the absence of Ca2+, vesicle diameter changed little over 6 months. Addition of Ca2+, however, immediately induced formation of vesicle aggregates with an irregular shape, as revealed by confocal fluorescence microscopy. Aggregation was reversible upon addition of EDTA; however, the reversibility varied with temperature as well as incubation time with Ca2+. Freeze-fracture electron microscopy showed that, after a long period of incubation (2 weeks) with Ca2+, the PLFE vesicles had not just aggregated, but had fused or coalesced. The initial rate of vesicle aggregation varied sigmoidally with Ca2+ concentration. At pH 6.6, the threshold calcium concentration (Cr) for vesicle aggregation at 25 and 40 degrees C was 11 and 17 mM, respectively. At pH 3.0, the Cr at 25 degrees C increased to 25 mM. The temperature dependence of Cr may be attributable to changes in membrane surface potential, which was -22.0 and -13.2 mV at 25 and 40 degrees C, respectively, at pH 6.6, as determined by 2-(p-toluidinyl)naphthalene-6-sulfonic acid fluorescence. The variation in surface potential with temperature is discussed in terms of changes in lipid conformation and membrane organization.

  19. Fabrication of aggregation-induced emission based fluorescent nanoparticles and their biological imaging application: recent progress and perspectives

    Directory of Open Access Journals (Sweden)

    Bin Yang

    2016-06-01

    Full Text Available Aggregation-induced emission (AIE dyes have received wide-spread concern since their inception. Several types of AIE-based fluorescent nanoparticle (FNP have been developed, and the potential applications of these FNPs have also been explored. Recent studies of AIE-based FNPs in biological areas have suggested that they show promise as bio-materials for cell imaging and other biomedical applications. This article reviews recent progress in the synthesis of AIE-based FNPs via non-covalent, covalent and novel one-pot strategies, and the subsequent cell-imaging of those AIE-based FNPs. Many successes have been achieved, and there is still plenty of space for the development of AIE-based FNPs as new bio-materials.

  20. Adenosine diphosphate-induced aggregation of human platelets in flow through tubes. I. Measurement of concentration and size of single platelets and aggregates.

    Science.gov (United States)

    Bell, D N; Spain, S; Goldsmith, H L

    1989-11-01

    A double infusion flow system and particle sizing technique were developed to study the effect of time and shear rate on adenosine diphosphate-induced platelet aggregation in Poiseuille flow. Citrated platelet-rich plasma, PRP, and 2 microM ADP were simultaneously infused into a 40-microliters cylindrical mixing chamber at a fixed flow ratio, PRP/ADP = 9:1. After rapid mixing by a rotating magnetic stirbar, the platelet suspension flowed through 1.19 or 0.76 mm i.d. polyethylene tubing for mean transit times, t, from 0.1 to 86 s, over a range of mean tube shear rate, G, from 41.9 to 1,000 s-1. Known volumes of suspension were collected into 0.5% buffered glutaraldehyde, and all particles in the volume range 1-10(5) microns 3 were counted and sized using a model ZM particle counter (Coulter Electronics Inc., Hialeah, FL) and a logarithmic amplifier. The decrease in the single platelet concentration served as an overall index of aggregation. The decrease in the total particle concentration was used to calculate the collision capture efficiency during the early stages of aggregation, and aggregate growth was followed by changes in the volume fraction of particles of successively increasing size. Preliminary results demonstrate that both collision efficiency and particle volume fraction reveal important aspects of the aggregation process not indicated by changes in the single platelet concentration alone.

  1. Effect of Thai medicinal plant extracts on cell aggregation of Escherichia coli O157: H7.

    Directory of Open Access Journals (Sweden)

    Limsuwan, S.

    2005-08-01

    Full Text Available Medicinal plants have been used for treating diarrhoea but the interference mechanisms are not clearly understood. One possible hypothesis is that of an effect on cell surface hydrophobicity of microbial cells. In this study, we examined cell aggregation affected by crude extracts of Thai medicinal plants on cell surface hydrophobicity of Escherichia coli strains by salt aggregation test. Correlation between minimal inhibitory concentration and cell aggregation was performed. Aqueous and ethanolic extracts of 8 medicinal plants including Acacia catechu, Holarrhena antidysenterica, Peltophorum pterocarpum, Piper sarmentosum, Psidium guajava, Punica granatum, Quercus infectoria, and Tamarindus indica were tested with E. coli O157: H7 and other E. coli strains isolated from human, porcine, and foods. Aqueous extracts of Peltophorum pterocarpum, Psidium guajava, and Punica granatum were highly effective against E. coli O157: H7 with the MIC values of 0.09 to 0.39, 0.19 to 0.78, and 0.09 to 1.56 mg/ml, respectively. Ethanolic extract of Quercus infectoria and Punica granatum demonstrated good MIC values of 0.09 to 0.78, and 0.19 to 0.78 mg/ml, respectively. It was established that aqueous extracts of Punica granatum and Piper sarmentosum at high concentration (25 mg/ml enhanced cell aggregation of almost all E. coli strains while aqueous and ethanolic extracts ofQuercus infectoria enhanced cell aggregation of some E. coli strains. Correlation between minimal inhibitory concentration and cell aggregation was not found in this study.

  2. Shear-induced reaction-limited aggregation kinetics of brownian particles at arbitrary concentrations.

    Science.gov (United States)

    Zaccone, Alessio; Gentili, Daniele; Wu, Hua; Morbidelli, Massimo

    2010-04-07

    The aggregation of interacting brownian particles in sheared concentrated suspensions is an important issue in colloid and soft matter science per se. Also, it serves as a model to understand biochemical reactions occurring in vivo where both crowding and shear play an important role. We present an effective medium approach within the Smoluchowski equation with shear which allows one to calculate the encounter kinetics through a potential barrier under shear at arbitrary colloid concentrations. Experiments on a model colloidal system in simple shear flow support the validity of the model in the concentration range considered. By generalizing Kramers' rate theory to the presence of shear and collective hydrodynamics, our model explains the significant increase in the shear-induced reaction-limited aggregation kinetics upon increasing the colloid concentration.

  3. Detection of Gold Nanoparticles Aggregation Growth Induced by Nucleic Acid through Laser Scanning Confocal Microscopy

    Science.gov (United States)

    Gary, Ramla; Carbone, Giovani; Petriashvili, Gia; De Santo, Maria Penelope; Barberi, Riccardo

    2016-01-01

    The gold nanoparticle (GNP) aggregation growth induced by deoxyribonucleic acid (DNA) is studied by laser scanning confocal and environmental scanning electron microscopies. As in the investigated case the direct light scattering analysis is not suitable, we observe the behavior of the fluorescence produced by a dye and we detect the aggregation by the shift and the broadening of the fluorescence peak. Results of laser scanning confocal microscopy images and the fluorescence emission spectra from lambda scan mode suggest, in fact, that the intruding of the hydrophobic moiety of the probe within the cationic surfactants bilayer film coating GNPs results in a Förster resonance energy transfer. The environmental scanning electron microscopy images show that DNA molecules act as template to assemble GNPs into three-dimensional structures which are reminiscent of the DNA helix. This study is useful to design better nanobiotechnological devices using GNPs and DNA. PMID:26907286

  4. Detection of Gold Nanoparticles Aggregation Growth Induced by Nucleic Acid through Laser Scanning Confocal Microscopy

    Directory of Open Access Journals (Sweden)

    Ramla Gary

    2016-02-01

    Full Text Available The gold nanoparticle (GNP aggregation growth induced by deoxyribonucleic acid (DNA is studied by laser scanning confocal and environmental scanning electron microscopies. As in the investigated case the direct light scattering analysis is not suitable, we observe the behavior of the fluorescence produced by a dye and we detect the aggregation by the shift and the broadening of the fluorescence peak. Results of laser scanning confocal microscopy images and the fluorescence emission spectra from lambda scan mode suggest, in fact, that the intruding of the hydrophobic moiety of the probe within the cationic surfactants bilayer film coating GNPs results in a Förster resonance energy transfer. The environmental scanning electron microscopy images show that DNA molecules act as template to assemble GNPs into three-dimensional structures which are reminiscent of the DNA helix. This study is useful to design better nanobiotechnological devices using GNPs and DNA.

  5. Phytoplankton aggregate formation: observations of patterns and mechanisms of cell sticking and the significance of exopolymeric material

    DEFF Research Database (Denmark)

    Kiørboe, Thomas; Hansen, Jorgen L. S.

    1993-01-01

    are sticky in themselves, and coagulation depends on cell-cell sticking and does not involve mucus. Aggregates are composed solely of cells. Cells of the diatom Chaetoceros affinis, on the other hand, are not in themselves sticky. Transparent exopolymeric particles (TEP), produced by the diatom, cause...... the cells to aggregate and coagulation depends on TEP-cell rather than cell-cell sticking. Aggregates are formed of a mixture of mucus and cells. We found several species of diatoms and one flagellate species to produce copious amounts of TEP. TEP from some species (e.g. Coscinodiscus sp.) is sticky and may...

  6. Submicrometer-scale ZnO Composite Aggregate Arrays Photoanodes for Dye-sensitized Solar Cells

    Institute of Scientific and Technical Information of China (English)

    Wei Jia; Suihu Dang; Hairui Liu; Zhuxia Zhang; Tianbao Li; Xuguang Liu; Bingshe Xu

    2013-01-01

    Submicrometer-scale ZnO composite aggregate arrays of nanorods and nanoparticles were prepared by simple wet-chemical route and studied as dye-sensitized solar cells (DSSCs) photoanodes.The ZnO composite aggregate arrays significantly improved the efficiency of DSSCs due to their relatively high surface area,fast electron transport,and enhanced light-scattering capability.A short current density (Jsc) of 11.7 mA/cm2 and an overall solar-to-electric energy conversion efficiency (η) of 3.17% were achieved for the ZnO composite aggregate DSSCs,which were much higher than those obtained for the monodisperse aggregate DSSCs (Jsc=6.9mA/cm2,η=1.51 %) and ZnO nanorod array DSSCs (Jsc =4.2 mA/cm2,η=0.61%).

  7. Gadolinium-functionalized aggregation-induced emission dots as dual-modality probes for cancer metastasis study.

    Science.gov (United States)

    Li, Kai; Ding, Dan; Prashant, Chandrasekharan; Qin, Wei; Yang, Chang-Tong; Tang, Ben Zhong; Liu, Bin

    2013-12-01

    Understanding the localization and engraftment of tumor cells at postintravasation stage of metastasis is of high importance in cancer diagnosis and treatment. Advanced fluorescent probes and facile methodologies for cell tracing play a key role in metastasis studies. In this work, we design and synthesize a dual-modality imaging dots with both optical and magnetic contrast through integration of a magnetic resonance imaging reagent, gadolinium(III), into a novel long-term cell tracing probe with aggregation-induced emission (AIE) in far-red/near-infrared region. The obtained fluorescent-magnetic AIE dots have both high fluorescence quantum yield (25%) and T1 relaxivity (7.91 mM(-1) s(-1) ) in aqueous suspension. After further conjugation with a cell membrane penetrating peptide, the dual-modality dots can be efficiently internalized into living cells. The gadolinium(III) allows accurate quantification of biodistribution of cancer cells via intraveneous injection, while the high fluorescence provides engraftment information of cells at single cellular level. The dual-modality AIE dots show obvious synergistic advantages over either single imaging modality and hold great promises in advanced biomedical studies.

  8. In-cell aggregation of a polyglutamine-containing chimera is a multistep process initiated by the flanking sequence.

    Science.gov (United States)

    Ignatova, Zoya; Thakur, Ashwani K; Wetzel, Ronald; Gierasch, Lila M

    2007-12-14

    Toxicity in amyloid diseases is intimately linked to the nature of aggregates, with early oligomeric species believed to be more cytotoxic than later fibrillar aggregates. Yet mechanistic understanding of how aggregating species evolve with time is currently lacking. We have explored the aggregation process of a chimera composed of a globular protein (cellular retinoic acid-binding protein, CRABP) and huntingtin exon 1 with polyglutamine tracts either above (Q53) or below (Q20) the pathological threshold using Escherichia coli cells as a model intracellular environment. Previously we showed that fusion of the huntingtin exon 1 sequence with >40Q led to structural perturbation and decreased stability of CRABP (Ignatova, Z., and Gierasch, L. M. (2006) J. Biol. Chem. 281, 12959-12967). Here we report that the Q53 chimera aggregates in cells via a multistep process: early stage aggregates are spherical and detergent-soluble, characteristics of prefibrillar aggregates, and appear to be dominated structurally by CRABP, in that they can promote aggregation of a CRABP variant but not oligoglutamine aggregation, and the CRABP domain is relatively sequestered based on its protection from proteolysis. Late stage aggregates appear to be dominated by polyGln; they are fibrillar, detergent-resistant, capable of seeding aggregation of oligoglutamine but not the CRABP variant, and show relative protection of the polyglutamine-exon1 domain from proteolysis. These results point to an evolution of the dominant sequences in intracellular aggregates and may provide molecular insight into origins of toxic prefibrillar aggregates.

  9. Analysis of red blood cell aggregation in cardio-pulmonary bypass (CPB) surgery.

    Science.gov (United States)

    Graaff, R; Gu, Y J; Boonstra, P W; van Oeveren, W; Rakhorst, G

    2004-06-01

    Not much is known about red cell aggregation during cardio-pulmonary bypass surgery (CPB). Blood samples from 19 patients undergoing CPB were anticoagulated with EDTA. Hematocrit was adjusted to 40%. A red blood cell aggregometer (LORCA) measured changes in light reflection from each blood sample after cessation of the rotation, and calculated an aggregation index (AI). Reflection measurements were stored. Because LORCA software failed for 87 of 171 samples, we developed new software, and applied it to the stored reflection measurements. This software failed only in 7 out of 171 cases and showed that all LORCA failures occurred for AI CPB and recovered to 37.1 +/- 13.5 at day 1. It is concluded that the new software can be used to study decreased red cell aggregation during CPB.

  10. A Voronoi Interface approach to cell aggregate electropermeabilization

    Science.gov (United States)

    Guittet, Arthur; Poignard, Clair; Gibou, Frederic

    2017-03-01

    We present a Voronoi Interface approach to the study of cell electropermeabilization. We consider the nonlinear electropermeabilization model of Poignard et al. [20], which takes into account the jump in the voltage potential across cells' membrane. The jump condition is imposed in a sharp manner, using the Voronoi Interface Method of Guittet et al. [14], while adaptive Quad/Oc-tree grids are employed to automatically refine near the cells boundary for increased accuracy. Numerical results are provided to illustrate the accuracy of the methods. We also carry out simulations in three spatial dimensions to investigate the influence of shadowing and of the cells shape on the degree of permeabilization.

  11. White-Light Electroluminescence with Tetraphenylethylene as Emitting Layer of Aggregation-Induced Emissions Enhancement

    Institute of Scientific and Technical Information of China (English)

    罗建芳; 王晓宏; 王筱梅; 苏文明; 陶绪堂; 陈志刚

    2012-01-01

    Tetraphenylethylene (TPE) based molecules with easy synthesis, good thermal stability, and especially their aggregation-induced emissions enhancement (AIEE) effect recently become attractive organic emitting materials due to their potentially practical application in OLEDs. Herein, the AIEE behaviors of tetraphenylethylene dyes (TMTPE and TBTPE) were investigated. Fabricated luminesent device using TMTPE dye as emitting layer displays two strong emitting bands: the blue emission coming from the first-step aggregation and the yellow emission attrib- uted to the second-step aggregation. Thus, it can be utilized to fabricate the white-light OLEDs (WOLEDs) of the single-emitting-component. A three-layer device with the brightness of 1200 cd·m^-2 and current efficiency of 0.78 cd·A^-1 emits the close to white light with the CIE coordinates of x=0.333 and y=0.358, when applied voltage from 8-13 V, verifying that the TPE-based dyes of AIEE effect can be effectively applied in single-emitting- component WOLEDs fabrication.

  12. CAG Expansions Are Genetically Stable and Form Nontoxic Aggregates in Cells Lacking Endogenous Polyglutamine Proteins

    Directory of Open Access Journals (Sweden)

    Ashley A. Zurawel

    2016-09-01

    Full Text Available Proteins containing polyglutamine (polyQ regions are found in almost all eukaryotes, albeit with various frequencies. In humans, proteins such as huntingtin (Htt with abnormally expanded polyQ regions cause neurodegenerative diseases such as Huntington’s disease (HD. To study how the presence of endogenous polyQ aggregation modulates polyQ aggregation and toxicity, we expressed polyQ expanded Htt fragments (polyQ Htt in Schizosaccharomyces pombe. In stark contrast to other unicellular fungi, such as Saccharomyces cerevisiae, S. pombe is uniquely devoid of proteins with more than 10 Q repeats. We found that polyQ Htt forms aggregates within S. pombe cells only with exceedingly long polyQ expansions. Surprisingly, despite the presence of polyQ Htt aggregates in both the cytoplasm and nucleus, no significant growth defect was observed in S. pombe cells. Further, PCR analysis showed that the repetitive polyQ-encoding DNA region remained constant following transformation and after multiple divisions in S. pombe, in contrast to the genetic instability of polyQ DNA sequences in other organisms. These results demonstrate that cells with a low content of polyQ or other aggregation-prone proteins can show a striking resilience with respect to polyQ toxicity and that genetic instability of repetitive DNA sequences may have played an important role in the evolutionary emergence and exclusion of polyQ expansion proteins in different organisms.

  13. Photoacoustic ultrasound spectroscopy for assessing red blood cell aggregation and oxygenation

    Science.gov (United States)

    Hysi, Eno; Saha, Ratan K.; Kolios, Michael C.

    2012-12-01

    Red blood cell (RBC) aggregation and oxygenation are important markers for a variety of blood disorders. No current technique is capable of simultaneously measuring aggregation/oxygenation levels noninvasively. We propose using photoacoustic ultrasound spectroscopy (PAUS) for assessing both phenomena. This technique relies on frequency-domain analysis of the PA signals by extracting parameters such as the ultrasound spectral slope and the midband fit. To investigate the effect of hematocrit, aggregation, and oxygenation levels on PAUS parameters, a Monte Carlo-based theoretical model and an experimental protocol using porcine RBCs were developed. The samples were illuminated at 750 and 1064 nm and changes in the PAUS parameters were compared to the oxygen-dependent optical absorption coefficients to assess the oxygenation level. Good agreement between the theoretical and experimental spectral parameters was obtained for the spectral slope of the nonaggregated spectra (˜0.3 dB/MHz). The experimental midband fit increased by ˜5 dB for the largest aggregate size. Based on the analysis of the PA signals, the oxygen saturation level of the most aggregated sample was >20% greater than the nonaggregated sample. The results provide a framework for using PA signals' spectroscopic parameters for monitoring the aggregation and oxygenation levels of RBCs.

  14. Measurement of interaction forces between red blood cells in aggregates by optical tweezers

    Energy Technology Data Exchange (ETDEWEB)

    Maklygin, A Yu; Priezzhev, A V; Karmenian, A; Nikitin, Sergei Yu; Obolenskii, I S; Lugovtsov, Andrei E; Kisun Li

    2012-06-30

    We have fabricated double-beam optical tweezers and demonstrated the possibility of their use for measuring the interaction forces between red blood cells (erythrocytes). It has been established experimentally that prolonged trapping of red blood cells in a tightly focused laser beam does not cause any visible changes in their shape or size. We have measured the interaction between red blood cells in the aggregate, deformed by optical tweezers.

  15. Reparative Dentinogenesis Induced by Mineral Trioxide Aggregate: A Review from the Biological and Physicochemical Points of View

    Directory of Open Access Journals (Sweden)

    Takashi Okiji

    2009-01-01

    Full Text Available This paper aims to review the biological and physicochemical properties of mineral trioxide aggregate (MTA with respect to its ability to induce reparative dentinogenesis, which involves complex cellular and molecular events leading to hard-tissue repair by newly differentiated odontoblast-like cells. Compared with that of calcium hydroxide-based materials, MTA is more efficient at inducing reparative dentinogenesis in vivo. The available literature suggests that the action of MTA is attributable to the natural wound healing process of exposed pulps, although MTA can stimulate hard-tissue-forming cells to induce matrix formation and mineralization in vitro. Physicochemical analyses have revealed that MTA not only acts as a “calcium hydroxide-releasing” material, but also interacts with phosphate-containing fluids to form apatite precipitates. MTA also shows better sealing ability and structural stability, but less potent antimicrobial activity compared with that of calcium hydroxide. The clinical outcome of direct pulp capping and pulpotomy with MTA appears quite favorable, although the number of controled prospective studies is still limited. Attempts are being conducted to improve the properties of MTA by the addition of setting accelerators and the development of new calcium silicate-based materials.

  16. The Evolution of Aggregative Multicellularity and Cell-Cell Communication in the Dictyostelia.

    Science.gov (United States)

    Du, Qingyou; Kawabe, Yoshinori; Schilde, Christina; Chen, Zhi-Hui; Schaap, Pauline

    2015-11-20

    Aggregative multicellularity, resulting in formation of a spore-bearing fruiting body, evolved at least six times independently amongst both eukaryotes and prokaryotes. Amongst eukaryotes, this form of multicellularity is mainly studied in the social amoeba Dictyostelium discoideum. In this review, we summarise trends in the evolution of cell-type specialisation and behavioural complexity in the four major groups of Dictyostelia. We describe the cell-cell communication systems that control the developmental programme of D. discoideum, highlighting the central role of cAMP in the regulation of cell movement and cell differentiation. Comparative genomic studies showed that the proteins involved in cAMP signalling are deeply conserved across Dictyostelia and their unicellular amoebozoan ancestors. Comparative functional analysis revealed that cAMP signalling in D. discoideum originated from a second messenger role in amoebozoan encystation. We highlight some molecular changes in cAMP signalling genes that were responsible for the novel roles of cAMP in multicellular development.

  17. Effect of molecular aggregation on the photo-induced anisotropy in amorphous polymethacrylate bearing an aminonitroazobenzene moiety

    CERN Document Server

    Kim, B J; Choi, D H

    2001-01-01

    We investigated H-type molecular aggregation in a simply spin-coated amorphous homopolymer film of polymethacrylate containing push-pull azobenzene moieties. It was found that the aggregate formation was strongly influenced by thermal treatment and that the aggregate created in the polymer film could be easily disrupted by irradiation of a linearly polarized light. In the first writing cycle of aggregated polymer film, photo-induced birefringence showed a steep increase to the highest value followed by a gradual decrease to the certain asymptotic value under longer irradiation of linearly polarized light. This unique behavior could be attributed to the cooperative motion and the disruption of the aggregated molecules under continuous irradiation of light.

  18. Bacillus thuringiensis Cyt2Aa2 toxin disrupts cell membranes by forming large protein aggregates

    Science.gov (United States)

    Tharad, Sudarat; Toca-Herrera, José L.; Promdonkoy, Boonhiang; Krittanai, Chartchai

    2016-01-01

    Bacillus thuringiensis (Bt) Cyt2Aa2 showed toxicity against Dipteran insect larvae and in vitro lysis activity on several cells. It has potential applications in the biological control of insect larvae. Although pore-forming and/or detergent-like mechanisms were proposed, the mechanism underlying cytolytic activity remains unclear. Analysis of the haemolytic activity of Cyt2Aa2 with osmotic stabilizers revealed partial toxin inhibition, suggesting a distinctive mechanism from the putative pore formation model. Membrane permeability was studied using fluorescent dye entrapped in large unilamellar vesicles (LUVs) at various protein/lipid molar ratios. Binding of Cyt2Aa2 monomer to the lipid membrane did not disturb membrane integrity until the critical protein/lipid molar ratio was reached, when Cyt2Aa2 complexes and cytolytic activity were detected. The complexes are large aggregates that appeared as a ladder when separated by agarose gel electrophoresis. Interaction of Cyt2Aa2 with Aedes albopictus cells was investigated by confocal microscopy and total internal reflection fluorescent microscopy (TIRF). The results showed that Cyt2Aa2 binds on the cell membrane at an early stage without cell membrane disruption. Protein aggregation on the cell membrane was detected later which coincided with cell swelling. Cyt2Aa2 aggregations on supported lipid bilayers (SLBs) were visualized by AFM. The AFM topographic images revealed Cyt2Aa2 aggregates on the lipid bilayer at low protein concentration and subsequently disrupts the lipid bilayer by forming a lesion as the protein concentration increased. These results supported the mechanism whereby Cyt2Aa2 binds and aggregates on the lipid membrane leading to the formation of non-specific hole and disruption of the cell membrane. PMID:27612497

  19. Promotion on Nucleation and Aggregation of Calcium Oxalate Crystals by Injured African Green Monkey Renal Epithelial Cells

    Institute of Scientific and Technical Information of China (English)

    张燊; 彭花; 姚秀琼; 苏泽轩; 欧阳健明

    2012-01-01

    The purpose of this work was to detect the properties of African green monkey renal epithelial cells (Vero) after oxidative injury and to study the mediation of the injured Vero on aggregation and formation of calcium oxalate crystals. This injury model was induced by 0.15 mmol/L H2O2 according to the pretest evaluation. The results suggested that H2O2 could injure Vero significantly and decrease cell viability in a time-dependent manner for exposure time of 0.5--2 h. After cell injury, the indexes connected with oxidative injury changed. The malondialdehyde (MDA) content and osteopontin (OPN) expression increased, while superoxide dismutase (SOD) level decreased. It resulted in the increase of both the amount of CaOxa crystals and the degree of crystal aggregation on the injured cells. This work indicated that injured cells promoted the formation of calcium oxalate monohydrate (COM) crystals, thus increased the risk of formation of urinary stone.

  20. Quantification of depletion-induced adhesion of Red Blood Cells

    CERN Document Server

    Steffen, Patrick; Wagner, Christian

    2012-01-01

    Red blood cells (RBC) are known to form aggregates in the forms of rouleaux due to the presence of plasma proteins under physiological conditions. Rouleaux formation can be also induced in vitro by the addition of macromolecules to the RBC solution. Current data on the adhesion strength between red blood cells in their natural discocyte shapes mostly rely on indirect measurements like flow chamber experiments, but on the single cell level data is lacking. Here we present measurements on the dextran induced aggregation of red blood cells by use of atomic force microscopy based single cell force spectroscopy (SCFS). The effects of dextran concentration and molecular weight on the interaction energy of adhering RBCs was determined. The results are in good agreement with a model based on the depletion effect and former experimental studies.

  1. Optimization and comparison of two different 3D culture methods to prepare cell aggregates as a bioink for organ printing.

    Science.gov (United States)

    Imani, Rana; Hojjati Emami, Shahriar; Fakhrzadeh, Hossein; Baheiraei, Nafiseh; Sharifi, Ali M

    2012-04-01

    The ultimate goal of tissue engineering is to design and fabricate functional human tissues that are similar to natural cells and are capable of regeneration. Preparation of cell aggregates is one of the important steps in 3D tissue engineering technology, particularly in organ printing. Two simple methods, hanging drop (HD) and conical tube (CT) were utilized to prepare cell aggregates. The size and viability of the aggregates obtained at different initial cell densities and pre-culture duration were compared. The proliferative ability of the cell aggregates and their ability to spread in culture plates were also investigated. In both methods, the optimum average size of the aggregates was less than 500 microm. CT aggregates were smaller than HD aggregates. 5,000 cells per drop HD aggregates showed a marked ability to attach and spread on the culture surface. The proliferative ability reduced when the initial cell density was increased. Comparing these methods, we found that the HD method having better size controlling ability as well as enhanced ability to maintain higher rates of viability, spreading, and proliferation. In conclusion, smaller HD aggregates might be a suitable choice as building blocks for making bioink particles in bioprinting technique.

  2. Synthesis and Properties of Gelators Derived from Tetraphenylethylene and Gallic Acid with Aggregation-Induced Emission

    Science.gov (United States)

    Luo, Miao; Zhou, Xie; Chi, Zhenguo; Ma, Chunping; Zhang, Yi; Liu, Siwei; Xu, Jiarui

    2013-09-01

    Two novel organogelators (TEG and TAG) based on tetraphenylethylene and 3,4,5-tris(dodecyloxy) benzoic acid were synthesized through ester bond and amido bond linkages, respectively. Compounds TEG and TAG were able to induce gelation in ethanol. Aggregation-induced enhanced emission was observed in these organogelator molecules, with increased fluorescence intensity from the solutions to the gels. The completely thermoreversible gelation occurred due to the aggregation of the organogelators. In the process, a fibrous network was formed by a combination of intermolecular hydrogen bonding, π-π stacking and van der Waals interactions. These phenomena were observed in the xerogels by field-emission scanning electron microscopy and Fourier-transform infrared spectroscopy. The results of differential scanning calorimetry and polarized optical microscopy indicated that compound TAG exhibited stable liquid crystalline phases over a wide temperature range. The linking groups have severe influence on the properties of the organogelators, which was mainly attributed to the hydrogen bonding interaction in compound TAG.

  3. Changes of conformation and aggregation state induced by binding of lanthanide ions to insulin

    Institute of Scientific and Technical Information of China (English)

    程驿; 李荣昌; 王夔

    2002-01-01

    To clarify the mechanism of lanthanide ions (Ln3+) on the across-membrane transport of insulin and subsequent reducing blood glucose, the interactions of Ln3+ with Zn-insulin and Zn-free insulin are investigated by spectroscopic methods. The results reveal that the binding of Ln3+ to insulin can induce its structure changes from secondary to quaternary structure, depending on the Ln3+ concentration. In the lower concentration, it triggers the conformational changes of insulin monomer in the binding region with insulin receptor (B(24-30)). It would affect insulin-insulin receptor interaction. Moreover, Ln3+ binding promotes the assembly of insulin monomer from dimer to polymer. The potency of Ln3+ in inducing insulin's aggregation is stronger than that of Zn2+. Furthermore, the aggregation can be reversed partly by EDTA-treatment, indicating that it is not due to denaturation. Similar to Zn2+ effect, Ln3+ can stabilize insulin hexamer in a certain range of concentration, but is stronger than the former.

  4. Suppression of Kasha's rule as a mechanism for fluorescent molecular rotors and aggregation-induced emission

    Science.gov (United States)

    Qian, Hai; Cousins, Morgan E.; Horak, Erik H.; Wakefield, Audrey; Liptak, Matthew D.; Aprahamian, Ivan

    2017-01-01

    Although there are some proposed explanations for aggregation-induced emission, a phenomenon with applications that range from biosensors to organic light-emitting diodes, current understanding of the quantum-mechanical origin of this photophysical behaviour is limited. To address this issue, we assessed the emission properties of a series of BF2-hydrazone-based dyes as a function of solvent viscosity. These molecules turned out to be highly efficient fluorescent molecular rotors. This property, in addition to them being aggregation-induced emission luminogens, enabled us to probe deeper into their emission mechanism. Time-dependent density functional theory calculations and experimental results showed that the emission is not from the S1 state, as predicted from Kasha's rule, but from a higher energy (>S1) state. Furthermore, we found that suppression of internal conversion to the dark S1 state by restricting the rotor rotation enhances fluorescence, which leads to the proposal that suppression of Kasha's rule is the photophysical mechanism responsible for emission in both viscous solution and the solid state.

  5. Changes of conformation and aggregation state induced by binding of lanthanide ions to insulin

    Institute of Scientific and Technical Information of China (English)

    程驿; 李荣昌; 王夔

    2002-01-01

    To clarify the mechanism of lanthanide ions (Ln3+) on the across-membrane transport of insulin and subsequent reducing blood glucose, the interactions of Ln3+with Zn-insulin and Zn-free insulin are investigated by spectroscopic methods. The results reveal that the binding of Ln3+ to insulin can induce its structure changes from secondary to quaternary structure, depending on the Ln3+ concentration. In the lower concentration, it triggers the conformational changes of insulin monomer in the binding region with insulin receptor (B(24-30)). It would affect insulin-insulin receptor interaction. Moreover, Ln3+ binding promotes the assembly of insulin monomer from dimer to polymer. The potency of Ln3+ in inducing insulin’s aggregation is stronger than that of Zn2+. Furthermore, the aggregation can be reversed partly by EDTA-treatment, indicating that it is not due to denaturation. Similar to Zn2+ effect, Ln3+ can stabilize insulin hexamer in a certain range of concentration, but is stronger than the former.

  6. Normal and system lupus erythematosus red blood cell interactions studied by double trap optical tweezers: direct measurements of aggregation forces

    Science.gov (United States)

    Khokhlova, Maria D.; Lyubin, Eugeny V.; Zhdanov, Alexander G.; Rykova, Sophia Yu.; Sokolova, Irina A.; Fedyanin, Andrey A.

    2012-02-01

    Direct measurements of aggregation forces in piconewton range between two red blood cells in pair rouleau are performed under physiological conditions using double trap optical tweezers. Aggregation and disaggregation properties of healthy and pathologic (system lupus erythematosis) blood samples are analyzed. Strong difference in aggregation speed and behavior is revealed using the offered method which is proposed to be a promising tool for SLE monitoring at single cell level.

  7. Elucidating double aggregation mechanisms in the morphology optimization of diketopyrrolopyrrole-based narrow bandgap polymer solar cells.

    Science.gov (United States)

    Gao, Jing; Chen, Wei; Dou, Letian; Chen, Chun-Chao; Chang, Wei-Hsuan; Liu, Yongsheng; Li, Gang; Yang, Yang

    2014-05-21

    The power conversion efficiency (PCE) of a DPP-based polymer solar cell is significantly improved by using DIO or DCB as processing additives. The discovery that DCB outperforms DIO with a significantly wider solvent mixture operation window suggests different optimization mechanisms. Although both solvent mixture systems involve double aggregation processes, including a similar solution-to-film aggregation, however, two distinct solution-stage aggregations are observed: relatively amorphous polymer aggregates form in the CF-DIO solution, while more crystalline polymer aggregates form in CF-DCB solution.

  8. Photometric measurements of red blood cell aggregation: light transmission versus light reflectance

    NARCIS (Netherlands)

    Baskurt, O.K.; Uyuklu, M.; Hardeman, M.R.; Meiselman, H.J.

    2009-01-01

    Red blood cell (RBC) aggregation is the reversible and regular clumping in the presence of certain macromolecules. This is a clinically important phenomenon, being significantly enhanced in the presence of acute phase reactants (e. g., fibrinogen). Both light reflection (LR) and light transmission (

  9. Alkali silica reaction in concrete induced by mortar adhered to recycled aggregate

    Directory of Open Access Journals (Sweden)

    Etxeberria, M.

    2010-02-01

    Full Text Available The durability of recycled concrete must be determined before this material can be used in construction. In this paper the alkali-silica reaction in recycled concrete is analyzed. The recycled concrete is made with recycled aggregates, composed by original limestone aggregates and adhered mortar with reactive silica sand, and high alkali content cement. Due to the manufacturing process used for concrete production and the high water absorption capacity of recycled aggregates, cement accumulation happens in the interface (ITZ. The concentration of alkalis on the surface of recycled aggregates- ITZ and the presence of reactive sand in the mortar adhering to the recycled aggregate induce an alkali-silica reaction in 6-month concrete. The existence of this reaction is confirmed by environmental scanning electron microscopy (ESEM and EDX analysis. The mechanical properties of 6-month recycled concrete were similar to those values at 28-days of curing.

    La durabilidad del hormigón fabricado con árido reciclado es necesario determinarla antes de su utilización como material de construcción. En este artículo se analiza la reacción álcali-sílice manifestada en el hormigón fabricado con árido reciclado procedente de hormigón (compuesto de árido original calizo y mortero adherido de arena sílice reactiva, y cemento de alto contenido en álcalis. Debido al proceso de fabricación del hormigón y la alta capacidad de absorción del árido reciclado se produce una acumulación del cemento en la Interfase (ITZ. Debido al contacto directo de los álcalis del cemento con la arena sílice reactiva se produce una reacción álcali sílice a los 6 meses de edad del hormigón. Se realiza un análisis mediante microscopio electrónico de barrido ambiental (ESEM y sistema analítico de EDX. Se determina que las propiedades mecánicas del hormigón reciclado a 6 meses son similares a las obtenidas a los 28 días de curado.

  10. The secondary and aggregation structural changes of BSA induced by trivalent chromium: A biophysical study

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Mingmao [Institute of Biomedical and Pharmaceutical Technology, Fuzhou University, Fuzhou 350002, Fujian (China); Liu, Yan, E-mail: liuyan@fjirsm.ac.cn [The State Key Lab of Structural Chemistry, Fujian Institute of Research on the Structure of Matter, Chinese Academy of Sciences, Fuzhou 350002, Fujian (China); Cao, Huan [Institute of Biomedical and Pharmaceutical Technology, Fuzhou University, Fuzhou 350002, Fujian (China); Song, Ling [The State Key Lab of Structural Chemistry, Fujian Institute of Research on the Structure of Matter, Chinese Academy of Sciences, Fuzhou 350002, Fujian (China); Zhang, Qiqing, E-mail: zhangqiq@126.com [Institute of Biomedical and Pharmaceutical Technology, Fuzhou University, Fuzhou 350002, Fujian (China)

    2015-02-15

    Trivalent chromium Cr(III), which was originally considered to be innocuous as a nutriment, has been suspected to induce some abnormalities in human body recently. In the present work, the effects of Cr(III) on the structural state of BSA were comprehensively investigated through a series of appropriate methods in combination, including X-ray photoelectron spectroscopy (XPS), fourier transform infrared spectroscopy (FTIR), circular dichroism (CD), UV–vis absorption, synchronous fluorescence, fluorescence lifetime analysis, resonance light scattering (RLS), dynamic light scattering (DLS) and excitation–emission matrix spectroscopy (EEMS) methods. XPS accurately described the binding activity of Cr(III) with protein C, N and O atoms. The structural analysis according to FTIR and CD methods showed that the Cr(III) binding altered BSA conformation with a major reduction of α-helix. RLS and DLS analyses demonstrated that the presence of Cr(III) with low concentration could induce the aggregation structural changes of BSA. UV–vis absorption, EEMS and synchronous fluorescence suggested that the interaction between Cr(III) and BSA induced a slight unfolding of the polypeptide backbone and altered the microenvironments of Trp and Tyr residues in BSA. This research is helpful for understanding the structure-function relationship involved in metal ion-protein bioconjugate process. - Highlights: • The effect of Cr(III) on the conformational state of BSA was comprehensively studied. • XPS described the binding activity of Cr(III) with protein C, N and O atoms. • FTIR and CD data revealed secondary structural alteration in BSA. • Cr(III) complexation induced microenvironmental changes of Trp and Tyr. • RLS, DLS and EEMS presented the aggregational states of Cr(III)–BSA complex.

  11. Interactions of Pluronic nanocarriers with 2D and 3D cell cultures: Effects of PEO block length and aggregation state.

    Science.gov (United States)

    Arranja, Alexandra; Denkova, Antonia G; Morawska, Karolina; Waton, Gilles; van Vlierberghe, Sandra; Dubruel, Peter; Schosseler, François; Mendes, Eduardo

    2016-02-28

    This work reveals how the physicochemical properties of Pluronic block copolymers influence significantly their interactions with cancer cells, whether in monolayer or spheroid cultures, and how different clinical applications can be foreseen. Two-dimensional (2D) and three-dimensional (3D) cell culture models were used to investigate the interactions of Pluronic carriers with different PEO block length and aggregation state (unimers versus cross-linked micelles) in HeLa and U87 cancer cells. Stabilized micelles of Pluronic P94 or F127 were obtained by polymerization of a crosslinking agent in the micelles hydrophobic core. Nanocarriers were functionalized with a fluorescent probe for visualization, and with a chelator for radiolabeling with Indium-111 and gamma-quantification. The 2D cell models revealed that the internalization pathways and ultimate cellular localization of the Pluronic nanocarriers depended largely on both the PEO block size and aggregation state of the copolymers. The smaller P94 unimers with an average radius of 2.1nm and the shortest PEO block mass (1100gmol(-1)) displayed the highest cellular uptake and retention. 3D tumor spheroids were used to assess the penetration capacity and toxicity potential of the nanocarriers. Results showed that cross-linked F127 micelles were more efficiently delivered across the tumor spheroids, and the penetration depth depends mostly on the transcellular transport of the carriers. The Pluronic P94-based carriers with the shortest PEO block length induced spheroid toxicity, which was significantly influenced by the spheroid cellular type.

  12. Characterization at the individual cell level and in whole blood samples of shear stress preventing red blood cells aggregation.

    Science.gov (United States)

    Lee, K; Kinnunen, M; Danilina, A V; Ustinov, V D; Shin, S; Meglinski, I; Priezzhev, A V

    2016-05-03

    The aggregation of red blood cells (RBC) is an intrinsic feature of blood that has a strong impact on its microcirculation. For a number of years it has been attracting a great attention in basic research and clinical studies. Here, we study a relationship between the RBC aggregation parameters measured at the individual cell level and in a whole blood sample. The home made optical tweezers were used to measure the aggregating and disaggregating forces for a pair of interacting RBCs, at the individual cell level, in order to evaluate the corresponding shear stresses. The RheoScan aggregometer was used for the measurements of critical shear stress (CSS) in whole blood samples. The correlation between CSS and the shear stress required to stop an RBC pair from aggregating was found. The shear stress required to disaggregate a pair of RBCs using the double channel optical tweezers appeared to be about 10 times higher than CSS. The correlation between shear stresses required to prevent RBCs from aggregation at the individual cell level and in whole blood samples was estimated and assessed quantitatively. The experimental approach developed has a high potential for advancing hemorheological studies.

  13. Role of pH-induced structural change in protein aggregation in foam fractionation of bovine serum albumin

    Directory of Open Access Journals (Sweden)

    Rui Li

    2016-03-01

    Full Text Available For reducing protein aggregation in foam fractionation, the role of pH-induced structural change in the interface-induced protein aggregation was analyzed using bovine serum albumin (BSA as a model protein. The results show that the decrease in pH from 7.0 to 3.0 gradually unfolded the BSA structure to increase the molecular size and the relative content of β-sheet and thus reduced the stability of BSA in the aqueous solution. At the isoelectric point (pH 4.7, BSA suffered the lowest level in protein aggregation induced by the gas–liquid interface. In the pH range from 7.0 to 4.7, most BSA aggregates were formed in the defoaming process while in the pH range from 4.7 to 3.0, the BSA aggregates were formed at the gas–liquid interface due to the unfolded BSA structure and they further aggregated to form insoluble ones in the desorption process.

  14. Simple and efficient production of mice derived from embryonic stem cells aggregated with tetraploid embryos.

    Science.gov (United States)

    Li, Xiangyun; Yu, Yuansong; Wei, Wei; Yong, Jun; Yang, Jie; You, Jiefang; Xiong, Xiaoran; Qing, Tingting; Deng, Hongkui

    2005-06-01

    Six newly derived hybrid mouse embryonic stem (ES) cell lines and two inbred ES cell lines were tested for their ability to produce completely ES cell-derived mice by aggregation of ES cells with tetraploid embryos. Forty-five ES cell-tetraploid pups were generated from six hybrid ES cell lines and no pups from two inbred ES cell lines. These pups were found to have increased embryonic and placental weights than control mice. Twenty-two pups survived to adulthood and produced normal offsprings, and the other 23 pups died of several reasons including respiratory distress, abdomen ulcer-like symptoms, and foster failure. The 22 adult ES cell-tetraploid mice were completely ES cell-derived as judged by coat color and germline transmission, only two of them was found to have tetraploid component in liver, blood, and lung as analyzed by microsatellite loci. Our data suggested that genetic heterozygosity is a crucial factor for postnatal survival of ES cell-tetraploid mice, and tetraploid embryo aggregation using hybrid ES cells is a simple and efficient procedure for immediate generation of targeted mouse mutants from genetically modified ES cell clones, in contrast to the standard protocol, which involves the production of chimeras and several breeding steps.

  15. Cationic Conjugated Polymers-Induced Quorum Sensing of Bacteria Cells.

    Science.gov (United States)

    Zhang, Pengbo; Lu, Huan; Chen, Hui; Zhang, Jiangyan; Liu, Libing; Lv, Fengting; Wang, Shu

    2016-03-15

    Bacteria quorum sensing (QS) has attracted significant interest for understanding cell-cell communication and regulating biological functions. In this work, we demonstrate that water-soluble cationic conjugated polymers (PFP-G2) can interact with bacteria to form aggregates through electrostatic interactions. With bacteria coated in the aggregate, PFP-G2 can induce the bacteria QS system and prolong the time duration of QS signal molecules (autoinducer-2 (AI-2)) production. The prolonged AI-2 can bind with specific protein and continuously regulate downstream gene expression. Consequently, the bacteria show a higher survival rate against antibiotics, resulting in decreased antimicrobial susceptibility. Also, AI-2 induced by PFP-G2 can stimulate 55.54 ± 12.03% more biofilm in E. coli. This method can be used to understand cell-cell communication and regulate biological functions, such as the production of signaling molecules, antibiotics, other microbial metabolites, and even virulence.

  16. Interplay between desolvation and secondary structure in mediating cosolvent and temperature induced alpha-synuclein aggregation

    Science.gov (United States)

    Anderson, V. L.; Webb, W. W.; Eliezer, D.

    2012-10-01

    Both increased temperature and moderate concentrations of fluorinated alcohols enhance aggregation of the Parkinson's disease-associated protein α-synuclein (αS). Here, we investigate the secondary structural rearrangements induced by heating and trifluoroethanol [TFE]. At low TFE concentrations, CD spectra feature a negative peak characteristic of disordered polypeptides near 200 nm and a slight shoulder around 220 nm suggesting some polyproline-II content. Upon heating, these peaks weaken, while a weak negative signal develops at 222 nm. At high TFE concentrations, the spectra show distinct minima at 208 and 222 nm, indicative of considerable α-helical structure, which diminish upon heating. We observe a crossover between the low-TFE and high-TFE behavior near 15% TFE, where we previously showed that a partially helical intermediate is populated. We postulate that the protein is well solvated by water at low TFE concentrations and by TFE at high TFE concentrations, but may become desolvated at the crossover point. We discuss the potential roles and interplay of desolvation and helical secondary structure in driving αS aggregation.

  17. Aggregation-induced emissive nanoparticles for fluorescence signaling in a low cost paper-based immunoassay.

    Science.gov (United States)

    Engels, Jan F; Roose, Jesse; Zhai, Demi Shuang; Yip, Ka Man; Lee, Mei Suet; Tang, Ben Zhong; Renneberg, Reinhard

    2016-07-01

    Low cost paper based immunoassays are receiving interest due to their fast performance and small amounts of biomolecules needed for developing an immunoassay complex. In this work aggregation-induced emissive (AIE) nanoparticles, obtained from a diastereoisomeric mixture of 1,2-di-(4-hydroxyphenyl)-1,2-diphenylethene (TPEDH) in a one-step top-down method, are characterized through Dynamic Light Scattering (DLS), Scanning Electron Microscopy (SEM), and Zeta potential. By measuring the Zeta potential before and after labeling the nanoparticles with antibodies we demonstrate that the colloidal system is stable in a wide pH-range. The AIE-active nanoparticles are deposited on chitosan and glutaraldehyde modified paper pads overcoming the common aggregation-caused quenching (ACQ) effect. Analyte concentrations from 1000ng and below are applied in a model immunocomplex using Goat anti-Rabbit IgG and Rabbit IgG. In the range of 7.81ng-250ng, linear trends with a high R(2) are observed, which leads to a strong increase of the blue fluorescence from the TPEDH nanoparticles.

  18. Cell Surface Binding and Internalization of Aβ Modulated by Degree of Aggregation

    Directory of Open Access Journals (Sweden)

    David A. Bateman

    2011-01-01

    Full Text Available The amyloid peptides, Aβ40 and Aβ42, are generated through endoproteolytic cleavage of the amyloid precursor protein. Here we have developed a model to investigate the interaction of living cells with various forms of aggregated Aβ40/42. After incubation at endosomal pH 6, we observed a variety of Aβ conformations after 3 (Aβ3, 24 (Aβ24, and 90 hours (Aβ90. Both Aβ4224 and Aβ4024 were observed to rapidly bind and internalize into differentiated PC12 cells, leading to accumulation in the lysosome. In contrast, Aβ40/4290 were both found to only weakly associate with cells, but were observed as the most aggregated using dynamic light scattering and thioflavin-T. Internalization of Aβ40/4224 was inhibited with treatment of monodansylcadaverine, an endocytosis inhibitor. These studies indicate that the ability of Aβ40/42 to bind and internalize into living cells increases with degree of aggregation until it reaches a maximum beyond which its ability to interact with cells diminishes drastically.

  19. The influence of the crystal structure on aggregation-induced luminescence of derivatives of aminobenzoic acid

    Science.gov (United States)

    Nosova, D. A.; Zarochentseva, E. P.; Vysotskaya, S. O.; Klemesheva, N. A.; Korotkov, V. I.

    2014-12-01

    The luminescence of three derivatives of 2-(phenylamino)-benzoic acid (N-phenylanthranilic, mefenamic, and niflumic acids) in benzene solution, in the polycrystalline state, and in the hexamethylbenzene matrix is studied. In the crystalline state, these compounds exhibit intense aggregation-induced luminescence. An increase in luminescence is also observed in the impurity crystal. The hexamethylbenzene crystal lattice restricts the mobility of molecules, thus ensuring the rigidity of the molecular structure of acids, which decreases the efficiency of nonradiative electron energy degradation. The main reason for the increase in the luminescence intensity in the case of fixation in a crystalline matrix is the formation of intramolecular hydrogen bonds and dimers of acid molecules.

  20. A Robust Damage-Reporting Strategy for Polymeric Materials Enabled by Aggregation-Induced Emission.

    Science.gov (United States)

    Robb, Maxwell J; Li, Wenle; Gergely, Ryan C R; Matthews, Christopher C; White, Scott R; Sottos, Nancy R; Moore, Jeffrey S

    2016-09-28

    Microscopic damage inevitably leads to failure in polymers and composite materials, but it is difficult to detect without the aid of specialized equipment. The ability to enhance the detection of small-scale damage prior to catastrophic material failure is important for improving the safety and reliability of critical engineering components, while simultaneously reducing life cycle costs associated with regular maintenance and inspection. Here, we demonstrate a simple, robust, and sensitive fluorescence-based approach for autonomous detection of damage in polymeric materials and composites enabled by aggregation-induced emission (AIE). This simple, yet powerful system relies on a single active component, and the general mechanism delivers outstanding performance in a wide variety of materials with diverse chemical and mechanical properties.

  1. Polyamines induce aggregation of LHC II and quenching of fluorescence in vitro.

    Science.gov (United States)

    Tsiavos, Theodoros; Ioannidis, Nikolaos E; Kotzabasis, Kiriakos

    2012-05-01

    Dissipation of excess excitation energy within the light-harvesting complex of Photosystem II (LHC II) is a main process in plants, which is measured as the non-photochemical quenching of chlorophyll fluorescence or qE. We showed in previous works that polyamines stimulate qE in higher plants in vivo and in eukaryotic algae in vitro. In the present contribution we have tested whether polyamines can stimulate quenching in trimeric LHC II and monomeric light-harvesting complex b proteins from higher plants. The tetramine spermine was the most potent quencher and induced aggregation of LHC II trimers, due to its highly cationic character. Two transients are evident at 100 μM and 350 μM for the fluorescence and absorbance signals of LHC II respectively. On the basis of observations within this work, some links between polyamines and the activation of qE in vivo is discussed.

  2. Tel, a Frequent Target of Leukemic Translocations, Induces Cellular Aggregation and Influences Expression of Extracellular Matrix Components

    Directory of Open Access Journals (Sweden)

    L. Van Rompaey

    1999-12-01

    Full Text Available Tel is an Ets transcription factor that is the target of chromosome translocations in lymphoid and myeloid leukemias and in solid tumors. It contains two functional domains, a pointed oligomerization domain and a DNAbinding domain. Retroviral transduction of a wild-type Tel cDNA into a clonal subline of NIH3T3 fibroblasts resulted in a striking morphologic change: at confluency, the cells reorganized into a specific “bridge-like” pattern over the entire surface of the culture dish, started migrating, thereby leaving circular holes in the monolayer. Thereafter, formation of cellular cords became apparent. This sequence of events was inhibited by coating the culture dishes with fibronectin and collagen IV. Retroviral transduction of Tel into MS1 endothelial cells reproduced the aggregation phenotype, but not the cellular cord formation. Tel -mutagenesis showed that both the pointed domain and the DNAbinding domain of Tel are required for the morphologic change. Other Ets family genes, Fli-1 and Ets-1 that are both endogenously expressed in endothelial cells, could not induce this morphologic change. Exogenous Tel expression is associated with transcriptional upregulation of entactin/nidogen, Smad5, Col3a1, CD44 and fibronectin, downregulation of Coliai and secretory leukocyte protease inhibitor. Interestingly, Tel, Smad5, fibronectin, Coliai and Col3a1 all have essential roles during vascular development.

  3. A novel fusion protein domain III-capsid from dengue-2, in a highly aggregated form, induces a functional immune response and protection in mice.

    Science.gov (United States)

    Valdés, Iris; Bernardo, Lidice; Gil, Lázaro; Pavón, Alekis; Lazo, Laura; López, Carlos; Romero, Yaremis; Menendez, Ivón; Falcón, Viviana; Betancourt, Lázaro; Martín, Jorge; Chinea, Glay; Silva, Ricardo; Guzmán, María G; Guillén, Gerardo; Hermida, Lisset

    2009-11-25

    Based on the immunogenicity of domain III from the Envelope protein of dengue virus as well as the proven protective capacity of the capsid antigen, we have designed a novel domain III-capsid chimeric protein with the goal of obtaining a molecule potentially able to induce both humoral and cell-mediated immunity (CMI). After expression of the recombinant gene in Escherichia coli, the domain III moiety retained its antigenicity as evaluated with anti-dengue sera. In order to explore alternatives for modulating the immunogenicity of the protein, it was mixed with oligodeoxynucleotides in order to obtain particulated aggregates and then immunologically evaluated in mice in comparison with non-aggregated controls. Although the humoral immune response induced by both forms of the protein was equivalent, the aggregated variant resulted in a much stronger CMI as measured by in vitro IFN-gamma secretion and protection experiments, mediated by CD4(+) and CD8(+) cells. The present work provides additional evidence in support for a crucial role of CMI in protection against dengue virus and describes a novel vaccine candidate against the disease based on a recombinant protein that can stimulate both arms of the acquired immune system.

  4. Bioengineering of injectable encapsulated aggregates of pluripotent stem cells for therapy of myocardial infarction

    Science.gov (United States)

    Zhao, Shuting; Xu, Zhaobin; Wang, Hai; Reese, Benjamin E.; Gushchina, Liubov V.; Jiang, Meng; Agarwal, Pranay; Xu, Jiangsheng; Zhang, Mingjun; Shen, Rulong; Liu, Zhenguo; Weisleder, Noah; He, Xiaoming

    2016-10-01

    It is difficult to achieve minimally invasive injectable cell delivery while maintaining high cell retention and animal survival for in vivo stem cell therapy of myocardial infarction. Here we show that pluripotent stem cell aggregates pre-differentiated into the early cardiac lineage and encapsulated in a biocompatible and biodegradable micromatrix, are suitable for injectable delivery. This method significantly improves the survival of the injected cells by more than six-fold compared with the conventional practice of injecting single cells, and effectively prevents teratoma formation. Moreover, this method significantly enhances cardiac function and survival of animals after myocardial infarction, as a result of a localized immunosuppression effect of the micromatrix and the in situ cardiac regeneration by the injected cells.

  5. Passage number affects the pluripotency of mouse embryonic stem cells as judged by tetraploid embryo aggregation.

    Science.gov (United States)

    Li, Xiang-Yun; Jia, Qing; Di, Ke-Qian; Gao, Shu-Min; Wen, Xiao-Hui; Zhou, Rong-Yan; Wei, Wei; Wang, Li-Ze

    2007-03-01

    The aim of this study was to determine whether the number of passages affected the developmental pluripotency of embryonic stem (ES) cells as measured by the attainment of adult fertile mice derived from embryonic stem (ES) cell/tetraploid embryo complementation. Thirty-six newborns were produced by the aggregation of tetraploid embryos and hybrid ES cells after various numbers of passages. These newborns were entirely derived from ES cells as judged by microsatellite DNA, coat-color phenotype, and germline transmission. Although 15 survived to adulthood, 17 died of respiratory failure, and four were eaten by their foster mother. From the 15 mice that reached adulthood and that could reproduce, none arose from ES cells at passage level 15 or more. All 15 arose from cells at passages 3-11. Our results demonstrate that the number of passages affects the developmental pluripotency of ES cells.

  6. Generation of chimeras by aggregation of embryonic stem cells with diploid or tetraploid mouse embryos.

    Science.gov (United States)

    Artus, Jérôme; Hadjantonakis, Anna-Katerina

    2011-01-01

    From the hybrid creatures of the Greek and Egyptian mythologies, the concept of the chimera has evolved and, in modern day biology, refers to an organism comprises of at least two populations of genetically distinct cells. Mouse chimeras have proven an invaluable tool for the generation of genetically modified strains. In addition, chimeras have been extensively used in developmental biology as a powerful tool to analyze the phenotype of specific mutations, to attribute function to gene products and to address the question of cell autonomy versus noncell autonomy of gene function. This chapter describes a simple and economical technique used to generate mouse chimeras by embryo aggregation. Multiple aggregation combinations are described each of which can be tailored to answer particular biological questions.

  7. Uplink Inter-Site Carrier Aggregation Between Macro and Small Cells in Heterogeneous Networks

    DEFF Research Database (Denmark)

    Wang, Hua; Rosa, Claudio; Pedersen, Klaus I.

    2014-01-01

    With uplink inter-site carrier aggregation (CA), it is possible to configure a user equipment (UE) to transmit on multiple layers (macro and small cells) simultaneously, each of which may exhibit different radio channel characteristics. This introduces new challenging issues such as how to config......With uplink inter-site carrier aggregation (CA), it is possible to configure a user equipment (UE) to transmit on multiple layers (macro and small cells) simultaneously, each of which may exhibit different radio channel characteristics. This introduces new challenging issues such as how...... deciding whether UEs should be configured with uplink inter-site CA or not. Simulation results show that with proper configuration of UEs to operate with uplink inter-site CA, good user throughput gain compared to the case without inter-site CA can be achieved at low load due to larger bandwidth...

  8. Phenotypic switch in blood: effects of pro-inflammatory cytokines on breast cancer cell aggregation and adhesion.

    Directory of Open Access Journals (Sweden)

    Yue Geng

    Full Text Available Hematogeneous metastasis can occur via a cascade of circulating tumor cell adhesion events to the endothelial lining of the vasculature, i.e. the metastatic cascade. Interestingly, the pro-inflammatory cytokines IL-6 and TNF-α, which play an important role in potentiating the inflammatory cascade, are significantly elevated in metastatic breast cancer (BCa patients. Despite their high metastatic potential, human breast carcinoma cells MDA-MB-231 lack interactions with E-selectin functionalized surfaces under physiological shear stresses. We hypothesized that human plasma, 3-D tumor spheroid culture, and cytokine-supplemented culture media could induce a phenotypic switch that allows BCa cells to interact with E-selectin coated surfaces under physiological flow. Flow cytometry, immunofluorescence imaging, and flow-based cell adhesion assay were utilized to investigate the phenotypic changes of MDA-MB-231 cells with various treatments. Our results indicate that plasma, IL-6, and TNF-α promote breast cancer cell growth as aggregates and induce adhesive recruitment of BCa cells on E-selectin coated surfaces under flow. 3-D tumor spheroid culture exhibits the most significant increases in the interactions between BCa and E-selectin coated surfaces by upregulating CD44V4 and sLe(x expression. Furthermore, we show that IL-6 and TNF-α concentrations in blood may regulate the recruitment of BCa cells to the inflamed endothelium. Finally, we propose a mechanism that could explain the invasiveness of 'triple-negative' breast cancer cell line MDA-MB-231 via a positive feedback loop of IL-6 secretion and maintenance. Taken together, our results suggest that therapeutic approaches targeting cytokine receptors and adhesion molecules on cancer cells may potentially reduce metastatic load and improve current cancer treatments.

  9. Modulation of invasive phenotype by interstitial pressure-driven convection in aggregates of human breast cancer cells.

    Directory of Open Access Journals (Sweden)

    Joe Tien

    Full Text Available This paper reports the effect of elevated pressure on the invasive phenotype of patterned three-dimensional (3D aggregates of MDA-MB-231 human breast cancer cells. We found that the directionality of the interstitial pressure profile altered the frequency of invasion by cells located at the surface of an aggregate. In particular, application of pressure at one end of an aggregate suppressed invasion at the opposite end. Experimental alteration of the configuration of cell aggregates and computational modeling of the resulting flow and solute concentration profiles revealed that elevated pressure inhibited invasion by altering the chemical composition of the interstitial fluid near the surface of the aggregate. Our data reveal a link between hydrostatic pressure, interstitial convection, and invasion.

  10. Modeling multivalent ligand-receptor interactions with steric constraints on configurations of cell surface receptor aggregates

    Energy Technology Data Exchange (ETDEWEB)

    Monine, Michael [Los Alamos National Laboratory; Posner, Richard [TRANSLATION GENOMICS RESAEARCH INSTITUTE; Savage, Paul [BYU; Faeder, James [UNIV OF PITTSBURGH; Hlavacek, William S [UNM

    2008-01-01

    Signal transduction generally involves multivalent protein-protein interactions, which can produce various protein complexes and post-translational modifications. The reaction networks that characterize these interactions tend to be so large as to challenge conventional simulation procedures. To address this challenge, a kinetic Monte Carlo (KMC) method has been developed that can take advantage of a model specification in terms of reaction rules for molecular interactions. A set of rules implicitly defines the reactions that can occur as a result of the interactions represented by the rules. With the rule-based KMC method, explicit generation of the underlying chemical reaction network implied by rules is avoided. Here, we apply and extend this method to characterize the interactions of a trivalent ligand with a bivalent cell-surface receptor. This system is also studied experimentally. We consider the following kinetic models: an equivalent-site model, an extension of this model, which takes into account steric constraints on the configurations of receptor aggregates, and finally, a model that accounts for cyclic receptor aggregates. Simulation results for the equivalent-site model are consistent with an equilibrium continuum model. Using these models, we investigate the effects of steric constraints and the formation of cyclic aggregates on the kinetics and equilibria of small and large aggregate formation and the percolation phase transition that occurs in this system.

  11. Indole and synthetic derivative activate chaperone expression to reduce polyQ aggregation in SCA17 neuronal cell and slice culture models

    Directory of Open Access Journals (Sweden)

    Kung PJ

    2014-10-01

    Full Text Available Pin-Jui Kung,1,* Yu-Chen Tao,1,* Ho-Chiang Hsu,1 Wan-Ling Chen,1 Te-Hsien Lin,1 Donala Janreddy,2 Ching-Fa Yao,2 Kuo-Hsuan Chang,3 Jung-Yaw Lin,1 Ming-Tsan Su,1 Chung-Hsin Wu,1 Guey-Jen Lee-Chen,1 Hsiu-Mei Hsieh-Li1 1Department of Life Science, 2Department of Chemistry, National Taiwan Normal University, Taipei, Taiwan; 3Department of Neurology, Chang Gung Memorial Hospital, Chang Gung University College of Medicine, Taipei, Taiwan *These authors contributed equally to this work Abstract: In spinocerebellar ataxia type 17 (SCA17, the expansion of a translated CAG repeat in the TATA box binding protein (TBP gene results in a long polyglutamine (polyQ tract in the TBP protein, leading to intracellular accumulation of aggregated TBP and cell death. The molecular chaperones act in preventing protein aggregation to ameliorate downstream harmful events. In this study, we used Tet-On SH-SY5Y cells with inducible SCA17 TBP/Q79-green fluorescent protein (GFP expression to test indole and synthetic derivative NC001-8 for neuroprotection. We found that indole and NC001-8 up-regulated chaperone expression to reduce polyQ aggregation in neuronal differentiated TBP/Q79 cells. The effects on promoting neurite outgrowth and on reduction of aggregation on Purkinje cells were also confirmed with cerebellar primary and slice cultures of SCA17 transgenic mice. Our results demonstrate how indole and derivative NC001-8 reduce polyQ aggregation to support their therapeutic potentials in SCA17 treatment. Keywords: spinocerebellar ataxia type 17, TATA box binding protein, polyQ aggregation, indole and derivative, therapeutics

  12. Regulator of G protein signaling 20 enhances cancer cell aggregation, migration, invasion and adhesion.

    Science.gov (United States)

    Yang, Lei; Lee, Maggie M K; Leung, Manton M H; Wong, Yung H

    2016-11-01

    Several RGS (regulator of G protein signaling) proteins are known to be upregulated in a variety of tumors but their roles in modulating tumorigenesis remain undefined. Since the expression of RGS20 is elevated in metastatic melanoma and breast tumors, we examined the effects of RGS20 overexpression and knockdown on the cell mobility and adhesive properties of different human cancer cell lines, including cervical cancer HeLa, breast adenocarcinoma MDA-MB-231, and non-small cell lung carcinoma H1299 and A549 cells. Expression of RGS20 enhanced cell aggregation, migration, invasion and adhesion as determined by hanging drop aggregation, wound healing, transwell chamber migration and invasion assays. Conversely, shRNA-mediated knockdown of endogenous RGS20 impaired these responses. In addition, RGS20 elevated the expression of vimentin (a mesenchymal cell marker) but down-regulated the expression of E-cadherin, two indicators commonly associated with metastasis. These results suggest that the expression of RGS20 may promote metastasis of tumor cells.

  13. Optimising cell aggregate expansion in a perfused hollow fibre bioreactor via mathematical modelling.

    KAUST Repository

    Chapman, Lloyd A C

    2014-08-26

    The need for efficient and controlled expansion of cell populations is paramount in tissue engineering. Hollow fibre bioreactors (HFBs) have the potential to meet this need, but only with improved understanding of how operating conditions and cell seeding strategy affect cell proliferation in the bioreactor. This study is designed to assess the effects of two key operating parameters (the flow rate of culture medium into the fibre lumen and the fluid pressure imposed at the lumen outlet), together with the cell seeding distribution, on cell population growth in a single-fibre HFB. This is achieved using mathematical modelling and numerical methods to simulate the growth of cell aggregates along the outer surface of the fibre in response to the local oxygen concentration and fluid shear stress. The oxygen delivery to the cell aggregates and the fluid shear stress increase as the flow rate and pressure imposed at the lumen outlet are increased. Although the increased oxygen delivery promotes growth, the higher fluid shear stress can lead to cell death. For a given cell type and initial aggregate distribution, the operating parameters that give the most rapid overall growth can be identified from simulations. For example, when aggregates of rat cardiomyocytes that can tolerate shear stresses of up to 0:05 Pa are evenly distributed along the fibre, the inlet flow rate and outlet pressure that maximise the overall growth rate are predicted to be in the ranges 2.75 x 10(-5) m(2) s(-1) to 3 x 10(-5) m(2) s(-1) (equivalent to 2.07 ml min(-1) to 2.26 ml min(-1)) and 1.077 x 10(5) Pa to 1.083 x 10(5) Pa (or 15.6 psi to 15.7 psi) respectively. The combined effects of the seeding distribution and flow on the growth are also investigated and the optimal conditions for growth found to depend on the shear tolerance and oxygen demands of the cells.

  14. Cell surface alterations during blood-storage characterized by artificial aggregation of washed red blood cells.

    Science.gov (United States)

    Hessel, E; Lerche, D

    1985-01-01

    Aggregation measurement of washed human erythrocytes (RBC) were carried out in a NaCl-PBS solution under laminar shear conditions. Artificial aggregation of fresh and stored erythrocytes was caused by decreased pH and reduced ionic strength and characterized by collision efficiency alpha. Generally, the collision efficiency alpha of stored erythrocytes rises with the increased storage time. Such an aggregation technique might be useful to detect and quantify changes of the membrane and/or the surface structure due to aging and/or storage.

  15. Orally given gastroprotective capsaicin does not modify aspirin-induced platelet aggregation in healthy male volunteers (human phase I examination).

    Science.gov (United States)

    Sandor, B; Papp, J; Mozsik, Gy; Szolcsanyi, J; Keszthelyi, Zs; Juricskay, I; Toth, K; Habon, Tamas

    2014-12-01

    Capsaicin is a well-known component of red pepper. Recent studies have shown that capsaicin could prevent gastric ulcer provoked by various NSAID-s like acetylsalicylic acid (ASA). Primary objective of this human clinical phase I trial was to investigate whether two different doses of capsaicin co-administered with ASA could alter the inhibitory effect of ASA on platelet aggregation. 15 healthy male subjects were involved in the study and treated orally with 400 μg capsaicin, 800 μg capsaicin, 500 mg ASA, 400 μg capsaicin+500 mg ASA and 800 μg capsaicin+500 mg ASA. Blood was drawn before and 1, 2, 6 and 24 hours after the drug administration. After that epinephrine induced platelet aggregation was measured by optical aggregometry. Between treatments, volunteers had a 6-day wash-out period. Our results showed that capsaicin had no effect on platelet aggregation, while as expected, ASA monotherapy resulted in a significant and clinically effective platelet aggregation inhibition (p ≤ 0.001). The combined ASA-capsaicin therapies reached equivalent effectiveness in platelet aggregation inhibition as ASA monotherapy. Our investigation proved that capsaicin did not influence the inhibitory effect of ASA on platelet aggregation, thus the capsaicin-ASA treatment would combine the antiplatelet effect of ASA with the possible gastroprotection of capsaicin.

  16. Finite-size corrections to scaling behavior in sorted cell aggregates.

    Science.gov (United States)

    Klopper, A V; Krens, G; Grill, S W; Heisenberg, C-P

    2010-10-01

    Cell sorting is a widespread phenomenon pivotal to the early development of multicellular organisms. In vitro cell sorting studies have been instrumental in revealing the cellular properties driving this process. However, these studies have as yet been limited to two-dimensional analysis of three-dimensional cell sorting events. Here we describe a method to record the sorting of primary zebrafish ectoderm and mesoderm germ layer progenitor cells in three dimensions over time, and quantitatively analyze their sorting behavior using an order parameter related to heterotypic interface length. We investigate the cell population size dependence of sorted aggregates and find that the germ layer progenitor cells engulfed in the final configuration display a relationship between total interfacial length and system size according to a simple geometrical argument, subject to a finite-size effect.

  17. Liquid-Crystalline Star-Shaped Supergelator Exhibiting Aggregation-Induced Blue Light Emission.

    Science.gov (United States)

    Pathak, Suraj Kumar; Pradhan, Balaram; Gupta, Monika; Pal, Santanu Kumar; Sudhakar, Achalkumar Ammathnadu

    2016-09-13

    A family of closely related star-shaped stilbene-based molecules containing an amide linkage are synthesized, and their self-assembly in liquid-crystalline and gel states was investigated. The number and position of the peripheral alkyl tails were systematically varied to understand the structure-property relation. Interestingly, one of the molecules with seven peripheral chains was bimesomorphic, exhibiting columnar hexagonal and columnar rectangular phases, whereas the rest of them stabilized the room-temperature columnar hexagonal phase. The self-assembly of these molecules in liquid-crystalline and organogel states is extremely sensitive to the position and number of alkoxy tails in the periphery. Two of the compounds with six and seven peripheral tails exhibited supergelation behavior in long-chain hydrocarbon solvents. One of these compounds with seven alkyl chains was investigated further, and it has shown higher stability and moldability in the gel state. The xerogel of the same compound was characterized with the help of extensive microscopic and X-ray diffraction studies. The nanofibers in the xerogel are found to consist of molecules arranged in a lamellar fashion. Furthermore, this compound shows very weak emission in solution but an aggregation-induced emission property in the gel state. Considering the dearth of solid-state blue-light-emitting organic materials, this molecular design is promising where the self-assembly and emission in the aggregated state can be preserved. The nonsymmetric design lowers the phase-transition temperatures.The presence of an amide bond helps to stabilize columnar packing over a long range because of its polarity and intermolecular hydrogen bonding in addition to promoting organogelation.

  18. DNA packaging induced by micellar aggregates: a novel in vitro DNA condensation system.

    Science.gov (United States)

    Ghirlando, R; Wachtel, E J; Arad, T; Minsky, A

    1992-08-11

    Evidence for a conceptually novel DNA packaging process is presented. X-ray scattering, electron microscopy, and circular dichroism measurements indicate that in the presence of positively charged micellar aggregates and flexible anionic polymers, such as negatively charged polypeptides or single-stranded RNA species, a complex is formed in which DNA molecules are partially embedded within a micellar scaffold and partially condensed into highly packed chiral structures. Based on studies of micelle-DNA and micelle-flexible anionic polymer systems, as well as on the known effects of a high charge density upon the micellar organization, a DNA packaging model is proposed. According to this model, the DNA induces the elongation of the micelles into rodlike aggregates, forming a closely packed matrix in which the DNA molecules are immobilized. In contrast, the flexible anionic polymers stabilize clusters of spherical micelles which are proposed to effect a capping of the rodlike micelles, thus arresting their elongation and creating surfactant-free segments of the DNA that are able to converge and collapse. Thus, unlike other in vitro DNA packaging systems, in which condensation follows encounters between charge-neutralized DNA molecules, a prepackaging phase where the DNA is immobilized within a matrix is proposed in this case. Cellular and nuclear membranes have been implicated in DNA packaging processes in vivo, and negatively charged polyelectrolytes were shown to be involved in the processes. These observations, combined with the basic tenets of the DNA condensation system described here, allow for the progression to the study of more elaborate model systems and thus might lead to insights into the nature and roles of the intricate in vivo DNA-membrane complexes.

  19. Design and application of anthracene derivative with aggregation-induced emission charateristics for visualization and monitoring of erythropoietin unfolding.

    Science.gov (United States)

    Sun, Binjie; Yang, Xiaojun; Ma, Lin; Niu, Caixia; Wang, Fangfang; Na, Na; Wen, Jiying; Ouyang, Jin

    2013-02-12

    Erythropoietin (EPO) is an attractive protein-unfolding/folding model because of its high degree of unfolding and folding reversibility and intermediate size. Due to its function for regulating red blood cell production by stimulating late erythroid precursor cells, EPO presents obvious values to biological research. A nonemissive anthracene derivative, that is 9,10-bis[4-(3-sulfonatopropoxyl)-styryl]anthracene sodium salt (BSPSA), with aggregation-induced emission (AIE) charateristics shows a novel phenomenon of AIE when EPO is added. The AIE biosensor for EPO shows the limit of detection is 1 × 10(-9) M. Utilizing the AIE feature of BSPSA, the unfolding process of EPO using guanidine hydrochloride is monitored, which indicates three steps for the folding structures of EPO to transform to random coil. Computational modeling suggests that the BSPSA luminogens prefer docking in the hydrophobic cavity in the EPO folding structures, and the assembly of BSPSA in this cavity makes the AIE available, making the monitoring of unfolding of EPO possible.

  20. Temperature induced structural transitions from native to unfolded aggregated states of tobacco etch virus protease

    Science.gov (United States)

    Zhu, Guo-Fei; Ren, Si-Yan; Xi, Lei; Du, Lin-Fang; Zhu, Xiao-Feng

    2015-02-01

    Tobacco etch virus protease (TEVp) is widely used to remove fusion tags from recombinant proteins because of its high and unique specificity. This work describes the conformational and the thermodynamic properties in the unfolding/refolding process of TEVp3M (three-point mutant: L56V/S135G/S219V) induced by temperature. With temperature increasing from 20 to 100 °C, the CD spectra showed a transition trend from α-helix to β-sheet, and the fluorescence emission, synchronous fluorescence, ANS and RLS spectroscopy consistently revealed that the temperature-induced unfolding process behaved in a three-state manner, for there was a relatively stable intermediate state observed around 50 °C. The reversibility of thermal unfolding of TEVp3M further showed that the transition from the native to the intermediate state was reversible (below 50 °C), however the transition from the intermediate to the unfolded state was irreversible (above 60 °C). Moreover, aggregates were observed above 60 °C as revealed by SDS-PAGE, Thioflavin-T fluorescence and Congo red absorbance.

  1. Aggregation of human recombinant monoclonal antibodies influences the capacity of dendritic cells to stimulate adaptive T-cell responses in vitro.

    Science.gov (United States)

    Rombach-Riegraf, Verena; Karle, Anette C; Wolf, Babette; Sordé, Laetitia; Koepke, Stephan; Gottlieb, Sascha; Krieg, Jennifer; Djidja, Marie-Claude; Baban, Aida; Spindeldreher, Sebastian; Koulov, Atanas V; Kiessling, Andrea

    2014-01-01

    Subvisible proteinaceous particles which are present in all therapeutic protein formulations are in the focus of intense discussions between health authorities, academics and biopharmaceutical companies in the context of concerns that such particles could promote unwanted immunogenicity via anti-drug antibody formation. In order to provide further understanding of the subject, this study closely examines the specific biological effects proteinaceous particles may exert on dendritic cells (DCs) as the most efficient antigen-presenting cell population crucial for the initiation of the adaptive immune response. Two different model IgG antibodies were subjected to three different types of exaggerated physical stress to generate subvisible particles in far greater concentrations than the ones typical for the currently marketed biotherapeutical antibodies. The aggregated samples were used in in vitro biological assays in order to interrogate the early DC-driven events that initiate CD4 T-cell dependent humoral adaptive immune responses--peptide presentation capacity and co-stimulatory activity of DCs. Most importantly, antigen presentation was addressed with a unique approach called MHC-associated Peptide Proteomics (MAPPs), which allows for identifying the sequences of HLA-DR associated peptides directly from human dendritic cells. The experiments demonstrated that highly aggregated solutions of two model mAbs generated under controlled conditions can induce activation of human monocyte-derived DCs as indicated by upregulation of typical maturation markers including co-stimulatory molecules necessary for CD4 T-cell activation. Additional data suggest that highly aggregated proteins could induce in vitro T-cell responses. Intriguingly, strong aggregation-mediated changes in the pattern and quantity of antigen-derived HLA-DR associated peptides presented on DCs were observed, indicating a change in protein processing and presentation. Increasing the amounts of subvisible

  2. Enzymatically degradable poly(ethylene glycol) hydrogels for the 3D culture and release of human embryonic stem cell derived pancreatic precursor cell aggregates.

    Science.gov (United States)

    Amer, Luke D; Holtzinger, Audrey; Keller, Gordon; Mahoney, Melissa J; Bryant, Stephanie J

    2015-08-01

    This study aimed to develop a three dimensional culture platform for aggregates of human embryonic stem cell (hESC)-derived pancreatic progenitors that enables long-term culture, maintains aggregate size and morphology, does not adversely affect differentiation and provides a means for aggregate recovery. A platform was developed with poly(ethylene glycol) hydrogels containing collagen type I, for cell-matrix interactions, and peptide crosslinkers, for facile recovery of aggregates. The platform was first demonstrated with RIN-m5F cells, showing encapsulation and subsequent release of single cells and aggregates without adversely affecting viability. Aggregates of hESC-derived pancreatic progenitors with an effective diameter of 82 (15)μm were either encapsulated in hydrogels or cultured in suspension for 28 days. At day 14, aggregate viability was maintained in the hydrogels, but significantly reduced (88%) in suspension culture. However by day 28, viability was reduced under both culture conditions. Aggregate size was maintained in the hydrogels, but in suspension was significantly higher (∼ 2-fold) by day 28. The ability to release aggregates followed by a second enzyme treatment to achieve single cells enabled assessment by flow cytometry. Prior to encapsulation, there were 39% Pdx1(+)/Nkx6.1(+) cells, key endocrine markers required for β-cell maturation. The fraction of doubly positive cells was not affected in hydrogels but was slightly and significantly lower in suspension culture by 28 days. In conclusion, we demonstrate that a MMP-sensitive PEG hydrogel containing collagen type I is a promising platform for hESC-derived pancreatic progenitors that maintains viable aggregates, aggregate size, and progenitor state and offers facile recovery of aggregates.

  3. Aggregates of mutant CFTR fragments in airway epithelial cells of CF lungs: new pathologic observations.

    Science.gov (United States)

    Du, Kai; Karp, Philip H; Ackerley, Cameron; Zabner, Joseph; Keshavjee, Shaf; Cutz, Ernest; Yeger, Herman

    2015-03-01

    Cystic fibrosis (CF) is caused by a mutation in the CF transmembrane conductance regulator (CFTR) gene resulting in a loss of Cl(-) channel function, disrupting ion and fluid homeostasis, leading to severe lung disease with airway obstruction due to mucus plugging and inflammation. The most common CFTR mutation, F508del, occurs in 90% of patients causing the mutant CFTR protein to misfold and trigger an endoplasmic reticulum based recycling response. Despite extensive research into the pathobiology of CF lung disease, little attention has been paid to the cellular changes accounting for the pathogenesis of CF lung disease. Here we report a novel finding of intracellular retention and accumulation of a cleaved fragment of F508del CFTR in concert with autophagic like phagolysosomes in the airway epithelium of patients with F508del CFTR. Aggregates consisting of poly-ubiquitinylated fragments of only the N-terminal domain of F508del CFTR but not the full-length molecule accumulate to appreciable levels. Importantly, these undegraded intracytoplasmic aggregates representing the NT-NBD1 domain of F508del CFTR were found in ciliated, in basal, and in pulmonary neuroendocrine cells. Aggregates were found in both native lung tissues and ex-vivo primary cultures of bronchial epithelial cells from CF donors, but not in normal control lungs. Our findings present a new, heretofore, unrecognized innate CF gene related cell defect and a potential contributing factor to the pathogenesis of CF lung disease. Mutant CFTR intracytoplasmic aggregates could be analogous to the accumulation of misfolded proteins in other degenerative disorders and in pulmonary "conformational protein-associated" diseases. Consequently, potential alterations to the functional integrity of airway epithelium and regenerative capacity may represent a critical new element in the pathogenesis of CF lung disease.

  4. Blockade of the formation of insoluble ubiquitinated protein aggregates by EGCG3"Me in the alloxan-induced diabetic kidney.

    Directory of Open Access Journals (Sweden)

    Shuxian Cai

    Full Text Available BACKGROUND: Renal accumulation of reactive carbonyl compounds (RCCs has been linked to the progression of diabetic nephropathy. We previously demonstrated that carbonyl stress induces the formation of amino-carbonyl cross-links and sharply increases the content of β-sheet-rich structures, which is the seed of insoluble aggregates formation, and tea catechin (--epigallocatechin 3-gallate (EGCG can reverse this process in vitro and in vivo. In this study, methylated derivative (--epigallocatechin-3-O-(3-O-methyl-gallate (EGCG3"Me was hypothesized to neutralize carbonyl stress mediating the formation of insoluble ubiquitinated protein (IUP aggregates, and reduce the early development of diabetic nephropathy. METHODS AND RESULTS: Diabetes was induced in mice by intraperitoneally injecting alloxan monohydrate (200 mg/kg/d twice and administering EGCG3"Me by gavage for 15 d. Reagent case and western blot results showed that, in diabetic kidneys, the carbonyl proteins in the serum increased; and in insoluble protein fraction, 4-hydroxynonenal-modified proteins, IUP aggregates and p62 accumulated; FT-IR study demonstrated that the lipid content, anti-parallel β-sheet structure and aggregates increased. EGCG3"Me treatment could effectively reverse this process, even better than the negative control treatment. CONCLUSIONS: EGCG3"Me exhibiting anti-β-sheet-rich IUP aggregate properties, maybe represents a new strategy to impede the progression of diabetic nephropathy and other diabetic complications.

  5. Symmetry breaking, germ layer specification and axial organisation in aggregates of mouse embryonic stem cells.

    Science.gov (United States)

    van den Brink, Susanne C; Baillie-Johnson, Peter; Balayo, Tina; Hadjantonakis, Anna-Katerina; Nowotschin, Sonja; Turner, David A; Martinez Arias, Alfonso

    2014-11-01

    Mouse embryonic stem cells (mESCs) are clonal populations derived from preimplantation mouse embryos that can be propagated in vitro and, when placed into blastocysts, contribute to all tissues of the embryo and integrate into the normal morphogenetic processes, i.e. they are pluripotent. However, although they can be steered to differentiate in vitro into all cell types of the organism, they cannot organise themselves into structures that resemble embryos. When aggregated into embryoid bodies they develop disorganised masses of different cell types with little spatial coherence. An exception to this rule is the emergence of retinas and anterior cortex-like structures under minimal culture conditions. These structures emerge from the cultures without any axial organisation. Here, we report that small aggregates of mESCs, of about 300 cells, self-organise into polarised structures that exhibit collective behaviours reminiscent of those that cells exhibit in early mouse embryos, including symmetry breaking, axial organisation, germ layer specification and cell behaviour, as well as axis elongation. The responses are signal specific and uncouple processes that in the embryo are tightly associated, such as specification of the anteroposterior axis and anterior neural development, or endoderm specification and axial elongation. We discuss the meaning and implications of these observations and the potential uses of these structures which, because of their behaviour, we suggest to call 'gastruloids'.

  6. Prefoldin Protects Neuronal Cells from Polyglutamine Toxicity by Preventing Aggregation Formation*

    Science.gov (United States)

    Tashiro, Erika; Zako, Tamotsu; Muto, Hideki; Itoo, Yoshinori; Sörgjerd, Karin; Terada, Naofumi; Abe, Akira; Miyazawa, Makoto; Kitamura, Akira; Kitaura, Hirotake; Kubota, Hiroshi; Maeda, Mizuo; Momoi, Takashi; Iguchi-Ariga, Sanae M. M.; Kinjo, Masataka; Ariga, Hiroyoshi

    2013-01-01

    Huntington disease is caused by cell death after the expansion of polyglutamine (polyQ) tracts longer than ∼40 repeats encoded by exon 1 of the huntingtin (HTT) gene. Prefoldin is a molecular chaperone composed of six subunits, PFD1–6, and prevents misfolding of newly synthesized nascent polypeptides. In this study, we found that knockdown of PFD2 and PFD5 disrupted prefoldin formation in HTT-expressing cells, resulting in accumulation of aggregates of a pathogenic form of HTT and in induction of cell death. Dead cells, however, did not contain inclusions of HTT, and analysis by a fluorescence correlation spectroscopy indicated that knockdown of PFD2 and PFD5 also increased the size of soluble oligomers of pathogenic HTT in cells. In vitro single molecule observation demonstrated that prefoldin suppressed HTT aggregation at the small oligomer (dimer to tetramer) stage. These results indicate that prefoldin inhibits elongation of large oligomers of pathogenic Htt, thereby inhibiting subsequent inclusion formation, and suggest that soluble oligomers of polyQ-expanded HTT are more toxic than are inclusion to cells. PMID:23720755

  7. Nonlinear Absorption Spectroscopy of Porphyrin J-aggregates in Aqueous Solution: Evidence for Control of Degree of Association by Light-Induced Force

    Science.gov (United States)

    Shirakawa, Masayuki; Nakata, Kazuaki; Suzuki, Masaya; Kobayashi, Takayoshi; Tokunaga, Eiji

    2017-04-01

    Spectroscopic evidence was obtained for molecular aggregation states to be controlled by the irradiation of light, which is off-resonant below the peak absorption energies of both monomers and well-grown J-aggregates. In low (undersaturated)-concentration aqueous solutions of porphyrin molecules (tetraphenyl porphyrin tetrasulfonic acid; TPPS) where the monomer absorbance dominates, irradiation with a 532 nm laser induces a decrease in the monomer absorbance and an increase in the aggregate absorbance. The increase in the absorbance of J-aggregates occurs in a broad spectral range associated with the increase in the number of not only variously sized oligomer aggregates but also aggregates structurally different from well-grown stable J-aggregates. In high-concentration solutions where the J-aggregate absorbance dominates, a blue shift of the absorption peak of J-aggregates is induced at the same 532 nm irradiation, corresponding to a decrease in the aggregation number or in the association energy. By contrast, for spin-coated polymer films of monomers and J-aggregates where molecules are immobile, these features are not observed. It is remarkable that the gradient force potential is smaller by more than seven orders of magnitude than the kinetic energy of the thermal motion of the molecule at room temperature, but the absorption change in solution indicating the increase in the number of aggregates is as large as ΔA ˜ 10-3 in magnitude.

  8. Chapter 3: A fluorescent window into protein folding and aggregation in cells.

    Science.gov (United States)

    Ignatova, Zoya; Gierasch, Lila M

    2008-01-01

    Evolutionary selective pressures have tuned the efficiency of the protein-folding reaction in the crowded complex environment in the cell. Nevertheless, the fidelity of folding is imperfect, leading to off-pathway intermolecular interactions that compete with proper folding and to consequent formation of thermodynamically stable aggregates. Such aggregates constitute the histopathological hallmarks of many neurodegenerative pathologies. Yet, most of the approaches to characterize protein folding and/or misfolding are limited to in vitro conditions. Here, we describe a strategy to directly monitor the behavior of a protein in prokaryotic and eukaryotic cells. The method is based on incorporation of structurally non-perturbing, specific binding motifs for a bis-arsenical fluoroscein dye, FlAsH, in sites that result in distinct dye fluorescence signals for the folded and unfolded states of the protein under study. Our approach has been developed using as a case study the predominantly beta-sheet intracellular lipid-binding protein, cellular retinoic acid-binding protein, alone or as a chimera fused to the exon 1-encoded fragment of huntingtin, which harbors a polyglutamine repeat tract. We have designed protocols to label this protein in vivo and to monitor the resulting fluorescence signal, which reports on any misfolding transition and formation of aggregates, yielding quantitatively interpretable data.

  9. Effect of the crude extract of Cestrum parqui on carrageenin-induced rat paw oedema and aggregation of human blood platelets.

    Science.gov (United States)

    Shehnaz, D; Hamid, F; Baqai, F T; Uddin Ahmad, V

    1999-08-01

    An extract of Cestrum parqui aerial parts in methanol:water (1:1) showed inhibition of carrageenin-induced oedema. The aggregation of human blood platelets induced by adenosine diphosphate and platelet activating factor was also inhibited (IC(50)s were 3 and 2 mg/mL, respectively). On the contrary, the extract did not inhibit arachidonic acid-mediated platelet aggregation.

  10. Glyceraldehyde 3-phosphate dehydrogenase augments the intercellular transmission and toxicity of polyglutamine aggregates in a cell model of Huntington disease.

    Science.gov (United States)

    Mikhaylova, Elena R; Lazarev, Vladimir F; Nikotina, Alina D; Margulis, Boris A; Guzhova, Irina V

    2016-03-01

    The common feature of Huntington disease is the accumulation of oligomers or aggregates of mutant huntingtin protein (mHTT), which causes the death of a subset of striatal neuronal populations. The cytotoxic species can leave neurons and migrate to other groups of cells penetrating and damaging them in a prion-like manner. We hypothesized that the glycolytic enzyme glyceraldehyde 3-phosphate dehydrogenase (GAPDH), previously shown to elevate the aggregation of mHTT, is associated with an increased efficiency of intercellular propagation of mHTT. GAPDH, on its own or together with polyglutamine species, was shown to be released into the extracellular milieu mainly from dying cells as assessed by a novel enzyme immunoassay, western blotting, and ultrafiltration. The conditioned medium of cells with growing GAPDH-polyQ aggregates was toxic to naïve cells, whereas depletion of the aggregates from the medium lowered this cytotoxicity. The GAPDH component of the aggregates was found to increase their toxicity by two-fold in comparison with polyQ alone. Furthermore, GAPDH-polyQ complexes were shown to penetrate acceptor cells and to increase the capacity of polyQ to prionize its intracellular homolog containing a repeat of 25 glutamine residues. Finally, inhibitors of intracellular transport showed that polyQ-GAPDH complexes, as well as GAPDH itself, penetrated cells using clathrin-mediated endocytosis. This suggested a pivotal role of the enzyme in the intercellular transmission of Huntington disease pathogenicity. In conclusion, GAPDH occurring in complexes with polyglutamine strengthens the prion-like activity and toxicity of the migrating aggregates. Aggregating polygluatmine tracts were shown to release from the cells over-expressing mutant huntingtin in a complex with glyceraldehyde 3-phosphate dehydrogenase (GAPDH). The enzyme enhances the intracellular transport of aggregates to healthy cells, prionization of normal cellular proteins and finally cell death, thus

  11. Type I collagen gel protects murine fibrosarcoma L929 cells from TNFα-induced cell death

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Hong-Ju; He, Wen-Qi; Chen, Ling; Liu, Wei-Wei; Xu, Qian; Xia, Ming-Yu; Hayashi, Toshihiko [China-Japan Research Institute of Medical and Pharmaceutical Sciences, Shenyang Pharmaceutical University, Shenyang 110016 (China); Fujisaki, Hitomi; Hattori, Shunji [Nippi Research Institute of Biomatrix, Toride, Ibaraki 302-0017 (Japan); Tashiro, Shin-ichi [Institute for Clinical and Biomedical Sciences, Kyoto 603-8072 (Japan); Onodera, Satoshi [Department of Clinical and Pharmaceutical Sciences, Showa Pharmaceutical University, Tokyo 194-8543 (Japan); Ikejima, Takashi, E-mail: ikejimat@vip.sina.com [China-Japan Research Institute of Medical and Pharmaceutical Sciences, Shenyang Pharmaceutical University, Shenyang 110016 (China)

    2015-02-20

    Murine fibrosarcoma L929 cells have been used to test efficacy of proinflammatory cytokine TNFα. In the present study, we reported on protective effect of type I collagen gel used as L929 cell culture. L929 cell grew and proliferated well on collagen gel. However, the L929 cells exhibited cobblestone-like morphology which was much different from the spread fusiform shape when cultured on conventional cell dishes as well as the cells tended to aggregate. On conventional cell culture dishes, the cells treated with TNFα became round in shape and eventually died in a necroptotic manner. The cells cultured on collagen gel, however, were completely unaffected. TNFα treatment was reported to induce autophagy in L929 cells on the plastic dish, and therefore we investigated the effect of collagen gel on induction of autophagy. The results indicated that autophagy induced by TNFα treatment was much reduced when the cells were cultured on collagen gel. In conclusion, type I collagen gel protected L929 cell from TNFα-induced cell death. - Highlights: • Collagen gel culture changed the morphology of L929 cells. • L929 cell cultured on collagen gel were resistant to TNFα-induced cell death. • Collagen gel culture inhibited TNFα-induced autophagy in L929 cells.

  12. Quantitation of soluble aggregates in recombinant monoclonal antibody cell culture by pH-gradient protein A chromatography.

    Science.gov (United States)

    Pan, Hai; Chen, Ken; Pulisic, Matt; Apostol, Izydor; Huang, Gang

    2009-05-15

    Monoclonal antibodies (mAbs) produced from mammalian cell culture may contain significant amounts of dimers and higher order aggregates. Quantitation of soluble aggregates in the cell culture is time-consuming and labor-intensive, usually involving a purification step to remove the impurities that interfere with the subsequent size exclusion chromatography (SEC) analysis. We have developed a novel pH-gradient protein A chromatography for rapid, non-size based separation of the aggregates in mAb cell culture samples. Our results demonstrate that this method has excellent correlation with SEC and can be applied to both human immunoglobulin gamma 1 (IgG1) and IgG2 antibodies. This approach can be useful in the quantitation of soluble aggregates in crude cell culture samples.

  13. Aggregation-induced fluorescence behavior of triphenylamine-based Schiff bases: the combined effect of multiple forces.

    Science.gov (United States)

    Yang, Mingdi; Xu, Dongling; Xi, Wengang; Wang, Lianke; Zheng, Jun; Huang, Jing; Zhang, Jingyan; Zhou, Hongping; Wu, Jieying; Tian, Yupeng

    2013-10-18

    Eight triphenylamine (TPA)-based Schiff bases that exhibit different aggregation-induced emission (AIE) or aggregation-caused quenching (ACQ) behavior in tetrahydrofuran (THF)/water mixtures have been synthesized and characterized. The photophysical properties in solution, aqueous suspension, film, and the crystalline state along with their relationships were comparatively investigated. The single-crystal structures of 1-8 indicate that compact π···π stacking or excimers induce fluorescence quenching of 1, 2, 5, and 7. However, the existence of J aggregates or multiple intra- and intermolecular interactions restrict the intramolecular vibration and rotation, enabling compounds 3, 4, 6, and 8 to exhibit good AIE character. The size and growth process of particles with different water fractions were studied using scanning electron microscopy, which demonstrated that smaller uniformly dispersed nanoparticles in the THF/water mixtures favor fluorescence emission. The above results suggest that the combined effects of multiple forces caused by structural variation have a great influence on their molecular packing, electronic structure, and aggregation-induced fluorescence properties. In addition, piezofluorochromic experiments verified the potential applications of 4 and 6.

  14. Inhibitory effects and mechanisms of high molecular-weight phlorotannins from Sargassum thunbergii on ADP-induced platelet aggregation

    Institute of Scientific and Technical Information of China (English)

    WEI Yuxi; WANG Changyun; LI Jing; GUO Qi; QI Hongtao

    2009-01-01

    We evaluated the effects of high molecular-weight phlorotannins from Sargassum thunbergii (STP) on ADP-induced platelet aggregation and arachidonic acid (AA) metabolism in New Zealand white rabbits and Wistar rats. The inhibition of STP on platelet aggregation was investigated using a turbidimetric method, and the levels of the terminal products of AA metabolism were measured using the corresponding kits for maleic dialdehyde (MDA), thromboxane B2 (TXB2) and 6-keto-prostaglandin F1α (6-keto-PGF1α) by colorimetry and radioimmunoassay, as appropriate. We found that STP could inhibit ADP-induced platelet aggregation, and the inhibitory ratio was 91.50% at the STP concentration of 4.0 mg/mL. Furthermore, STP markedly affected AA metabolism by decreasing the synthesis of MDA (P<0.01) and increasing the synthesis of 6-keto-PGF1α, thus changing the plasma TXB2/6-keto-PGF1α balance when the platelets were activated (P<0.01). Therefore, STP altered AA metabolism and these findings partly revealed the molecular mechanism by which STP inhibits ADP-induced platelet aggregation.

  15. Putative porcine embryonic stem cell lines derived from aggregated four-celled cloned embryos produced by oocyte bisection cloning.

    Science.gov (United States)

    Siriboon, Chawalit; Lin, Yu-Hsuan; Kere, Michel; Chen, Chun-Da; Chen, Lih-Ren; Chen, Chien-Hong; Tu, Ching-Fu; Lo, Neng-Wen; Ju, Jyh-Cherng

    2015-01-01

    We attempted to isolate ES cell lines using inner cell masses from high-quality cloned porcine blastocysts. After being seeded onto feeders, embryos had better (P cloned embryos (62.8, 42.6 and 12.8% vs. 76.2, 55.2 and 26.2%, respectively) compared to the non-aggregated group (41.6, 23.4 and 3.9%). Effects of feeder types (STO vs. MEF) and serum sources (FBS vs. KSR) on extraction of cloned embryo-derived porcine ES cells were examined. More (17.1%) ntES cell lines over Passage 3 were generated in the MEF/KSR group. However, ntES cells cultured in KSR-supplemented medium had a low proliferation rate with defective morphology, and eventually underwent differentiation or apoptosis subsequently. Approximately 26.1, 22.7 and 35.7% of primary colonies were formed after plating embryos in DMEM, DMEM/F12 and α-MEM media, respectively. Survival rates of ntES cells cultured in α-MEM, DMEM and DMEM/F12 were 16.7, 4.3 and 6.8%, respectively (P > 0.05). We further examined the beneficial effect of TSA treatment of 3× aggregated cloned embryos on establishment of ntES cell lines. Primary colony numbers and survival rates of ntES cells beyond passage 3 were higher (P cloned embryos produced by embryo aggregation, and optimized the ES cell culture system suitable for establishing and maintaining ntES cell lines in undifferentiated state.

  16. Reaction-induced fracturing in a hot pressed calcite-periclase aggregate

    Science.gov (United States)

    Kuleci, H.; Ulven, O. I.; Rybacki, E.; Wunder, B.; Abart, R.

    2017-01-01

    The chemo-mechanical feedbacks associated with hydration of periclase immersed in a calcite matrix were investigated experimentally. Dense calcite-periclase aggregates with calcite to periclase ratio of 90/10 and 95/5 by volume were prepared by hot isostatic pressing. Subsequent hydration experiments were performed in a hydrothermal apparatus at temperatures of 580-610 °C and a pressure of 200 MPa for run durations of 5-60 min. The rate of the periclase to brucite transformation was primarily controlled by the access of fluid. Where fluid was present, the reaction was too fast for the associated positive volume increase of the solids of about 100% to be accommodated by creep of the calcite matrix, and fracturing was induced. The newly formed cracks greatly enhanced the access of fluid leading to a positive feedback between hydration and fracturing. Mostly the newly formed cracks follow pre-existing grain boundaries in the calcite matrix. Comparison of experimental results with numerical 2D discrete element modelling (DEM) of crack formation revealed that the geometry of the crack pattern around a reacting particle depends on the shape of the original periclase particle, on the mechanical strength of the particle-matrix interface and on the mechanical strength and arrangement of grain boundaries in the calcite matrix in the immediate vicinity of the swelling particle.

  17. Synthesis, Aggregation Induced Emission and Mechanochromic Luminescence of New β-Diketone Derivatives Bearing Tetraphenylene Moieties.

    Science.gov (United States)

    Shi, Haijie; Liu, Rui; Zhu, Senqiang; Gong, Qiqi; Shi, Hong; Zhu, Xiaolin; Zhu, Hongjun

    2016-11-01

    A series of β-diketone derivatives bearing tetraphenylene (TPE) moieties were synthesized and characterized. Their photophysical properties were investigated systematically via spectroscopic and theoretical methods. All compounds exhibit broad absorption bands between 300 and 450 nm, which are assigned to the (1)π-π* transition of the conjugated system mixed intramolecular charge-transfer (ICT) transitions. Meanwhile, the emission of these compounds in solution at room temperature (λ em = 458 ~ 509 nm) can be attributed to the (1)π,π*/(1)ICT state. Introduction of freely rotatable TPE to conventional β-diketone luminophors quenches their light emissions in the solutions, but endows these molecules with aggregation-induced emission (AIE) characteristics in the condensed phase due to the restriction of intramolecular rotation. The spectroscopic studies and theoretical calculations indicate that the photophysical properties of these β-diketone derivatives can be tuned by the appended substituents, which would be useful for rational design of AIE compounds with high solid state luminescence performance. Furthermore, these AIE-active compounds exhibited distinct piezofluorochromic properties and switched reversibly upon grinding-fuming. Their photophysical properties have been investigated with the aim to provide a basis for elucidating the structure-property correlations and developing new multi-stimuli responsive luminescent materials.

  18. Astrocyte-derived tissue Transglutaminase affects fibronectin deposition, but not aggregation, during cuprizone-induced demyelination

    Science.gov (United States)

    Espitia Pinzon, Nathaly; Sanz-Morello, Berta; Brevé, John J. P.; Bol, John G. J. M.; Drukarch, Benjamin; Bauer, Jan; Baron, Wia; van Dam, Anne-Marie

    2017-01-01

    Astrogliosis as seen in Multiple Sclerosis (MS) develops into astroglial scarring, which is beneficial because it seals off the site of central nervous system (CNS) damage. However, astroglial scarring also forms an obstacle that inhibits axon outgrowth and (re)myelination in brain lesions. This is possibly an important cause for incomplete remyelination in the CNS of early stage MS patients and for failure in remyelination when the disease progresses. In this study we address whether under demyelinating conditions in vivo, tissue Transglutaminase (TG2), a Ca2+ -dependent enzyme that catalyses posttranslational modification of proteins, contributes to extracellular matrix (ECM) deposition and/or aggregation. We used the cuprizone model for de- and remyelination. TG2 immunoreactivity and enzymatic activity time-dependently appeared in astrocytes and ECM, respectively, in the corpus callosum of cuprizone-treated mice. Enhanced presence of soluble monomeric and multimeric fibronectin was detected during demyelination, and fibronectin immunoreactivity was slightly decreased in cuprizone-treated TG2−/− mice. In vitro TG2 overexpression in astrocytes coincided with more, while knock-down of TG2 with less fibronectin production. TG2 contributes, at least partly, to fibronectin production, and may play a role in fibronectin deposition during cuprizone-induced demyelination. Our observations are of interest in understanding the functional implications of TG2 during astrogliosis. PMID:28128219

  19. Depletion induced encapsulation by dumbbell-shaped patchy colloids stabilize microspheres against aggregation.

    Science.gov (United States)

    Wolters, Joost Robert; Verweij, Joanne E; Avvisati, Guido; Dijkstra, Marjolein; Kegel, Willem K

    2017-03-08

    In this paper, we demonstrate the stabilization of polystyrene microspheres by encapsulating them with dumbbell-shaped colloids with a sticky and a non-sticky lobe. Upon adding a depletant, an effective short ranged attraction is induced between the microspheres and the smaller, smooth lobes of the dumbbells, making those specifically sticky, whereas the interaction with the larger lobes of the dumbbells is considerably less attractive due to their rough surface, which reduces the overlap volume and leaves them non-sticky. The encapsulation of the microspheres by these rough-smooth patchy dumbbells is investigated using a combination of experiments and computer simulations, both resulting in partial coverage of the template particles. For larger microspheres, the depletion attraction is stronger, resulting in a larger fraction of dumbbells that are attached with both lobes to the surface of microspheres. We thus find a template curvature dependent orientation of the dumbbells. In the Monte Carlo simulations, the introduction of such a small, curvature dependent attraction between the rough lobes of the dumbbells resulted in an increased coverage. However, kinetic constraints imposed by the dumbbell geometry seem to prevent optimal packing of the dumbbells on the template particles under all investigated conditions in experiments and simulations. Despite the incomplete coverage, the encapsulation by dumbbell particles does prevent aggregation of the microspheres, thus acting as a colloid-sized steric stabilizer.

  20. Brain Aggregates: An Effective In Vitro Cell Culture System Modeling Neurodegenerative Diseases.

    Science.gov (United States)

    Ahn, Misol; Kalume, Franck; Pitstick, Rose; Oehler, Abby; Carlson, George; DeArmond, Stephen J

    2016-03-01

    Drug discovery for neurodegenerative diseases is particularly challenging because of the discrepancies in drug effects between in vitro and in vivo studies. These discrepancies occur in part because current cell culture systems used for drug screening have many limitations. First, few cell culture systems accurately model human aging or neurodegenerative diseases. Second, drug efficacy may differ between dividing and stationary cells, the latter resembling nondividing neurons in the CNS. Brain aggregates (BrnAggs) derived from embryonic day 15 gestation mouse embryos may represent neuropathogenic processes in prion disease and reflect in vivo drug efficacy. Here, we report a new method for the production of BrnAggs suitable for drug screening and suggest that BrnAggs can model additional neurological diseases such as tauopathies. We also report a functional assay with BrnAggs by measuring electrophysiological activities. Our data suggest that BrnAggs could serve as an effective in vitro cell culture system for drug discovery for neurodegenerative diseases.

  1. Comparison of Tooth Discoloration Induced by Calcium-Enriched Mixture and Mineral Trioxide Aggregate

    Science.gov (United States)

    Rouhani, Armita; Akbari, Majid; Farhadi-faz, Aida

    2016-01-01

    Introduction: The aim of this in vitro study was to evaluate the tooth discoloration induced by calcium-enriched mixture (CEM) cement and mineral trioxide aggregate (MTA). Methods and Materials: Forty five endodontically treated human maxillary central incisors were selected and divided into three groups (n=15) after removing the coronal 3 mm of the obturating materials. In the MTA group, white MTA plug was placed in pulp chamber and coronal zone of the root canal. In CEM cement group, CEM plug was placed in the tooth in the same manner. In both groups, a wet cotton pellet was placed in the access cavity and the teeth were temporarily sealed. After 24 h the teeth were restored with resin composite. In the negative control group the teeth were also restored with resin composite. The color change in the cervical third of teeth was measured with a colorimeter and was repeated 3 times for each specimen. The teeth were kept in artificial saliva for 6 months. After this period, the color change was measured again. Data were collected by Commission International de I'Eclairage's L*a*b color values, and corresponding ΔE values were calculated. The results were analyzed using the one-way ANOVA and post-hoc Tukey’s test with the significance level defined as 0.05. Results: There was no significant differences between CEM group and control group in mean discoloration. The mean tooth discoloration in MTA group was significantly greater than CEM and control groups (P<0.05). Conclusion: According to the result of the present study CEM cement did not induce tooth discoloration after six months. Therefore it can be used in vital pulp therapy of esthetically sensitive teeth. PMID:27471526

  2. Rosin Surfactant QRMAE Can Be Utilized as an Amorphous Aggregate Inducer: A Case Study of Mammalian Serum Albumin.

    Directory of Open Access Journals (Sweden)

    Mohd Ishtikhar

    Full Text Available Quaternary amine of diethylaminoethyl rosin ester (QRMAE, chemically synthesized biocompatible rosin based cationic surfactant, has various biological applications including its use as a food product additive. In this study, we examined the amorphous aggregation behavior of mammalian serum albumins at pH 7.5, i.e., two units above their isoelectric points (pI ~5.5, and the roles played by positive charge and hydrophobicity of exogenously added rosin surfactant QRMAE. The study was carried out on five mammalian serum albumins, using various spectroscopic methods, dye binding assay, circular dichroism and electron microscopy. The thermodynamics of the binding of mammalian serum albumins to cationic rosin modified surfactant were established using isothermal titration calorimetry (ITC. It was observed that a suitable molar ratio of protein to QRMAE surfactant enthusiastically induces amorphous aggregate formation at a pH above two units of pI. Rosin surfactant QRMAE-albumins interactions revealed a unique interplay between the initial electrostatic and the subsequent hydrophobic interactions that play an important role towards the formation of hydrophobic interactions-driven amorphous aggregate. Amorphous aggregation of proteins is associated with varying diseases, from the formation of protein wine haze to the expansion of the eye lenses in cataract, during the expression and purification of recombinant proteins. This study can be used for the design of novel biomolecules or drugs with the ability to neutralize factor(s responsible for the aggregate formation, in addition to various other industrial applications.

  3. Milk protein suspensions enriched with three essential minerals: Physicochemical characterization and aggregation induced by a novel enzymatic pool.

    Science.gov (United States)

    Lombardi, Julia; Spelzini, Darío; Corrêa, Ana Paula Folmer; Brandelli, Adriano; Risso, Patricia; Boeris, Valeria

    2016-04-01

    Structural changes of casein micelles and their aggregation induced by a novel enzymatic pool isolated from Bacillus spp. in the presence of calcium, magnesium or zinc were investigated. The effect of cations on milk protein structure was studied using fluorescence and dynamic light scattering. In the presence of cations, milk protein structure rearrangements and larger casein micelle size were observed. The interaction of milk proteins with zinc appears to be of a different nature than that with calcium or magnesium. Under the experimental conditions assayed, the affinity of each cation for some groups present in milk proteins seems to play an important role, besides electrostatic interaction. On the other hand, the lowest aggregation times were achieved at the highest calcium and zinc concentrations (15 mM and 0.25 mM, respectively). The study found that the faster the aggregation of casein micelles, the less compact the gel matrix obtained. Cation concentrations affected milk protein aggregation kinetics and the structure of the aggregates formed.

  4. Effect of carbon sources on the aggregation of photo fermentative bacteria induced by L-cysteine for enhancing hydrogen production.

    Science.gov (United States)

    Xie, Guo-Jun; Liu, Bing-Feng; Ding, Jie; Wang, Qilin; Ma, Chao; Zhou, Xu; Ren, Nan-Qi

    2016-12-01

    Poor flocculation of photo fermentative bacteria resulting in continuous biomass washout from photobioreactor is a critical challenge to achieve rapid and stable hydrogen production. In this work, the aggregation of Rhodopseudomonas faecalis RLD-53 was successfully developed in a photobioreactor and the effects of different carbon sources on hydrogen production and aggregation ability were investigated. Extracellular polymeric substances (EPS) production by R. faecalis RLD-53 cultivated using different carbon sources were stimulated by addition of L-cysteine. The absolute ζ potentials of R. faecalis RLD-53 were considerably decreased with addition of L-cysteine, and aggregation barriers based on DLVO dropped to 15-43 % of that in control groups. Thus, R. faecalis RLD-53 flocculated effectively, and aggregation abilities of strain RLD-53 cultivated with acetate, propionate, lactate and malate reached 29.35, 32.34, 26.07 and 24.86 %, respectively. In the continuous test, hydrogen-producing activity was also promoted and reached 2.45 mol H2/mol lactate, 3.87 mol H2/mol propionate and 5.10 mol H2/mol malate, respectively. Therefore, the aggregation of R. faecalis RLD-53 induced by L-cysteine is independent on the substrate types, which ensures the wide application of this technology to enhance hydrogen recovery from wastewater dominated by different organic substrates.

  5. Rosin Surfactant QRMAE Can Be Utilized as an Amorphous Aggregate Inducer: A Case Study of Mammalian Serum Albumin.

    Science.gov (United States)

    Ishtikhar, Mohd; Chandel, Tajjali Ilm; Ahmad, Aamir; Ali, Mohd Sajid; Al-Lohadan, Hamad A; Atta, Ayman M; Khan, Rizwan Hasan

    2015-01-01

    Quaternary amine of diethylaminoethyl rosin ester (QRMAE), chemically synthesized biocompatible rosin based cationic surfactant, has various biological applications including its use as a food product additive. In this study, we examined the amorphous aggregation behavior of mammalian serum albumins at pH 7.5, i.e., two units above their isoelectric points (pI ~5.5), and the roles played by positive charge and hydrophobicity of exogenously added rosin surfactant QRMAE. The study was carried out on five mammalian serum albumins, using various spectroscopic methods, dye binding assay, circular dichroism and electron microscopy. The thermodynamics of the binding of mammalian serum albumins to cationic rosin modified surfactant were established using isothermal titration calorimetry (ITC). It was observed that a suitable molar ratio of protein to QRMAE surfactant enthusiastically induces amorphous aggregate formation at a pH above two units of pI. Rosin surfactant QRMAE-albumins interactions revealed a unique interplay between the initial electrostatic and the subsequent hydrophobic interactions that play an important role towards the formation of hydrophobic interactions-driven amorphous aggregate. Amorphous aggregation of proteins is associated with varying diseases, from the formation of protein wine haze to the expansion of the eye lenses in cataract, during the expression and purification of recombinant proteins. This study can be used for the design of novel biomolecules or drugs with the ability to neutralize factor(s) responsible for the aggregate formation, in addition to various other industrial applications.

  6. Tunneling electron induced molecular electroluminescence from individual porphyrin J-aggregates

    Energy Technology Data Exchange (ETDEWEB)

    Meng, Qiushi; Zhang, Chao; Zhang, Yang, E-mail: zhyangnano@ustc.edu.cn, E-mail: zcdong@ustc.edu.cn; Zhang, Yao; Liao, Yuan; Dong, Zhenchao, E-mail: zhyangnano@ustc.edu.cn, E-mail: zcdong@ustc.edu.cn [Hefei National Laboratory for Physical Sciences at the Microscale and Synergetic Innovation Center of Quantum Information and Quantum Physics, University of Science and Technology of China, Hefei, Anhui 230026 (China)

    2015-07-27

    We investigate molecular electroluminescence from individual tubular porphyrin J-aggregates on Au(111) by tunneling electron excitations in an ultrahigh-vacuum scanning tunneling microscope (STM). High-resolution STM images suggest a spiral tubular structure for the porphyrin J-aggregate with highly ordered “brickwork”-like arrangements. Such aggregated nanotube is found to behave like a self-decoupled molecular architecture and shows red-shifted electroluminescence characteristics of J-aggregates originated from the delocalized excitons. The positions of the emission peaks are found to shift slightly depending on the excitation sites, which, together with the changes in the observed spectral profiles with vibronic progressions, suggest a limited exciton coherence number within several molecules. The J-aggregate electroluminescence is also found unipolar, occurring only at negative sample voltages, which is presumably related to the junction asymmetry in the context of molecular excitations via the carrier injection mechanism.

  7. An induced junction photovoltaic cell

    Science.gov (United States)

    Call, R. L.

    1974-01-01

    Silicon solar cells operating with induced junctions rather than diffused junctions have been fabricated and tested. Induced junctions were created by forming an inversion layer near the surface of the silicon by supplying a sheet of positive charge above the surface. Measurements of the response of the inversion layer cell to light of different wavelengths indicated it to be more sensitive to the shorter wavelengths of the sun's spectrum than conventional cells. The greater sensitivity occurs because of the shallow junction and the strong electric field at the surface.

  8. Direct contact with endoderm-like cells efficiently induces cardiac progenitors from mouse and human pluripotent stem cells.

    Directory of Open Access Journals (Sweden)

    Hideki Uosaki

    Full Text Available RATIONALE: Pluripotent stem cell-derived cardiac progenitor cells (CPCs have emerged as a powerful tool to study cardiogenesis in vitro and a potential cell source for cardiac regenerative medicine. However, available methods to induce CPCs are not efficient or require high-cost cytokines with extensive optimization due to cell line variations. OBJECTIVE: Based on our in-vivo observation that early endodermal cells maintain contact with nascent pre-cardiac mesoderm, we hypothesized that direct physical contact with endoderm promotes induction of CPCs from pluripotent cells. METHOD AND RESULT: To test the hypothesis, we cocultured mouse embryonic stem (ES cells with the endodermal cell line End2 by co-aggregation or End2-conditioned medium. Co-aggregation resulted in strong induction of Flk1(+ PDGFRa(+ CPCs in a dose-dependent manner, but the conditioned medium did not, indicating that direct contact is necessary for this process. To determine if direct contact with End2 cells also promotes the induction of committed cardiac progenitors, we utilized several mouse ES and induced pluripotent (iPS cell lines expressing fluorescent proteins under regulation of the CPC lineage markers Nkx2.5 or Isl1. In agreement with earlier data, co-aggregation with End2 cells potently induces both Nkx2.5(+ and Isl1(+ CPCs, leading to a sheet of beating cardiomyocytes. Furthermore, co-aggregation with End2 cells greatly promotes the induction of KDR(+ PDGFRa(+ CPCs from human ES cells. CONCLUSIONS: Our co-aggregation method provides an efficient, simple and cost-effective way to induce CPCs from mouse and human pluripotent cells.

  9. P2Y2 nucleotide receptor activation enhances the aggregation and self-organization of dispersed salivary epithelial cells.

    Science.gov (United States)

    El-Sayed, Farid G; Camden, Jean M; Woods, Lucas T; Khalafalla, Mahmoud G; Petris, Michael J; Erb, Laurie; Weisman, Gary A

    2014-07-01

    Hyposalivation resulting from salivary gland dysfunction leads to poor oral health and greatly reduces the quality of life of patients. Current treatments for hyposalivation are limited. However, regenerative medicine to replace dysfunctional salivary glands represents a revolutionary approach. The ability of dispersed salivary epithelial cells or salivary gland-derived progenitor cells to self-organize into acinar-like spheres or branching structures that mimic the native tissue holds promise for cell-based reconstitution of a functional salivary gland. However, the mechanisms involved in salivary epithelial cell aggregation and tissue reconstitution are not fully understood. This study investigated the role of the P2Y2 nucleotide receptor (P2Y2R), a G protein-coupled receptor that is upregulated following salivary gland damage and disease, in salivary gland reconstitution. In vitro results with the rat parotid acinar Par-C10 cell line indicate that P2Y2R activation with the selective agonist UTP enhances the self-organization of dispersed salivary epithelial cells into acinar-like spheres. Other results indicate that the P2Y2R-mediated response is dependent on epidermal growth factor receptor activation via the metalloproteases ADAM10/ADAM17 or the α5β1 integrin/Cdc42 signaling pathway, which leads to activation of the MAPKs JNK and ERK1/2. Ex vivo data using primary submandibular gland cells from wild-type and P2Y2R(-/-) mice confirmed that UTP-induced migratory responses required for acinar cell self-organization are mediated by the P2Y2R. Overall, this study suggests that the P2Y2R is a promising target for salivary gland reconstitution and identifies the involvement of two novel components of the P2Y2R signaling cascade in salivary epithelial cells, the α5β1 integrin and the Rho GTPase Cdc42.

  10. Alpha-synuclein-induced aggregation of cytoplasmic vesicles in Saccharomyces cerevisiae.

    Science.gov (United States)

    Soper, James H; Roy, Subhojit; Stieber, Anna; Lee, Eliza; Wilson, Robert B; Trojanowski, John Q; Burd, Christopher G; Lee, Virginia M-Y

    2008-03-01

    Aggregated alpha-synuclein (alpha-syn) fibrils form Lewy bodies (LBs), the signature lesions of Parkinson's disease (PD) and related synucleinopathies, but the pathogenesis and neurodegenerative effects of LBs remain enigmatic. Recent studies have shown that when overexpressed in Saccharomyces cerevisiae, alpha-syn localizes to plasma membranes and forms cytoplasmic accumulations similar to human alpha-syn inclusions. However, the exact nature, composition, temporal evolution, and underlying mechanisms of yeast alpha-syn accumulations and their relevance to human synucleinopathies are unknown. Here we provide ultrastructural evidence that alpha-syn accumulations are not comprised of LB-like fibrils, but are associated with clusters of vesicles. Live-cell imaging showed alpha-syn initially localized to the plasma membrane and subsequently formed accumulations in association with vesicles. Imaging of truncated and mutant forms of alpha-syn revealed the molecular determinants and vesicular trafficking pathways underlying this pathological process. Because vesicular clustering is also found in LB-containing neurons of PD brains, alpha-syn-mediated vesicular accumulation in yeast represents a model system to study specific aspects of neurodegeneration in PD and related synucleinopathies.

  11. TTYH2, a human homologue of the Drosophila melanogaster gene tweety, is up-regulated in colon carcinoma and involved in cell proliferation and cell aggregation

    Institute of Scientific and Technical Information of China (English)

    Yuji Toiyama; Akira Mizoguchi; Kazushi Kimura; Junichirou Hiro; Yasuhiro Inoue; Tomonari Tutumi; Chikao Miki; Masato Kusunoki

    2007-01-01

    AIM: To investigate the expression patterns of TTYH2 in the human colon cancer and colon cancer cell lines and to evaluate the inhibitory effect of small interfering RNA (siRIMA) on the expression of TTYH2 in colon cancer cell lines.METHODS: We investigated the expression patterns of TTYH2 in colon cancer, adjacent non-tumorous colon mucosa, and cancer cell lines (DLD-1, caco-2, and Lovo) by RT-PCR. Furthermore, a siRNA plasmid expression vector against TTYH2 was constructed and transfected into DLD-1 and Caco-2 with LipofectamineTM 2000. The down regulation of TTYH2 expression was detected by RT-PCR and the role of siRNA in inducing cell proliferation and cell aggregation was evaluated by MTT and aggregation assay.RESULTS: TTYH2 gene expression in colon cancer tissue was significantly up-regulated compared with normal colonic mucosa (1.23 ± 0.404 vs 0.655 ± 0.373, P=0.0103). Colon cancer derived cell lines including DLD-1, Caco-2, and Lovo also expressed high levels of TTYH2. In contrast, transfection with siRNA-TTYH2 significantly inhibited both proliferation and scattering of these cancer cell lines.CONCLUSION: The present work demonstrates, for the first time, that the TTYH2 gene expression is significantly up-regulated in colon cancer. The TTYH2 gene may play an important role in regulating both proliferating and metastatic potentials of colorectal cancer.

  12. A Diffusion Model of Field-Induced Aggregation in Ferrofluid Film

    Institute of Scientific and Technical Information of China (English)

    FANG Wen-Xiao; HE Zhen-Hui; CHEN Di-Hu; ZHAO Yan-E

    2008-01-01

    By introducing Arrhenius behaviour to the ferroparticles on the surface of the aggregated columnar structure in a diffusion model, equilibrium equations are set up. The solution of the equations shows that to keep the aggregated structures stable, a characteristic fleld is needed. The aggregation is enhanced by magnetic fields, yet suppressed as the temperature increases. Analysing the influence of the magnetic field on the interaction energy between the dipolar particles, we estimate the portion of the diffusing particles, and provide the agreeable ratio of the column radius over the centre-to-centre spacing between columns in a hexagonal columnar structure formed under a perpendicular magnetic field.

  13. Evaluation of bioactivity of octacalcium phosphate using osteoblastic cell aggregates on a spheroid culture device

    Directory of Open Access Journals (Sweden)

    Takahisa Anada

    2016-03-01

    Full Text Available Much attention has been paid to three-dimensional cell culture systems in the field of regenerative medicine, since three-dimensional cellular aggregates, or spheroids, are thought to better mimic the in vivo microenvironments compared to conventional monolayer cultured cells. Synthetic calcium phosphate (CaP materials are widely used as bone substitute materials in orthopedic and dental surgeries. Here we have developed a technique for constructing a hybrid spheroid consisting of mesenchymal stem cells (MSCs and synthetic CaP materials using a spheroid culture device. We found that the device is able to generate uniform-sized CaP/cell hybrid spheroids rapidly and easily. The results showed that the extent of osteoblastic differentiation from MSCs was different when cells were grown on octacalcium phosphate (OCP, hydroxyapatite (HA, or β-tricalcium phosphate (β-TCP. OCP showed the greatest ability to increase the alkaline phosphatase activity of the spheroid cells. The results suggest that the spheroids with incorporated OCP may be an effective implantable hybrid consisting of scaffold material and cells for bone regeneration. It is also possible that this CaP–cell spheroid system may be used as an in vitro method for assessing the osteogenic induction ability of CaP materials.

  14. Influence of erythrocyte aggregation at pathological levels on cell-free marginal layer in a narrow circular tube.

    Science.gov (United States)

    Namgung, Bumseok; Sakai, Hiromi; Kim, Sangho

    2015-01-01

    Human red blood cells (RBCs) were perfused in a circular micro-tube (inner diameter of 25 μm) to examine the dynamic changes of cell-free marginal region at both physiological (normal) and pathophysiological (hyper) levels of RBC aggregation. The cell-free area (CFA) was measured to provide additional information on the cell-free layer (CFL) width changes in space and time domains. A prominent enhancement in the mean CFL width was found in hyper-aggregating conditions as compared to that in non-aggregating conditions (P <  0.001). The frequent contacts between RBC and the tube wall were observed and the contact frequency was greatly decreased when the aggregation level was increased from none to normal (P <  0.05) and to hyper (P <  0.001) levels. In addition, the enhanced aggregation from none to hyper levels significantly enlarged the CFA (P <  0.01). We concluded that the RBC aggregation at pathophysiological levels could promote not only the CFL width (one-dimensional parameter) but also the spatiotemporal variation of CFA (two-dimensional parameter).

  15. Kinetics of self-induced aggregation of Brownian particles: non-Markovian and non-Gaussian features

    CERN Document Server

    Ghosh, Pulak Kumar; Bag, Bidhan Chandra

    2012-01-01

    In this paper we have studied a model for self-induced aggregation in Brownian particle incorporating the non-Markovian and non-Gaussian character of the associated random noise process. In this model the time evolution of each individual is guided by an over-damped Langevin equation of motion with a non-local drift resulting from the local unbalance distributions of the other individuals. Our simulation result shows that colored nose can induce the cluster formation even at large noise strength. Another observation is that critical noise strength grows very rapidly with increase of noise correlation time for Gaussian noise than non Gaussian one. However, at long time limit the cluster number in aggregation process decreases with time following a power law. The exponent in the power law increases remarkable for switching from Markovian to non Markovian noise process.

  16. Development of a valve-based cell printer for the formation of human embryonic stem cell spheroid aggregates.

    Science.gov (United States)

    Faulkner-Jones, Alan; Greenhough, Sebastian; King, Jason A; Gardner, John; Courtney, Aidan; Shu, Wenmiao

    2013-03-01

    In recent years, the use of a simple inkjet technology for cell printing has triggered tremendous interest and established the field of biofabrication. A key challenge has been the development of printing processes which are both controllable and less harmful, in order to preserve cell and tissue viability and functions. Here, we report on the development of a valve-based cell printer that has been validated to print highly viable cells in programmable patterns from two different bio-inks with independent control of the volume of each droplet (with a lower limit of 2 nL or fewer than five cells per droplet). Human ESCs were used to make spheroids by overprinting two opposing gradients of bio-ink; one of hESCs in medium and the other of medium alone. The resulting array of uniform sized droplets with a gradient of cell concentrations was inverted to allow cells to aggregate and form spheroids via gravity. The resulting aggregates have controllable and repeatable sizes, and consequently they can be made to order for specific applications. Spheroids with between 5 and 140 dissociated cells resulted in spheroids of 0.25-0.6 mm diameter. This work demonstrates that the valve-based printing process is gentle enough to maintain stem cell viability, accurate enough to produce spheroids of uniform size, and that printed cells maintain their pluripotency. This study includes the first analysis of the response of human embryonic stem cells to the printing process using this valve-based printing setup.

  17. Design of small molecules that target metal-A{beta} species and regulate metal-induced A{beta} aggregation and neurotoxicity.

    Science.gov (United States)

    Choi, Jung-Suk; Braymer, Joseph J; Nanga, Ravi P R; Ramamoorthy, Ayyalusamy; Lim, Mi Hee

    2010-12-21

    The accumulation of metal ions and amyloid-β (Aβ) aggregates found in the brain of patients with Alzheimer's disease (AD) has been suggested to be involved in AD pathogenesis. To investigate metal-Aβ-associated pathways in AD, development of chemical tools to target metal-Aβ species is desired. Only a few efforts, however, have been reported. Here, we report bifunctional small molecules, N-(pyridin-2-ylmethyl)aniline (L2-a) and N(1),N(1)-dimethyl-N(4)-(pyridin-2-ylmethyl)benzene-1,4-diamine (L2-b) that can interact with both metal ions and Aβ species, as determined by spectroscopic methods including high-resolution NMR spectroscopy. Using the bifunctional compound L2-b, metal-inducedaggregation and neurotoxicity were modulated in vitro as well as in human neuroblastoma cells. Furthermore, treatment of human AD brain tissue homogenates containing metal ions and Aβ species with L2-b showed disassembly of Aβ aggregates. Therefore, our studies presented herein demonstrate the value of bifunctional compounds as chemical tools for investigating metal-Aβ-associated events and their mechanisms in the development and pathogenesis of AD and as potential therapeutics.

  18. Stabilization of tetanus and diphtheria toxoids against moisture-induced aggregation.

    OpenAIRE

    Schwendeman, S P; Costantino, H R; Gupta, R.K.; Siber, G R; Klibanov, A M; Langer, R.

    1995-01-01

    The progress toward single-dose vaccines has been limited by the poor solid-state stability of vaccine antigens within controlled-release polymers, such as poly(lactide-co-glycolide). For example, herein we report that lyophilized tetanus toxoid aggregates during incubation at 37 degrees C and elevated humidity--i.e., conditions relevant to its release from such systems. The mechanism and extent of this aggregation are dependent on the moisture level in the solid protein, with maximum aggrega...

  19. Protein folding, unfolding and aggregation. Pressure induced intermediate states on the refolding pathway of horseradish peroxidase

    Science.gov (United States)

    Smeller, László; Fidy, Judit; Heremans, Karel

    2004-04-01

    We studied the refolding and aggregation of pressure unfolded proteins. Horseradish peroxidase was found to be very stable and no partially folded intermediates were populated during the refolding. However, the removal of the haem group or the Ca2+ ions or reduction of the disulfide bridge destabilized the protein, resulting in a significant amount of aggregation prone intermediate conformation. Substitution of the haem for fluorescent porphyrin however did not influence the refolding of the protein.

  20. Ectoine and hydroxyectoine inhibit thermal-induced aggregation and increase thermostability of recombinant human interferon Alfa2b.

    Science.gov (United States)

    Salmannejad, Faranak; Nafissi-Varcheh, Nastaran

    2017-01-15

    This study is to investigate whether ectoines (ectoine and hydroxyectoine) can reduce aggregation of rhIFNα2b in aqueous solutions on thermal stress. The effect of thermal stress condition on the stability was therefore investigated using size exclusion-high performance liquid chromatography (SE-HPLC), different spectroscopic measurements, dynamic light scattering (DLS), electrophoresis, and differential scanning calorimetry (DSC). All experiments were performed in a sodium phosphate buffer system (100mM, pH7). The protein samples (100μg/ml) were incubated at 50°C for 14days in the absence or presence (1, 10, 20, and 100mM) of ectoines. In summary, thermal-induced aggregation was reduced in the presence of ectoines, regardless of the ectoines concentration in different periods of incubation time by analyzing with SE-HPLC and turbidity measurement. The inhibitory effect of ectoines on the aggregation was shown by other techniques used. The optimal ectoines concentration was 10mM for aggregation reduction, so samples containing of 10mM of ectoines were selected for further evaluation. Secondary structural and conformational stability increased in presence of ectoines as measured by far-UV circular dichroism and fluorescence spectroscopy, respectively. DSC showed slight increase in Tm of interferon in the presence of ectoines. Hydroxyectoine had superior protein-stabilizing properties than ectoine. In conclusion, this study demonstrates that ectoine and hydroxyectoine are highly effective excipients which can significantly reduce the thermal-induced aggregation of rhIFNα2b at low concentration.

  1. Priming effects in aggregate size fractions induced by glucose addition and grinding

    Science.gov (United States)

    Tian, Jing; Blagodatskaya, Evgenia; Pausch, Johanna; Kuzaykov, Yakov

    2014-05-01

    It is widely recognized that soil organic matter (SOM) mineralization can be accelerated (positive priming) or retarded (negative priming) by addition of easily available substrates to soil. SOM is a heterogeneous mixture, which contains numerous compounds with different degradability and turnover rates times. Nevertheless, so far, there is still lack of knowledge on identifying single fractions of the SOM as the source of C and N released by priming effects. The aim of this study was to determine the priming effect as related to different aggregate fractions, aggregate disruption and the amounts of substrate. In a 49 days incubation experiment, the soil samples were separated into three aggregate fractions (>2 mm, 2-0.25 mm and 0.25 mm) than in the microaggregates (priming effect increased as added glucose increased in all intact aggregate size, and highest priming effect was observed in >2 mm fraction. However, the magnitude of priming effect response to glucose addition depended on the aggregate size after grinding. This study demonstrates that substrate amounts, aggregate fractionation and grinding can have obvious impacts on priming effect, indicating important implications for understanding SOM cycling and stability.

  2. Production Pattern of Ajmalicine in Catharanthus roseus (L. G. Don. Cell Aggregates Culture in the Airlift Bioreactor

    Directory of Open Access Journals (Sweden)

    RIZKITA RACHMI ESYANTI

    2006-12-01

    Full Text Available A research has been conducted to optimize the rate of aeration and initial weight of cell aggregates in the production of ajmalicine in Catharanthus roseus cell culture in airlift bioreactor. Catharanthus roseus culture were grown in Zenk medium with the addition of 2.50 x 10-6 M naphthalene acetic acid (NAA and 10-5 M benzyl amino purine (BAP. Cell aggregates were sub-cultured two times before transferring 20 and 30 g/fw of cell aggregates into bioreactor, respectively, and aerated with the rate of 0.25 l min-1 and 0.34 l min-1, respectively. The pattern of ajmalicine production in bioreactor were observed in every three days within 24 days. Qualitative and quantitative analysis were conducted using HPLC connected to Cromatopac CL-7A Plus. The results showed that the cell aggregates and medium contain ajmalicine. The highest concentration was obtained in combination of 30 g/fw and 0.34 l min-1 aeration compare to 20 g/fw - 0.25 l min-1, 20 g/fw - 0.34 l min-1, as well as 30 g/fw – 0.25 l min-1. The highest ajmalicine content in cell aggregates was obtained on the 12 days (79.23 µg g-1 whilst in medium was obtained in the 18th days (981.15 µg l-1.

  3. Effect of particle collisions and aggregation on red blood cell passage through a bifurcation.

    Science.gov (United States)

    Chesnutt, J K W; Marshall, J S

    2009-12-01

    Blood flow through bifurcating vessels in the microvasculature leads to uneven distribution of red blood cells (RBC) in the downstream channels when the channels have different sizes or the flow rates through the channels are different. This phenomenon, known as plasma skimming, is responsible for the large variation in hematocrit throughout the circulatory system. Furthermore, the strong streamline curvature present within bifurcations leads to frequent collisions between the blood elements (red and white blood cells and platelets) and the vessel walls, as well as a rearrangement of the distribution of the blood elements in the channels downstream of the bifurcation. Computational models of bifurcation flows typically neglect collision and adhesion of RBCs with each other. In this paper, we use a new type of discrete-element model to investigate the effect of particle-particle collisions and RBC aggregation on modeling of plasma skimming in bifurcations. Cases are examined with and without RBC adhesion and for different hematocrit values, including validation against previous computational results. Results show significant plasma skimming in the low-flow daughter branch and an increase in fractional particle flux with increased hematocrit, as in experimental studies. Accounting for particle-particle collisions leads to a considerable increase in collision rate of particles with the vessel wall. Particles are found to approximately follow fluid velocity streamlines, and consequently particle-particle collisions and aggregation do not significantly affect plasma skimming.

  4. Arginine-aromatic interactions and their effects on arginine-induced solubilization of aromatic solutes and suppression of protein aggregation

    KAUST Repository

    Shah, Dhawal

    2011-09-21

    We examine the interaction of aromatic residues of proteins with arginine, an additive commonly used to suppress protein aggregation, using experiments and molecular dynamics simulations. An aromatic-rich peptide, FFYTP (a segment of insulin), and lysozyme and insulin are used as model systems. Mass spectrometry shows that arginine increases the solubility of FFYTP by binding to the peptide, with the simulations revealing the predominant association of arginine to be with the aromatic residues. The calculations further show a positive preferential interaction coefficient, Γ XP, contrary to conventional thinking that positive Γ XP\\'s indicate aggregation rather than suppression of aggregation. Simulations with lysozyme and insulin also show arginine\\'s preference for aromatic residues, in addition to acidic residues. We use these observations and earlier results reported by us and others to discuss the possible implications of arginine\\'s interactions with aromatic residues on the solubilization of aromatic moieties and proteins. Our results also highlight the fact that explanations based purely on Γ XP, which measures average affinity of an additive to a protein, could obscure or misinterpret the underlying molecular mechanisms behind additive-induced suppression of protein aggregation. © 2011 American Institute of Chemical Engineers (AIChE).

  5. Aggregation-induced reversal of transport distances of soil organic matter: are our balances correct?

    Science.gov (United States)

    Hu, Yaxian; Kuhn, Nikolaus

    2014-05-01

    The effect of soil erosion on global carbon cycling, especially as a source or sink of green-house gases (GHGs), is the subject of intense debate. The controversy arises mostly from the lack of information on the fate of eroded soil organic carbon (SOC) as it moves from the site of erosion to the site of longer-term deposition. This requires improved understanding the transport distances of eroded SOC, which is principally related to the settling velocities of sediment fractions that carry the eroded SOC. For aggregated soils, settling velocities are affected by their actual aggregate size rather than the mineral grain size distribution. Aggregate stability is, in turn, strongly influenced by soil organic matter. This study aims at identifying the effect of aggregation on the transport distances of eroded SOC and its susceptibility to mineralization after transport and deposition. A rainfall simulation was carried out on a silty loam soil. The eroded sediments were fractionated by a settling tube apparatus into six different size classes according to their settling velocities and likely transport distances. Weight, SOC concentration and instantaneous respiration rates of the fractions of the six classes were measured. Our results show that: 1) 41% of the eroded SOC was transported with coarse aggregates that would be likely re-distributed across landscapes; 2) erosion was prone to accelerate the mineralization of eroded organic carbon immediately after erosion, compared to undisturbed aggregates; 3) erosion might make a higher contribution to atmospheric CO2 than the estimation made without considering the effects of aggregation and extra SOC mineralization during transport.

  6. Post-assembly atomic layer deposition of ultrathin metal-oxide coatings enhances the performance of an organic dye-sensitized solar cell by suppressing dye aggregation.

    Science.gov (United States)

    Son, Ho-Jin; Kim, Chul Hoon; Kim, Dong Wook; Jeong, Nak Cheon; Prasittichai, Chaiya; Luo, Langli; Wu, Jinsong; Farha, Omar K; Wasielewski, Michael R; Hupp, Joseph T

    2015-03-11

    Dye aggregation and concomitant reduction of dye excited-state lifetimes and electron-injection yields constitute a significant mechanism for diminution of light-to-electrical energy conversion efficiencies in many dye-sensitized solar cells (DSCs). For TiO2-based DSCs prepared with an archetypal donor-acceptor organic dye, (E)-2-cyano-3-(5'-(5''-(p-(diphenylamino)phenyl)-thiophen-2''-yl)thiophen-2'-yl)acrylic acid (OrgD), we find, in part via ultrafast spectroscopy measurements, that postdye-adsorption atomic layer deposition (ALD) of ultrathin layers of either TiO2 or Al2O3 effectively reverses residual aggregation. Notably, the ALD treatment is significantly more effective than the widely used aggregation-inhibiting coadsorbent, chenodeoxycholic acid. Primarily because of reversal of OrgD aggregation, and resulting improved injection yields, ALD post-treatment engenders a 30+% increase in overall energy conversion efficiency. A secondary contributor to increased currents and efficiencies is an ALD-induced attenuation of the rate of interception of injected electrons, resulting in slightly more efficient charge collection.

  7. Multiscale modeling of bacterial colonies: how pili mediate the dynamics of single cells and cellular aggregates

    Science.gov (United States)

    Pönisch, Wolfram; Weber, Christoph A.; Juckeland, Guido; Biais, Nicolas; Zaburdaev, Vasily

    2017-01-01

    Neisseria gonorrhoeae is the causative agent of one of the most common sexually transmitted diseases, gonorrhea. Over the past two decades there has been an alarming increase of reported gonorrhea cases where the bacteria were resistant to the most commonly used antibiotics thus prompting for alternative antimicrobial treatment strategies. The crucial step in this and many other bacterial infections is the formation of microcolonies, agglomerates consisting of up to several thousands of cells. The attachment and motility of cells on solid substrates as well as the cell-cell interactions are primarily mediated by type IV pili, long polymeric filaments protruding from the surface of cells. While the crucial role of pili in the assembly of microcolonies has been well recognized, the exact mechanisms of how they govern the formation and dynamics of microcolonies are still poorly understood. Here, we present a computational model of individual cells with explicit pili dynamics, force generation and pili-pili interactions. We employ the model to study a wide range of biological processes, such as the motility of individual cells on a surface, the heterogeneous cell motility within the large cell aggregates, and the merging dynamics and the self-assembly of microcolonies. The results of numerical simulations highlight the central role of pili generated forces in the formation of bacterial colonies and are in agreement with the available experimental observations. The model can quantify the behavior of multicellular bacterial colonies on biologically relevant temporal and spatial scales and can be easily adjusted to include the geometry and pili characteristics of various bacterial species. Ultimately, the combination of the microbiological experimental approach with the in silico model of bacterial colonies might provide new qualitative and quantitative insights on the development of bacterial infections and thus pave the way to new antimicrobial treatments.

  8. Alginate encapsulation parameters influence the differentiation of microencapsulated embryonic stem cell aggregates.

    Science.gov (United States)

    Wilson, Jenna L; Najia, Mohamad Ali; Saeed, Rabbia; McDevitt, Todd C

    2014-03-01

    Pluripotent embryonic stem cells (ESCs) have tremendous potential as tools for regenerative medicine and drug discovery, yet the lack of processes to manufacture viable and homogenous cell populations of sufficient numbers limits the clinical translation of current and future cell therapies. Microencapsulation of ESCs within microbeads can shield cells from hydrodynamic shear forces found in bioreactor environments while allowing for sufficient diffusion of nutrients and oxygen through the encapsulation material. Despite initial studies examining alginate microbeads as a platform for stem cell expansion and directed differentiation, the impact of alginate encapsulation parameters on stem cell phenotype has not been thoroughly investigated. Therefore, the objective of this study was to systematically examine the effects of varying alginate compositions on microencapsulated ESC expansion and phenotype. Pre-formed aggregates of murine ESCs were encapsulated in alginate microbeads composed of a high or low ratio of guluronic to mannuronic acid residues (High G and High M, respectively), with and without a poly-L-lysine (PLL) coating, thereby providing four distinct alginate bead compositions for analysis. Encapsulation in all alginate compositions was found to delay differentiation, with encapsulation within High G alginate yielding the least differentiated cell population. The addition of a PLL coating to the High G alginate prevented cell escape from beads for up to 14 days. Furthermore, encapsulation within High M alginate promoted differentiation toward a primitive endoderm phenotype. Taken together, the findings of this study suggest that distinct ESC expansion capacities and differentiation trajectories emerge depending on the alginate composition employed, indicating that encapsulation material physical properties can be used to control stem cell fate.

  9. Aggregation of soy protein-isoflavone complexes and gel formation induced by glucono-δ-lactone in soymilk

    Science.gov (United States)

    Hsia, Sheng-Yang; Hsiao, Yu-Hsuan; Li, Wen-Tai; Hsieh, Jung-Feng

    2016-10-01

    This study investigated the glucono-δ-lactone (GDL)-induced aggregation of isoflavones and soy proteins in soymilk. High-performance liquid chromatography (HPLC) analysis indicated that isoflavones mixed with β-conglycinin (7S) and glycinin (11S) proteins formed 7S-isoflavone and 11S-isoflavone complexes in soymilk supernatant fraction (SSF). Most of the soy protein-isoflavone complexes then precipitated into the soymilk pellet fraction (SPF) following the addition of 4 mM GDL, whereupon the pH value of the soymilk dropped from 6.6 to 5.9. Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) and HPLC analysis suggest that the addition of 4 mM GDL induced the aggregation of most 7S (α’, α and β subunits), 11S acidic and 11S basic proteins as well as isoflavones, including most aglycones, including daidzein, glycitein, genistein and a portion of glucosides, including daidzin, glycitin, genistin, malonyldaidzin and malonylgenistin. These results provide an important reference pertaining to the effects of GDL on the aggregation of soy protein-isoflavone complexes and could benefit future research regarding the production of tofu from soymilk.

  10. Nitric oxide inhibited the melanophore aggregation induced by extracellular calcium concentration in snakehead fish, Channa punctatus.

    Science.gov (United States)

    Biswas, Saikat P; Palande, Nikhil V; Jadhao, Arun G

    2011-12-01

    We studied the role of nitric oxide (NO) and extra-cellular Ca(2+) on the melanophores in Indian snakehead teleost, Channa punctatus. Increase of Ca(2+) level in the external medium causes pigment aggregation in melanophores. This pigment-aggregating effect was found to be inhibited when the external medium contained spontaneous NO donor, sodium nitro prusside (SNP) at all the levels of concentration tested. Furthermore, it has been observed that SNP keeps the pigment in dispersed state even after increasing the amount of Ca(2+). In order to test whether NO donor SNP causes dispersion of pigments or not is checked by adding the inhibitor of nitric oxide synthase, N-omega-Nitro-L-arginine (L-NNA) in the medium. It has been noted that the inhibitor L-NNA blocked the effect of NO donor SNP causing aggregation of pigments. In that way NO is inhibiting the effect of extracellular Ca(2+), keeping the pigment dispersed.

  11. Role of amorphous and aggregate phases on field-induced exciton dissociation in a conjugated polymer

    Science.gov (United States)

    Mróz, Marta M.; Lüer, Larry; Houarner-Rassin, Coralie; Anderson, Harry L.; Cabanillas-Gonzalez, Juan

    2013-01-01

    We have applied electric field assisted pump-probe spectroscopy in order to unravel the interplay of amorphous and aggregate phases on the polaron-pair photogeneration process in a conjugated porphyrin polymer. We find that excitons photogenerated in both phases are precursors for polaron pairs with different yields. Kinetic modeling indicates a substantially larger barrier for exciton dissociation in aggregates compared to amorphous areas. The majority of polaron pairs are however formed in aggregate phases due to efficient energy transfer from the amorphous phase. Based on the change in the Stark shift associated with the photogenerated polaron density, we provide a picture of the motion of polaron pairs under the external electric field.

  12. Aggregation-Induced Emission Active Metal-Free Chemosensing Platform for Highly Selective Turn-On Sensing and Bioimaging of Pyrophosphate Anion.

    Science.gov (United States)

    Gogoi, Abhijit; Mukherjee, Sandipan; Ramesh, Aiyagari; Das, Gopal

    2015-07-07

    We report the synthesis of a metal-free chemosensor for highly selective sensing of pyrophosphate (PPi) anion in physiological medium. The novel phenylbenzimidazole functionalized imine containing chemosensor (L; [2,6-bis(((4-(1H-benzo[d]imidazol-2-yl)phenyl)imino) methyl)-4 methyl phenol]) could sense PPi anion through "turn-on" colorimetric and fluorimetric responses in a very competitive environment. The overall sensing mechanism is based on the aggregation-induced emission (AIE) phenomenon. Moreover, a real time in-field device application was demonstrated by sensing PPi in paper strips coated with L. Interestingly, detection of intracellular PPi ions in model human cells could also be possible by fluorescence microscopic studies without any toxicity to these cells.

  13. Influence of heat and shear induced protein aggregation on the in vitro digestion rate of whey proteins.

    Science.gov (United States)

    Singh, Tanoj K; Øiseth, Sofia K; Lundin, Leif; Day, Li

    2014-11-01

    Protein intake is essential for growth and repair of body cells, the normal functioning of muscles, and health related immune functions. Most food proteins are consumed after undergoing various degrees of processing. Changes in protein structure and assembly as a result of processing impact the digestibility of proteins. Research in understanding to what extent the protein structure impacts the rate of proteolysis under human physiological conditions has gained considerable interest. In this work, four whey protein gels were prepared using heat processing at two different pH values, 6.8 and 4.6, with and without applied shear. The gels showed different protein network microstructures due to heat induced unfolding (at pH 6.8) or lack of unfolding, thus resulting in fine stranded protein networks. When shear was applied during heating, particulate protein networks were formed. The differences in the gel microstructures resulted in considerable differences in their rheological properties. An in vitro gastric and intestinal model was used to investigate the resulting effects of these different gel structures on whey protein digestion. In addition, the rate of digestion was monitored by taking samples at various time points throughout the in vitro digestion process. The peptides in the digesta were profiled using SDS-polyacrylamide gel electrophoresis, reversed-phase-HPLC and LC-MS. Under simulated gastric conditions, whey proteins in structured gels were hydrolysed faster than native proteins in solution. The rate of peptides released during in vitro digestion differed depending on the structure of the gels and extent of protein aggregation. The outcomes of this work highlighted that changes in the network structure of the protein can influence the rate and pattern of its proteolysis under gastrointestinal conditions. Such knowledge could assist the food industry in designing novel food formulations to control the digestion kinetics and the release of biologically

  14. Aqueous Extract of Paeonia lactiflora and Paeoniflorin as Aggregation Reducers Targeting Chaperones in Cell Models of Spinocerebellar Ataxia 3

    Directory of Open Access Journals (Sweden)

    Kuo-Hsuan Chang

    2013-01-01

    Full Text Available Spinocerebellar ataxia (SCA types 1, 2, 3, 6, 7, and 17 as well as Huntington’s disease are a group of neurodegenerative disorders caused by expanded CAG repeats encoding a long polyglutamine (polyQ tract in the respective proteins. Evidence has shown that the accumulation of intranuclear and cytoplasmic misfolded polyQ proteins leads to apoptosis and cell death. Thus suppression of aggregate formation is expected to inhibit a wide range of downstream pathogenic events in polyQ diseases. In this study, we established a high-throughput aggregation screening system using 293 ATXN3/Q75-GFP cells and applied this system to test the aqueous extract of Paeonia lactiflora (P. lactiflora and its constituents. We found that the aggregation can be significantly prohibited by P. lactiflora and its active compound paeoniflorin. Meanwhile, P. lactiflora and paeoniflorin upregulated HSF1 and HSP70 chaperones in the same cell models. Both of them further reduced the aggregation in neuronal differentiated SH-SY5Y ATXN3/Q75-GFP cells. Our results demonstrate how P. lactiflora and paeoniflorin are likely to work on polyQ-aggregation reduction and provide insight into the possible working mechanism of P. lactiflora in SCA3. We anticipate our paper to be a starting point for screening more potential herbs for the treatment of SCA3 and other polyQ diseases.

  15. Experiment on ``discovery'' STS 51-C: Aggregation of red cells and thrombocytes in heart disease, hyperlipidaemia and other conditions

    Science.gov (United States)

    Dintenfass, L.

    The aim of this experiment was to study aggregation of red cells in the blood of patients with ischaemic heart disease, diabetes, hyperlipidaemia, and (silent) cancer, and in two normal donors. Reconstituted blood using IgG was also used. The instrument, the automated slit-capillary photo-viscometer (100 kg weight) was set on the middeck of the Space Shuttle. An analogous instrument was at the Kennedy Space Center. Blood was obtained from donors, anticoagulated, and adjusted to haematocrit of 30% using native plasma. Experiments took place at 25°C, during which blood was forced to flow in the slit formed by two parallel glass plates. Macro and microphotography was carried out at specific intervals controlled by a computer. During stasis, lasting 6 minutes, aggregates (or clumps) of the red cells were formed. Results indicated that red cell aggregates do form under zero-G; that such aggregates are smaller than the ones obtained at one-G; that morphology is different, the zero-G showing rouleaux while one-G showing usual sludge-like clumps of red cells in all severe disorders. Platelets appeared to remain monodisperse under zero-G. Assuming that these data can be confirmed, one could suggest that zero-G affects cell-cell interaction, and may consequently influence the internal microstructure of the cell membrane and of the receptors, as well as their activity. Gravitational studies may thus open a new door on immunology and haematology in general.

  16. Natural killer cell activity, lymphocyte proliferation, and cytokine profile in tumor-bearing mice treated with MAPA, a magnesium aggregated polymer from Aspergillus oryzae.

    Science.gov (United States)

    Justo, G Z; Durán, N; Queiroz, M L S

    2003-08-01

    The present study examined the effects of MAPA, an antitumor aggregated polymer of protein magnesium ammonium phospholinoleate-palmitoleate anhydride, isolated from Aspergillus oryzae, on concanavalin A (Con A)-induced spleen cell proliferation, cytokine production and on natural killer (NK) cell activity in Ehrlich ascites tumor-bearing mice. The Ehrlich ascites tumor (EAT) growth led to diminished mitogen-induced expansion of spleen cell populations and total NK activity. This was accompanied by striking spleen enlargement, with a marked increase in total cell counts. Moreover, a substantial enhancement in IL-10 levels, paralleled by a significant decrease in IL-2 was observed, while production of IL-4 and interferon-gamma (IFN-gamma) was not altered. Treatment of mice with 5 mg/kg MAPA for 7 days promoted spleen cell proliferation, IL-2 production and NK cell activity regardless of tumor outgrowth. In addition, MAPA treatment markedly enhanced IFN-gamma levels and reduced IL-10 production relative to EAT mice. A 35% reduction in splenomegaly with normal number of nucleated cells was also found. Altogether, our results suggest that MAPA directly and/or indirectly modulates immune cell activity, and probably disengages tumor-induced suppression of these responses. Clearly, MAPA has an impact and may delay tumor outgrowth through immunotherapeutic mechanisms.

  17. Biological and biophysics aspects of metformin-induced effects: cortex mitochondrial dysfunction and promotion of toxic amyloid pre-fibrillar aggregates

    Science.gov (United States)

    Picone, Pasquale; Vilasi, Silvia; Librizzi, Fabio; Contardi, Marco; Nuzzo, Domenico; Caruana, Luca; Baldassano, Sara; Amato, Antonella; Mulè, Flavia; San Biagio, Pier Luigi; Giacomazza, Daniela; Di Carlo, Marta

    2016-01-01

    The onset of Alzheimer disease (AD) is influenced by several risk factors comprising diabetes. Within this context, antidiabetic drugs, including metformin, are investigated for their effect on AD. We report that in the C57B6/J mice, metformin is delivered to the brain where activates AMP-activated kinase (AMPK), its molecular target. This drug affects the levels of β-secretase (BACE1) and β-amyloid precursor protein (APP), promoting processing and aggregation of β-amyloid (Aβ), mainly in the cortex region. Moreover, metformin induces mitochondrial dysfunction and cell death by affecting the level and conformation of Translocase of the Outer Membrane 40 (TOM40), voltage-dependent anion-selective channels 1 (VDAC1) and hexokinase I (HKI), proteins involved in mitochondrial transport of molecules, including Aβ. By using biophysical techniques we found that metformin is able to directly interact with Aβ influencing its aggregation kinetics and features. These findings indicate that metformin induces different adverse effects, leading to an overall increase of the risk of AD onset. PMID:27509335

  18. Embryo aggregation does not improve the development of interspecies somatic cell nuclear transfer embryos in the horse.

    Science.gov (United States)

    Gambini, Andrés; De Stéfano, Adrián; Jarazo, Javier; Buemo, Carla; Karlanian, Florencia; Salamone, Daniel Felipe

    2016-09-01

    The low efficiency of interspecies somatic cell nuclear transfer (iSCNT) makes it necessary to investigate new strategies to improve embryonic developmental competence. Embryo aggregation has been successfully applied to improve cloning efficiency in mammals, but it remains unclear whether it could also be beneficial for iSCNT. In this study, we first compared the effect of embryo aggregation over in vitro development and blastocyst quality of porcine, bovine, and feline zona-free (ZF) parthenogenetic (PA) embryos to test the effects of embryo aggregation on species that were later used as enucleated oocytes donors in our iSCNT study. We then assessed whether embryo aggregation could improve the in vitro development of ZF equine iSCNT embryos after reconstruction with porcine, bovine, and feline ooplasm. Bovine- and porcine-aggregated PA blastocysts had significantly larger diameters compared with nonaggregated embryos. On the other hand, feline- and bovine-aggregated PA embryos had higher blastocyst cell number. Embryo aggregation of equine-equine SCNT was found to be beneficial for embryo development as we have previously reported, but the aggregation of three ZF reconstructed embryos did not improve embryo developmental rates on iSCNT. In vitro embryo development of nonaggregated iSCNT was predominantly arrested around the stage when transcriptional activation of the embryonic genome is reported to start on the embryo of the donor species. Nevertheless, independent of embryo aggregation, equine blastocyst-like structures could be obtained in our study using domestic feline-enucleated oocytes. Taken together, these results reported that embryo aggregation enhance in vitro PA embryo development and embryo quality but effects vary depending on the species. Embryo aggregation also improves, as expected, the in vitro embryo development of equine-equine SCNT embryos; however, we did not observe positive effects on equine iSCNT embryo development. Among oocytes

  19. Porphyrin Dye-Sensitized Zinc Oxide Aggregated Anodes for Use in Solar Cells

    Directory of Open Access Journals (Sweden)

    Yu-Kai Syu

    2016-08-01

    Full Text Available Porphyrin YD2-o-C8-based dyes were employed to sensitize room-temperature (RT chemical-assembled ZnO aggregated anodes for use in dye-sensitized solar cells (DSSCs. To reduce the acidity of the YD2-o-C8 dye solution, the proton in the carboxyl group of a porphyrin dye was replaced with tetrabuthyl ammonium (TBA+ in this work. The short-circuit current density (Jsc of the YD2-o-C8-TBA-sensitized ZnO DSSCs is higher than that of the YD2-o-C8-sensitized cells, resulting in the improvement of the efficiency of the YD2-o-C8-based ZnO DSSCs. With an appropriate incorporation of chenodeoxycholic acid (CDCA as coadsorbate, the Jsc and efficiency of the YD2-o-C8-TBA-sensitized ZnO DSSC are enhanced due to the improvement of the incident-photon-to-current efficiency (IPCE values in the wavelength range of 400–450 nm. Moreover, a considerable increase in Jsc is achieved by the addition of a light scattering layer in the YD2-o-C8-TBA-sensitized ZnO photoanodes. Significant IPCE enhancement in the range 475–600 nm is not attainable by tuning the YD2-o-C8-TBA sensitization processes for the anodes without light scattering layers. Using the RT chemical-assembled ZnO aggregated anode with a light scattering layer, an efficiency of 3.43% was achieved in the YD2-o-C8-TBA-sensitized ZnO DSSC.

  20. Roles of autophagy in MPP+-induced neurotoxicity in vivo: the involvement of mitochondria and α-synuclein aggregation.

    Directory of Open Access Journals (Sweden)

    Kai-Chih Hung

    Full Text Available Macroautophagy (also known as autophagy is an intracellular self-eating mechanism and has been proposed as both neuroprotective and neurodestructive in the central nervous system (CNS neurodegenerative diseases. In the present study, the role of autophagy involving mitochondria and α-synuclein was investigated in MPP+ (1-methyl-4-phenylpyridinium-induced oxidative injury in chloral hydrate-anesthetized rats in vivo. The oxidative mechanism underlying MPP+-induced neurotoxicity was identified by elevated lipid peroxidation and heme oxygenase-1 levels, a redox-regulated protein in MPP+-infused substantia nigra (SN. At the same time, MPP+ significantly increased LC3-II levels, a hallmark protein of autophagy. To block MPP+-induced autophagy in rat brain, Atg7siRNA was intranigrally infused 4 d prior to MPP+ infusion. Western blot assay showed that in vivo Atg7siRNA transfection not only reduced Atg7 levels in the MPP+-infused SN but attenuated MPP+-induced elevation in LC3-II levels, activation of caspase 9 and reduction in tyrosine hydroxylase levels, indicating that autophagy is pro-death. The immunostaining study demonstrated co-localization of LC3 and succinate dehydrogenase (a mitochondrial complex II as well as LC3 and α-synuclein, suggesting that autophagy may engulf mitochondria and α-synuclein. Indeed, in vivo Atg7siRNA transfection mitigated MPP+-induced reduction in cytochrome c oxidase. In addition, MPP+-induced autophagy differentially altered the α-synuclein aggregates in the infused SN. In conclusion, autophagy plays a prodeath role in the MPP+-induced oxidative injury by sequestering mitochondria in the rat brain. Moreover, our data suggest that the benefits of autophagy depend on the levels of α-synuclein aggregates in the nigrostriatal dopaminergic system of the rat brain.

  1. Weak glycolipid binding of a microdomain-tracer peptide correlates with aggregation and slow diffusion on cell membranes.

    Directory of Open Access Journals (Sweden)

    Tim Lauterbach

    Full Text Available Organized assembly or aggregation of sphingolipid-binding ligands, such as certain toxins and pathogens, has been suggested to increase binding affinity of the ligand to the cell membrane and cause membrane reorganization or distortion. Here we show that the diffusion behavior of the fluorescently tagged sphingolipid-interacting peptide probe SBD (Sphingolipid Binding Domain is altered by modifications in the construction of the peptide sequence that both result in a reduction in binding to ganglioside-containing supported lipid membranes, and at the same time increase aggregation on the cell plasma membrane, but that do not change relative amounts of secondary structural features. We tested the effects of modifying the overall charge and construction of the SBD probe on its binding and diffusion behavior, by Surface Plasmon Resonance (SPR; Biacore analysis on lipid surfaces, and by Fluorescence Correlation Spectroscopy (FCS on live cells, respectively. SBD binds preferentially to membranes containing the highly sialylated gangliosides GT1b and GD1a. However, simple charge interactions of the peptide with the negative ganglioside do not appear to be a critical determinant of binding. Rather, an aggregation-suppressing amino acid composition and linker between the fluorophore and the peptide are required for optimum binding of the SBD to ganglioside-containing supported lipid bilayer surfaces, as well as for interaction with the membrane. Interestingly, the strength of interactions with ganglioside-containing artificial membranes is mirrored in the diffusion behavior by FCS on cell membranes, with stronger binders displaying similar characteristic diffusion profiles. Our findings indicate that for aggregation-prone peptides, aggregation occurs upon contact with the cell membrane, and rather than giving a stronger interaction with the membrane, aggregation is accompanied by weaker binding and complex diffusion profiles indicative of heterogeneous

  2. PEG-stabilized core-shell nanoparticles: impact of linear versus dendritic polymer shell architecture on colloidal properties and the reversibility of temperature-induced aggregation.

    Science.gov (United States)

    Gillich, Torben; Acikgöz, Canet; Isa, Lucio; Schlüter, A Dieter; Spencer, Nicholas D; Textor, Marcus

    2013-01-22

    Superparamagnetic iron oxide nanoparticles (SPIONs) have been widely used experimentally and also clinically tested in diverse areas of biology and medicine. Applications include magnetic resonance imaging, cell sorting, drug delivery, and hyperthermia. Physicochemical surface properties are particularly relevant in the context of achieving high colloidal nanoparticle (NP) stability and preventing agglomeration (particularly challenging in biological fluids), increasing blood circulation time, and possibly targeting specific cells or tissues through the presentation of bioligands. Traditionally, NP surfaces are sterically stabilized with hydrophilic polymeric matrices, such as dextran or linear poly(ethylene glycol) brushes. While dendrimers have found applications as drug carriers, dispersants with dendritic ("dendrons") or hyperbranched structures have been comparatively neglected despite their unique properties, such as a precisely defined molecular structure and the ability to present biofunctionalities at high density at the NP periphery. This work covers the synthesis of SPIONs and their stabilization based on poly(ethylene glycol) (PEG) and oligo(ethylene glycol) (OEG) chemistry and compares the physicochemical properties of NPs stabilized with linear and dendritic macromolecules of comparable molecular weight. The results highlight the impact of the polymeric interface architecture on solubility, colloidal stability, hydrodynamic radius, and thermoresponsive behavior. Dendron-stabilized NPs were found to provide excellent colloidal stability, despite a smaller hydrodynamic radius and lower degree of soft shell hydration compared to linear PEG analogues. Moreover, for the same grafting density and molecular weight of the stabilizers, OEG dendron-stabilized NPs show a reversible temperature-induced aggregation behavior, in contrast to the essentially irreversible aggregation and sedimentation observed for the linear PEG analogues. This new class of

  3. Reinjury risk of nano-calcium oxalate monohydrate and calcium oxalate dihydrate crystals on injured renal epithelial cells: aggravation of crystal adhesion and aggregation

    Science.gov (United States)

    Gan, Qiong-Zhi; Sun, Xin-Yuan; Bhadja, Poonam; Yao, Xiu-Qiong; Ouyang, Jian-Ming

    2016-01-01

    Background Renal epithelial cell injury facilitates crystal adhesion to cell surface and serves as a key step in renal stone formation. However, the effects of cell injury on the adhesion of nano-calcium oxalate crystals and the nano-crystal-induced reinjury risk of injured cells remain unclear. Methods African green monkey renal epithelial (Vero) cells were injured with H2O2 to establish a cell injury model. Cell viability, superoxide dismutase (SOD) activity, malonaldehyde (MDA) content, propidium iodide staining, hematoxylin–eosin staining, reactive oxygen species production, and mitochondrial membrane potential (Δψm) were determined to examine cell injury during adhesion. Changes in the surface structure of H2O2-injured cells were assessed through atomic force microscopy. The altered expression of hyaluronan during adhesion was examined through laser scanning confocal microscopy. The adhesion of nano-calcium oxalate monohydrate (COM) and calcium oxalate dihydrate (COD) crystals to Vero cells was observed through scanning electron microscopy. Nano-COM and COD binding was quantitatively determined through inductively coupled plasma emission spectrometry. Results The expression of hyaluronan on the cell surface was increased during wound healing because of Vero cell injury. The structure and function of the cell membrane were also altered by cell injury; thus, nano-crystal adhesion occurred. The ability of nano-COM to adhere to the injured Vero cells was higher than that of nano-COD crystals. The cell viability, SOD activity, and Δψm decreased when nano-crystals attached to the cell surface. By contrast, the MDA content, reactive oxygen species production, and cell death rate increased. Conclusion Cell injury contributes to crystal adhesion to Vero cell surface. The attached nano-COM and COD crystals can aggravate Vero cell injury. As a consequence, crystal adhesion and aggregation are enhanced. These findings provide further insights into kidney stone

  4. Effect of phosphate buffer on aggregation kinetics of citrate-coated silver nanoparticles induced by monovalent and divalent electrolytes.

    Science.gov (United States)

    Afshinnia, K; Baalousha, M

    2017-03-01

    The attachment efficiency (α) is an important parameter that can be used to characterize nanoparticle (NPs) aggregation behavior and has been a topic of discussion of several papers in the past few years. The importance of α is because it is one of the key parameters that can be used to model NP environmental fate and behavior. This study uses UV-vis and laser Doppler electrophoresis to monitor the aggregation behavior of citrate-coated silver nanoparticles (cit-AgNPs) induced by Na(+) and Ca(2+) as counter ions in the presence and absence of Suwannee River fulvic acid (SRFA) as a surrogate of natural organic matter and different concentrations of phosphate buffer (0-1mM). Results demonstrate that phosphate buffer, which serves to maintain pH nearly constant over the course of a reaction, is an important determinant of NP aggregation behavior. Increasing phosphate buffer concentration results in a decrease in the critical coagulation concentrations (CCC) of cit-AgNPs to lower counter ion concentration and an increase of α at the same counter ion concentration, both in the absence and presence of SRFA. SRFA stabilizes AgNPs and increases the CCC to higher counter ion concentrations. The outcome of this study can be used to rationalize the variation in α and CCC values reported in the literature for NPs with similar physicochemical properties, where different α and CCC values are reported when different types of buffers and buffer concentrations are used in different studies.

  5. Simulation of Deformation and Aggregation of Two Red Blood Cells in a Stenosed Microvessel by Dissipative Particle Dynamics.

    Science.gov (United States)

    Xiao, Lanlan; Liu, Yang; Chen, Shuo; Fu, Bingmei

    2016-12-01

    The motion of two red blood cells in a stenosed microvessel was simulated using dissipative particle dynamics. The effects of intercellular interaction, red blood cell deformability and the initial cell orientation on the deformation and aggregation of the RBCs and on the flow resistance were investigated. The red blood cell membrane was treated as a three-dimensional coarse-grained network model and the intercellular interaction was modeled by the Morse potential based on a depletion-mediated assumption. It is shown that the flow resistance increases dramatically when the red blood cells enter into the stenosis and decreases rapidly as RBCs move away from the stenosis. Particularly, for a pair of stiffer red blood cells with the initial inclination angle of 90°, the maximum value of the flow resistance is larger; while a higher flow resistance can also come from a stronger aggregation. For a pair of stiffer red blood cells moving parallel to the main flow, when their positions are closer to the vessel wall at the upstream of the stenosis, the flow resistance increases due to the migration to the vessel center at the stenosis. In addition, for a pair of red blood cells with the initial inclination angle of 0°, the flow resistance from the aggregate formed by a pair of red blood cells with a larger deformation is higher.

  6. The Kinetics of Dislocation Loop Formation in Ferritic Alloys Through the Aggregation of Irradiation Induced Defects

    Science.gov (United States)

    Kohnert, Aaron Anthony

    The mechanical properties of materials are often degraded over time by exposure to irradiation environments, a phenomenon that has hindered the development of multiple nuclear reactor design concepts. Such property changes are the result of microstructural changes induced by the collision of high energy particles with the atoms in a material. The lattice defects generated in these recoil events migrate and interact to form extended damage structures. This study has used theoretical models based on the mean field chemical reaction rate theory to analyze the aggregation of isolated lattice defects into larger microstructural features that are responsible for long term property changes, focusing on the development of black dot damage in ferritic iron based alloys. The purpose of such endeavors is two-fold. Primarily, such models explain and quantify the processes through which these microstructures form. Additionally, models provide insight into the behavior and properties of the point defects and defect clusters which drive general microstructural evolution processes. The modeling effort presented in this work has focused on physical fidelity, drawing from a variety of sources of information to characterize the unobservable defect generation and agglomeration processes that give rise to the observable features reported in experimental data. As such, the models are based not solely on isolated point defect creation, as is the case with many older rate theory approaches, but instead on realistic estimates of the defect cluster population produced in high energy cascade damage events. Experimental assessments of the microstructural changes evident in transmission electron microscopy studies provide a means to measure the efficacy of the kinetic models. Using common assumptions of the mobility of defect clusters generated in cascade damage conditions, an unphysically high density of damage features develops at the temperatures of interest with a temperature dependence

  7. Zinc induces unfolding and aggregation of dimeric arginine kinase by trapping reversible unfolding intermediate.

    Science.gov (United States)

    Liu, Taotao; Wang, Xicheng

    2010-11-01

    Arginine kinase plays an important role in the cellular energy metabolism of invertebrates. Dimeric arginine kinase (dAK) is unique in some marine invertebrates. The effects of Zn²(+) on the unfolding and aggregation of dAK from the sea cucumber Stichopus japonicus were investigated. Our results indicated that Zn²(+) caused dAK inactivation accompanied by conformational unfolding, the exposure of hydrophobic surface, and aggregation. Kinetic studies showed the inactivation and unfolding of dAK followed biphasic kinetic courses. Zn²(+) can affect unfolding and refolding of dAK by trapping the reversible intermediate. Our study provides important information regarding the effect of Zn²(+) on metabolic enzymes in marine invertebrates.

  8. α-Synuclein-induced Aggregation of Cytoplasmic Vesicles in Saccharomyces cerevisiae

    OpenAIRE

    Soper, James H.; Roy, Subhojit; Stieber, Anna; Lee, Eliza; Wilson, Robert B.; Trojanowski, John Q.; Burd, Christopher G.; Lee, Virginia M.-Y.

    2008-01-01

    Aggregated α-synuclein (α-syn) fibrils form Lewy bodies (LBs), the signature lesions of Parkinson's disease (PD) and related synucleinopathies, but the pathogenesis and neurodegenerative effects of LBs remain enigmatic. Recent studies have shown that when overexpressed in Saccharomyces cerevisiae, α-syn localizes to plasma membranes and forms cytoplasmic accumulations similar to human α-syn inclusions. However, the exact nature, composition, temporal evolution, and underlying mechanisms of ye...

  9. Role of Au nanoparticle aggregation in laser induced anisotropy of ITO transparent substrates

    Energy Technology Data Exchange (ETDEWEB)

    Kityk, I.V., E-mail: iwank74@gmail.com [Electrical Engineering Department, Czetochowa University Technology, Armii Krajowej 17, Czestochowa (Poland); Ebothe, J.; Bercu, N-B. [Laboratoire de Recherche en Nanosciences, E.A. 4682, Université de Reims, 21 rue Clément Ader, 51685 Reims cedex 02 (France); Aziz, Md. Abdul [Center of Research Excellence in Nanotechnology, King Fahd University of Petroleum and Minerals, Dhahran 31261 (Saudi Arabia); Oyama, Munetaka [Department of Material Chemistry, Graduate School of Engineering, Kyoto University, Nishikyo-ku, Kyoto 615-8520 (Japan)

    2014-02-05

    Highlights: • Principal role of Au NP surface aggregation on the output photoinduced birefringence is shown. • The higher changes are obtained for the samples with sizes 30 nm. • The process is slowly relaxed. -- Abstract: A principal possibility to achieve the photoinduced anisotropy in the Au NP deposited onto the ITO substrate is experimentally shown. The sizes of the Au NP forming the corresponding nanocomposites were 20 nm, 30 nm and 40 nm. As a photoinducing light source we have used two coherent beam originating from the Er:glass laser generating at 1540 nm with frequency repetition 15 Hz as well as its second harmonic doubled frequency signal at 770 nm. The effect is sensitive to the angle between the two laser beams as well as to the Au NP sizes, inter-particle distances and topology connected with their aggregation. The effect shows slow relaxation to the initial state. The optimal conditions are achieved for nanocomposites formed by 30 nm despite the expected 20 nm. This one may be caused by crucial role of the partial aggregation which even changes the effective grain sizes. The contribution of the dipole–dipole as well as quadrupole–dipole interactions to the changes of the anisotropy is discussed. The excitation is far from the resonance which allow to predict that effective role play overlap with nanotrapping levels. So principal role may belongs to surface topology and which is studied using the birefringence directly connected with the anisotropy.

  10. Amino acid induced fractal aggregation of gold nanoparticles: Why and how.

    Science.gov (United States)

    Doyen, Matthieu; Goole, Jonathan; Bartik, Kristin; Bruylants, Gilles

    2016-02-15

    Gold colloids are the object of many studies as they are reported to have potential biological sensing, imaging and drug delivery applications. In the presence of certain amino acids the aggregation of the gold nanoparticles into linear structures is observed, as highlighted by the appearance of a second plasmon band in the UV-Vis spectra of the colloid. The mechanism behind this phenomenon is still under debate. In order to help elucidate this issue, the interaction between gold colloids and different amino acids, modified amino acids and molecules mimicking their side-chain was monitored by UV-Vis absorption, DLS and TEM. The results show that phenomenon can be rationalized in terms of the Diffusion Limited Colloid Aggregation (DLCA) model which gives rise to the fractal aggregation colloids. The global charge of the compound, which influences the ionic strength of the solution, and the ease with which the compound can interact with the GNPs and affect their surface potential, are, the two parameters which control the DLCA regime. Calculations based on the Derjaguin, Landau, Verwey and Overbeek (DLVO) theory confirm all the experimental observations.

  11. Predictive response surface model for heat-induced rheological changes and aggregation of whey protein concentrate.

    Science.gov (United States)

    Alvarez, Pedro A; Emond, Charles; Gomaa, Ahmed; Remondetto, Gabriel E; Subirade, Muriel

    2015-02-01

    Whey proteins are now far more than a by-product of cheese processing. In the last 2 decades, food manufacturers have developed them as ingredients, with the dairy industry remaining as a major user. For many applications, whey proteins are modified (denatured) to alter their structure and functional properties. The objective of this research was to study the influence of 85 to 100 °C, with protein concentration of 8% to 12%, and treatment times of 5 to 30 min, while measuring rheological properties (storage modulus, loss modulus, and complex viscosity) and aggregation (intermolecular beta-sheet formation) in dispersions of whey protein concentrate (WPC). A Box-Behnken Response Surface Methodology modeled the heat denaturation of liquid sweet WPC at 3 variables and 3 levels. The model revealed a very significant fit for viscoelastic properties, and a lesser fit for protein aggregation, at temperatures not previously studied. An exponential increase of rheological parameters was governed by protein concentration and temperature, while a modest linear relationship of aggregation was governed by temperature. Models such as these can serve as valuable guides to the ingredient and dairy industries to develop target products, as whey is a major ingredient in many functional foods.

  12. Varicella-zoster virus induces the formation of dynamic nuclear capsid aggregates

    Energy Technology Data Exchange (ETDEWEB)

    Lebrun, Marielle [University of Liege (ULg), GIGA-Infection Immunity and Inflammation, Laboratory of Virology and Immunology, Liege (Belgium); Thelen, Nicolas; Thiry, Marc [University of Liege (ULg), GIGA-Neurosciences, Laboratory of Cellular and Tissular Biology, Liege (Belgium); Riva, Laura; Ote, Isabelle; Condé, Claude; Vandevenne, Patricia [University of Liege (ULg), GIGA-Infection Immunity and Inflammation, Laboratory of Virology and Immunology, Liege (Belgium); Di Valentin, Emmanuel [University of Liege (ULg), GIGA-Viral Vectors Platform, Liege (Belgium); Bontems, Sébastien [University of Liege (ULg), GIGA-Infection Immunity and Inflammation, Laboratory of Virology and Immunology, Liege (Belgium); Sadzot-Delvaux, Catherine, E-mail: csadzot@ulg.ac.be [University of Liege (ULg), GIGA-Infection Immunity and Inflammation, Laboratory of Virology and Immunology, Liege (Belgium)

    2014-04-15

    The first step of herpesviruses virion assembly occurs in the nucleus. However, the exact site where nucleocapsids are assembled, where the genome and the inner tegument are acquired, remains controversial. We created a recombinant VZV expressing ORF23 (homologous to HSV-1 VP26) fused to the eGFP and dually fluorescent viruses with a tegument protein additionally fused to a red tag (ORF9, ORF21 and ORF22 corresponding to HSV-1 UL49, UL37 and UL36). We identified nuclear dense structures containing the major capsid protein, the scaffold protein and maturing protease, as well as ORF21 and ORF22. Correlative microscopy demonstrated that the structures correspond to capsid aggregates and time-lapse video imaging showed that they appear prior to the accumulation of cytoplasmic capsids, presumably undergoing the secondary egress, and are highly dynamic. Our observations suggest that these structures might represent a nuclear area important for capsid assembly and/or maturation before the budding at the inner nuclear membrane. - Highlights: • We created a recombinant VZV expressing the small capsid protein fused to the eGFP. • We identified nuclear dense structures containing capsid and procapsid proteins. • Correlative microscopy showed that the structures correspond to capsid aggregates. • Procapsids and partial capsids are found within the aggregates of WT and eGFP-23 VZV. • FRAP and FLIP experiments demonstrated that they are dynamic structures.

  13. Effect of alginate on the aggregation kinetics of copper oxide nanoparticles (CuO NPs): bridging interaction and hetero-aggregation induced by Ca(2.).

    Science.gov (United States)

    Miao, Lingzhan; Wang, Chao; Hou, Jun; Wang, Peifang; Ao, Yanhui; Li, Yi; Lv, Bowen; Yang, Yangyang; You, Guoxiang; Xu, Yi

    2016-06-01

    The stability of CuO nanoparticles (NPs) is expected to play a key role in the environmental risk assessment of nanotoxicity in aquatic systems. In this study, the effect of alginate (model polysaccharides) on the stability of CuO NPs in various environmentally relevant ionic strength conditions was investigated by using time-resolved dynamic light scattering. Significant aggregation of CuO NPs was observed in the presence of both monovalent and divalent cations. The critical coagulation concentrations (CCC) were 54.5 and 2.9 mM for NaNO3 and Ca(NO3)2, respectively. The presence of alginate slowed nano-CuO aggregation rates over the entire NaNO3 concentration range due to the combined electrostatic and steric effect. High concentrations of Ca(2+) (>6 mM) resulted in stronger adsorption of alginate onto CuO NPs; however, enhanced aggregation of CuO NPs occurred simultaneously under the same conditions. Spectroscopic analysis revealed that the bridging interaction of alginate with Ca(2+) might be an important mechanism for the enhanced aggregation. Furthermore, significant coagulation of the alginate molecules was observed in solutions of high Ca(2+) concentrations, indicating a hetero-aggregation mechanism between the alginate-covered CuO NPs and the unabsorbed alginate. These results suggested a different aggregation mechanism of NPs might co-exist in aqueous systems enriched with natural organic matter, which should be taken into consideration in future studies. Graphical abstract Hetero-aggregation mechanism of CuO nanoparticles and alginate under high concentration of Ca(2.)

  14. Conservation of root regeneration potential of cell aggregates from horseradish hairy roots used as artificial seeds

    Energy Technology Data Exchange (ETDEWEB)

    Repunte, V.; Taya, M.; Tone, S. [Osaka University, Osaka (Japan). Faculty of Engineering Science

    1996-10-20

    The effects of water content in agar gel used as a medium and oxygen level in the gas phase on the adventitious root regeneration of cell aggregates (CA) derived from horseradish hairy roots were investigated in cultures at 25{degree}C. The number of roots emerging from CA was highly dependent on water content of the agar gel and no root regeneration was observed at a gel water content of 66% during culture time of 20 days. CA root regeneration was suppressed when the CA were kept for 20 days in an atmosphere oxygen composition of 10%, but was restored upon transfer of the CA to normal atmosphere of 21% oxygen. On the basis of these findings, an artificial seed was proposed using the CA as a cell inclusion material encapsulated in alginate gels covered with coats. From the perspective of conserving the root regeneration potential of the CA by preventing the drying of alginate gel while keeping oxygen availability to the CA, different coating materials of ethylene vinyl acetate acrylic acid terpolymer, paraffin and polyorganosiloxane were tested. Paraffin was selected as a suitable coating material because of its efficient drying tolerance and adequate permeability to oxygen. A regeneration efficiency of 90% could be obtained from the CA, stored in alginate gel covered with a paraffin coating of 0.40 mm thickness at 25{degree}C for 60 days in air, when sucrose concentration in the gel was over 240 mol m{sup -3}. 25 refs., 8 figs.

  15. A novel method for study of the aggregation of protein induced by metal ion aluminum(III) using resonance Rayleigh scattering technique

    Science.gov (United States)

    Long, Xiufen; Zhang, Caihua; Cheng, Jiongjia; Bi, Shuping

    2008-01-01

    We present a novel method for the study of the aggregation of protein induced by metal ion aluminum(III) using resonance Rayleigh scattering (RRS) technique. In neutral Tris-HCl medium, the effect of this aggregation of protein results in the enhancement of RRS intensity and the relationship between the enhancement of the RRS signal and the Al concentration is nonlinear. On this basis, we established a new method for the determination of the critical induced-aggregation concentrations ( CCIAC) of metal ion Al(III) inducing the protein aggregation. Our results show that many factors, such as, pH value, anions, salts, temperature and solvents have obvious effects. We also studied the extent of aggregation and structural changes using ultra-violet spectrometry, protein intrinsic fluorescence and circular dichroism to further understand the exact mechanisms of the aggregation characteristics of proteins induced by metal ion Al(III) at the molecular level, to help us to develop effective methods to investigate the toxicity of metal ion Al, and to provide theoretical and quantitative evidences for the development of appropriate treatments for neurodementia such as Parkinson's disease, Alzheimer's disease and dementia related to dialysis.

  16. Physical enviroment of 2-D animal cell aggregates formed in a short pathlength ultrasound standing wave trap.

    Science.gov (United States)

    Bazou, Despina; Kuznetsova, Larisa A; Coakley, W Terence

    2005-03-01

    2-D mammalian cell aggregates can be formed and levitated in a 1.5 MHz single half wavelength ultrasound standing wave trap. The physical environment of cells in such a trap has been examined. Attention was paid to parameters such as temperature, acoustic streaming, cavitation and intercellular forces. The extent to which these factors might be intrusive to a neural cell aggregate levitated in the trap was evaluated. Neural cells were exposed to ultrasound at a pressure amplitude of 0.54 MPa for 30 s; a small aggregate had been formed at the center of the trap. The pressure amplitude was then decreased to 0.27 MPa for 2 min, at which level the aggregation process continued at a slower rate. The pressure amplitude was then decreased to 0.06 MPa for 1 h. Temperature measurements that were conducted in situ with a 200 microm thermocouple over a 30 min period showed that the maximum temperature rise was less than 0.5 K. Acoustic streaming was measured by the particle image velocimetry method (PIV). It was shown that the hydrodynamic stress imposed on cells by acoustic streaming is less than that imposed by gentle preparative centrifugation procedures. Acoustic spectrum analysis showed that cavitation activity does not occur in the cell suspensions sonicated at the above pressures. White noise was detected only at a pressure amplitude of 1.96 MPa. Finally, it was shown that the attractive acoustic force between ultrasonically agglomerated cells is small compared with the normal attractive van der Waals force that operates at close cell surface separations. It is concluded that the standing wave trap operates only to concentrate cells locally, as in tissue, and does not modify the in vitro expression of surface receptor interactions.

  17. Studies on the photophysical properties of 1,8-naphthalimide derivative and aggregation induced emission recognition for casein

    Energy Technology Data Exchange (ETDEWEB)

    Sun, Yang, E-mail: 66160692@qq.com [Department of Chemistry and Chemical Engineering, Xi' an University of Arts and Science, No. 168, Taibai South Road, Xi' an, Shaanxi 710065 (China); Liang, Xuhua; Fan, Jun [School of Chemical Engineering, Northwest University, No. 229, Taibai North Road, Xi' an, Shaanxi 710069 (China); Han, Quan, E-mail: xahanq@hotmail.com [Department of Chemistry and Chemical Engineering, Xi' an University of Arts and Science, No. 168, Taibai South Road, Xi' an, Shaanxi 710065 (China)

    2013-09-15

    A novel water-soluble 1,8-naphthalimide derivative 1, bearing two acetic carboxylic groups, exhibited fluorescent turn-on recognition for casein micelle based on the aggregation induced emission (AIE) character. The photophysical properties of 1 consisting of donor and acceptor units were investigated by the solvation effect. Changing from polar to non-polar solvent increased the solvent interaction; both the excitation and emission spectra were shifted to shorter wavelength and intensity decreased through taking advantage of twisted intramolecular charge transfer (TICT) and self-association fluorescence emission. Moreover, the red-shift and quenching in protic solvent were caused by the excited-state hydrogen bond strengthening effect. The density functional theory (DFT) and time dependent density functional theory (TDDFT) were used to obtain the most stable structure, electronic excitation energy, dipole moments and charge distribution. The AIE mechanism of 1 with casein micelle was due to 1 docked in the hydrophobic cavity between sub-micelles and bound with amino acid residues, resulting in the aggregation of 1 on the casein micelle surface and emission enhancement, based on which, a novel casein assay method was developed. The proposed method exhibited a good linear range from 0.1 to 10.5 μg mL{sup −1}, with the detection limit of 3.0 ng mL{sup −1}. Satisfactory reproducibility, reversibility and a short response time were realized. This method was applied for the determination of casein in milk powder samples, avoiding the interferences from other components and illegal additives in milk. -- Highlights: • A water-soluble 1,8-naphthalimide-based fluorescent probe 1 was synthesized. • Photophysical characterization of 1 was studied. • Aggregation induced emission enhancement of 1 with casein was investigated. • A novel casein quantification method was developed.

  18. Magnetic engineering of stable rod-shaped stem cell aggregates: circumventing the pitfall of self-bending.

    Science.gov (United States)

    Du, V; Fayol, D; Reffay, M; Luciani, N; Bacri, J-C; Gay, C; Wilhelm, C

    2015-02-01

    A current challenge for tissue engineering while restoring the function of diseased or damaged tissue is to customize the tissue according to the target area. Scaffold-free approaches usually yield spheroid shapes with the risk of necrosis at the center due to poor nutrient and oxygen diffusion. Here, we used magnetic forces developed at the cellular scale by miniaturized magnets to create rod-shaped aggregates of stem cells that subsequently matured into a tissue-like structure. However, during the maturation process, the tissue-rods spontaneously bent and coiled into sphere-like structures, triggered by the increasing cell-cell adhesion within the initially non-homogeneous tissue. Optimisation of the intra-tissular magnetic forces successfully hindered the transition, in order to produce stable rod-shaped stem cells aggregates.

  19. Creation of reduced fat foods: influence of calcium-induced droplet aggregation on microstructure and rheology of mixed food dispersions.

    Science.gov (United States)

    Wu, Bi-cheng; Degner, Brian; McClements, David Julian

    2013-12-15

    The impact of calcium-induced fat droplet aggregation on the microstructure and physicochemical properties of model mixed colloidal dispersions was investigated. These systems consisted of 2 wt% whey protein-coated fat droplets and 4 wt% modified starch granules heated to induce starch swelling (pH 7). Optical and confocal microscopy showed that the fat droplets were dispersed within the interstitial region between the swollen starch granules. The structural organisation of the fat droplets within these interstitial regions could be modulated by controlling the calcium concentration: (i) at a low calcium concentration the droplets were evenly distributed; (ii) at an intermediate calcium concentration they formed a layer around the starch granules; (iii) at a high calcium concentration they formed a network of aggregated droplets. Paste-like materials were produced when the fat droplets formed a three-dimensional network in the interstitial region. The properties of fat droplet-starch granule suspensions can be modulated by altering the electrostatic interactions to alter microstructure.

  20. In vitro platelet activation, aggregation and platelet-granulocyte complex formation induced by surface modified single-walled carbon nanotubes.

    Science.gov (United States)

    Fent, János; Bihari, Péter; Vippola, Minnamari; Sarlin, Essi; Lakatos, Susan

    2015-08-01

    Surface modification of single-walled carbon nanotubes (SWCNTs) such as carboxylation, amidation, hydroxylation and pegylation is used to reduce the nanotube toxicity and render them more suitable for biomedical applications than their pristine counterparts. Toxicity can be manifested in platelet activation as it has been shown for SWCNTs. However, the effect of various surface modifications on the platelet activating potential of SWCNTs has not been tested yet. In vitro platelet activation (CD62P) as well as the platelet-granulocyte complex formation (CD15/CD41 double positivity) in human whole blood were measured by flow cytometry in the presence of 0.1mg/ml of pristine or various surface modified SWCNTs. The effect of various SWCNTs was tested by whole blood impedance aggregometry, too. All tested SWCNTs but the hydroxylated ones activate platelets and promote platelet-granulocyte complex formation in vitro. Carboxylated, pegylated and pristine SWCNTs induce whole blood aggregation as well. Although pegylation is preferred from biomedical point of view, among the samples tested by us pegylated SWCNTs induced far the most prominent activation and a well detectable aggregation of platelets in whole blood.

  1. Dlgh1 coordinates actin polymerization, synaptic T cell receptor and lipid raft aggregation, and effector function in T cells.

    Science.gov (United States)

    Round, June L; Tomassian, Tamar; Zhang, Min; Patel, Viresh; Schoenberger, Stephen P; Miceli, M Carrie

    2005-02-07

    Lipid raft membrane compartmentalization and membrane-associated guanylate kinase (MAGUK) family molecular scaffolds function in establishing cell polarity and organizing signal transducers within epithelial cell junctions and neuronal synapses. Here, we elucidate a role for the MAGUK protein, Dlgh1, in polarized T cell synapse assembly and T cell function. We find that Dlgh1 translocates to the immune synapse and lipid rafts in response to T cell receptor (TCR)/CD28 engagement and that LckSH3-mediated interactions with Dlgh1 control its membrane targeting. TCR/CD28 engagement induces the formation of endogenous Lck-Dlgh1-Zap70-Wiskott-Aldrich syndrome protein (WASp) complexes in which Dlgh1 acts to facilitate interactions of Lck with Zap70 and WASp. Using small interfering RNA and overexpression approaches, we show that Dlgh1 promotes antigen-induced actin polymerization, synaptic raft and TCR clustering, nuclear factor of activated T cell activity, and cytokine production. We propose that Dlgh1 coordinates TCR/CD28-induced actin-driven T cell synapse assembly, signal transduction, and effector function. These findings highlight common molecular strategies used to regulate cell polarity, synapse assembly, and transducer organization in diverse cellular systems.

  2. Early stages of salmon calcitonin aggregation: effect induced by ageing and oxidation processes in water and in the presence of model membranes.

    Science.gov (United States)

    Gaudiano, Maria Cristina; Colone, Marisa; Bombelli, Cecilia; Chistolini, Pietro; Valvo, Luisa; Diociaiuti, Marco

    2005-06-30

    The natural ageing- and hydrogen peroxide-induced aggregation of salmon calcitonin were studied in water and in the presence of dipalmitoylphosphatidylcholine (DPPC) liposomes. The early stages of the aggregation process at low protein concentration were investigated by means of Circular Dichroism spectroscopy (CD) and conventional and immunogold labelling Transmission Electron Microscopy (TEM). In buffered water solution, salmon calcitonin showed a two-stage conformational variation related to fibril formation and phase-separation of larger aggregates. A first stage, characterised by small conformational changes but a decrease in dichroic band intensity, was followed by a second stage, 6 days after, leading to higher conformational variations and aggregations. Salmon calcitonin showed a distinct modification in the secondary structure and aggregate morphology in the presence of hydrogen peroxide with respect to natural ageing, indicating that the two aggregation processes (natural and chemical-induced) followed a distinct mechanism. The oxidised forms of the peptide were separated by liquid chromatography. The same study was performed in the presence of DPPC liposomes. The results obtained by conventional and immunogold labelling TEM evidenced that salmon calcitonin in buffered water solution essentially does not enter the liposomes but forms around them a fibril network characterised by the same conformational changes after 6 days. The oxidised sample in the presence of liposomes showed a "fibrils hank", separated from liposomes. The presence of liposomes did not affect either the aggregation or the conformational modifications yet observed by TEM and CD in water solution.

  3. Root Cause Analysis of Tungsten-Induced Protein Aggregation in Pre-filled Syringes.

    Science.gov (United States)

    Liu, Wei; Swift, Rob; Torraca, Gianni; Nashed-Samuel, Yasser; Wen, Zai-Qing; Jiang, Yijia; Vance, Aylin; Mire-Sluis, Anthony; Freund, Erwin; Davis, Janice; Narhi, Linda

    2010-01-01

    Particles isolated from a pre-filled syringe containing a protein-based solution were identified as aggregated protein and tungsten. The origin of the tungsten was traced to the tungsten pins used in the supplier's syringe barrel forming process. A tungsten recovery study showed that the vacuum stopper placement process has a significant impact on the total amount of tungsten in solutions. The air gap formed in the syringe funnel area (rich in residual tungsten) becomes accessible to solutions when the vacuum is pulled. Leachable tungsten deposits that were not removed by the supplier's wash process are concentrated in this small area. Extraction procedures used to measure residual tungsten in empty syringes would under-report the tungsten quantity unless the funnel area is wetted during the extraction. Improved syringe barrel forming and washing processes at the supplier have lowered the residual tungsten content and significantly reduced the risk of protein aggregate formation. This experience demonstrates that packaging component manufacturing processes, which are outside the direct control of drug manufacturers, can have an impact on the drug product quality. Thus close technical communication with suppliers of product contact components plays an important role in making a successful biotherapeutic.

  4. Influence of Mechanical Cell Salvage on Red Blood Cell Aggregation, Deformability, and 2,3-Diphosphoglycerate in Patients Undergoing Cardiac Surgery With Cardiopulmonary Bypass

    NARCIS (Netherlands)

    Gu, Y. John; Vermeijden, Wytze J.; de Vries, Adrianus J.; Hagenaars, J. Ans M.; Graaff, Reindert; van Oeveren, Willem

    2008-01-01

    Background. Mechanical cell salvage is increasingly used during cardiac surgery. Although this procedure is considered safe, it is unknown whether it affects the red blood cell (RBC) function, especially the RBC aggregation, deformability, and the contents of 2,3-diphosphoglycerate (2,3-DPG). This s

  5. From single cell mechanics and intercellular forces to collective aggregate dynamics Individual cell-based modeling of cell cultures for tissue engineering

    OpenAIRE

    Smeets, Bart

    2016-01-01

    The mechanisms that biological cells exploit to organize themselves into multicellular aggregates and tissue-like structures are based on fundamental physical principles. Yet, the natural emergence of complexity in biological systems, while of great importance for many applications in biology and medicine, is still poorly understood. For this, mathematical models can be of great help by identifying key components and mechanisms that govern a system, and based on these, predict the inception o...

  6. The use of quartz crystal microbalance with dissipation (QCM-D for studying nanoparticle-induced platelet aggregation

    Directory of Open Access Journals (Sweden)

    Santos-Martinez MJ

    2012-01-01

    Full Text Available Maria Jose Santos-Martinez1–3, Iwona Inkielewicz-Stepniak1,4, Carlos Medina1, Kamil Rahme5,6, Deirdre M D'Arcy1, Daniel Fox3, Justin D Holmes3,5, Hongzhou Zhang3, Marek Witold Radomski3,51School of Pharmacy and Pharmaceutical Sciences, 2School of Medicine, 3Center for Research on Adaptive Nanostructures and Nanodevices, Trinity College Dublin, Dublin, Ireland; 4Department of Medicinal Chemistry, Medical University of Gdansk, Gdansk, Poland; 5Materials and Supercritical Fluids Group, Department of Chemistry and the Tyndall National Institute, University College Cork, Cork, Ireland; 6Department of Sciences, Faculty of Natural and Applied Science, Notre Dame University, Zouk Mosbeh, LebanonAbstract: Interactions between blood platelets and nanoparticles have both pharmacological and toxicological significance and may lead to platelet activation and aggregation. Platelet aggregation is usually studied using light aggregometer that neither mimics the conditions found in human microvasculature nor detects microaggregates. A new method for the measurement of platelet microaggregation under flow conditions using a commercially available quartz crystal microbalance with dissipation (QCM-D has recently been developed. The aim of the current study was to investigate if QCM-D could be used for the measurement of nanoparticle-platelet interactions. Silica, polystyrene, and gold nanoparticles were tested. The interactions were also studied using light aggregometry and flow cytometry, which measured surface abundance of platelet receptors. Platelet activation was imaged using phase contrast and scanning helium ion microscopy. QCM-D was able to measure nanoparticle-induced platelet microaggregation for all nanoparticles tested at concentrations that were undetectable by light aggregometry and flow cytometry. Microaggregates were measured by changes in frequency and dissipation, and the presence of platelets on the sensor surface was confirmed and imaged by

  7. Bromelain proteases reduce human platelet aggregation in vitro, adhesion to bovine endothelial cells and thrombus formation in rat vessels in vivo.

    Science.gov (United States)

    Metzig, C; Grabowska, E; Eckert, K; Rehse, K; Maurer, H R

    1999-01-01

    The thiol protease, bromelain, an extract from pineapple stem, was suggested to have antithrombotic and anticoagulant activities in vivo. We studied the effects of bromelain on cell size distribution of isolated human platelets in vitro by Coulter Counter measurements. Preincubation of platelets with bromelain (10 micrograms/mL) completely prevented the thrombin (0.2 U/mL) induced platelet aggregation. Papain was less active in preventing platelet aggregation. In vitro, bromelain (0.1 microgram/mL) reduced the adhesion of bound, thrombin stimulated, fluorescent labeled platelets to bovine aorta endothelial cells. In addition, preincubation of platelets with bromelain, prior to thrombin, activation, reduced the platelet adhesion to the endothelial cells to the low binding value of unstimulated platelets. On the basis of mass concentrations, the proteases papain and trypsin were as effective as bromelain. Using a laser thrombosis model, the in vivo effects of orally and intraveneously applied bromelain on thrombus formation in rat mesenteric vessels were studied. Bromelain, orally applied at 60 mg/kg body weight, inhibited the thrombus formation in a time dependent manner, the maximum being after 2 hours in 11% of arterioles and 6% of venoles. Intravenous application at 30 mg/kg was slightly more active in reducing thrombus formation in arterioles (13%) and venoles (5%), suggesting that orally applied bromelain is biologically active. These results may help to explain some of the clinical effects observed after bromelain treatment in patients with thrombosis and related diseases.

  8. Reversal of a full-length mutant huntingtin neuronal cell phenotype by chemical inhibitors of polyglutamine-mediated aggregation

    Directory of Open Access Journals (Sweden)

    MacDonald Marcy E

    2005-01-01

    Full Text Available Abstract Background Huntington's disease (HD is an inherited neurodegenerative disorder triggered by an expanded polyglutamine tract in huntingtin that is thought to confer a new conformational property on this large protein. The propensity of small amino-terminal fragments with mutant, but not wild-type, glutamine tracts to self-aggregate is consistent with an altered conformation but such fragments occur relatively late in the disease process in human patients and mouse models expressing full-length mutant protein. This suggests that the altered conformational property may act within the full-length mutant huntingtin to initially trigger pathogenesis. Indeed, genotype-phenotype studies in HD have defined genetic criteria for the disease initiating mechanism, and these are all fulfilled by phenotypes associated with expression of full-length mutant huntingtin, but not amino-terminal fragment, in mouse models. As the in vitro aggregation of amino-terminal mutant huntingtin fragment offers a ready assay to identify small compounds that interfere with the conformation of the polyglutamine tract, we have identified a number of aggregation inhibitors, and tested whether these are also capable of reversing a phenotype caused by endogenous expression of mutant huntingtin in a striatal cell line from the HdhQ111/Q111 knock-in mouse. Results We screened the NINDS Custom Collection of 1,040 FDA approved drugs and bioactive compounds for their ability to prevent in vitro aggregation of Q58-htn 1–171 amino terminal fragment. Ten compounds were identified that inhibited aggregation with IC50 HdhQ111/Q111 striatal cells. Conclusions At least some compounds identified as aggregation inhibitors also prevent a neuronal cellular phenotype caused by full-length mutant huntingtin, suggesting that in vitro fragment aggregation can act as a proxy for monitoring the disease-producing conformational property in HD. Thus, identification and testing of compounds that

  9. pH-Induced aggregated melanin nanoparticles for photoacoustic signal amplification

    Science.gov (United States)

    Ju, Kuk-Youn; Kang, Jeeun; Pyo, Jung; Lim, Joohyun; Chang, Jin Ho; Lee, Jin-Kyu

    2016-07-01

    We present a new melanin-like nanoparticle (MelNP) and its performance evaluation results. This particle is proposed as an exogenous contrast agent for photoacoustic (PA) imaging. Conventional PA contrast agents are based on non-biological materials. In contrast, the MelNPs are organic nanoparticles inspired by natural melanin. Melanin is an endogenous chromophore that has the ability to produce a PA signal in vivo. The developed MelNPs are capable of aggregating with one another under mildly acidic conditions after introducing hydrolysis-susceptible citraconic amide on the surface of bare MelNPs. We ascertained that the physical aggregation of the MelNPs resulted in an increased PA signal strength in the near-infrared window of biological tissue (i.e., 700 nm) without absorption tuning. This phenomenon is likely because of the overlapping thermal fields of the developed MelNPs. The PA signal produced from the developed MelNPs, after exposure to mildly acidic conditions (i.e., pH 6), is 8.1 times stronger than under neutral conditions. This unique characteristic found in this study can be utilized in a practical strategy for highly sensitive in vivo cancer target imaging in response to its acidic microenvironment. This approach to amplify the PA response of MelNPs in clusters could accelerate the use of MelNPs as an alternative to non-biological nanoprobes, so that MelNPs may be applicable in PA imaging and functional PA imaging such as stimuli sensitive, multimodal, and theranostic imaging.We present a new melanin-like nanoparticle (MelNP) and its performance evaluation results. This particle is proposed as an exogenous contrast agent for photoacoustic (PA) imaging. Conventional PA contrast agents are based on non-biological materials. In contrast, the MelNPs are organic nanoparticles inspired by natural melanin. Melanin is an endogenous chromophore that has the ability to produce a PA signal in vivo. The developed MelNPs are capable of aggregating with one

  10. MULTICELLULAR-MEDIATED EXPRESSION OF P-GP AND MRP AND RELATIONSHIP WITH CELL CYCLE PROFILES IN HUMAN OVARIAN CANCER SK-OV-3ip1 MULTICELLULAR AGGREGATES

    Institute of Scientific and Technical Information of China (English)

    陈建利; 丰有吉; 张琴

    2001-01-01

    To investigate the expression of P-glycoprotein (P-gp) and multidrug resistance- associated protein (MRP) and the relationship with cell cycle profiles in ovarian cancer SK-OV-3ip1 multicellular aggregates. Methods: Liquid overlay system was employed to obtain multicellular aggregates. Expression of P-gp and MRP was detected with flow cytometry (FCM). Outer, intermediate and inner cells from multicellular aggregates were collected by layer-trypsinized method. Cell cycle profiles were also analyzed by FCM. Results: Compared with control cells, no expression of P-gp and MRP was detected in monolyer cells (P=0.128 and P=0.604), but expression of P-gp and MRP in aggregate cells was significantly elevated (P<0.01). P-gp expression in every layer cells was also obviously increased (P<0.01). Furthermore, P-gp expression in every layer cells was also obviously increased (P=0.071). Tendency to increased G0-G1 phase and reduced S phase cells existed from outer through intermediate to inner layers in multicellular aggregates but with no statistical difference. Cell percentages in G2-M phase also had no difference. However, compared with monolayer cells, cells in G0-G1 phase increased and cells in S and G2-M phases lowered significantly in every layer and in the whole multicellular aggregates. Expression elevation of P-gp and MRP was consistent with increased G0-G1 percentage in aggregate cells. Conclusion: Expression of P-gp and MRP increases in cells of SK-OV-3ip1 multicellular aggregates and is consistent with increased G0-G1 percentage, which implies the possible relationship between them and the possible role in multicellular-mediated drug resistance.

  11. The use of quartz crystal microbalance with dissipation (QCM-D) for studying nanoparticle-induced platelet aggregation

    Science.gov (United States)

    Santos-Martinez, Maria Jose; Inkielewicz-Stepniak, Iwona; Medina, Carlos; Rahme, Kamil; D’Arcy, Deirdre M; Fox, Daniel; Holmes, Justin D; Zhang, Hongzhou; Radomski, Marek Witold

    2012-01-01

    Interactions between blood platelets and nanoparticles have both pharmacological and toxicological significance and may lead to platelet activation and aggregation. Platelet aggregation is usually studied using light aggregometer that neither mimics the conditions found in human microvasculature nor detects microaggregates. A new method for the measurement of platelet microaggregation under flow conditions using a commercially available quartz crystal microbalance with dissipation (QCM-D) has recently been developed. The aim of the current study was to investigate if QCM-D could be used for the measurement of nanoparticle-platelet interactions. Silica, polystyrene, and gold nanoparticles were tested. The interactions were also studied using light aggregometry and flow cytometry, which measured surface abundance of platelet receptors. Platelet activation was imaged using phase contrast and scanning helium ion microscopy. QCM-D was able to measure nanoparticle-induced platelet microaggregation for all nanoparticles tested at concentrations that were undetectable by light aggregometry and flow cytometry. Microaggregates were measured by changes in frequency and dissipation, and the presence of platelets on the sensor surface was confirmed and imaged by phase contrast and scanning helium ion microscopy. PMID:22275839

  12. Boron-doped graphene quantum dots for selective glucose sensing based on the "abnormal" aggregation-induced photoluminescence enhancement.

    Science.gov (United States)

    Zhang, Li; Zhang, Zhi-Yi; Liang, Ru-Ping; Li, Ya-Hua; Qiu, Jian-Ding

    2014-05-06

    A hydrothermal approach for the cutting of boron-doped graphene (BG) into boron-doped graphene quantum dots (BGQDs) has been proposed. Various characterizations reveal that the boron atoms have been successfully doped into graphene structures with the atomic percentage of 3.45%. The generation of boronic acid groups on the BGQDs surfaces facilitates their application as a new photoluminescence (PL) probe for label free glucose sensing. It is postulated that the reaction of the two cis-diol units in glucose with the two boronic acid groups on the BGQDs surfaces creates structurally rigid BGQDs-glucose aggregates, restricting the intramolecular rotations and thus resulting in a great boost in the PL intensity. The present unusual "aggregation-induced PL increasing" sensing process excludes any saccharide with only one cis-diol unit, as manifested by the high specificity of BGQDs for glucose over its close isomeric cousins fructose, galactose, and mannose. It is believed that the doping of boron can introduce the GQDs to a new kind of surface state and offer great scientific insights to the PL enhancement mechanism with treatment of glucose.

  13. Drosophila UNC-45 prevents heat-induced aggregation of skeletal muscle myosin and facilitates refolding of citrate synthase

    Energy Technology Data Exchange (ETDEWEB)

    Melkani, Girish C.; Lee, Chi F.; Cammarato, Anthony [Department of Biology and the Molecular Biology Institute, San Diego State University, San Diego, CA 92182-4614 (United States); Bernstein, Sanford I., E-mail: sbernst@sciences.sdsu.edu [Department of Biology and the Molecular Biology Institute, San Diego State University, San Diego, CA 92182-4614 (United States)

    2010-05-28

    UNC-45 belongs to the UCS (UNC-45, CRO1, She4p) domain protein family, whose members interact with various classes of myosin. Here we provide structural and biochemical evidence that Escherichia coli-expressed Drosophila UNC-45 (DUNC-45) maintains the integrity of several substrates during heat-induced stress in vitro. DUNC-45 displays chaperone function in suppressing aggregation of the muscle myosin heavy meromyosin fragment, the myosin S-1 motor domain, {alpha}-lactalbumin and citrate synthase. Biochemical evidence is supported by electron microscopy, which reveals the first structural evidence that DUNC-45 prevents inter- or intra-molecular aggregates of skeletal muscle heavy meromyosin caused by elevated temperatures. We also demonstrate for the first time that UNC-45 is able to refold a denatured substrate, urea-unfolded citrate synthase. Overall, this in vitro study provides insight into the fate of muscle myosin under stress conditions and suggests that UNC-45 protects and maintains the contractile machinery during in vivo stress.

  14. Men with Sickle Cell Anemia and Priapism Exhibit Increased Hemolytic Rate, Decreased Red Blood Cell Deformability and Increased Red Blood Cell Aggregate Strength.

    Directory of Open Access Journals (Sweden)

    Kizzy-Clara Cita

    Full Text Available To investigate the association between priapism in men with sickle cell anemia (SCA and hemorheological and hemolytical parameters.Fifty-eight men with SCA (median age: 38 years were included; 28 who had experienced priapism at least once during their life (priapism group and 30 who never experienced this complication (control group. Twenty-two patients were treated with hydroxycarbamide, 11 in each group. All patients were at steady state at the time of inclusion. Hematological and biochemical parameters were obtained through routine procedures. The Laser-assisted Optical Rotational Cell Analyzer was used to measure red blood cell (RBC deformability at 30 Pa (ektacytometry and RBC aggregation properties (laser backscatter versus time. Blood viscosity was measured at a shear rate of 225 s-1 using a cone/plate viscometer. A principal component analysis was performed on 4 hemolytic markers (i.e., lactate dehydrogenase (LDH, aspartate aminotransferase (ASAT, total bilirubin (BIL levels and reticulocyte (RET percentage to calculate a hemolytic index.Compared to the control group, patients with priapism exhibited higher ASAT (p = 0.01, LDH (p = 0.03, RET (p = 0.03 levels and hemolytic indices (p = 0.02. Higher RBC aggregates strength (p = 0.01 and lower RBC deformability (p = 0.005 were observed in patients with priapism compared to controls. After removing the hydroxycarbamide-treated patients, RBC deformability (p = 0.01 and RBC aggregate strength (p = 0.03 were still different between the two groups, and patients with priapism exhibited significantly higher hemolytic indices (p = 0.01 than controls.Our results confirm that priapism in SCA is associated with higher hemolytic rates and show for the first time that this complication is also associated with higher RBC aggregate strength and lower RBC deformability.

  15. Soluble aggregates of the amyloid-β peptide are trapped by serum albumin to enhance amyloid-β activation of endothelial cells

    Directory of Open Access Journals (Sweden)

    Gonzalez-Velasquez Francisco J

    2009-04-01

    Full Text Available Abstract Background Self-assembly of the amyloid-β peptide (Aβ has been implicated in the pathogenesis of Alzheimer's disease (AD. As a result, synthetic molecules capable of inhibiting Aβ self-assembly could serve as therapeutic agents and endogenous molecules that modulate Aβ self-assembly may influence disease progression. However, increasing evidence implicating a principal pathogenic role for small soluble Aβ aggregates warns that inhibition at intermediate stages of Aβ self-assembly may prove detrimental. Here, we explore the inhibition of Aβ1–40 self-assembly by serum albumin, the most abundant plasma protein, and the influence of this inhibition on Aβ1–40 activation of endothelial cells for monocyte adhesion. Results It is demonstrated that serum albumin is capable of inhibiting in a dose-dependent manner both the formation of Aβ1–40 aggregates from monomeric peptide and the ongoing growth of Aβ1–40 fibrils. Inhibition of fibrillar Aβ1–40 aggregate growth is observed at substoichiometric concentrations, suggesting that serum albumin recognizes aggregated forms of the peptide to prevent monomer addition. Inhibition of Aβ1–40 monomer aggregation is observed down to stoichiometric ratios with partial inhibition leading to an increase in the population of small soluble aggregates. Such partial inhibition of Aβ1–40 aggregation leads to an increase in the ability of resulting aggregates to activate endothelial cells for adhesion of monocytes. In contrast, Aβ1–40 activation of endothelial cells for monocyte adhesion is reduced when more complete inhibition is observed. Conclusion These results demonstrate that inhibitors of Aβ self-assembly have the potential to trap small soluble aggregates resulting in an elevation rather than a reduction of cellular responses. These findings provide further support that small soluble aggregates possess high levels of physiological activity and underscore the importance of

  16. Detection of adenosine triphosphate through polymerization-induced aggregation of actin-conjugated gold/silver nanorods

    Science.gov (United States)

    Liao, Yu-Ju; Shiang, Yen-Chun; Chen, Li-Yi; Hsu, Chia-Lun; Huang, Chih-Ching; Chang, Huan-Tsung

    2013-11-01

    We have developed a simple and selective nanosensor for the optical detection of adenosine triphosphate (ATP) using globular actin-conjugated gold/silver nanorods (G-actin-Au/Ag NRs). By simply mixing G-actin and Au/Ag NRs (length ˜56 nm and diameter ˜12 nm), G-actin-Au/Ag NRs were prepared which were stable in physiological solutions (25 mM Tris-HCl, 150 mM NaCl, 5.0 mM KCl, 3.0 mM MgCl2 and 1.0 mM CaCl2; pH 7.4). Introduction of ATP into the G-actin-Au/Ag NR solutions in the presence of excess G-actin induced the formation of filamentous actin-conjugated Au/Ag NR aggregates through ATP-induced polymerization of G-actin. When compared to G-actin-modified spherical Au nanoparticles having a size of 13 nm or 56 nm, G-actin-Au/Ag NRs provided better sensitivity for ATP, mainly because the longitudinal surface plasmon absorbance of the Au/Ag NR has a more sensitive response to aggregation. This G-actin-Au/Ag NR probe provided high sensitivity (limit of detection 25 nM) for ATP with remarkable selectivity (>10-fold) over other adenine nucleotides (adenosine, adenosine monophosphate and adenosine diphosphate) and nucleoside triphosphates (guanosine triphosphate, cytidine triphosphate and uridine triphosphate). It also allowed the determination of ATP concentrations in plasma samples without conducting tedious sample pretreatments; the only necessary step was simple dilution. Our experimental results are in good agreement with those obtained from a commercial luciferin-luciferase bioluminescence assay. Our simple, sensitive and selective approach appears to have a practical potential for the clinical diagnosis of diseases (e.g. cystic fibrosis) associated with changes in ATP concentrations.

  17. Latent heat induced rotation limited aggregation in 2D ice nanocrystals

    CERN Document Server

    Bampoulis, Pantelis; Kooij, E Stefan; Lohse, Detlef; Zandvliet, Harold J W; Poelsema, Bene

    2016-01-01

    The basic science responsible for the fascinating shapes of ice crystals and snowflakes is still not understood. Insufficient knowledge of the interaction potentials and the lack of relevant experimental access to the growth process are to blame for this failure. Here, we study the growth of fractal nanostructures in a two-dimensional (2D) system, intercalated between mica and graphene. Based on our Scanning Tunneling Spectroscopy (STS) data we provide compelling evidence that these fractals are 2D ice. They grow while they are in material contact with the atmosphere at 20 $^{\\circ}$C and without significant thermal contact to the ambient. The growth is studied in-situ, in real time and space at the nanoscale. We find that the growing 2D ice nanocrystals assume a fractal shape, which is conventionally attributed to Diffusion Limited Aggregation (DLA). However, DLA requires a low mass density mother phase, in contrast to the actual currently present high mass density mother phase. Latent heat effects and conse...

  18. HIV transcription is induced in dying cells

    Energy Technology Data Exchange (ETDEWEB)

    Woloschak, G.E.; Chang-Liu, Chin-Mei [Argonne National Lab., IL (United States); Schreck, S. [Argonne National Lab., IL (United States)]|[Univ. of South Carolina, Columbia, SC (United States). Dept. of Chemistry; Panozzo, J. [Loyola Univ. Medical Center, Maywood, IL (United States); Libertin, C.R. [Loyola Univ. Medical Center, Maywood, IL (United States)

    1996-02-01

    Using HeLa cells stably transfected with an HIV-LTR-CAT construct, we demonstrated a peak in CAT induction that occurs in viable (but not necessarily cell-division-competent) cells 24 h following exposure to some cell-killing agents. {gamma} rays were the only cell-killing agent which did not induce HIV transcription; this can be attributed to the fact that {gamma}-ray-induced apoptotic death requires functional p53, which is not present in HeLa cells. For all other agents, HIV-LTR induction was dose-dependent and correlated with the amount of cell killing that occurred in the culture. Doses which caused over 99% cell killing induced HIV-LTR transcription maximally, demonstrating that cells that will go on to die by 14 days are the cells expressing HIV-LTR-CAT.

  19. Cell Communication during Aggregation and Development of the Cellular Slime Mould Distyostelium discoideum.

    Science.gov (United States)

    1985-01-01

    assistance, and love. With special feelings, I thank my beautiful daughters, Jennifer and Renata, for their many hugs and words of encouragement during times...cellularly throughout development (Bonner et al., 1969; Malkinson and Ashworth , 1972) and it has been suggested that cAMP may induce cells to enter...Axenic Liquid Media (Watts and Ashworth , 1970) Oxoid Bacteriological Peptone 14.3g Oxoid Yeast Extract 7.15g D-Glucose 15.4g Na .12H 20 1.28g KHt2P:O4

  20. Controlling solution-phase polymer aggregation with molecular weight and solvent additives to optimize polymer-fullerene bulk heterojunction solar cells

    KAUST Repository

    Bartelt, Jonathan A.

    2014-03-20

    The bulk heterojunction (BHJ) solar cell performance of many polymers depends on the polymer molecular weight (M n) and the solvent additive(s) used for solution processing. However, the mechanism that causes these dependencies is not well understood. This work determines how M n and solvent additives affect the performance of BHJ solar cells made with the polymer poly(di(2-ethylhexyloxy)benzo[1,2-b:4,5-b\\']dithiophene-co- octylthieno[3,4-c]pyrrole-4,6-dione) (PBDTTPD). Low M n PBDTTPD devices have exceedingly large fullerene-rich domains, which cause extensive charge-carrier recombination. Increasing the M n of PBDTTPD decreases the size of these domains and significantly improves device performance. PBDTTPD aggregation in solution affects the size of the fullerene-rich domains and this effect is linked to the dependency of PBDTTPD solubility on M n. Due to its poor solubility high M n PBDTTPD quickly forms a fibrillar polymer network during spin-casting and this network acts as a template that prevents large-scale phase separation. Furthermore, processing low M n PBDTTPD devices with a solvent additive improves device performance by inducing polymer aggregation in solution and preventing large fullerene-rich domains from forming. These findings highlight that polymer aggregation in solution plays a significant role in determining the morphology and performance of BHJ solar cells. The performance of poly(di(2-ethylhexyloxy) benzo[1,2-b:4,5-b\\']dithiophene-co-octylthieno[3,4-c]pyrrole-4,6-dione) (PBDTTPD) bulk heterojunction solar cells strongly depends on the polymer molecular weight, and processing these bulk heterojunctions with a solvent additive preferentially improves the performance of low molecular weight devices. It is demonstrated that polymer aggregation in solution significantly impacts the thin-film bulk heterojunction morphology and is vital for high device performance. © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  1. Glycosylated asterisks are among the most potent low valency inducers of Concanavalin A aggregation.

    Science.gov (United States)

    Sleiman, Mazen; Varrot, Annabelle; Raimundo, Jean-Manuel; Gingras, Marc; Goekjian, Peter G

    2008-12-28

    A new class of sulfurated, semi-rigid, radial and low-valent glycosylated asterisk ligands with potential dual function as ligand and probe has some of the highest inhibition potencies of Con A-induced hemagglutination, by using a cross-linking mechanism of Con A which amplifies the enhancement to near nanomolar concentrations with the alpha-d-mannose asterisk.

  2. UV-inducible cellular aggregation of the hyperthermophilic archaeon Sulfolobus solfataricus is mediated by pili formation

    NARCIS (Netherlands)

    Froels, Sabrina; Ajon, Malgorzata; Wagner, Michaela; Teichmann, Daniela; Zolghadr, Behnam; Folea, Mihaela; Boekema, Egbert J.; Driessen, Arnold J. M.; Schleper, Christa; Albers, Sonja-Verena

    2008-01-01

    The hyperthermophilic archaeon Sulfolobus solfataricus has been shown to exhibit a complex transcriptional response to UV irradiation involving 55 genes. Among the strongest UV-induced genes was a putative pili biogenesis operon encoding a potential secretion ATPase, two pre-pilins, a putative trans

  3. Aggregation induced Raman scattering of squaraine dye: Implementation in diagnosis of cervical cancer dysplasia by SERS imaging.

    Science.gov (United States)

    Narayanan, Nisha; Karunakaran, Varsha; Paul, Willi; Venugopal, Karunakaran; Sujathan, K; Kumar Maiti, Kaustabh

    2015-08-15

    The extent of squaraine dye aggregation that reflects on surface enhanced Raman signal scattering (SERS) intensity upon adsorption on nano-roughened gold surface has been investigated. Here we have synthesized a serious of six squaraine dyes consisting of two different electron donor moiety i.e. 1,1,2-trimethyl-1H-benzo[e]indole and 2-methylbenzo[d]thiazole which modulates the chemisorptions and hydrophobicity being designated as SQ1, SQ2, SQ3, SQ4, SQ5 and SQ6. Interestingly, SQ2 (mono lipoic acid appended), SQ5 and SQ6 (conjugated with hexyl and dodecyl side chain) squaraine derivatives having more tendency of aggregation in DMSO-water mixed solvent showed significant increase of Raman scattering in the fingerprint region when chemisorbed on spherical gold nanoparticles. Two sets of SERS nanotags were prepared with colloidal gold nanoparticle (Au-NPs size: 40 nm) by incorporating Raman reporters SQ2 and SQ5 followed by thiolated PEG encapsulation (SH-PEG, SH-PEG-COOH) denoted as AuNPs-SQ2-PEG and AuNPs-SQ5-PEG. Further conjugation of these nanotag with monoclonal antibodies specific to over expressed receptors, EGFR and p16/Ki-67 in cervical cancer cell, HeLa showed prominent SERS mapping intensity and selectivity towards cell surface and nucleus. The fast and accurate recognition obtained by antibody triggered SERS-nanotag has been compared with conventional time consuming immunocytochemistry technique which prompted us to extend further investigation using real patient cervical smear sample for a non-invasive, ultrafast and accurate diagnosis.

  4. Pulsed electric field (PEF)-induced aggregation between lysozyme, ovalbumin and ovotransferrin in multi-protein system.

    Science.gov (United States)

    Wu, Li; Zhao, Wei; Yang, Ruijin; Yan, Wenxu

    2015-05-15

    The aggregation of multi-proteins is of great interest in food processing and a good understanding of the formation of aggregates during PEF processing is needed for the application of the process to pasteurize protein-based foods. The aggregates formation of a multi-protein system (containing ovalbumin, ovotransferrin and lysozyme) was studied through turbidity, size exclusion chromatography and SDS-PAGE patterns for interaction studies and binding forces. Results from size exclusion chromatography indicated that there was no soluble aggregates formed during PEF processing. The existence of lysozyme was important to form insoluble aggregates in the chosen ovalbumin solution. The results of SDS-PAGE patterns indicated that lysozyme was prone to precipitate, and was relatively the higher component of aggregates. Citric acid could be effective in inhibiting lysozyme from interacting with other proteins during PEF processing. Blocking the free sulphydryl by N-ethylmaleimide (NEM) did not affect aggregation inhibition.

  5. Reinjury risk of nano-calcium oxalate monohydrate and calcium oxalate dihydrate crystals on injured renal epithelial cells: aggravation of crystal adhesion and aggregation

    Directory of Open Access Journals (Sweden)

    Gan QZ

    2016-06-01

    Full Text Available Qiong-Zhi Gan,1,2 Xin-Yuan Sun,1,2 Poonam Bhadja,1,2 Xiu-Qiong Yao,1,2 Jian-Ming Ouyang1,2 1Department of Chemistry, Jinan University, Guangzhou, People’s Republic of China; 2Institute of Biomineralization and Lithiasis Research, Jinan University, Guangzhou, People’s Republic of China Background: Renal epithelial cell injury facilitates crystal adhesion to cell surface and serves as a key step in renal stone formation. However, the effects of cell injury on the adhesion of nano-calcium oxalate crystals and the nano-crystal-induced reinjury risk of injured cells remain unclear.Methods: African green monkey renal epithelial (Vero cells were injured with H2O2 to establish a cell injury model. Cell viability, superoxide dismutase (SOD activity, malonaldehyde (MDA content, propidium iodide staining, hematoxylin–eosin staining, reactive oxygen species production, and mitochondrial membrane potential (ΔΨm were determined to examine cell injury during adhesion. Changes in the surface structure of H2O2-injured cells were assessed through atomic force microscopy. The altered expression of hyaluronan during adhesion was examined through laser scanning confocal microscopy. The adhesion of nano-calcium oxalate monohydrate (COM and calcium oxalate dihydrate (COD crystals to Vero cells was observed through scanning electron microscopy. Nano-COM and COD binding was quantitatively determined through inductively coupled plasma emission spectrometry.Results: The expression of hyaluronan on the cell surface was increased during wound healing because of Vero cell injury. The structure and function of the cell membrane were also altered by cell injury; thus, nano-crystal adhesion occurred. The ability of nano-COM to adhere to the injured Vero cells was higher than that of nano-COD crystals. The cell viability, SOD activity, and ΔΨm decreased when nano-crystals attached to the cell surface. By contrast, the MDA content, reactive oxygen species production

  6. Metformin induces apoptosis of pancreatic cancer cells

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    AIM: To assess the role and mechanism of mefformin in inducing apoptosis of pancreatic cancer cells. METHODS: The human pancreatic cancer cell lines ASPC-1, BxPc-3, PANC-1 and SW1990 were exposed to mefformin. The inhibition of cell proliferation and colony formation via apoptosis induction and S phase arrest in pancreatic cancer cell lines of mefformin was tested.RESULTS: In each pancreatic cancer cell line tested, metformin inhibited cell proliferation in a dose dependent manner in MTS (3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium assays). Flow cytometric analysis showed that metformin reduced the number of cells in G1 and increased the percentage of cells in S phase as well as the apoptotic fraction. Enzymelinked immunosorbent assay (EUSA) showed that metformin induced apaptosis in all pancreatic cancer cell lines. In Western blot studies, metformin induced oly-ADP-ribose polymerase(PARP) cleavage (an indicator of aspase activation) in all pancreatic cancer cell lines. The general caspase inhibitor (VAD-fmk) completely abolished metformin-induced PARP cleavage and apoptosis in ASPC-1 BxPc-3 and PANC-1, the caspase-8 specific inhibitor (IETD-fmk) and the caspase-9 specific inhibitor (LEHD-fmk) only partially abrogated metformin-induced apoptosis and PARP cleavage in BxPc-3 and PANC-1 cells. We also observed that metformin treatment ramatically reduced epidermal growth factor receptor (EGFR) and phosphorylated mitogen activated protein kinase (P-MAPK) in both a time- and dose-dependent manner in all cell lines tested.CONCLUSION: Metformin significantly inhibits cell proliferation and apoptosis in all pancreatic cell lines. And the metformin-induced apoptosis is associated with PARP leavage, activation of caspase-3, -8, and -9 in a time- and dose-dependent manner. Hence, both caspase-8 and -9-initiated apoptotic signaling pathways contribute to metforrnin-induced apoptosis in pancreatic cell lines.

  7. Symmetry recovery of cell-free layer after bifurcations of small arterioles in reduced flow conditions: effect of RBC aggregation.

    Science.gov (United States)

    Ng, Yan Cheng; Namgung, Bumseok; Tien, Sim Leng; Leo, Hwa Liang; Kim, Sangho

    2016-08-01

    Heterogeneous distribution of red blood cells (RBCs) in downstream vessels of arteriolar bifurcations can be promoted by an asymmetric formation of cell-free layer (CFL) in upstream vessels. Consequently, the CFL widths in subsequent downstream vessels become an important determinant for tissue oxygenation (O2) and vascular tone change by varying nitric oxide (NO) availability. To extend our previous understanding on the formation of CFL in arteriolar bifurcations, this study investigated the formation of CFL widths from 2 to 6 vessel-diameter (2D-6D) downstream of arteriolar bifurcations in the rat cremaster muscle (D = 51.5 ± 1.3 μm). As the CFL widths are highly influenced by RBC aggregation, the degree of aggregation was adjusted to simulate levels seen during physiological and pathological states. Our in vivo experimental results showed that the asymmetry of CFL widths persists along downstream vessels up to 6D from the bifurcating point. Moreover, elevated levels of RBC aggregation appeared to retard the recovery of CFL width symmetry. The required length of complete symmetry recovery was estimated to be greater than 11D under reduced flow conditions, which is relatively longer than interbifurcation distances of arterioles for vessel diameter of ∼50 μm. In addition, our numerical prediction showed that the persistent asymmetry of CFL widths could potentially result in a heterogeneous vasoactivity over the entire arteriolar network in such abnormal flow conditions.

  8. Extract from Aronia melanocarpa fruits potentiates the inhibition of platelet aggregation in the presence of endothelial cells

    Science.gov (United States)

    Luzak, Boguslawa; Golanski, Jacek; Rozalski, Marek; Krajewska, Urszula; Olas, Beata

    2010-01-01

    Introduction Some polyphenolic compounds extracted from Aronia melanocarpa fruits (AM) have been reported to be cardioprotective agents. In this study we evaluated the ability of AM extract to increase the efficacy of human umbilical vein endothelial cells (HUVECs) to inhibit platelet functions in vitro. Material and methods This study encompasses two models of monitoring platelet reactivity: optical aggregation and platelet degranulation (monitored as the surface CD62P expression) in PRP upon the stimulation with ADP. Results We observed that only at low concentrations (5 µg/ml) did AM extract significantly improve antiplatelet action of HUVECs towards ADP-activated platelets in the aggregation test. Conclusions It is concluded that the potentiating effect of AM extract on the endothelial cell-mediated inhibition of platelet aggregation clearly depends on the used concentrations of Aronia-derived active compounds. Therefore, despite these encouraging preliminary outcomes on the beneficial effects of AM extract polyphenols, more profound dose-effect studies should certainly be considered before the implementation of Aronia-originating compounds in antiplatelet therapy and the prevention of cardiovascular diseases. PMID:22371737

  9. Effect of Aggregated β-Amyloid (1-42 on Synaptic Plasticity of Hippocampal Dentate Gyrus Granule Cells in Vivo

    Directory of Open Access Journals (Sweden)

    Shirin Babri

    2012-12-01

    Full Text Available Introduction: Alzheimer’s disease (AD is a common neurodegenerative disorder in elderly people with an impairment of cognitive decline and memory loss. β-amyloid (Aβ as a potent neurotoxic peptide has a pivotal role in the pathogenesis of AD. This disease begins with impairment in synaptic functions before developing into later neuro­degeneration and neuronal loss. The aim of this study was to evaluate the synaptic plasticity and electrophysiological function of granule cells in hippocampal dentate gyrus (DG after intracerebroventricular (i.c.v. administration of aggregated Aβ (1-42 peptide in vivo. Methods: Animals were divided to control and Aβ (1-42 groups. Long-term potentia­tion (LTP in perforant path-DG synapses was assessed in order to investigate the effect of aggregated Aβ (1-42 on synaptic plasticity. Field excitatory post-synaptic potential (fEPSP slope and population spike (PS amplitude were measured. Results: Administration of Aβ (1-42 significantly decreased fEPSP slope and PS amplitude in Aβ (1-42 group comparing with the control group and had no effect on baseline activity of neurons. Conclusion: The present study indicates that administration of aggregated form of Aβ (1-42 into the lateral ventricle effectively inhibits LTP in granular cells of the DG in hippocampus in vivo.

  10. From the test tube to the cell: exploring the folding and aggregation of a beta-clam protein.

    Science.gov (United States)

    Ignatova, Zoya; Krishnan, Beena; Bombardier, Jeffrey P; Marcelino, Anna Marie C; Hong, Jiang; Gierasch, Lila M

    2007-01-01

    A crucial challenge in present biomedical research is the elucidation of how fundamental processes like protein folding and aggregation occur in the complex environment of the cell. Many new physico-chemical factors like crowding and confinement must be considered, and immense technical hurdles must be overcome in order to explore these processes in vivo. Understanding protein misfolding and aggregation diseases and developing therapeutic strategies to these diseases demand that we gain mechanistic insight into behaviors and misbehaviors of proteins as they fold in vivo. We have developed a fluorescence approach using FlAsH labeling to study the thermodynamics of folding of a model beta-rich protein, cellular retinoic acid binding protein (CRABP) in Escherichia coli cells. The labeling approach has also enabled us to follow aggregation of a modified version of CRABP and chimeras between CRABP and huntingtin exon 1 with its glutamine repeat tract. In this article, we review our recent results using FlAsH labeling to study in-vivo folding and present new observations that hint at fundamental differences between the thermodynamics and kinetics of protein folding in vivo and in vitro.

  11. Some Induced Correlated Aggregating Operators with Interval Grey Uncertain Linguistic Information and Their Application to Multiple Attribute Group Decision Making

    Directory of Open Access Journals (Sweden)

    Zu-Jun Ma

    2013-01-01

    Full Text Available We propose the interval grey uncertain linguistic correlated ordered arithmetic averaging (IGULCOA operator and the induced interval grey uncertain linguistic correlated ordered arithmetic averaging (I-IGULCOA operator based on the correlation properties of the Choquet integral and the interval grey uncertain linguistic variables to investigate the multiple attribute group decision making (MAGDM problems, in which both the attribute weights and the expert weights are correlative. Firstly, the relative concepts of interval grey uncertain linguistic variables are defined and the operation rules between the two interval grey uncertain linguistic variables are established. Then, two new aggregation operators: the interval grey uncertain linguistic correlated ordered arithmetic averaging (IGULCOA operator and the induced interval grey uncertain linguistic correlated ordered arithmetic averaging (I-IGULCOA operator are developed and some desirable properties of the I-IGULCOA operator are studied, such as commutativity, idempotency, monotonicity, and boundness. Furthermore, the IGULCOA and I-IGULCOA operators based approach is developed to solve the MAGDM problems, in which both the attribute weights and the expert weights are correlative and the attribute values take the form of the interval grey uncertain linguistic variables. Finally, an illustrative example is given to verify the developed approach and to demonstrate its practicality and effectiveness.

  12. Effect of extraction pH on heat-induced aggregation, gelation and microstructure of protein isolate from quinoa (Chenopodium quinoa Willd)

    NARCIS (Netherlands)

    Ruiz, Geraldine Avila; Xiao, Wukai; Boekel, van Tiny; Minor, Marcel; Stieger, Markus

    2016-01-01

    The aim of this study was to determine the influence of extraction pH on heat-induced aggregation, gelation and microstructure of suspensions of protein isolates extracted from quinoa (Chenopodium quinoa Willd). Quinoa seed protein was extracted by alkaline treatment at various pH values (pH 8 (E

  13. Synthesis of huaicarbon A/B and their activating effects on platelet glycoprotein VI receptor to mediate collagen-induced platelet aggregation

    Science.gov (United States)

    Yu, Hongli; Chen, Yeqing; Wu, Hao; Wang, Kuilong; Liu, Liping; Zhang, Xingde

    2017-01-01

    Quercetin and rhamnose were efficiently converted into huaicarbon A/B by heating at 250°C for 10-15 min or at 200°C for 25-30 min. With the optimum molar ratio of quercetin/rhamnose (1:3), huaicarbon A and B yields reached 25% and 16% respectively after heating at 250°C, with 55% quercetin conversion. Huaicarbon A/B both promoted washed platelet aggregation dose-dependently, which was antagonized by an inhibitor of glycoprotein VI (GPVI) receptor. Similarly, they both promoted collagen-induced platelet aggregation in platelet-rich plasma in dose-dependent manners. According to the S type dose-response model, EC50 values of huaicarbon A and huaicarbon B were calculated as 33.48 μM and 48.73 μM respectively. They induced intracellular Ca2+ accumulation that was specifically blocked by GPVI antagonist. Huaicarbon A/B enhanced intracellular Ca2+ accumulation and facilitated collagen-induced platelet aggregation, which were blocked by GPVI antagonist. They were conducive to collagen-induced platelet aggregation by activating platelet GPVI receptor. PMID:28337278

  14. Small-angle X-ray scattering studies of metastable intermediates of beta-lactoglobulin isolated after heat-induced aggregation

    DEFF Research Database (Denmark)

    Carrotta, R.; Arleth, L.; Pedersen, J.S.

    2003-01-01

    Small-angle x-ray scattering was used for studying intermediate species, isolated after heat-induced aggregation of the A variant of bovine P-lactoglobulin. The intermediates were separated in two fractions, the heated metastable dimer and heated metastable oligomers larger than the dimer. The pa...

  15. Incorporation of an aggregation-induced-emissive tetraphenylethene derivative into cationic gene delivery vehicles manifested the nuclear translocation of uncomplexed DNA.

    Science.gov (United States)

    Han, Xiongqi; Chen, Qixian; Lu, Hongguang; Guo, Pan; Li, Wei; Wu, Guolin; Ma, Jianbiao; Gao, Hui

    2016-03-11

    A fluorophore displaying aggregation-induced emission was introduced at the terminus of branched polyethylenimine (PEI). The formulated polyplex not only demonstrated an improved safety profile and preserved transfection activity but also importantly indicated that the uncomplexed naked DNA rather than the polyplexes translocated into the nucleus.

  16. Arsenite-induced autophagy is associated with proteotoxicity in human lymphoblastoid cells

    Energy Technology Data Exchange (ETDEWEB)

    Bolt, Alicia M.; Zhao, Fei; Pacheco, Samantha; Klimecki, Walter T., E-mail: klimecki@pharmacy.arizona.edu

    2012-10-15

    Epidemiological studies of arsenic-exposed populations have provided evidence that arsenic exposure in humans is associated with immunosuppression. Previously, we have reported that arsenite-induced toxicity is associated with the induction of autophagy in human lymphoblastoid cell lines (LCL). Autophagy is a cellular process that functions in the degradation of damaged cellular components, including protein aggregates formed by misfolded or damaged proteins. Accumulation of misfolded or damaged proteins in the endoplasmic reticulum (ER) lumen causes ER stress and activates the unfolded protein response (UPR). In an effort to investigate the mechanism of autophagy induction by arsenite in the LCL model, we examined the potential contribution of ER stress and activation of the UPR. LCL exposed to sodium arsenite for 8-days induced expression of UPR-activated genes, including CHOP and GRP78, at the RNA and the protein level. Evidence for activation of the three arms of the UPR was observed. The arsenite-induced activation of the UPR was associated with an accumulation of protein aggregates containing p62 and LC3, proteins with established roles in the sequestration and autophagic clearance of protein aggregates. Taken together, these data provide evidence that arsenite-induced autophagy is associated with the generation of ER stress, activation of the UPR, and formation of protein aggregates that may be targeted to the lysosome for degradation. -- Highlights: ► Arsenite induces endoplasmic reticulum stress and the unfolded protein response. ► Arsenite induces the formation of protein aggregates that contain p62 and LC3-II. ► Time-course data suggests that arsenite-induced autophagy precedes ER stress.

  17. Resveratrol, Acetyl-Resveratrol, and Polydatin Exhibit Antigrowth Activity against 3D Cell Aggregates of the SKOV-3 and OVCAR-8 Ovarian Cancer Cell Lines

    OpenAIRE

    Hogg, Simon J.; Kenny Chitcholtan; Wafaa Hassan; Peter H. Sykes; Ashley Garrill

    2015-01-01

    Resveratrol has aroused significant scientific interest as it has been claimed that it exhibits a spectrum of health benefits. These include effects as an anti-inflammatory and an antitumour compound. The purpose of this study was to investigate and compare any potential antigrowth effects of resveratrol and two of its derivatives, acetyl-resveratrol and polydatin, on 3D cell aggregates of the EGFR/Her-2 positive and negative ovarian cancer cell lines SKOV-3 and OVCAR-8, respectively. Results...

  18. A computer-assisted 3D model for analyzing the aggregation of tumorigenic cells reveals specialized behaviors and unique cell types that facilitate aggregate coalescence.

    Directory of Open Access Journals (Sweden)

    Amanda Scherer

    Full Text Available We have developed a 4D computer-assisted reconstruction and motion analysis system, J3D-DIAS 4.1, and applied it to the reconstruction and motion analysis of tumorigenic cells in a 3D matrix. The system is unique in that it is fast, high-resolution, acquires optical sections using DIC microscopy (hence there is no associated photoxicity, and is capable of long-term 4D reconstruction. Specifically, a z-series at 5 μm increments can be acquired in less than a minute on tissue samples embedded in a 1.5 mm thick 3D Matrigel matrix. Reconstruction can be repeated at intervals as short as every minute and continued for 30 days or longer. Images are converted to mathematical representations from which quantitative parameters can be derived. Application of this system to cancer cells from established lines and fresh tumor tissue has revealed unique behaviors and cell types not present in non-tumorigenic lines. We report here that cells from tumorigenic lines and tumors undergo rapid coalescence in 3D, mediated by specific cell types that we have named "facilitators" and "probes." A third cell type, the "dervish", is capable of rapid movement through the gel and does not adhere to it. These cell types have never before been described. Our data suggest that tumorigenesis in vitro is a developmental process involving coalescence facilitated by specialized cells that culminates in large hollow spheres with complex architecture. The unique effects of select monoclonal antibodies on these processes demonstrate the usefulness of the model for analyzing the mechanisms of anti-cancer drugs.

  19. Nanoscale analysis of caspofungin-induced cell surface remodelling in Candida albicans

    Science.gov (United States)

    El-Kirat-Chatel, Sofiane; Beaussart, Audrey; Alsteens, David; Jackson, Desmond N.; Lipke, Peter N.; Dufrêne, Yves F.

    2013-01-01

    The advent of fungal pathogens that are resistant to the classic repertoire of antifungal drugs has increased the need for new therapeutic agents. A prominent example of such a novel compound is caspofungin, known to alter cell wall biogenesis by inhibiting β-1,3-d-glucan synthesis. Although much progress has been made in understanding the mechanism of action of caspofungin, little is known about its influence on the biophysical properties of the fungal cells. Here, we use atomic force microscopy (AFM) to demonstrate that caspofungin induces major remodelling of the cell surface properties of Candida albicans. Caspofungin causes major morphological and structural alterations of the cells, which correlate with a decrease of the cell wall mechanical strength. Moreover, we find that the drug induces the massive exposure of the cell adhesion protein Als1 on the cell surface and leads to increased cell surface hydrophobicity, two features that trigger cell aggregation. This behaviour is not observed in yeast species lacking Als1, demonstrating the key role that the protein plays in determining the aggregation phenotype of C. albicans. The results show that AFM opens up new avenues for understanding the molecular bases of microbe-drug interactions and for developing new therapeutic agents.The advent of fungal pathogens that are resistant to the classic repertoire of antifungal drugs has increased the need for new therapeutic agents. A prominent example of such a novel compound is caspofungin, known to alter cell wall biogenesis by inhibiting β-1,3-d-glucan synthesis. Although much progress has been made in understanding the mechanism of action of caspofungin, little is known about its influence on the biophysical properties of the fungal cells. Here, we use atomic force microscopy (AFM) to demonstrate that caspofungin induces major remodelling of the cell surface properties of Candida albicans. Caspofungin causes major morphological and structural alterations of the

  20. The genetic heterozygosity and fitness of tetraploid embryos and embryonic stem cells are crucial parameters influencing survival of mice derived from embryonic stem cells by tetraploid embryo aggregation.

    Science.gov (United States)

    Li, Xiangyun; Wei, Wei; Yong, Jun; Jia, Qing; Yu, Yuansong; Di, Keqian

    2005-07-01

    The aim of this paper was to determine whether the genetic background of tetraploid embryos contributed to the survival of mice derived from embryonic stem (ES) cells by tetraploid embryo complementation. Twenty-five newborns were produced by aggregation of hybrid ES cells and tetraploid embryos with different genetic backgrounds. These newborns were entirely derived from ES cells judged by microsatellite DNA (A specific sequence of DNA bases or nucleotides that contains mono, di, tri or tetra repeats) and coat colour phenotype and germline transmission. Fifteen survived to adulthood while seven died of respiratory failure. All newborns were derived from outbred or hybrid tetraploid aggregates and no newborns were from the inbreds. Our results demonstrate that the genetic heterozygosity, fitness of tetraploid embryos and fitness of ES cells are crucial parameters influencing survival of mice derived from ES cells by tetraploid embryo aggregation. In addition, this method represents a simple and efficient procedure for immediate generation of targeted mouse mutants from genetically modified ES cell clones, in contrast to the standard protocol, which involves the production of chimeras and several breeding steps.

  1. Multifunctional 8-hydroxyquinoline-appended cyclodextrins as new inhibitors of metal-induced protein aggregation.

    Science.gov (United States)

    Oliveri, Valentina; Attanasio, Francesco; Puglisi, Antonino; Spencer, John; Sgarlata, Carmelo; Vecchio, Graziella

    2014-07-14

    Mounting evidence suggests a pivotal role of metal imbalances in protein misfolding and amyloid diseases. As such, metal ions represent a promising therapeutic target. In this context, the synthesis of chelators that also contain complementary functionalities to combat the multifactorial nature of neurodegenerative diseases is a highly topical issue. We report two new 8-hydroxyquinoline-appended cyclodextrins and highlight their multifunctional properties, including their Cu(II) and Zn(II) binding abilities, and capacity to act as antioxidants and metal-induced antiaggregants. In particular, the latter property has been applied in the development of an effective assay that exploits the formation of amyloid fibrils when β-lactoglobulin A is heated in the presence of metal ions.

  2. Cholesterol impairment contributes to neuroserpin aggregation

    Science.gov (United States)

    Giampietro, Costanza; Lionetti, Maria Chiara; Costantini, Giulio; Mutti, Federico; Zapperi, Stefano; La Porta, Caterina A. M.

    2017-03-01

    Intraneural accumulation of misfolded proteins is a common feature of several neurodegenerative pathologies including Alzheimer’s and Parkinson’s diseases, and Familial Encephalopathy with Neuroserpin Inclusion Bodies (FENIB). FENIB is a rare disease due to a point mutation in neuroserpin which accelerates protein aggregation in the endoplasmic reticulum (ER). Here we show that cholesterol depletion induced either by prolonged exposure to statins or by inhibiting the sterol reg-ulatory binding-element protein (SREBP) pathway also enhances aggregation of neuroserpin proteins. These findings can be explained considering a computational model of protein aggregation under non-equilibrium conditions, where a decrease in the rate of protein clearance improves aggregation. Decreasing cholesterol in cell membranes affects their biophysical properties, including their ability to form the vesicles needed for protein clearance, as we illustrate by a simple mathematical model. Taken together, these results suggest that cholesterol reduction induces neuroserpin aggregation, even in absence of specific neuroserpin mutations. The new mechanism we uncover could be relevant also for other neurodegenerative diseases associated with protein aggregation.

  3. Cholesterol impairment contributes to neuroserpin aggregation

    Science.gov (United States)

    Giampietro, Costanza; Lionetti, Maria Chiara; Costantini, Giulio; Mutti, Federico; Zapperi, Stefano; La Porta, Caterina A. M.

    2017-01-01

    Intraneural accumulation of misfolded proteins is a common feature of several neurodegenerative pathologies including Alzheimer’s and Parkinson’s diseases, and Familial Encephalopathy with Neuroserpin Inclusion Bodies (FENIB). FENIB is a rare disease due to a point mutation in neuroserpin which accelerates protein aggregation in the endoplasmic reticulum (ER). Here we show that cholesterol depletion induced either by prolonged exposure to statins or by inhibiting the sterol reg-ulatory binding-element protein (SREBP) pathway also enhances aggregation of neuroserpin proteins. These findings can be explained considering a computational model of protein aggregation under non-equilibrium conditions, where a decrease in the rate of protein clearance improves aggregation. Decreasing cholesterol in cell membranes affects their biophysical properties, including their ability to form the vesicles needed for protein clearance, as we illustrate by a simple mathematical model. Taken together, these results suggest that cholesterol reduction induces neuroserpin aggregation, even in absence of specific neuroserpin mutations. The new mechanism we uncover could be relevant also for other neurodegenerative diseases associated with protein aggregation. PMID:28255164

  4. Cellulose synthase complexes act in a concerted fashion to synthesize highly aggregated cellulose in secondary cell walls of plants.

    Science.gov (United States)

    Li, Shundai; Bashline, Logan; Zheng, Yunzhen; Xin, Xiaoran; Huang, Shixin; Kong, Zhaosheng; Kim, Seong H; Cosgrove, Daniel J; Gu, Ying

    2016-10-04

    Cellulose, often touted as the most abundant biopolymer on Earth, is a critical component of the plant cell wall and is synthesized by plasma membrane-spanning cellulose synthase (CESA) enzymes, which in plants are organized into rosette-like CESA complexes (CSCs). Plants construct two types of cell walls, primary cell walls (PCWs) and secondary cell walls (SCWs), which differ in composition, structure, and purpose. Cellulose in PCWs and SCWs is chemically identical but has different physical characteristics. During PCW synthesis, multiple dispersed CSCs move along a shared linear track in opposing directions while synthesizing cellulose microfibrils with low aggregation. In contrast, during SCW synthesis, we observed swaths of densely arranged CSCs that moved in the same direction along tracks while synthesizing cellulose microfibrils that became highly aggregated. Our data support a model in which distinct spatiotemporal features of active CSCs during PCW and SCW synthesis contribute to the formation of cellulose with distinct structure and organization in PCWs and SCWs of Arabidopsis thaliana This study provides a foundation for understanding differences in the formation, structure, and organization of cellulose in PCWs and SCWs.

  5. Protein oxidation and aggregation in UVA-irradiated Escherichia coli cells as signs of accelerated cellular senescence.

    Science.gov (United States)

    Bosshard, Franziska; Riedel, Kathrin; Schneider, Thomas; Geiser, Carina; Bucheli, Margarete; Egli, Thomas

    2010-11-01

    Solar disinfection (SODIS) is a simple drinking water treatment method that improves microbiological water quality where other means are unavailable. It makes use of the deleterious effect of solar irradiation on pathogenic microbes and viruses. A positive impact on health has been documented in several epidemiological studies. However, the molecular mechanisms damaging cells during this simple treatment are not yet fully understood. Here we show that protein damage is crucial in the process of inactivation by sunlight. Protein damages in UVA-irradiated Escherichia coli cells have been evaluated by an immunoblot method for carbonylated proteins and an aggregation assay based on semi-quantitative proteomics. A wide spectrum of structural and enzymatic proteins within the cell is affected by carbonylation and aggregation. Vital cellular functions like the transcription and translation apparatus, transport systems, amino acid synthesis and degradation, respiration, ATP synthesis, glycolysis, the TCA cycle, chaperone functions and catalase are targeted by UVA irradiation. The protein damage pattern caused by SODIS strongly resembles the pattern caused by reactive oxygen stress. Hence, sunlight probably accelerates cellular senescence and leads to the inactivation and finally death of UVA-irradiated cells.

  6. EDTA enhances high-throughput two-dimensional bioprinting by inhibiting salt scaling and cell aggregation at the nozzle surface.

    Science.gov (United States)

    Parzel, Cheryl A; Pepper, Matthew E; Burg, Timothy; Groff, Richard E; Burg, Karen J L

    2009-06-01

    Tissue-engineering strategies may be employed in the development of in vitro breast tissue models for use in testing regimens of drug therapies and vaccines. The physical and chemical interactions that occur among cells and extracellular matrix components can also be elucidated with these models to gain an understanding of the progression of transformed epithelial cells into tumours and the ultimate metastases of tumour cells. The modified inkjet printer may be a useful tool for creating three-dimensional (3D) in vitro models, because it offers an inexpensive and high-throughput solution to microfabrication, and because the printer can be easily manipulated to produce varying tissue attributes. We hypothesized, however, that when ink is replaced with a biologically based fluid (i.e. a 'bio-ink'), specifically a serum-free cell culture medium, printer nozzle failure can result from salt scale build-up as fluid evaporates on the printhead surface. In this study, ethylene diamine tetra-acetic acid (EDTA) was used as a culture medium additive to prevent salt scaling and cell aggregation during the bioprinting process. The results showed that EDTA, at a concentration typically found in commercially available trypsin solutions (0.53 mM), prevented nozzle failure when a serum-free culture medium was printed from a nozzle at 1000 drops/s. Furthermore, increasing concentrations of EDTA appeared to mildly decrease aggregation of 4T07 cells. Cell viability studies were performed to demonstrate that addition of EDTA did not result in significant cell death. In conclusion, it is recommended that EDTA be incorporated into bio-ink solutions containing salts that could lead to nozzle failure.

  7. Mitochondrial trafficking through Rhot1 is involved in the aggregation of germinal granule components during primordial germ cell formation in Xenopus embryos.

    Science.gov (United States)

    Tada, Haru; Taira, Yuya; Morichika, Keisuke; Kinoshita, Tsutomu

    2016-10-01

    In many animals, the germ plasm is sufficient and necessary for primordial germ cell (PGC) formation. It contains germinal granules and abundant mitochondria (germline-Mt). However, the role of germline-Mt in germ cell formation remains poorly understood. In Xenopus, the germ plasm is distributed as many small islands at the vegetal pole, which gradually aggregates to form a single large mass in each of the four vegetal pole cells at the early blastula stage. Polymerized microtubules and the adapter protein kinesin are required for the aggregation of germ plasm. However, it remains unknown whether germline-Mt trafficking is important for the cytoplasmic transport of germinal granules during germ plasm aggregation. In this study, we focused on the mitochondrial small GTPase protein Rhot1 to inhibit mitochondrial trafficking during the germ plasm aggregation. Expression of Rhot1ΔC, which lacks the C-terminal mitochondrial transmembrane domain, inhibited the aggregation of germline-Mt during early development. In Rhot1-inhibited embryos, germinal granule components did not aggregate during cleavage stages, which reduced the number of PGCs on the genital ridge at tail-bud stage. These results suggest that mitochondrial trafficking is involved in the aggregation of germinal granule components, which are essential for the formation of PGCs.

  8. Chaperone effects on prion and nonprion aggregates.

    Science.gov (United States)

    Rikhvanov, Eugene G; Romanova, Nina V; Chernoff, Yury O

    2007-01-01

    Exposure to high temperature or other stresses induces a synthesis of heat shock proteins. Many of these proteins are molecular chaperones, and some of them help cells to cope with heat-induced denaturation and aggregation of other proteins. In the last decade, chaperones have received increased attention in connection with their role in maintenance and propagation of the Saccharomyces cerevisiae prions, infectious or heritable agents transmitted at the protein level. Recent data suggest that functioning of the chaperones in reactivation of heat-damaged proteins and in propagation of prions is based on the same molecular mechanisms but may lead to different consequences depending on the type of aggregate. In both cases the concerted and balanced action of "chaperones' team," including Hsp104, Hsp70, Hsp40 and possibly other proteins, determines whether a misfolded protein is to be incorporated into an aggregate, rescued to the native state or targeted for degradation.

  9. Effect of the surfactant tween 80 on the detachment and dispersal of Salmonella enterica serovar Thompson single cells and aggregates from cilantro leaves as revealed by image analysis.

    Science.gov (United States)

    Brandl, Maria T; Huynh, Steven

    2014-08-01

    Salmonella enterica has the ability to form biofilms and large aggregates on produce surfaces, including on cilantro leaves. Aggregates of S. enterica serovar Thompson that remained attached to cilantro leaves after rigorous washing and that were present free or bound to dislodged leaf tissue in the wash suspension were observed by confocal microscopy. Measurement of S. Thompson population sizes in the leaf washes by plate counts failed to show an effect of 0.05% Tween 80 on the removal of the pathogen from cilantro leaves 2 and 6 days after inoculation. On the contrary, digital image analysis of micrographs of single cells and aggregates of green fluorescent protein (GFP)-S. Thompson present in cilantro leaf washes revealed that single cells represented 13.7% of the cell assemblages in leaf washes containing Tween 80, versus 9.3% in those without the surfactant. Moreover, Tween 80 decreased the percentage of the total S. Thompson cell population located in aggregates equal to or larger than 64 cells from 9.8% to 4.4% (P Tween 80 showed that the surfactant promoted the dispersal of cells from large aggregates into smaller ones and into single cells (P < 0.05). Our study underlines the importance of investigating bacterial behavior at the scale of single cells in order to uncover trends undetectable at the population level by bacterial plate counts. Such an approach may provide valuable information to devise strategies aimed at enhancing the efficacy of produce sanitization treatments.

  10. Induced Pluripotent Stem Cells and Periodontal Regeneration

    OpenAIRE

    Du, Mi; Duan, Xuejing; Yang, Pishan

    2015-01-01

    Periodontitis is a chronic inflammatory disease which leads to destruction of both the soft and hard tissues of the periodontium. Tissue engineering is a therapeutic approach in regenerative medicine that aims to induce new functional tissue regeneration via the synergistic combination of cells, biomaterials, and/or growth factors. Advances in our understanding of the biology of stem cells, including embryonic stem cells and mesenchymal stem cells, have provided opportunities for periodontal ...

  11. Method for Producing Non-Neoplastic, Three Dimensional, Mammalian Tissue and Cell Aggregates Under Microgravity Culture Conditions and the Products Produced Therefrom

    Science.gov (United States)

    Goodwin, Thomas J. (Inventor); Wolf, David A. (Inventor); Spaulding, Glenn F. (Inventor); Prewett, Tracey L. (Inventor)

    1996-01-01

    Normal mammalian tissue and the culturing process has been developed for the three groups of organ, structural, and blood tissue. The cells are grown in vitro under microgravity culture conditions and form three dimensional cells aggregates with normal cell function. The microgravity culture conditions may be microgravity or simulated microgravity created in a horizontal rotating wall culture vessel.

  12. Acid-Induced Cold Gelation of Globular Proteins: Effects of Protein Aggregate Characteristics and Disulfide Bonding on Rheological Properties

    NARCIS (Netherlands)

    Alting, A.C.; Weijers, M.; Hoog, E.H.A. de; Pijpekamp, A.M. van de; Cohen Stuart, M.A.; Hamer, R.J.; Kruif, C.G. de; Visschers, R.W.

    2004-01-01

    The process of cold gelation of ovalbumin and the properties of the resulting cold-set gels were compared to those of whey protein isolate. Under the chosen heating conditions, most protein was organized in aggregates. For both protein preparations, the aggregates consisted of covalently linked mono

  13. Acid-induced cold gelation of globular proteins: effects of protein aggregate characteristics ans disulfide bonding on Rheological properties.

    NARCIS (Netherlands)

    Alting, A.C.; Weijers, M.; Hoog, de E.H.A.; Pijpekamp, A.M.; Cohen Stuart, M.A.; Hamer, R.J.; Kruif, de C.G.; Visschers, R.W.

    2004-01-01

    The process of cold gelation of ovalbumin and the properties of the resulting cold-set gels were compared to those of whey protein isolate. Under the chosen heating conditions, most protein was organized in aggregates. For both protein preparations, the aggregates consisted of covalently linked mono

  14. Smart Cu(II)-aptamer complexes based gold nanoplatform for tumor micro-environment triggered programmable intracellular prodrug release, photodynamic treatment and aggregation induced photothermal therapy of hepatocellular carcinoma

    Science.gov (United States)

    Zhang, Da; Zheng, Aixian; Li, Juan; Wu, Ming; Wu, Lingjie; Wei, Zuwu; Liao, Naishun; Zhang, Xiaolong; Cai, Zhixiong; Yang, Huanghao; Liu, Gang; Liu, Xiaolong; Liu, Jingfeng

    2017-01-01

    This study describes smart Cu(II)-aptamer complexes based gold nanoplatform for tumor micro-environment triggered programmable prodrug release, in demand photodynamic therapy and aggregation induced photothermal ablation of hepatocellular carcinoma. The nanoplatform is consist of monodispersed gold nanoparticle (GNP) that is binding to HCC cell specific targeting aptamers (TLS11a) through Au-S bond; the aptamer is labeled with Ce6 at the 5'end and coordinated with Cu(II) through (GA)10 repeating bases to load AQ4N at the 3' end. In normal physiological conditions, the fluorescence and ROS generation ability of Ce6 are quenched by GNPs via RET; but in cancerous cells, the fluorescence and the ROS generation of Ce6 could be recovered by cleavage of Au-S bond through high level of intracellular GSH for real-time imaging and in demand PDT. Meanwhile, the prodrug AQ4N release could be triggered by acid-cleavage of coordination bonds, then accompanied by a release of Cu(II) that would induce the electrostatic aggregation of GNPs for photo-thermal ablation; furthermore, the significantly enhanced chemotherapy efficiency could be achieved by PDT produced hypoxia to convert AQ4N into AQ4. In summary, here described nanoplatform with tumor cell specific responsive properties and programmable PDT/PTT/chemotherapy functions, might be an interesting synergistic strategy for HCC treatment. PMID:28042325

  15. Induced pluripotent stem cell lines derived from human somatic cells.

    Science.gov (United States)

    Yu, Junying; Vodyanik, Maxim A; Smuga-Otto, Kim; Antosiewicz-Bourget, Jessica; Frane, Jennifer L; Tian, Shulan; Nie, Jeff; Jonsdottir, Gudrun A; Ruotti, Victor; Stewart, Ron; Slukvin, Igor I; Thomson, James A

    2007-12-21

    Somatic cell nuclear transfer allows trans-acting factors present in the mammalian oocyte to reprogram somatic cell nuclei to an undifferentiated state. We show that four factors (OCT4, SOX2, NANOG, and LIN28) are sufficient to reprogram human somatic cells to pluripotent stem cells that exhibit the essential characteristics of embryonic stem (ES) cells. These induced pluripotent human stem cells have normal karyotypes, express telomerase activity, express cell surface markers and genes that characterize human ES cells, and maintain the developmental potential to differentiate into advanced derivatives of all three primary germ layers. Such induced pluripotent human cell lines should be useful in the production of new disease models and in drug development, as well as for applications in transplantation medicine, once technical limitations (for example, mutation through viral integration) are eliminated.

  16. Molecular Level Insights into Thermally Induced [alpha]-Chymotrypsinogen A Amyloid Aggregation Mechanism and Semiflexible Protofibril Morphology

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Aming; Jordan, Jacob L.; Ivanova, Magdalena I.; Weiss, IV., William F.; Roberts, Christopher J.; Fernandez, Erik J. (UCLA); (Delaware); (UV)

    2010-12-07

    Understanding nonnative protein aggregation is critical not only to a number of amyloidosis disorders but also for the development of effective and safe biopharmaceuticals. In a series of previous studies [Weiss et al. (2007) Biophys. J. 93, 4392-4403; Andrews et al. (2007) Biochemistry 46, 7558-7571; Andrews et al. (2008) Biochemistry 47, 2397-2403], {alpha}-chymotrypsinogen A (aCgn) and bovine granulocyte colony stimulating factor (bG-CSF) have been shown to exhibit the kinetic and morphological features of other nonnative aggregating proteins at low pH and ionic strength. In this study, we investigated the structural mechanism of aCgn aggregation. The resultant aCgn aggregates were found to be soluble and exhibited semiflexible filamentous aggregate morphology under transmission electron microscopy. In addition, the filamentous aggregates were demonstrated to possess amyloid characteristics by both Congo red binding and X-ray diffraction. Peptide level hydrogen exchange (HX) analysis suggested that a buried native {beta}-sheet comprised of three peptide segments (39-46, 51-64, and 106-114) reorganizes into the cross-{beta} amyloid core of aCgn aggregates and that at least 50% of the sequence adopts a disordered structure in the aggregates. Furthermore, the equimolar, bimodal HX labeling distribution observed for three reported peptides (65-102, 160-180, and 229-245) suggested a heterogeneous assembly of two molecular conformations in aCgn aggregates. This demonstrates that extended {beta}-sheet interactions typical of the amyloid are sufficiently strong that a relatively small fraction of polypeptide sequence can drive formation of filamentous aggregates even under conditions favoring colloidal stability.

  17. In vitro generation of three-dimensional substrate-adherent embryonic stem cell-derived neural aggregates for application in animal models of neurological disorders.

    Science.gov (United States)

    Hargus, Gunnar; Cui, Yi-Fang; Dihné, Marcel; Bernreuther, Christian; Schachner, Melitta

    2012-05-01

    In vitro-differentiated embryonic stem (ES) cells comprise a useful source for cell replacement therapy, but the efficiency and safety of a translational approach are highly dependent on optimized protocols for directed differentiation of ES cells into the desired cell types in vitro. Furthermore, the transplantation of three-dimensional ES cell-derived structures instead of a single-cell suspension may improve graft survival and function by providing a beneficial microenvironment for implanted cells. To this end, we have developed a new method to efficiently differentiate mouse ES cells into neural aggregates that consist predominantly (>90%) of postmitotic neurons, neural progenitor cells, and radial glia-like cells. When transplanted into the excitotoxically lesioned striatum of adult mice, these substrate-adherent embryonic stem cell-derived neural aggregates (SENAs) showed significant advantages over transplanted single-cell suspensions of ES cell-derived neural cells, including improved survival of GABAergic neurons, increased cell migration, and significantly decreased risk of teratoma formation. Furthermore, SENAs mediated functional improvement after transplantation into animal models of Parkinson's disease and spinal cord injury. This unit describes in detail how SENAs are efficiently derived from mouse ES cells in vitro and how SENAs are isolated for transplantation. Furthermore, methods are presented for successful implantation of SENAs into animal models of Huntington's disease, Parkinson's disease, and spinal cord injury to study the effects of stem cell-derived neural aggregates in a disease context in vivo.

  18. HIV transcription is induced in dying cells

    Energy Technology Data Exchange (ETDEWEB)

    Woloschak, G.E.; Chang-Liu, Chin-Mei [Argonne National Lab., IL (United States); Schreck, S. [Argonne National Lab., IL (United States)]|[Univ. of South Carolina, Columbia, SC (United States). Dept. of Chemistry

    1995-06-01

    Using HeLa cells stably transfected with an HIV-LTR-CAT construct, we demonstrated a peak in CAT induction that occurs in viable (but not necessarily cell-division-competent) cells 24 h following exposure to some cell-killing agents. {gamma} rays were the only cell-killing agent which did not induce HIV transcription; this can be attributed to the fact that {gamma}-ray-induced apoptotic death requires functional p53, which is not present in HeLa cells. For all other agents, HIV-LTR induction was dose-dependent and correlated with the amount of cell killing that occurred in the culture. 14 refs., 4 figs., 1 tab.

  19. Disulfide scrambling in superoxide dismutase 1 reduces its cytotoxic effect in cultured cells and promotes protein aggregation.

    Directory of Open Access Journals (Sweden)

    Lina Leinartaitė

    Full Text Available Mutations in the gene coding for superoxide dismutase 1 (SOD1 are associated with familiar forms of the neurodegenerative disease amyotrophic lateral sclerosis (ALS. These mutations are believed to result in a "gain of toxic function", leading to neuronal degeneration. The exact mechanism is still unknown, but misfolding/aggregation events are generally acknowledged as important pathological events in this process. Recently, we observed that demetallated apoSOD1, with cysteine 6 and 111 substituted for alanine, is toxic to cultured neuroblastoma cells. This toxicity depended on an intact, high affinity Zn(2+ site. It was therefor contradictory to discover that wild-type apoSOD1 was not toxic, despite of its high affinity for Zn(2+. This inconsistency was hypothesized to originate from erroneous disulfide formation involving C6 and C111. Using high resolution non-reducing SDS-PAGE, we have in this study demonstrated that the inability of wild-type apoSOD1 to cause cell death stems from formation of non-native intra-molecular disulfides. Moreover, monomeric apoSOD1 variants capable of such disulfide scrambling aggregated into ThT positive oligomers under physiological conditions without agitation. The oligomers were stabilized by inter-molecular disulfides and morphologically resembled what has in other neurodegenerative diseases been termed protofibrils. Disulfide scrambling thus appears to be an important event for misfolding and aggregation of SOD1, but may also be significant for protein function involving cysteines, e.g. mitochondrial import and copper loading.

  20. HIF induces human embryonic stem cell markers in cancer cells.

    Science.gov (United States)

    Mathieu, Julie; Zhang, Zhan; Zhou, Wenyu; Wang, Amy J; Heddleston, John M; Pinna, Claudia M A; Hubaud, Alexis; Stadler, Bradford; Choi, Michael; Bar, Merav; Tewari, Muneesh; Liu, Alvin; Vessella, Robert; Rostomily, Robert; Born, Donald; Horwitz, Marshall; Ware, Carol; Blau, C Anthony; Cleary, Michele A; Rich, Jeremy N; Ruohola-Baker, Hannele

    2011-07-01

    Low oxygen levels have been shown to promote self-renewal in many stem cells. In tumors, hypoxia is associated with aggressive disease course and poor clinical outcomes. Furthermore, many aggressive tumors have been shown to display gene expression signatures characteristic of human embryonic stem cells (hESC). We now tested whether hypoxia might be responsible for the hESC signature observed in aggressive tumors. We show that hypoxia, through hypoxia-inducible factor (HIF), can induce an hESC-like transcriptional program, including the induced pluripotent stem cell (iPSC) inducers, OCT4, NANOG, SOX2, KLF4, cMYC, and microRNA-302 in 11 cancer cell lines (from prostate, brain, kidney, cervix, lung, colon, liver, and breast tumors). Furthermore, nondegradable forms of HIFα, combined with the traditional iPSC inducers, are highly efficient in generating A549 iPSC-like colonies that have high tumorigenic capacity. To test potential correlation between iPSC inducers and HIF expression in primary tumors, we analyzed primary prostate tumors and found a significant correlation between NANOG-, OCT4-, and HIF1α-positive regions. Furthermore, NANOG and OCT4 expressions positively correlated with increased prostate tumor Gleason score. In primary glioma-derived CD133 negative cells, hypoxia was able to induce neurospheres and hESC markers. Together, these findings suggest that HIF targets may act as key inducers of a dynamic state of stemness in pathologic conditions.

  1. Dissipative particle dynamics simulations of deformation and aggregation of healthy and diseased red blood cells in a tube flow

    Energy Technology Data Exchange (ETDEWEB)

    Ye, Ting; Phan-Thien, Nhan, E-mail: Nhan@nus.edu.sg; Khoo, Boo Cheong; Lim, Chwee Teck [Department of Mechanical Engineering, National University of Singapore, Singapore 119260 (Singapore)

    2014-11-15

    In this paper, we report simulation results assessing the deformation and aggregation of mixed healthy and malaria-infected red blood cells (RBCs) in a tube flow. A three dimensional particle model based on Dissipative Particle Dynamics (DPD) is developed to predict the tube flow containing interacting cells. The cells are also modelled by DPD, with a Morse potential to characterize the cell-cell interaction. As validation tests, a single RBC in a tube flow and two RBCs in a static flow are simulated to examine the cell deformation and intercellular interaction, respectively. The study of two cells, one healthy and the other malaria-infected RBCs in a tube flow demonstrates that the malaria-infected RBC (in the leading position along flow direction) has different effects on the healthy RBC (in the trailing position) at the different stage of parasite development or at the different capillary number. With parasitic development, the malaria-infected RBC gradually loses its deformability, and in turn the corresponding trailing healthy RBC also deforms less due to the intercellular interaction. With increasing capillary number, both the healthy and malaria-infected RBCs are likely to undergo an axisymmetric motion. The minimum intercellular distance becomes small enough so that rouleaux is easily formed, i.e., the healthy and malaria-infected RBCs are difficultly disaggregated.

  2. Abrogation of E-cadherin-mediated cellular aggregation allows proliferation of pluripotent mouse embryonic stem cells in shake flask bioreactors.

    Directory of Open Access Journals (Sweden)

    Lisa Mohamet

    Full Text Available BACKGROUND: A fundamental requirement for the exploitation of embryonic stem (ES cells in regenerative medicine is the ability to reproducibly derive sufficient numbers of cells of a consistent quality in a cost-effective manner. However, undifferentiated ES cells are not ideally suited to suspension culture due to the formation of cellular aggregates, ultimately limiting scalability. Significant advances have been made in recent years in the culture of ES cells, including automated adherent culture and suspension microcarrier or embryoid body bioreactor culture. However, each of these methods exhibits specific disadvantages, such as high cost, additional downstream processes or reduced cell doubling times. METHODOLOGY/PRINCIPAL FINDINGS: Here we show that abrogation of the cell surface protein E-cadherin, using either gene knockout (Ecad-/- or the neutralising antibody DECMA-1 (EcadAb, allows culture of mouse ES cells as a near-single cell suspension in scalable shake flask culture over prolonged periods without additional media supplements. Both Ecad-/- and EcadAb ES cells exhibited adaptation phases in suspension culture, with optimal doubling times of 7.3 h±0.9 and 15.6 h±4.7 respectively and mean-fold increase in viable cell number of 95.1±2.0 and 16±0.9-fold over 48 h. EcadAb ES cells propagated as a dispersed cell suspension for 15 d maintained expression of pluripotent markers, exhibited a normal karyotype and high viability. Subsequent differentiation of EcadAb ES cells resulted in expression of transcripts and proteins associated with the three primary germ layers. CONCLUSIONS/SIGNIFICANCE: This is the first demonstration of the culture of pluripotent ES cells as a near-single cell suspension in a manual fed-batch shake flask bioreactor and represents a significant improvement on current ES cell culture techniques. Whilst this proof-of-principle method would be useful for the culture of human ES and iPS cells, further steps are

  3. Inhibitory effects of Tabebuia impetiginosa inner bark extract on platelet aggregation and vascular smooth muscle cell proliferation through suppressions of arachidonic acid liberation and ERK1/2 MAPK activation.

    Science.gov (United States)

    Son, Dong-Ju; Lim, Yong; Park, Young-Hyun; Chang, Sung-Keun; Yun, Yeo-Pyo; Hong, Jin-Tae; Takeoka, Gary R; Lee, Kwang-Geun; Lee, Sung-Eun; Kim, Mi-Ran; Kim, Jeong-Han; Park, Byeoung-Soo

    2006-11-03

    The antiplatelet and antiproliferative activities of extract of Tabebuia impetiginosa inner bark (taheebo) were investigated using washed rabbit platelets and cultured rat aortic vascular smooth muscle cells (VSMCs) in vitro. n-Hexane, chloroform and ethyl acetate fractions showed marked and selective inhibition of platelet aggregation induced by collagen and arachidonic acid (AA) in a dose-dependent manner. These fractions, especially the chloroform fraction, also significantly suppressed AA liberation induced by collagen in [(3)H]AA-labeled rabbit platelets. The fractions, especially the chloroform fraction, potently inhibited cell proliferation and DNA synthesis induced by platelet derived growth factor (PDGF)-BB, and inhibited the levels of phosphorylated extracellular signal regulated kinase (ERK1/2) mitogen activated protein kinase (MAPK) stimulated by PDGF-BB, in the same concentration range that inhibits VSMC proliferation and DNA synthesis.

  4. Lactose-Functionalized Dendrimers Arbitrate the Interaction of Galectin-3/MUC1 Mediated Cancer Cellular Aggregation

    Science.gov (United States)

    Michel, Anna K.; Nangia-Makker, Pratima; Raz, Avraham

    2015-01-01

    By using lactose-functionalized poly(amidoamine) dendrimers as a tunable multivalent platform, we studied cancer cell aggregation in three different cell lines (A549, DU-145, and HT-1080) with galectin-3. We found that small lactose-functionalized G(2)-dendrimer 1 inhibited galectin-3-induced aggregation of the cancer cells. In contrast, dendrimer 4 (a larger, generation 6 dendrimer with 100 carbohydrate end groups) caused cancer cells to aggregate through a galectin-3 pathway. This study indicates that inhibition of cellular aggregation occurred because 1 provided competitive binding sites for galectin-3 (compared to its putative cancer cell ligand, TF-antigen on MUC1). Dendrimer 4, in contrast, provided an excess of ligands for galectin-3 binding; this caused crosslinking and aggregation of cells to be increased. PMID:25138772

  5. [Induction of native platelets aggregation by incubation media of the UV irradiated leukocytes: possible role of the photo-induced ADP release].

    Science.gov (United States)

    Anosov, A K; Gorbach, M M

    2014-01-01

    It is shown that during incubation after UV irradiation (22-24 hours at 7-9 degrees C) irradiated isolated rabbit leukocytes release the compound(s) which induces platelets aggregation in the native platelet rich plasma. Treatment of the incubation media of irradiated leukocytes by heat (5 minutes at 100 degrees C) does not significantly change its pro-aggregation activity. Treatment of the platelet-rich plasma by the incubation media of irradiated leukocytes without stirring induces the refractoriness of platelets to ADP. The platelets treated by ADP without stirring do not react to the incubation media of irradiated leukocytes. The absorption spectrum of the incubation media of irradiated leukocytes has the maximum at 260 nm similar to that of the absorption spectra of ADP. It is possible that UVradiation induces the ADP release from leukocytes during post-irradiation incubation. Accumulation of this substance in the incubation media may be the cause of its pro-aggregation activity for native blood platelets.

  6. Fluorescence characterization of co-immobilization-induced multi-enzyme aggregation in a polymer matrix using Förster resonance energy transfer (FRET): toward the metabolon biomimic.

    Science.gov (United States)

    Wu, Fei; Minteer, Shelley D

    2013-08-12

    Sequential metabolic enzymes can form supramolecular complexes named metabolons in vivo through enzyme-enzyme association or aggregation to facilitate efficient substrate channeling. By separately labeling enzymes with lysine-targeting carboxylic acid succinimidyl ester fluorophores of distinct excitation wavelengths, this research presents a quantitative study of polymer-entrapment-induced in vitro multi-enzyme aggregation from three Krebs cycle enzymes using Förster resonance energy transfer (FRET) to find potential polymer materials for immobilizing enzyme cascades and inducing the metabolon biomimic formation on electrodes. The effect of hydrophobic modification of linear polyethylenimine, Nafion, and chitosan polymers on metabolon formation has been investigated through photobleaching FRET imaging in addition to traditional steady-state fluorescence spectroscopy. By partially destroying FRET acceptors of longer excitation wavelength, increased fluorescence from dequenched donors of shorter excitation wavelength was measured and enzyme interactions in terms of energy-transfer efficiencies were mapped point by point. Results show that trimethyloctadecylammonium-modified Nafion works best in inducing multi-enzyme aggregation and exhibits a promising future in immobilized metabolon biomimics with the most uniform enzyme organization, as indicated by the protein distance distribution.

  7. In vivo venous assessment of red blood cell aggregate sizes in diabetic patients with a quantitative cellular ultrasound imaging method: proof of concept.

    Directory of Open Access Journals (Sweden)

    Julien Tripette

    Full Text Available Diabetic patients present higher level of red blood cell (RBC aggregation contributing to the development of vascular complications. While it has been suggested that this hematology/rheology parameter could bring additional prognostic information for the management of those patients, RBC aggregation screening is not included as a clinical practice. Most medical centers are not equipped to measure properly this parameter, although sedimentation tests can bring some indication. Here, we aimed at evaluating the feasibility of using ultrasound to assess in-vivo hyper-aggregation in type 2 diabetic patients.Seventeen diabetic patients and 15 control subjects underwent ultrasound measurements of RBC aggregation in both cephalic and great saphenous veins. Non-invasive in-vivo ultrasound measurements were performed using a newly developed cellular imaging technique, the structure factor size and attenuation estimator (SFSAE. Comparisons with an ex-vivo gold standard rheometry technique were done, along with measurements of pro-aggregating plasma molecule concentrations.In-vivo RBC aggregation was significantly higher in diabetic patients compared with controls for cephalic vein measurements, while a trend (p = 0.055 was noticed in the great saphenous vein. SFSAE measurements were correlated with gold standard in-vitro measures, fibrinogen and C-reactive protein plasma concentrations.RBC aggregation can be measured in-vivo in diabetic patients using ultrasound. Prospective studies are needed to determine whether the SFSAE method could help clinicians in the early management of vascular complications in this patient population.

  8. Glyceraldehyde-3-phosphate Dehydrogenase Aggregates Accelerate Amyloid-β Amyloidogenesis in Alzheimer Disease*

    Science.gov (United States)

    Itakura, Masanori; Nakajima, Hidemitsu; Kubo, Takeya; Semi, Yuko; Kume, Satoshi; Higashida, Shusaku; Kaneshige, Akihiro; Kuwamura, Mitsuru; Harada, Naoki; Kita, Akinori; Azuma, Yasu-Taka; Yamaji, Ryoichi; Inui, Takashi; Takeuchi, Tadayoshi

    2015-01-01

    Alzheimer disease (AD) is a progressive neurodegenerative disorder characterized by loss of neurons and formation of pathological extracellular deposits induced by amyloid-β peptide (Aβ). Numerous studies have established Aβ amyloidogenesis as a hallmark of AD pathogenesis, particularly with respect to mitochondrial dysfunction. We have previously shown that glycolytic glyceraldehyde-3-phosphate dehydrogenase (GAPDH) forms amyloid-like aggregates upon exposure to oxidative stress and that these aggregates contribute to neuronal cell death. Here, we report that GAPDH aggregates accelerate Aβ amyloidogenesis and subsequent neuronal cell death both in vitro and in vivo. Co-incubation of Aβ40 with small amounts of GAPDH aggregates significantly enhanced Aβ40 amyloidogenesis, as assessed by in vitro thioflavin-T assays. Similarly, structural analyses using Congo red staining, circular dichroism, and atomic force microscopy revealed that GAPDH aggregates induced Aβ40 amyloidogenesis. In PC12 cells, GAPDH aggregates augmented Aβ40-induced cell death, concomitant with disruption of mitochondrial membrane potential. Furthermore, mice injected intracerebroventricularly with Aβ40 co-incubated with GAPDH aggregates exhibited Aβ40-induced pyramidal cell death and gliosis in the hippocampal CA3 region. These observations were accompanied by nuclear translocation of apoptosis-inducing factor and cytosolic release of cytochrome c from mitochondria. Finally, in the 3×Tg-AD mouse model of AD, GAPDH/Aβ co-aggregation and mitochondrial dysfunction were consistently detected in an age-dependent manner, and Aβ aggregate formation was attenuated by GAPDH siRNA treatment. Thus, this study suggests that GAPDH aggregates accelerate Aβ amyloidogenesis, subsequently leading to mitochondrial dysfunction and neuronal cell death in the pathogenesis of AD. PMID:26359500

  9. Thiophene functionalized silicon-containing aggregation-induced emission enhancement materials: applications as fluorescent probes for the detection of nitroaromatic explosives in aqueous-based solutions.

    Science.gov (United States)

    Wang, Xuefeng; Bian, Jiangyan; Xu, Lichao; Wang, Hua; Feng, Shengyu

    2015-12-28

    Two novel aggregation-induced emission enhancement (AIEE) molecules, namely, 3,4-diphenyl-2,5-di(2-thienyl)phenyltrimethylsilane (DPTB-TMS) and bis[3,4-diphenyl- 2,5-di(2-thienyl)phenyl]methylphenylsilane (DPTB-TMS) were designed and synthesized. The optical properties of the two silanes were completely opposite to the traditional luminescent materials. Unlike the aggregation caused quenching, they all emit faint fluorescence in the dispersed state, while emission intensity increased sharply in aggregate states. Fluorescence spectra showed that the two compounds exhibited AIEE properties and that is due to the weak π-π stacking caused by the restriction of intramolecular rotations of dye segments, particularly the -SiMe3 and thienyl groups in the aggregate state. As fluorescent (FL) probes, the fluorescence quenching behavior was further investigated. Thanks to the richer-electron thiophene groups, both compounds showed good performance in detecting nitroaromatics, especially picric acid (PA). The two AIEE FL probes exhibited better quenching efficiency in aqueous-based than in organic-based solutions. For DPTB-MPS, the addition of 80 μM nitrobenzene, 60 μM m-nitrobenzene and 40 μM PA resulted in about 50% quenching in aqueous solutions. The quenching mechanism would be electron transfer from silanes to nitroaromatics. This work provides a basis for designing organic-silanes with "abnormal" but useful optical properties and FL probes with AIEE properties for the detection of nitroaromatics.

  10. Protein-peptide interaction: study of heat-induced aggregation and gelation of β-lactoglobulin in the presence of two peptides from its own hydrolysate.

    Science.gov (United States)

    Kosters, Hans A; Wierenga, Peter A; de Vries, Renko; Gruppen, Harry

    2013-05-08

    Two peptides, [f135-158] and [f135-162]-SH, were used to study the binding of the peptides to native β-lactolobulin, as well as the subsequent effects on aggregation and gelation of β-lactoglobulin. The binding of the peptide [f135-158] to β-lactoglobulin at room temperature was confirmed by SELDI-TOF-MS. It was further illustrated by increased turbidity of mixed solutions of peptide and protein (at pH 7), indicating association of proteins and peptides in larger complexes. At pH below the isoelectric point of the protein, the presence of peptides did not lead to an increased turbidity, showing the absence of complexation. The protein-peptide complexes formed at pH 7 were found to dissociate directly upon heating. After prolonged heating, extensive aggregation was observed, whereas no aggregation was seen for the pure protein or pure peptide solutions. The presence of the free sulfhydryl group in [f135-162]-SH resulted in a 10 times increase in the amount of aggregation of β-lactoglobulin upon heating, illustrating the additional effect of the free sulfhydryl group. Subsequent studies on the gel strength of heat-induced gels also showed a clear difference between these two peptides. The replacement of additional β-lactoglobulin by [f135-158] resulted in a decrease in gel strength, whereas replacement by peptide [f135-162]-SH increased gel strength.

  11. Acetylsalicylic acid induces programmed cell death in Arabidopsis cell cultures.

    Science.gov (United States)

    García-Heredia, José M; Hervás, Manuel; De la Rosa, Miguel A; Navarro, José A

    2008-06-01

    Acetylsalicylic acid (ASA), a derivative from the plant hormone salicylic acid (SA), is a commonly used drug that has a dual role in animal organisms as an anti-inflammatory and anticancer agent. It acts as an inhibitor of cyclooxygenases (COXs), which catalyze prostaglandins production. It is known that ASA serves as an apoptotic agent on cancer cells through the inhibition of the COX-2 enzyme. Here, we provide evidences that ASA also behaves as an agent inducing programmed cell death (PCD) in cell cultures of the model plant Arabidopsis thaliana, in a similar way than the well-established PCD-inducing agent H(2)O(2), although the induction of PCD by ASA requires much lower inducer concentrations. Moreover, ASA is herein shown to be a more efficient PCD-inducing agent than salicylic acid. ASA treatment of Arabidopsis cells induces typical PCD-linked morphological and biochemical changes, namely cell shrinkage, nuclear DNA degradation, loss of mitochondrial membrane potential, cytochrome c release from mitochondria and induction of caspase-like activity. However, the ASA effect can be partially reverted by jasmonic acid. Taking together, these results reveal the existence of common features in ASA-induced animal apoptosis and plant PCD, and also suggest that there are similarities between the pathways of synthesis and function of prostanoid-like lipid mediators in animal and plant organisms.

  12. SIRT1 modulates aggregation and toxicity through deacetylation of the androgen receptor in cell models of SBMA.

    Science.gov (United States)

    Montie, Heather L; Pestell, Richard G; Merry, Diane E

    2011-11-30

    Posttranslational protein modifications can play a major role in disease pathogenesis; phosphorylation, sumoylation, and acetylation modulate the toxicity of a variety of proteotoxic proteins. The androgen receptor (AR) is substantially modified, in response to hormone binding, by phosphorylation, sumoylation, and acetylation; these modifications might thus contribute to DHT-dependent polyglutamine (polyQ)-expanded AR proteotoxicity in spinal and bulbar muscular atrophy (SBMA). SIRT1, a nuclear protein and deacetylase of the AR, is neuroprotective in many neurodegenerative disease models. Our studies reveal that SIRT1 also offers protection against polyQ-expanded AR by deacetylating the AR at lysines 630/632/633. This finding suggested that nuclear AR acetylation plays a role in the aberrant metabolism and toxicity of polyQ-expanded AR. Subsequent studies revealed that the polyQ-expanded AR is hyperacetylated and that pharmacologic reduction of acetylation reduces mutant AR aggregation. Moreover, genetic mutation to inhibit polyQ-expanded AR acetylation of lysines 630/632/633 substantially decreased its aggregation and completely abrogated its toxicity in cell lines and motor neurons. Our studies also reveal one means by which the AR acetylation state likely modifies polyQ-expanded AR metabolism and toxicity, through its effect on DHT-dependent AR stabilization. Overall, our findings reveal a neuroprotective function of SIRT1 that operates through its deacetylation of polyQ-expanded AR and highlight the potential of both SIRT1 and AR acetylation as powerful therapeutic targets in SBMA.

  13. Estimation of size of red blood cell aggregates using backscattering property of high-frequency ultrasound: In vivo evaluation

    Science.gov (United States)

    Kurokawa, Yusaku; Taki, Hirofumi; Yashiro, Satoshi; Nagasawa, Kan; Ishigaki, Yasushi; Kanai, Hiroshi

    2016-07-01

    We propose a method for assessment of the degree of red blood cell (RBC) aggregation using the backscattering property of high-frequency ultrasound. In this method, the scattering property of RBCs is extracted from the power spectrum of RBC echoes normalized by that from the posterior wall of a vein. In an experimental study using a phantom, employing the proposed method, the sizes of microspheres 5 and 20 µm in diameter were estimated to have mean values of 4.7 and 17.3 µm and standard deviations of 1.9 and 1.4 µm, respectively. In an in vivo experimental study, we compared the results between three healthy subjects and four diabetic patients. The average estimated scatterer diameters in healthy subjects at rest and during avascularization were 7 and 28 µm, respectively. In contrast, those in diabetic patients receiving both antithrombotic therapy and insulin therapy were 11 and 46 µm, respectively. These results show that the proposed method has high potential for clinical application to assess RBC aggregation, which may be related to the progress of diabetes.

  14. Curcumin reduces α-synuclein induced cytotoxicity in Parkinson's disease cell model

    Directory of Open Access Journals (Sweden)

    Emadi Sharareh

    2010-04-01

    Full Text Available Abstract Background Overexpression and abnormal accumulation of aggregated α-synuclein (αS have been linked to Parkinson's disease (PD and other synucleinopathies. αS can misfold and adopt a variety of morphologies but recent studies implicate oligomeric forms as the most cytotoxic species. Both genetic mutations and chronic exposure to neurotoxins increase αS aggregation and intracellular reactive oxygen species (ROS, leading to mitochondrial dysfunction and oxidative damage in PD cell models. Results Here we show that curcumin can alleviate αS-induced toxicity, reduce ROS levels and protect cells against apoptosis. We also show that both intracellular overexpression of αS and extracellular addition of oligomeric αS increase ROS which induces apoptosis, suggesting that aggregated αS may induce similar toxic effects whether it is generated intra- or extracellulary. Conclusions Since curcumin is a natural food pigment that can cross the blood brain barrier and has widespread medicinal uses, it has potential therapeutic value for treating PD and other neurodegenerative disorders.

  15. Induced pluripotent stem cells for regenerative medicine.

    Science.gov (United States)

    Hirschi, Karen K; Li, Song; Roy, Krishnendu

    2014-07-11

    With the discovery of induced pluripotent stem (iPS) cells, it is now possible to convert differentiated somatic cells into multipotent stem cells that have the capacity to generate all cell types of adult tissues. Thus, there is a wide variety of applications for this technology, including regenerative medicine, in vitro disease modeling, and drug screening/discovery. Although biological and biochemical techniques have been well established for cell reprogramming, bioengineering technologies offer novel tools for the reprogramming, expansion, isolation, and differentiation of iPS cells. In this article, we review these bioengineering approaches for the derivation and manipulation of iPS cells and focus on their relevance to regenerative medicine.

  16. TALEN-Induced Translocations in Human Cells.

    Science.gov (United States)

    Piganeau, Marion; Renouf, Benjamin; Ghezraoui, Hind; Brunet, Erika

    2016-01-01

    Induction of chromosomal translocations in human cells is of a great interest to study tumorigenesis and genome instability. Here, we explain in detail a method to induce translocations using the transcription activator-like effector nucleases (TALENs). We describe how to detect translocation formation by PCR, calculate translocation frequency by 96-well PCR screen, and analyze breakpoint junctions. When inducing cancer translocations, it is also possible to detect the fusion gene by FISH analysis or western blot.

  17. Identification of a small, naked virus in tumor-like aggregates in cell lines derived from a green turtle, Chelonia mydas, with fibropapillomas

    Science.gov (United States)

    Lu, Y.; Aguirre, A.A.; Work, T.M.; Balazs, G.H.; Nerurkar, V.R.; Yanagihara, R.

    2000-01-01

    Serial cultivation of cell lines derived from lung, testis, periorbital and tumor tissues of a green turtle (Chelonia mydas) with fibropapillomas resulted in the in vitro formation of tumor-like cell aggregates, ranging in size from 0.5 to 2.0 mm in diameter. Successful induction of tumor-like aggregates was achieved in a cell line derived from lung tissue of healthy green turtles, following inoculation with cell-free media from these tumor-bearing cell lines, suggesting the presence of a transmissible agent. Thin-section electron microscopy of the cell aggregates revealed massive collagen deposits and intranuclear naked viral particles, measuring 50??5 nm in diameter. These findings, together with the morphological similarity between these tumor-like cell aggregates and the naturally occurring tumor, suggest a possible association between this novel virus and the disease. Further characterization of this small naked virus will clarify its role in etiology of green turtle fibropapilloma, a life-threatening disease of this endangered marine species. Copyright (C) 2000 Elsevier Science B.V.

  18. Optically-Induced Cell Fusion on Cell Pairing Microstructures

    Science.gov (United States)

    Yang, Po-Fu; Wang, Chih-Hung; Lee, Gwo-Bin

    2016-02-01

    Cell fusion is a critical operation for numerous biomedical applications including cell reprogramming, hybridoma formation, cancer immunotherapy, and tissue regeneration. However, unstable cell contact and random cell pairings have limited efficiency and yields when utilizing traditional methods. Furthermore, it is challenging to selectively perform cell fusion within a group of cells. This study reports a new approach called optically-induced cell fusion (OICF), which integrates cell-pairing microstructures with an optically-induced, localized electrical field. By projecting light patterns onto a photoconductive film (hydrogen-rich, amorphous silicon) coated on an indium-tin-oxide (ITO) glass while an alternating current electrical field was applied between two such ITO glass slides, “virtual” electrodes could be generated that could selectively fuse pairing cells. At 10 kHz, a 57% cell paring rate and an 87% fusion efficiency were successfully achieved at a driving voltage of 20  Vpp, suggesting that this new technology could be promising for selective cell fusion within a group of cells.

  19. Induced pluripotent stem cells and neurodegenerative diseases.

    Science.gov (United States)

    Chen, Chao; Xiao, Shi-Fu

    2011-04-01

    Neurodegenerative diseases, including Parkinson's disease, Alzheimer's disease and Amyotrophic Lateral Sclerosis, are characterized by idiopathic neuron loss in different regions of the central nervous system, which contributes to the relevant dysfunctions in the patients. The application of cell replacement therapy using human embryonic stem (hES) cells, though having attracted much attention, has been hampered by the intrinsic ethical problems. It has been demonstrated that adult somatic cells can be reprogrammed into the embryonic state, called induced pluripotent stem (iPS) cells. It is soon realized that iPS cells may be an alternative source for cell replacement therapy, because it raises no ethical problems and using patient-specific iPS cells for autologous transplantation will not lead to immunological rejection. What's more, certain types of neurons derived from patient-specific iPS cells may display disease-relevant phenotypes. Thus, patient-specific iPS cells can provide a unique opportunity to directly investigate the pathological properties of relevant neural cells in individual patient, and to study the vulnerability of neural cells to pathogenic factors in vitro, which may help reveal the pathogenesis of many neurodegenerative diseases. In this review, the recent development in cellular treatment of neurodegenerative diseases using iPS cells was summarized, and the potential value of iPS cells in the modeling of neurodegenerative disease was discussed.

  20. Priming by chemokines restricts lateral mobility of the adhesion receptor LFA-1 and restores adhesion to ICAM-1 nano-aggregates on human mature dendritic cells.

    Directory of Open Access Journals (Sweden)

    Kyra J E Borgman

    Full Text Available LFA-1 is a leukocyte specific β2 integrin that plays a major role in regulating adhesion and migration of different immune cells. Recent data suggest that LFA-1 on mature dendritic cells (mDCs may function as a chemokine-inducible anchor during homing of DCs through the afferent lymphatics into the lymph nodes, by transiently switching its molecular conformational state. However, the role of LFA-1 mobility in this process is not yet known, despite that the importance of lateral organization and dynamics for LFA-1-mediated adhesion regulation is broadly recognized. Using single particle tracking approaches we here show that LFA-1 exhibits higher mobility on resting mDCs compared to monocytes. Lymphoid chemokine CCL21 stimulation of the LFA-1 high affinity state on mDCs, led to a significant reduction of mobility and an increase on the fraction of stationary receptors, consistent with re-activation of the receptor. Addition of soluble monomeric ICAM-1 in the presence of CCL21 did not alter the diffusion profile of LFA-1 while soluble ICAM-1 nano-aggregates in the presence of CCL21 further reduced LFA-1 mobility and readily bound to the receptor. Overall, our results emphasize the importance of LFA-1 lateral mobility across the membrane on the regulation of integrin activation and its function as adhesion receptor. Importantly, our data show that chemokines alone are not sufficient to trigger the high affinity state of the integrin based on the strict definition that affinity refers to the adhesion capacity of a single receptor to its ligand in solution. Instead our data indicate that nanoclustering of the receptor, induced by multi-ligand binding, is required to maintain stable cell adhesion once LFA-1 high affinity state is transiently triggered by inside-out signals.

  1. Priming by chemokines restricts lateral mobility of the adhesion receptor LFA-1 and restores adhesion to ICAM-1 nano-aggregates on human mature dendritic cells.

    Science.gov (United States)

    Borgman, Kyra J E; van Zanten, Thomas S; Manzo, Carlo; Cabezón, Raquel; Cambi, Alessandra; Benítez-Ribas, Daniel; Garcia-Parajo, Maria F

    2014-01-01

    LFA-1 is a leukocyte specific β2 integrin that plays a major role in regulating adhesion and migration of different immune cells. Recent data suggest that LFA-1 on mature dendritic cells (mDCs) may function as a chemokine-inducible anchor during homing of DCs through the afferent lymphatics into the lymph nodes, by transiently switching its molecular conformational state. However, the role of LFA-1 mobility in this process is not yet known, despite that the importance of lateral organization and dynamics for LFA-1-mediated adhesion regulation is broadly recognized. Using single particle tracking approaches we here show that LFA-1 exhibits higher mobility on resting mDCs compared to monocytes. Lymphoid chemokine CCL21 stimulation of the LFA-1 high affinity state on mDCs, led to a significant reduction of mobility and an increase on the fraction of stationary receptors, consistent with re-activation of the receptor. Addition of soluble monomeric ICAM-1 in the presence of CCL21 did not alter the diffusion profile of LFA-1 while soluble ICAM-1 nano-aggregates in the presence of CCL21 further reduced LFA-1 mobility and readily bound to the receptor. Overall, our results emphasize the importance of LFA-1 lateral mobility across the membrane on the regulation of integrin activation and its function as adhesion receptor. Importantly, our data show that chemokines alone are not sufficient to trigger the high affinity state of the integrin based on the strict definition that affinity refers to the adhesion capacity of a single receptor to its ligand in solution. Instead our data indicate that nanoclustering of the receptor, induced by multi-ligand binding, is required to maintain stable cell adhesion once LFA-1 high affinity state is transiently triggered by inside-out signals.

  2. Sodium Valproate Induces Cell Senescence in Human Hepatocarcinoma Cells

    Directory of Open Access Journals (Sweden)

    Hong-Mei An

    2013-12-01

    Full Text Available Hepatocarcinogenesis is associated with epigenetic changes, including histone deacetylases (HDACs. Epigenetic modulation by HDAC inhibition is a potentially valuable approach for hepatocellular carcinoma treatment. In present study, we evaluated the anticancer effects of sodium valproate (SVP, a known HDAC inhibitor, in human hepatocarcinoma cells. The results showed SVP inhibited the proliferation of Bel-7402 cells in a dose-dependent manner. Low dose SVP treatment caused a large and flat morphology change, positive SA-β-gal staining, and G0/G1 phase cell cycle arrest in human hepatocarcinoma cells. Low dose SVP treatment also increased acetylation of histone H3 and H4 on p21 promoter, accompanied by up-regulation of p21 and down-regulation of RB phosphorylation. These observations suggested that a low dose of SVP could induce cell senescence in hepatocarcinoma cells, which might correlate with hyperacetylation of histone H3 and H4, up-regulation of p21, and inhibition of RB phosphorylation. Since the effective concentration inducing cell senescence in hepatocarcinoma cells is clinically available, whether a clinical dose of SVP could induce cell senescence in clinical hepatocarcinoma is worthy of further study.

  3. Kit- and Fc epsilonRI-induced differential phosphorylation of the transmembrane adaptor molecule NTAL/LAB/LAT2 allows flexibility in its scaffolding function in mast cells

    DEFF Research Database (Denmark)

    Iwaki, Shoko; Spicka, Jiri; Tkaczyk, Christine;

    2008-01-01

    The transmembrane adaptor protein (TRAP), NTAL, is phosphorylated in mast cells following FcvarepsilonRI aggregation whereby it cooperates with LAT to induce degranulation. The Kit ligand, stem cell factor (SCF), enhances antigen-induced degranulation and this also appears to be NTAL......-knock down-human mast cells. The observations reported herein support the conclusion that NTAL may be differentially utilized by specific receptors for relaying alternative signals and this suggests a flexibility in the function of TRAPs not previously appreciated....

  4. Adenylyl cyclase localization to the uropod of aggregating Dictyostelium cells requires RacC

    Science.gov (United States)

    Wang, C.; Jung, D.; Cao, Z.; Chung, C. Y.

    2015-01-01

    The localization of adenylyl cyclase A (ACA) to uropod of cells is required for the stream formation during Dictyostelium development. RacC is a Dictyostelium orthologue of Cdc42. We identified a streaming defect of racC− cells as they are clearly less polarized and form smaller and fragmented streams. ACA-YFP is mainly associated with intracellular vesicular structures, but not with the plasma membrane in racC− cells. racC− cells have a slightly higher number of vesicles than Ax3 cells, suggesting that the defect of ACA trafficking is not simply due to the lack of vesicle formation. While the ACA-YFP vesicles traveled with an average velocity of 9.1 µm/min in Ax3 cells, a slow and diffusional movement without direction with an average velocity of 4 µm/min was maintained in racC− cells. Images acquired by using total internal reflection fluorescence (TIRF) microscopy and fluorescence recovery after photobleaching (FRAP) analysis revealed that a significantly decreased number of ACA-YFP vesicles appeared near the cell membrane, indicating a defect in ACA-YFP vesicle trafficking. These results suggest an important role of RacC in the rapid and directional movements of ACA vesicles on microtubules to the plasma membrane, especially to the back of polarized cell. PMID:26315268

  5. Failure of cell cleavage induces senescence in tetraploid primary cells.

    Science.gov (United States)

    Panopoulos, Andreas; Pacios-Bras, Cristina; Choi, Justin; Yenjerla, Mythili; Sussman, Mark A; Fotedar, Rati; Margolis, Robert L

    2014-10-15

    Tetraploidy can arise from various mitotic or cleavage defects in mammalian cells, and inheritance of multiple centrosomes induces aneuploidy when tetraploid cells continue to cycle. Arrest of the tetraploid cell cycle is therefore potentially a critical cellular control. We report here that primary rat embryo fibroblasts (REF52) and human foreskin fibroblasts become senescent in tetraploid G1 after drug- or small interfering RNA (siRNA)-induced failure of cell cleavage. In contrast, T-antigen-transformed REF52 and p53+/+ HCT116 tumor cells rapidly become aneuploid by continuing to cycle after cleavage failure. Tetraploid primary cells quickly become quiescent, as determined by loss of the Ki-67 proliferation marker and of the fluorescent ubiquitination-based cell cycle indicator/late cell cycle marker geminin. Arrest is not due to DNA damage, as the γ-H2AX DNA damage marker remains at control levels after tetraploidy induction. Arrested tetraploid cells finally become senescent, as determined by SA-β-galactosidase activity. Tetraploid arrest is dependent on p16INK4a expression, as siRNA suppression of p16INK4a bypasses tetraploid arrest, permitting primary cells to become aneuploid. We conclude that tetraploid primary cells can become senescent without DNA damage and that induction of senescence is critical to tetraploidy arrest.

  6. Abiotic factors in colony formation: effects of nutrition and light on extracellular polysaccharide production and cell aggregates of Microcystis aeruginosa

    Science.gov (United States)

    Yang, Zhen; Kong, Fanxiang

    2013-07-01

    Colony morphology is important for Microcystis to sustain a competitive advantage in eutrophic lakes. The mechanism of colony formation in Microcystis is currently unclear. Extracellular polysaccharide (EPS) has been reported to play an important role in cell aggregate formation of some phytoplankton. Microcystis aeruginosa was cultivated under varied abiotic conditions, including different nutrient, light, and temperature conditions, to investigate their effects on EPS production and morphological change. The results show that nutrient concentration and light intensity have great effects on EPS productionin M. aeruginosa. There was a considerable increase in EPS production after M. aeruginosa was cultivated in adjusted culture conditions similar to those present in the field (28.9 mg C/L, 1.98 mg N/L, 0.65 mg P/L, light intensity: 100 μmol/(m2 · s)). These results indicate that abiotic factors might be one of the triggers for colony formation in Microcystis.

  7. Mucin biopolymers prevent bacterial aggregation by retaining cells in the free-swimming state

    Science.gov (United States)

    Caldara, Marina; Friedlander, Ronn S.; Kavanaugh, Nicole L.; Aizenberg, Joanna; Foster, Kevin R.; Ribbeck, Katharina

    2013-01-01

    Summary Many species of bacteria form surface-attached communities known as biofilms. Surrounded in secreted polymers, these aggregates are difficult to both prevent and eradicate, posing problems for medicine and industry [1, 2]. Humans play host to hundreds of trillions of microbes that live adjacent to our epithelia and we are typically able to prevent harmful colonization. Mucus, the hydrogel overlying all wet epithelia in the body, can prevent bacterial contact with the underlying tissue. The digestive tract, for example, is lined by a firmly adherent mucus layer that is typically devoid of bacteria, followed by a second, loosely adherent layer that contains numerous bacteria [3]. Here, we investigate mucus's role as a principle arena for host-microbe interactions. Using defined in vitro assays, we found that mucin biopolymers, the main functional constituents of mucus, promote the motility of planktonic bacteria, and prevent their adhesion to underlying surfaces. The deletion of motility genes, however, allows Pseudomonas aeruginosa to overcome the dispersive effects of mucus and form suspended antibiotic-resistant flocs, which mirror the clustered morphology of immotile natural isolates found in the cystic fibrosis lung mucus [4, 5]. Mucus may offer new strategies to target bacterial virulence, such as the design of anti-biofilm coatings for implants. PMID:23142047

  8. Chirality-driven intrinsic spin-glass ordering and field-induced ferromagnetism in Ni3Al nanoparticle aggregates

    Science.gov (United States)

    Kaul, S. N.; Messala, Umasankar

    2016-03-01

    Weak itinerant-electron ferromagnet Ni3Al is driven to magnetic instability (quantum critical point, QCP, where the long-range ferromagnetic order of the bulk ceases to exist) by reducing the average crystallite size to d=50 nm. 'Zero-field' (H=0) linear and nonlinear ac-susceptibilities, measured on Ni3Al nanoparticle aggregates, with d=50 nm (S1) and d=5 nm (S2), provide strong evidence for two spin glass (SG)-like thermodynamic phase transitions: one at Ti(H = 0) ≃ 30 K (Ti† (H = 0) ≃ 230 K) and the other at a lower temperature Tp(H = 0) ≃ 8 K (Th(H = 0) ≃ 52 K) in S1 (S2). 'In-field' (H ≠ 0) linear ac-susceptibility and dc magnetization demonstrate that the thermodynamic nature of these transitions is preserved in finite fields. The presently determined H-T phase diagrams for the samples S1 and S2 are compared with those predicted by the Kotliar-Sompolinsky and Gabay-Toulouse mean-field models and Monte Carlo simulations, based on the chirality-driven spin glass (SG) ordering scenario, for a three-dimensional nearest-neighbor Heisenberg SG system with or without weak random anisotropy. Such a detailed comparison permits us to unambiguously identify various 'zero-field' and 'in-field' SG phase transitions as: (i) the simultaneous paramagnetic (PM)-chiral glass (CG) and PM-SG phase transitions at Ti(H), (ii) the PM-CG transition at Ti† (H), (iii) the replica symmetry-breaking SG transition at Tp(H), and (iv) the continuous spin-rotation symmetry-breaking SG transition at Th(H). In the presence of random anisotropy, magnetization fails to saturate even at 90 kOe in S1 whereas negligibly small anisotropy allows even fields as weak as 1 kOe to saturate magnetization and induce ferromagnetism in S2. Due to the proximity to CG/SG-QCP, magnetization and susceptibility both exhibit non-Fermi liquid behavior over a wide range at low temperatures.

  9. Fate of bone marrow mesenchymal stem cells following the allogeneic transplantation of cartilaginous aggregates into osteochondral defects of rabbits.

    Science.gov (United States)

    Yoshioka, Tomokazu; Mishima, Hajime; Kaul, Zeenia; Ohyabu, Yoshimi; Sakai, Shinsuke; Ochiai, Naoyuki; Kaul, Sunil C; Wadhwa, Renu; Uemura, Toshimasa

    2011-06-01

    The purpose of this study was to track mesenchymal stem cells (MSCs) labelled with internalizing quantum dots (i-QDs) in the reparative tissues, following the allogeneic transplantation of three-dimensional (3D) cartilaginous aggregates into the osteochondral defects of rabbits. QDs were conjugated with a unique internalizing antibody against a heat shock protein-70 (hsp70) family stress chaperone, mortalin, which is upregulated and expressed on the surface of dividing cells. The i-QDs were added to the culture medium for 24 h. Scaffold-free cartilaginous aggregates formed from i-QD-labelled MSCs (i-MSCs), using a 3D culture system with chondrogenic supplements for 1 week, were transplanted into osteochondral defects of rabbits. At 4, 8 and 26 weeks after the transplantation, the reparative tissues were evaluated macroscopically, histologically and fluoroscopically. At as early as 4 weeks, the defects were covered with a white tissue resembling articular cartilage. In histological appearance, the reparative tissues resembled hyaline cartilage on safranin-O staining throughout the 26 weeks. In the deeper portion, subchondral bone and bone marrow were well remodelled. On fluoroscopic evaluation, QDs were tracked mainly in bone marrow stromata, with some signals detected in cartilage and the subchondral bone layer. We showed that the labelling of rabbit MSCs with anti-mortalin antibody-conjugated i-QDs is a tolerable procedure and provides a stable fluorescence signal during the cartilage repair process for up to 26 weeks after transplantation. The results suggest that i-MSCs did not inhibit, and indeed contributed to, the regeneration of osteochondral defects.

  10. Nicotine promotes Streptococcus mutans extracellular polysaccharide synthesis, cell aggregation and overall lactate dehydrogenase activity.

    Science.gov (United States)

    Huang, R; Li, M; Gregory, R L

    2015-08-01

    Several epidemiology studies have reported a positive relationship between smoking and dental caries. Nicotine, an alkaloid component of tobacco, has been demonstrated to stimulate biofilm formation and metabolic activity of Streptococcus mutans, one of the most important pathogens of dental caries. The first aim of the present study was to explore the possible mechanisms leading to increased biofilm by nicotine treatment from three aspects, extracellular polysaccharides (EPS) synthesis, glucosyltransferase (Gtf) synthesis and glucan-binding protein (Gbp) synthesis at the mRNA and protein levels. The second aim was to investigate how nicotine affects S. mutans virulence, particular in lactate dehydrogenase (LDH) activity. Confocal laser scanning microscopy results demonstrated that both biofilm bacterial cell numbers and EPS were increased by nicotine. Gtf and GbpA protein expression of S. mutans planktonic cells were upregulated while GbpB protein expression of biofilm cells were downregulated by nicotine. The mRNA expression trends of those genes were mostly consistent with results on protein level but not statistically significant, and gtfD and gbpD of biofilm cells were inhibited. Nicotine was not directly involved in S. mutans LDH activity. However, since it increases the total number of bacterial cells in biofilm, the overall LDH activity of S. mutans biofilm is increased. In conclusion, nicotine stimulates S. mutans planktonic cell Gtf and Gbp expression. This leads to more planktonic cells attaching to the dental biofilm. Increased cell numbers within biofilm results in higher overall LDH activity. This contributes to caries development in smokers.

  11. Temperature-induced aggregation in aqueous solutions of pluronic F68 triblock copolymer containing small amount of o-xylene

    DEFF Research Database (Denmark)

    Borbely, S.; Pedersen, J.S.

    2000-01-01

    The temperature- and concentration-dependent aggregation of EO(78)PO(30)EO(78) triblock copolymer in aqueous solutions has been studied in the concentration range from 5 to 200 g/dm(3) at 25 degrees C, 45 degrees C and 60 degrees C. The influence of o-xylene on the micellization was measured at v...... at various (from 5 to 40) molar ratio of additive to polymer. The aggregates were described as polydisperse spheres with tethered Gaussian polymer chains. The results of the least-squares fit is briefly discussed. (C) 2000 Elsevier Science B.V. All rights reserved....

  12. Cell Death Mechanisms Induced by Cytotoxic Lymphocytes

    Institute of Scientific and Technical Information of China (English)

    Ch(a)vez-Gal(a)n L; Arenas-Del Angel MC; Zenteno E; Ch(a)vez R; Lascurain R

    2009-01-01

    One of the functions of the immune system is to recognize and destroy abnormal or infected cells to maintain homeostasis. This is accomplished by cytotoxic lymphocytes. Cytotoxicity is a highly organized multifactor process. Here, we reviewed the apoptosis pathways induced by the two main cytotoxic lymphocyte subsets, natural killer (NK) cells and CD8+T cells. In base to recent experimental evidence, we reviewed NK receptors involved in recognition of target-cell, as well as lytic molecules such as perforin, granzymes-A and -B, and granulysin. In addition, we reviewed the Fas-FasL intercellular linkage mediated pathway, and briefly the cross-linking of tumor necrosis factor (TNF) and TNF receptor pathway. We discussed three models of possible molecular interaction between lyric molecules from effector cytotoxic cells and target-cell membrane to induction of apoptosis.

  13. Induced Pluripotent Stem Cells in Cardiovascular Medicine

    Directory of Open Access Journals (Sweden)

    Toru Egashira

    2011-01-01

    Full Text Available Induced pluripotent stem (iPS cells are generated by reprogramming human somatic cells through the forced expression of several embryonic stem (ES cell-specific transcription factors. The potential of iPS cells is having a significant impact on regenerative medicine, with the promise of infinite self-renewal, differentiation into multiple cell types, and no problems concerning ethics or immunological rejection. Human iPS cells are currently generated by transgene introduction principally through viral vectors, which integrate into host genomes, although the associated risk of tumorigenesis is driving research into nonintegration methods. Techniques for pluripotent stem cell differentiation and purification to yield cardiomyocytes are also advancing constantly. Although there remain some unsolved problems, cardiomyocyte transplantation may be a reality in the future. After those problems will be solved, applications of human iPS cells in human cardiovascular regenerative medicine will be envisaged for the future. Furthermore, iPS cell technology has generated new human disease models using disease-specific cells. This paper summarizes the progress of iPS cell technology in cardiovascular research.

  14. Inhibition of glutamate receptors reduces the homocysteine-induced whole blood platelet aggregation but does not affect superoxide anion generation or platelet membrane fluidization.

    Science.gov (United States)

    Karolczak, Kamil; Pieniazek, Anna; Watala, Cezary

    2017-01-01

    Homocysteine (Hcy) is an excitotoxic amino acid. It is potentially possible to prevent Hcy-induced toxicity, including haemostatic impairments, by antagonizing glutaminergic receptors. Using impedance aggregometry with arachidonate and collagen as platelet agonists, we tested whether the blockade of platelet NMDA (N-methyl-D-aspartate), AMPA (α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid) and kainate receptors with their inhibitors: MK-801 (dizocilpine hydrogen maleate, [5R,10S]-[+]-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine), CNQX (7-nitro-2,3-dioxo-1,4-dihydroquinoxaline-6-carbonitrile) and UBP-302 (2-{[3-[(2S)-2-amino-2-carboxyethyl]-2,6-dioxo-3,6-dihydropyrimidin 1(2H)-yl]methyl}benzoic acid) may hamper Hcy-dependent platelet aggregation. All the tested compounds significantly inhibited Hcy-augmented aggregation of blood platelets stimulated either with arachidonate or collagen. Hcy stimulated the generation of superoxide anion in whole blood samples in a concentration-dependent manner; however, this process appeared as independent on ionotropic glutamate receptors, as well as on NADPH oxidase and protein kinase C, and was not apparently associated with the extent of either arachidonate- or collagen-dependent platelet aggregation. Moreover, Hcy acted as a significant fluidizer of surface (more hydrophilic) and inner (more hydrophobic) regions of platelet membrane lipid bilayer, when used at the concentration range from 10 to 50 µmol/l. However, this effect was independent on the Hcy action through glutamate ionotropic receptors, since there was no effects of MK-801, CNQX or UBP-302 on Hcy-mediated membrane fluidization. In conclusion, Hcy-induced changes in whole blood platelet aggregation are mediated through the ionotopic excitotoxic receptors, although the detailed mechanisms underlying such interactions remain to be elucidated.

  15. Trophoblast lineage cells derived from human induced pluripotent stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Ying, E-mail: ying.chen@hc.msu.edu [Department of Obstetrics, Gynecology and Reproductive Biology, Michigan State University, 333 Bostwick NE, Grand Rapids, MI 49503 (United States); Wang, Kai; Chandramouli, Gadisetti V.R. [Department of Obstetrics, Gynecology and Reproductive Biology, Michigan State University, 333 Bostwick NE, Grand Rapids, MI 49503 (United States); Knott, Jason G. [Developmental Epigenetics Laboratory, Department of Animal Science, Michigan State University (United States); Leach, Richard, E-mail: Richard.leach@hc.msu.edu [Department of Obstetrics, Gynecology and Reproductive Biology, Michigan State University, 333 Bostwick NE, Grand Rapids, MI 49503 (United States); Department of Obstetrics, Gynecology and Women’s Health, Spectrum Health Medical Group (United States)

    2013-07-12

    Highlights: •Epithelial-like phenotype of trophoblast lineage cells derived from human iPS cells. •Trophoblast lineage cells derived from human iPS cells exhibit trophoblast function. •Trophoblasts from iPS cells provides a proof-of-concept in regenerative medicine. -- Abstract: Background: During implantation, the blastocyst trophectoderm attaches to the endometrial epithelium and continues to differentiate into all trophoblast subtypes, which are the major components of a placenta. Aberrant trophoblast proliferation and differentiation are associated with placental diseases. However, due to ethical and practical issues, there is almost no available cell or tissue source to study the molecular mechanism of human trophoblast differentiation, which further becomes a barrier to the study of the pathogenesis of trophoblast-associated diseases of pregnancy. In this study, our goal was to generate a proof-of-concept model for deriving trophoblast lineage cells from induced pluripotency stem (iPS) cells from human fibroblasts. In future studies the generation of trophoblast lineage cells from iPS cells established from patient’s placenta will be extremely useful for studying the pathogenesis of individual trophoblast-associated diseases and for drug testing. Methods and results: Combining iPS cell technology with BMP4 induction, we derived trophoblast lineage cells from human iPS cells. The gene expression profile of these trophoblast lineage cells was distinct from fibroblasts and iPS cells. These cells expressed markers of human trophoblasts. Furthermore, when these cells were differentiated they exhibited invasive capacity and placental hormone secretive capacity, suggesting extravillous trophoblasts and syncytiotrophoblasts. Conclusion: Trophoblast lineage cells can be successfully derived from human iPS cells, which provide a proof-of-concept tool to recapitulate pathogenesis of patient placental trophoblasts in vitro.

  16. P2Y6 Receptor Antagonist MRS2578 Inhibits Neutrophil Activation and Aggregated Neutrophil Extracellular Trap Formation Induced by Gout-Associated Monosodium Urate Crystals.

    Science.gov (United States)

    Sil, Payel; Hayes, Craig P; Reaves, Barbara J; Breen, Patrick; Quinn, Shannon; Sokolove, Jeremy; Rada, Balázs

    2017-01-01

    Human neutrophils (polymorphonuclear leukocytes [PMNs]) generate inflammatory responses within the joints of gout patients upon encountering monosodium urate (MSU) crystals. Neutrophil extracellular traps (NETs) are found abundantly in the synovial fluid of gout patients. The detailed mechanism of MSU crystal-induced NET formation remains unknown. Our goal was to shed light on possible roles of purinergic signaling and neutrophil migration in mediating NET formation induced by MSU crystals. Interaction of human neutrophils with MSU crystals was evaluated by high-throughput live imaging using confocal microscopy. We quantitated NET levels in gout synovial fluid supernatants and detected enzymatically active neutrophil primary granule enzymes, myeloperoxidase, and human neutrophil elastase. Suramin and PPADS, general P2Y receptor blockers, and MRS2578, an inhibitor of the purinergic P2Y6 receptor, blocked NET formation triggered by MSU crystals. AR-C25118925XX (P2Y2 antagonist) did not inhibit MSU crystal-stimulated NET release. Live imaging of PMNs showed that MRS2578 represses neutrophil migration and blocked characteristic formation of MSU crystal-NET aggregates called aggregated NETs. Interestingly, the store-operated calcium entry channel inhibitor (SK&F96365) also reduced MSU crystal-induced NET release. Our results indicate that the P2Y6/store-operated calcium entry/IL-8 axis is involved in MSU crystal-induced aggregated NET formation, but MRS2578 could have additional effects affecting PMN migration. The work presented in the present study could lead to a better understanding of gouty joint inflammation and help improve the treatment and care of gout patients.

  17. Heat induces gene amplification in cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Yan, Bin, E-mail: yanbin@mercyhealth.com [Department of Radiation Oncology, University of Mississippi Medical Center, Jackson, MS 39213 (United States); Mercy Cancer Center, Mercy Medical Center-North Iowa, Mason City, IA 50401 (United States); Ouyang, Ruoyun [Department of Respiratory Medicine, The Second Xiangya Hospital, Xinagya School of Medicine, Central South University, Changsha 410011 (China); Huang, Chenghui [Department of Radiation Oncology, University of Mississippi Medical Center, Jackson, MS 39213 (United States); Department of Oncology, The Third Xiangya Hospital, Xinagya School of Medicine, Central South University, Changsha 410013 (China); Liu, Franklin [Department of Radiation Oncology, Duke University Medical Center, Durham, NC 27710 (United States); Neill, Daniel [Department of Radiation Oncology, University of Mississippi Medical Center, Jackson, MS 39213 (United States); Li, Chuanyuan [Dermatology, Duke University Medical Center, Durham, NC 27710 (United States); Dewhirst, Mark [Department of Radiation Oncology, Duke University Medical Center, Durham, NC 27710 (United States)

    2012-10-26

    Highlights: Black-Right-Pointing-Pointer This study discovered that heat exposure (hyperthermia) results in gene amplification in cancer cells. Black-Right-Pointing-Pointer Hyperthermia induces DNA double strand breaks. Black-Right-Pointing-Pointer DNA double strand breaks are considered to be required for the initiation of gene amplification. Black-Right-Pointing-Pointer The underlying mechanism of heat-induced gene amplification is generation of DNA double strand breaks. -- Abstract: Background: Hyperthermia plays an important role in cancer therapy. However, as with radiation, it can cause DNA damage and therefore genetic instability. We studied whether hyperthermia can induce gene amplification in cancer cells and explored potential underlying molecular mechanisms. Materials and methods: (1) Hyperthermia: HCT116 colon cancer cells received water-submerged heating treatment at 42 or 44 Degree-Sign C for 30 min; (2) gene amplification assay using N-(phosphoacetyl)-L-aspartate (PALA) selection of cabamyl-P-synthetase, aspartate transcarbarmylase, dihydro-orotase (cad) gene amplified cells; (3) southern blotting for confirmation of increased cad gene copies in PALA-resistant cells; (4) {gamma}H2AX immunostaining to detect {gamma}H2AX foci as an indication for DNA double strand breaks. Results: (1) Heat exposure at 42 or 44 Degree-Sign C for 30 min induces gene amplification. The frequency of cad gene amplification increased by 2.8 and 6.5 folds respectively; (2) heat exposure at both 42 and 44 Degree-Sign C for 30 min induces DNA double strand breaks in HCT116 cells as shown by {gamma}H2AX immunostaining. Conclusion: This study shows that heat exposure can induce gene amplification in cancer cells, likely through the generation of DNA double strand breaks, which are believed to be required for the initiation of gene amplification. This process may be promoted by heat when cellular proteins that are responsible for checkpoints, DNA replication, DNA repair and

  18. Combined Protein A and size exclusion high performance liquid chromatography for the single-step measurement of mAb, aggregates and host cell proteins.

    Science.gov (United States)

    Gjoka, Xhorxhi; Schofield, Mark; Cvetkovic, Aleksandar; Gantier, Rene

    2014-12-01

    Quantification of monoclonal antibody (mAb) monomer, mAb aggregates, and host cell proteins (HCPs) is critical for the optimization of the mAb production process. The present work describes a single high throughput analytical tool capable of tracking the concentration of mAb, mAb aggregate and HCPs in a growing cell culture batch. By combining two analytical HPLC methods, Protein A affinity and size-exclusion chromatography (SEC), it is possible to detect a relative increase or decrease in the concentration of all three entities simultaneously. A comparison of the combined Protein A-SEC assay to SEC alone was performed, demonstrating that it can be useful tool for the quantification of mAb monomer along with trending data for mAb aggregate and HCP. Furthermore, the study shows that the Protein A-SEC method is at least as accurate as other commonly used analytical methods such as ELISA and Bradford.

  19. Ionizing radiation induces stemness in cancer cells.

    Directory of Open Access Journals (Sweden)

    Laura Ghisolfi

    Full Text Available The cancer stem cell (CSC model posits the presence of a small number of CSCs in the heterogeneous cancer cell population that are ultimately responsible for tumor initiation, as well as cancer recurrence and metastasis. CSCs have been isolated from a variety of human cancers and are able to generate a hierarchical and heterogeneous cancer cell population. CSCs are also resistant to conventional chemo- and radio-therapies. Here we report that ionizing radiation can induce stem cell-like properties in heterogeneous cancer cells. Exposure of non-stem cancer cells to ionizing radiation enhanced spherogenesis, and this was accompanied by upregulation of the pluripotency genes Sox2 and Oct3/4. Knockdown of Sox2 or Oct3/4 inhibited radiation-induced spherogenesis and increased cellular sensitivity to radiation. These data demonstrate that ionizing radiation can activate stemness pathways in heterogeneous cancer cells, resulting in the enrichment of a CSC subpopulation with higher resistance to radiotherapy.

  20. Resveratrol induces apoptosis in pancreatic cancer cells

    Institute of Scientific and Technical Information of China (English)

    ZHOU Jia-hua; CHENG Hai-yan; YU Ze-qian; HE Dao-wei; PAN Zheng; YANG De-tong

    2011-01-01

    Background Pancreatic cancer is one of the most lethal human cancers with a very low survival rate of 5 years.Conventional cancer treatments including surgery, radiation, chemotherapy or combinations of these show little effect on this disease. Several proteins have been proved critical to the development and the progression of pancreatic cancer.The aim of this study was to investigate the effect of resveratrol on apoptosis in pancreatic cancer cells.Methods Several pancreatic cancer cell lines were screened by resveratrol, and its toxicity was tested by normal pancreatic cells. Western blotting was then performed to analyze the molecular mechanism of resveratrol induced apoptosis of pancreatic cancer cell lines.Results In the screened pancreatic cancer cell lines, capan-2 and colo357 showed high sensitivity to resveratrol induced apoptosis. Resveratrol exhibited insignificant toxicity to normal pancreatic cells. In resveratrol sensitive cells,capan-2 and colo357, the activation of caspase-3 was detected and showed significant caspase-3 activation upon resveratrol treatment; p53 and p21 were also detected up-regulated upon resveratrol treatment.Conclusion Resveratrol provides a promising anti-tumor stratagy to fight against pancreatic cancer.

  1. Active site cysteine-null glyceraldehyde-3-phosphate dehydrogenase (GAPDH) rescues nitric oxide-induced cell death.

    Science.gov (United States)

    Kubo, Takeya; Nakajima, Hidemitsu; Nakatsuji, Masatoshi; Itakura, Masanori; Kaneshige, Akihiro; Azuma, Yasu-Taka; Inui, Takashi; Takeuchi, Tadayoshi

    2016-02-29

    Glyceraldehyde-3-phosphate dehydrogenase (GAPDH), a homotetrameric enzyme involved in a key step of glycolysis, also has a role in mediating cell death under nitrosative stress. Our previous reports suggest that nitric oxide-induced intramolecular disulfide-bonding GAPDH aggregation, which occurs through oxidation of the active site cysteine (Cys-152), participates in a mechanism to account for nitric oxide-induced death signaling in some neurodegenerative/neuropsychiatric disorders. Here, we demonstrate a rescue strategy for nitric oxide-induced cell death accompanied by GAPDH aggregation in a mutant with a substitution of Cys-152 to alanine (C152A-GAPDH). Pre-incubation of purified wild-type GAPDH with C152A-GAPDH under exposure to nitric oxide inhibited wild-type GAPDH aggregation in a concentration-dependent manner in vitro. Several lines of structural analysis revealed that C152A-GAPDH extensively interfered with nitric oxide-induced GAPDH-amyloidogenesis. Overexpression of doxycycline-inducible C152A-GAPDH in SH-SY5Y neuroblastoma significantly rescued nitric oxide-induced death, concomitant with the decreased formation of GAPDH aggregates. Further, both co-immunoprecipitation assays and simulation models revealed a heterotetramer composed of one dimer each of wild-type GAPDH and C152A-GAPDH. These results suggest that the C152A-GAPDH mutant acts as a dominant-negative molecule against GAPDH aggregation via the formation of this GAPDH heterotetramer. This study may contribute to a new therapeutic approach utilizing C152A-GAPDH against brain damage in nitrosative stress-related disorders.

  2. Resveratrol, Acetyl-Resveratrol, and Polydatin Exhibit Antigrowth Activity against 3D Cell Aggregates of the SKOV-3 and OVCAR-8 Ovarian Cancer Cell Lines

    Directory of Open Access Journals (Sweden)

    Simon J. Hogg

    2015-01-01

    Full Text Available Resveratrol has aroused significant scientific interest as it has been claimed that it exhibits a spectrum of health benefits. These include effects as an anti-inflammatory and an antitumour compound. The purpose of this study was to investigate and compare any potential antigrowth effects of resveratrol and two of its derivatives, acetyl-resveratrol and polydatin, on 3D cell aggregates of the EGFR/Her-2 positive and negative ovarian cancer cell lines SKOV-3 and OVCAR-8, respectively. Results showed that resveratrol and acetyl-resveratrol reduced cell growth in the SKOV-3 and OVCAR-8 in a dose-dependant manner. The growth reduction was mediated by the induction of apoptosis via the cleavage of poly(ADP-ribose polymerase (PARP-1. At lower concentrations, 5 and 10 µM, resveratrol, acetyl-resveratrol, and polydatin were less effective than higher concentrations, 50 and 100 µM. In SKOV-3 line, at higher concentrations, resveratrol and polydatin significantly reduced the phosphorylation of Her-2 and EGFR and the expression of Erk. Acetyl-resveratrol, on the other hand, did not change the activation of Her-2 and EGFR. Resveratrol, acetyl-resveratrol, and polydatin suppressed the secretion of VEGF in a dose-dependant fashion. In the OVCAR-8 cell line, resveratrol and acetyl-resveratrol at 5 and 10 µM increased the activation of Erk. Above these concentrations they decreased activation. Polydatin did not produce this effect. This study demonstrates that resveratrol and its derivatives may inhibit growth of 3D cell aggregates of ovarian cancer cell lines via different signalling molecules. Resveratrol and its derivatives, therefore, warrant further in vivo evaluation to assess their potential clinical utility.

  3. Small steps to maturity. Fuel cell heating aggregates; Muehsames Herantasten. Brennstoffzellen-Heizgeraete

    Energy Technology Data Exchange (ETDEWEB)

    Schmid, Wolfgang

    2009-07-15

    By 2015 at the latest, fuel cell heating systems are expected to be available to the end user after practical tests have been completed. The ultimate breakthrough is to be achieved by a funding project of several thousand million Euros of the German Federal Minister of Transportation, Construction and Urban Development (BMVBS). But the 2009 Hanover Fair showed that the development of fuel cell heating systems for serial production is a process of small steps taken one by one. Also, it becomes clear that the optimistic forecasts of a mass market that were sometimes voiced will not become reality. (orig.)

  4. Erythrocyte aggregation: Basic aspects and clinical importance

    OpenAIRE

    Oğuz K. Başkurt; Meiselman, Herbert J.

    2013-01-01

    Red blood cells (RBC) aggregate to form two- and three-dimensional structures when suspended in aqueous solutions containing large plasma proteins or polymers; this aggregation is reversible and shear dependent (i.e., dispersed at high shear and reformed at low or stasis). The extent of aggregation is the main determinant of low shear blood viscosity, thus predicting an inverse relationship between aggregation and in vivo blood flow. However, the effects of aggregation on hemodynamic mechanis...

  5. Effect of extraction pH on heat-induced aggregation, gelation and microstructure of protein isolate from quinoa (Chenopodium quinoa Willd).

    Science.gov (United States)

    Ruiz, Geraldine Avila; Xiao, Wukai; van Boekel, Martinus; Minor, Marcel; Stieger, Markus

    2016-10-15

    The aim of this study was to determine the influence of extraction pH on heat-induced aggregation, gelation and microstructure of suspensions of protein isolates extracted from quinoa (Chenopodium quinoa Willd). Quinoa seed protein was extracted by alkaline treatment at various pH values (pH 8 (E8), 9 (E9), 10 (E10) and 11 (E11)), followed by acid precipitation. The obtained protein isolates were freeze dried. The protein isolates E8 and E9 resulted in a lower protein yield as well as less protein denaturation. These isolates also had a higher protein purity, more protein bands at higher molecular weights, and a higher protein solubility in the pH range of 3-4.5, compared to the isolates E10 and E11. Heating the 10%w/w protein isolate suspensions E8 and E9 led to increased aggregation, and semi-solid gels with a dense microstructure were formed. The isolate suspensions E10 and E11, on the other hand, aggregated less, did not form self-supporting gels and had loose particle arrangements. We conclude that extraction pH plays an important role in determining the functionality of quinoa protein isolates.

  6. Sulindac Sulfide Induces the Formation of Large Oligomeric Aggregates of the Alzheimer's Disease Amyloid-β Peptide Which Exhibit Reduced Neurotoxicity.

    Science.gov (United States)

    Prade, Elke; Barucker, Christian; Sarkar, Riddhiman; Althoff-Ospelt, Gerhard; Lopez del Amo, Juan Miguel; Hossain, Shireen; Zhong, Yifei; Multhaup, Gerd; Reif, Bernd

    2016-03-29

    Alzheimer's disease is characterized by deposition of the amyloid β-peptide (Aβ) in brain tissue of affected individuals. In recent years, many potential lead structures have been suggested that can potentially be used for diagnosis and therapy. However, the mode of action of these compounds is so far not understood. Among these small molecules, the nonsteroidal anti-inflammatory drug (NSAID) sulindac sulfide received a lot of attention. In this manuscript, we characterize the interaction between the monomeric Aβ peptide and the NSAID sulindac sulfide. We find that sulindac sulfide efficiently depletes the pool of toxic oligomers by enhancing the rate of fibril formation. In vitro, sulindac sulfide forms colloidal particles which catalyze the formation of fibrils. Aggregation is immediate, presumably by perturbing the supersaturated Aβ solution. We find that sulindac sulfide inducedaggregates are structurally homogeneous. The C-terminal part of the peptide adopts a β-sheet structure, whereas the N-terminus is disordered. The salt bridge between D23 and K28 is present, similar as in wild type fibril structures. (13)C-(19)F transferred echo double resonance experiments suggest that sulindac sulfide colocalizes with the Aβ peptide in the aggregate.

  7. Stable and Size-Tunable Aggregation-Induced Emission Nanoparticles Encapsulated with Nanographene Oxide and Applications in Three-Photon Fluorescence Bioimaging.

    Science.gov (United States)

    Zhu, Zhenfeng; Qian, Jun; Zhao, Xinyuan; Qin, Wei; Hu, Rongrong; Zhang, Hequn; Li, Dongyu; Xu, Zhengping; Tang, Ben Zhong; He, Sailing

    2016-01-26

    Organic fluorescent dyes with high quantum yield are widely applied in bioimaging and biosensing. However, most of them suffer from a severe effect called aggregation-caused quenching (ACQ), which means that their fluorescence is quenched at high molecular concentrations or in the aggregation state. Aggregation-induced emission (AIE) is a diametrically opposite phenomenon to ACQ, and luminogens with this feature can effectively solve this problem. Graphene oxide has been utilized as a quencher for many fluorescent dyes, based on which biosensing can be achieved. However, using graphene oxide as a surface modification agent of fluorescent nanoparticles is seldom reported. In this article, we used nanographene oxide (NGO) to encapsulate fluorescent nanoparticles, which consisted of a type of AIE dye named TPE-TPA-FN (TTF). NGO significantly improved the stability of nanoparticles in aqueous dispersion. In addition, this method could control the size of nanoparticles' flexibly as well as increase their emission efficiency. We then used the NGO-modified TTF nanoparticles to achieve three-photon fluorescence bioimaging. The architecture of ear blood vessels in mice and the distribution of nanoparticles in zebrafish could be observed clearly. Furthermore, we extended this method to other AIE luminogens and showed it was widely feasible.

  8. Chlorin e6 Prevents ADP-Induced Platelet Aggregation by Decreasing PI3K-Akt Phosphorylation and Promoting cAMP Production

    Directory of Open Access Journals (Sweden)

    Ji Young Park

    2013-01-01

    Full Text Available A number of reagents that prevent thrombosis have been developed but were found to have serious side effects. Therefore, we sought to identify complementary and alternative medicinal materials that are safe and have long-term efficacy. In the present studies, we have assessed the ability of chlorine e6 (CE6 to inhibit ADP-induced aggregation of rat platelets and elucidated the underlying mechanism. CE6 inhibited platelet aggregation induced by 10 µM ADP in a concentration-dependent manner and decreased intracellular calcium mobilization and granule secretion (i.e., ATP and serotonin release. Western blotting revealed that CE6 strongly inhibited the phosphorylations of PI3K, Akt, c-Jun N-terminal kinase (JNK, and different mitogen-activated protein kinases (MAPKs including extracellular signal-regulated kinase 1/2 (ERK1/2 as well as p38-MAPK. Our study also demonstrated that CE6 significantly elevated intracellular cAMP levels and decreased thromboxane A2 formation in a concentration-dependent manner. Furthermore, we determined that CE6 initiated the activation of PKA, an effector of cAMP. Taken together, our findings indicate that CE6 may inhibit ADP-induced platelet activation by elevating cAMP levels and suppressing PI3K/Akt activity. Finally, these results suggest that CE6 could be developed as therapeutic agent that helps prevent thrombosis and ischemia.

  9. One-Pot Click Access to a Cyclodextrin Dimer-Based Novel Aggregation Induced Emission Sensor and Monomer-Based Chiral Stationary Phase

    Directory of Open Access Journals (Sweden)

    Xiaoli Li

    2016-11-01

    Full Text Available A ‘two birds, one stone’ strategy was developed via a one-pot click reaction to simultaneously prepare a novel cyclodextrin (CD dimer based aggregation induced emission (AIE sensor (AIE-DCD and a monomer based chiral stationary phase (CSP-MCD for chiral high performance liquid chromatography (CHPLC. AIE-DCD was found to afford satisfactory AIE response for specific detection of Zn2+ with a detection limit of 50 nM. CSP-MCD exhibits excellent enantioseparation ability toward dansyl amino acids, where the resolution of dansyl amino leucine reaches 5.43.

  10. Submicron-Scale Heterogeneities in Nickel Sorption of Various Cell-Mineral Aggregates Formed by Fe(II)-Oxidizing Bacteria.

    Science.gov (United States)

    Schmid, Gregor; Zeitvogel, Fabian; Hao, Likai; Ingino, Pablo; Adaktylou, Irini; Eickhoff, Merle; Obst, Martin

    2016-01-01

    Fe(II)-oxidizing bacteria form biogenic cell-mineral aggregates (CMAs) composed of microbial cells, extracellular organic compounds, and ferric iron minerals. CMAs are capable of immobilizing large quantities of heavy metals, such as nickel, via sorption processes. CMAs play an important role for the fate of heavy metals in the environment, particularly in systems characterized by elevated concentrations of dissolved metals, such as mine drainage or contaminated sediments. We applied scanning transmission (soft) X-ray microscopy (STXM) spectrotomography for detailed 3D chemical mapping of nickel sorbed to CMAs on the submicron scale. We analyzed different CMAs produced by phototrophic or nitrate-reducing microbial Fe(II) oxidation and, in addition, a twisted stalk structure obtained from an environmental biofilm. Nickel showed a heterogeneous distribution and was found to be preferentially sorbed to biogenically precipitated iron minerals such as Fe(III)-(oxyhydr)oxides and, to a minor extent, associated with organic compounds. Some distinct nickel accumulations were identified on the surfaces of CMAs. Additional information obtained from scatter plots and angular distance maps, showing variations in the nickel-iron and nickel-organic carbon ratios, also revealed a general correlation between nickel and iron. Although a high correlation between nickel and iron was observed in 2D maps, 3D maps revealed this to be partly due to projection artifacts. In summary, by combining different approaches for data analysis, we unambiguously showed the heterogeneous sorption behavior of nickel to CMAs.

  11. Induced pluripotent stem cells for cardiac repair.

    Science.gov (United States)

    Zwi-Dantsis, Limor; Gepstein, Lior

    2012-10-01

    Myocardial stem cell therapies are emerging as novel therapeutic paradigms for myocardial repair, but are hampered by the lack of sources for autologous human cardiomyocytes. An exciting development in the field of cardiovascular regenerative medicine is the ability to reprogram adult somatic cells into pluripotent stem cell lines (induced pluripotent stem cells, iPSCs) and to coax their differentiation into functional cardiomyocytes. This technology holds great promise for the emerging disciplines of personalized and regenerative medicine, because of the ability to derive patient-specific iPSCs that could potentially elude the immune system. The current review describes the latest techniques of generating iPSCs as well as the methods used to direct their differentiation towards the cardiac lineage. We then detail the unique potential as well as the possible hurdles on the road to clinical utilizing of the iPSCs derived cardiomyocytes in the emerging field of cardiovascular regenerative medicine.

  12. Sodium phenylacetate (NaPa) induces modifications of the proliferation, the adhesion and the cell cycle of tumoral epithelial breast cells.

    Science.gov (United States)

    Thibout, D; Kraemer, M; Di Benedetto, M; Saffar, L; Gattegno, L; Derbin, C; Crépin, M

    1999-01-01

    Sodium phenylacetate (NaPa), a physiological product of phenylalanine metabolism, present in micromolar concentrations in human plasma, has been shown to induce in vivo and in vitro cytostatic antiproliferative effects at millimolar concentrations. Cadherin molecules are powerful invasion suppressor molecules and the reduction of E-cadherin expression plays an important role in the invasion and metastasis of human breast cancer. In this study, we demonstrated, on one hand, that NaPa stimulated aggregation by increasing the expression of E-cadherin at the surface of breast cancer MCF-7ras cells transformed by Ha-ras oncogene and inhibited its expression in MCF-7 cells. We demonstrated that NaPa increased the formation of MCF-7ras cell aggregates and did not alter the formation of MCF-7 cell aggregates. By Northern blot, we demonstrated that the E-cadherin expression was not regulated at the transcriptional level. On the other hand, we analyzed the cell cycle of these 2 cell lines after NaPa treatment and showed that NaPa induced arrest at the G1/S phase in both MCF-7 and MCF-7ras cells. bFGF increased the growth of MCF-7 cells, but inhibited MCF-7ras cell proliferation. NaPa treatment suppressed the stimulation of MCF-7 cell proliferation and increased MCF-7ras cell growth inhibition. We have demonstrated a new target of NaPa action in blocking the cell cycle of tumor cells in G0/G1. We suggest that the anti-proliferative effect of NaPa associated to the restoration of the cadherin function in human mammary carcinoma cells indicates that NaPa could be a novel therapeutic agent in breast cancer.

  13. Idarubicin induces mTOR-dependent cytotoxic autophagy in leukemic cells

    Energy Technology Data Exchange (ETDEWEB)

    Ristic, Biljana [Institute of Microbiology and Immunology, School of Medicine, University of Belgrade, Dr. Subotica 1, 11000 Belgrade (Serbia); Bosnjak, Mihajlo [Institute of Histology and Embryology, School of Medicine, University of Belgrade, Belgrade (Serbia); Arsikin, Katarina [Institute of Microbiology and Immunology, School of Medicine, University of Belgrade, Dr. Subotica 1, 11000 Belgrade (Serbia); Mircic, Aleksandar; Suzin-Zivkovic, Violeta [Institute of Histology and Embryology, School of Medicine, University of Belgrade, Belgrade (Serbia); Bogdanovic, Andrija [Clinic for Hematology, Clinical Centre of Serbia, School of Medicine, University of Belgrade, Belgrade (Serbia); Perovic, Vladimir [Institute of Microbiology and Immunology, School of Medicine, University of Belgrade, Dr. Subotica 1, 11000 Belgrade (Serbia); Martinovic, Tamara; Kravic-Stevovic, Tamara; Bumbasirevic, Vladimir [Institute of Histology and Embryology, School of Medicine, University of Belgrade, Belgrade (Serbia); Trajkovic, Vladimir, E-mail: vtrajkovic@med.bg.ac.rs [Institute of Microbiology and Immunology, School of Medicine, University of Belgrade, Dr. Subotica 1, 11000 Belgrade (Serbia); Harhaji-Trajkovic, Ljubica, E-mail: buajk@yahoo.com [Institute for Biological Research, University of Belgrade, Belgrade, Despot Stefan Blvd. 142, 11000 Belgrade (Serbia)

    2014-08-01

    We investigated if the antileukemic drug idarubicin induces autophagy, a process of programmed cellular self-digestion, in leukemic cell lines and primary leukemic cells. Transmission electron microscopy and acridine orange staining demonstrated the presence of autophagic vesicles and intracellular acidification, respectively, in idarubicin-treated REH leukemic cell line. Idarubicin increased punctuation/aggregation of microtubule-associated light chain 3B (LC3B), enhanced the conversion of LC3B-I to autophagosome-associated LC3B-II in the presence of proteolysis inhibitors, and promoted the degradation of the selective autophagic target p62, thus indicating the increase in autophagic flux. Idarubicin inhibited the phosphorylation of the main autophagy repressor mammalian target of rapamycin (mTOR) and its downstream target p70S6 kinase. The treatment with the mTOR activator leucine prevented idarubicin-mediated autophagy induction. Idarubicin-induced mTOR repression was associated with the activation of the mTOR inhibitor AMP-activated protein kinase and down-regulation of the mTOR activator Akt. The suppression of autophagy by pharmacological inhibitors or LC3B and beclin-1 genetic knockdown rescued REH cells from idarubicin-mediated oxidative stress, mitochondrial depolarization, caspase activation and apoptotic DNA fragmentation. Idarubicin also caused mTOR inhibition and cytotoxic autophagy in K562 leukemic cell line and leukocytes from chronic myeloid leukemia patients, but not healthy controls. By demonstrating mTOR-dependent cytotoxic autophagy in idarubicin-treated leukemic cells, our results warrant caution when considering combining idarubicin with autophagy inhibitors in leukemia therapy. - Highlights: • Idarubicin induces autophagy in leukemic cell lines and primary leukemic cells. • Idarubicin induces autophagy by inhibiting mTOR in leukemic cells. • mTOR suppression by idarubicin is associated with AMPK activation and Akt blockade.

  14. Ectopic expression of cyclase associated protein CAP restores the streaming and aggregation defects of adenylyl cyclase a deficient Dictyostelium discoideum cells

    OpenAIRE

    Sultana Hameeda; Neelakanta Girish; Rivero Francisco; Blau-Wasser Rosemarie; Schleicher Michael; Noegel Angelika A

    2012-01-01

    Abstract Background Cell adhesion, an integral part of D. discoideum development, is important for morphogenesis and regulated gene expression in the multicellular context and is required to trigger cell-differentiation. G-protein linked adenylyl cyclase pathways are crucially involved and a mutant lacking the aggregation specific adenylyl cyclase ACA does not undergo multicellular development. Results Here, we have investigated the role of cyclase-associated protein (CAP), an important regul...

  15. Cartilage tissue engineering using pre-aggregated human articular chondrocytes

    Directory of Open Access Journals (Sweden)

    F Wolf

    2008-12-01

    Full Text Available In this study, we first aimed at determining whether human articular chondrocytes (HAC proliferate in aggregates in the presence of strong chondrocyte mitogens. We then investigated if the aggregated cells have an enhanced chondrogenic capacity as compared to cells cultured in monolayer. HAC from four donors were cultured in tissue culture dishes either untreated or coated with 1% agarose in the presence of TGFb-1, FGF-2 and PDGF-BB. Proliferation and stage of differentiation were assessed by measuring respectively DNA contents and type II collagen mRNA. Expanded cells were induced to differentiate in pellets or in Hyaff®-11 meshes and the formed tissues were analysed biochemically for glycosaminoglycans (GAG and DNA, and histologically by Safranin O staining. The amount of DNA in aggregate cultures increased significantly from day 2 to day 6 (by 3.2-fold, but did not further increase with additional culture time. Expression of type II collagen mRNA was about two orders of magnitude higher in aggregated HAC as compared to monolayer expanded cells. Pellets generated by aggregated HAC were generally more intensely stained for GAG than those generated by monolayer-expanded cells. Scaffolds seeded with aggregates accumulated more GAG (1.3-fold than scaffolds seeded with monolayer expanded HAC. In conclusion, this study showed that HAC culture in aggregates does not support a relevant degree of expansion. However, aggregation of expanded HAC prior to loading into a porous scaffold enhances the quality of the resulting tissues and could thus be introduced as an intermediate culture phase in the manufacture of engineered cartilage grafts.

  16. Embryonic stem cells remain highly pluripotent following long term expansion as aggregates in suspension bioreactors.

    Science.gov (United States)

    zur Nieden, Nicole I; Cormier, Jaymi T; Rancourt, Derrick E; Kallos, Michael S

    2007-05-01

    Increasing attention has been drawn towards pluripotent embryonic stem cells (ESCs) and their potential use as the primary material in various tissue engineering applications. Successful clinical implementation of this technology would require a quality controlled reproducible culture system for the expansion of the cells to be used in the generation of functional tissues. Recently, we showed that suspension bioreactors could be used in the regulated large-scale expansion of highly pluripotent murine ESCs. The current study illustrates that these bioreactor protocols can be adapted for long term culture and that murine ESC cultures remain highly undifferentiated, when serially passaged in suspension bioreactors for extended periods. Flow cytometry analysis and gene expression profiles of several pluripotency markers, in addition to colony and embryoid body (EB) formation tests were conducted at the start and end of the experiment and all showed that the ESC cultures remained highly undifferentiated over extended culture time in suspension. In vivo teratoma formation and in vitro differentiation into neural, cardiomyocyte, osteoblast and chondrocyte lineages, performed at the end of the long term culture, further supported the presence of functional and undifferentiated ESCs in the expanded population. Overall, this system enables the controlled expansion of highly pluripotent murine ESC populations.

  17. Protein aggregation and lyophilization: Protein structural descriptors as predictors of aggregation propensity

    OpenAIRE

    Roughton, Brock C.; Iyer, Lavanya K.; Bertelsen, Esben; Topp, Elizabeth M.; Camarda, Kyle V.

    2013-01-01

    Lyophilization can induce aggregation in therapeutic proteins, but the relative importance of protein structure, formulation and processing conditions are poorly understood. To evaluate the contribution of protein structure to lyophilization-induced aggregation, fifteen proteins were co-lyophilized with each of five excipients. Extent of aggregation following lyophilization, measured using size-exclusion chromatography, was correlated with computational and biophysical protein structural desc...

  18. Aggregate resources in the Netherlands

    NARCIS (Netherlands)

    Meulen, M.J. van der; Gessel, S.F. van; Veldkamp, J.G.

    2005-01-01

    We have built a 3D lithological model of the Netherlands, for the purpose of mapping on-land aggregate resources down to 50 m below the surface. The model consists of voxel cells (1000 · 1000 · 1 m), with lithological composition and aggregate content estimates as primary attributes. These attribute

  19. Preliminary Study on in Vitro Induced Differentiation of Embryonic Stem Cells into Neurons

    Institute of Scientific and Technical Information of China (English)

    JianGe; ShunongLi; 等

    2002-01-01

    Purpose;To study preliminarily in vitro induced differentiation of embryonic stem cells into neurons for further investigation of an alternative for the treatment of glaumatous neuropathy.Materials and methods:Supernatant of cultured Buffalo rat liver cells (buffalorat liver cell-conditioned medium,BRL-CM)was used for culturing embryonic stem cells(ES-D3 cell line)..Morphological features of undifferentiated ES cells were studied by HE staining and electron microscopy.Based on the methods used by Bain et al,we modified the methods and used retinoic,acid(RA) as an inducer to differentiate ES-D3 cells and cytosine arabinoside(Ara-C) as inhibitor of proliferative cells.The growth of the cells was observed under phase contrast microscope.Reuslts:ES-D3 cells cultured by BRL-CM grew in aggregates and remained undifferentiated.Electromicroscopy showed large nucleus and a large amount of mitochondria in undifferentiated ES cells and many processes on the surfaces.In the first day after the adding of retinoic acid,some neuron-like cells with one,two or more processes were present.In the second day after adding RA and the first day after the plus of 10μm Ara-C,a large amount of neuron-like cells appeared,with the formation of neuron-like networks.Conclusions:Combined use of RA and Ara-C can induce ES cells to different into neuron-like cells.Our present preliminary study might provide insights into an alternative for the treatment of glaucomatous neuropathy by the transplantation of embryonic stem cells.Eye Science 2000;16:1-6.

  20. On the observation of the need for an unusually high concentration of cysteine and homocysteine to induce aggregation of polymer-stabilized gold nano particles

    Energy Technology Data Exchange (ETDEWEB)

    Radhakumary, C.; Sreenivasan, K., E-mail: sreeni@sctimst.ac.in [Sree Chitra Tirunal Institute for Medical Sciences and Technology, Laboratory for Polymer Analysis, Biomedical Technology Wing (India)

    2013-02-15

    This study reports the interaction of chitosan-stabilized gold nanoparticles (CH-AuNPs) with cysteine (Cys) and homocysteine (Hcys) in aqueous media at pH 1.4. Since the polymer precipitates at higher pH, and the amino acids Cys and HCys are soluble at acidic pH, we kept the pH around 1.4 for stabilizing the particles. Zeta potential of CH-AuNPs was found to be positive and it is reasonable to assume that +ve Cys or Hcys at pH 1.4 will experience repulsive force. However, TEM images and absorption spectra indicated formation of aggregates including rod-like assembly. An interesting observation was the need for unusually high concentration of analytes (Cys and Hcys) to induce the assembly of CH-AuNPs. We also found time bound variation of the optical properties probably indicating the interaction is kinetically controlled and only a fraction of the analyte molecules having sufficient energy can bind onto the particles. We observed that at elevated temperature, the reaction was faster with a lower concentration of Cys or Hcys. These observations were supported by the classical Derjaguin-Landau-Verwey-Overbeek (DLVO) theory which describes the interparticle interaction and the colloidal stability in solution. Only molecules possessing enough energy to cross this force barrier can cause the aggregation. We also noted a time lag between Cys and Hcys to influence optical properties reflecting the possibility of using this simple approach to discriminate these two clinically relevant molecules. Our observation shows that simple sensing as well as generation of novel nanostructures could be manipulated by a judicious choice of conditions such as stabilizing agents, pH, etc.Graphical AbstractMore energetic ones cross the barrier to induce aggregation.

  1. UV-Induced Cell Death in Plants

    Directory of Open Access Journals (Sweden)

    Chang Ho Kang

    2013-01-01

    Full Text Available Plants are photosynthetic organisms that depend on sunlight for energy. Plants respond to light through different photoreceptors and show photomorphogenic development. Apart from Photosynthetically Active Radiation (PAR; 400–700 nm, plants are exposed to UV light, which is comprised of UV-C (below 280 nm, UV-B (280–320 nm and UV-A (320–390 nm. The atmospheric ozone layer protects UV-C radiation from reaching earth while the UVR8 protein acts as a receptor for UV-B radiation. Low levels of UV-B exposure initiate signaling through UVR8 and induce secondary metabolite genes involved in protection against UV while higher dosages are very detrimental to plants. It has also been reported that genes involved in MAPK cascade help the plant in providing tolerance against UV radiation. The important targets of UV radiation in plant cells are DNA, lipids and proteins and also vital processes such as photosynthesis. Recent studies showed that, in response to UV radiation, mitochondria and chloroplasts produce a reactive oxygen species (ROS. Arabidopsis metacaspase-8 (AtMC8 is induced in response to oxidative stress caused by ROS, which acts downstream of the radical induced cell death (AtRCD1 gene making plants vulnerable to cell death. The studies on salicylic and jasmonic acid signaling mutants revealed that SA and JA regulate the ROS level and antagonize ROS mediated cell death. Recently, molecular studies have revealed genes involved in response to UV exposure, with respect to programmed cell death (PCD.

  2. Construction aggregates

    Science.gov (United States)

    Nelson, T.I.; Bolen, W.P.

    2007-01-01

    Construction aggregates, primarily stone, sand and gravel, are recovered from widespread naturally occurring mineral deposits and processed for use primarily in the construction industry. They are mined, crushed, sorted by size and sold loose or combined with portland cement or asphaltic cement to make concrete products to build roads, houses, buildings, and other structures. Much smaller quantities are used in agriculture, cement manufacture, chemical and metallurgical processes, glass production and many other products.

  3. Autophagy regulates chlorpyrifos-induced apoptosis in SH-SY5Y cells

    Energy Technology Data Exchange (ETDEWEB)

    Park, Jae Hyeon [Department of Pharmacology, College of Medicine, Hanyang University (Korea, Republic of); Hanyang Biomedical Research Institute, Seoul (Korea, Republic of); Graduate School of Biomedical Science and Engineering, Hanyang University, Seoul (Korea, Republic of); Lee, Jeong Eun [Department of Pharmacology, College of Medicine, Hanyang University (Korea, Republic of); Hanyang Biomedical Research Institute, Seoul (Korea, Republic of); Shin, In Chul [Department of Pharmacology, College of Medicine, Hanyang University (Korea, Republic of); Koh, Hyun Chul, E-mail: hckoh@hanyang.ac.kr [Department of Pharmacology, College of Medicine, Hanyang University (Korea, Republic of); Hanyang Biomedical Research Institute, Seoul (Korea, Republic of); Graduate School of Biomedical Science and Engineering, Hanyang University, Seoul (Korea, Republic of)

    2013-04-01

    Recent studies have shown that up-regulation of autophagy may be a tractable therapeutic intervention for clearing disease-causing proteins, including α-synuclein, ubiquitin, and other misfolded or aggregated proteins in pesticide-induced neurodegeneration. In a previous study, we reported that chlorpyrifos (CPF)-induced mitochondria-dependent apoptosis is mediated through reactive oxygen species in SH-SY5Y cells. In this study, we explored a novel pharmacotherapeutic approach to prevent CPF neurotoxicity involving the regulation of autophagy. We investigated the modulation of CPF-induced apoptosis according to autophagy regulation. We found that CPF induced apoptosis in SH-SY5Y cells, as demonstrated by the activation of caspase-3 and nuclear condensation. In addition, we observed that cells treated with CPF underwent autophagic cell death by monitoring the expression of LC3-II and p62. Pretreatment with the autophagy inducer rapamycin significantly enhanced the cell viability of CPF-exposed cells, and the enhancement of cell viability was partially due to alleviation of CPF-induced apoptosis via a decrease in levels of cleaved caspase-3. Specifically, rapamycin pretreatment decreased Bax and increased Bcl-2 expression in mitochondria. In addition, rapamycin significantly decreased cytochrome c release in from mitochondria into the cytosol. However, pretreatment of cells with the autophagy inhibitor, 3-methyladenine (3MA), remarkably increased CPF toxicity in these cells; this with correlated with increased expression of Bax and decreased expression of Bcl-2 in mitochondria. Our results suggest that CPF-induced cytotoxicity is modified by autophagy regulation and that rapamycin protects against CPF-induced apoptosis by enhancing autophagy. Pharmacologic induction of autophagy by rapamycin may be a useful treatment strategy in neurodegenerative disorders. - Highlights: ► Chlorpyrifos (CPF) is cytotoxic to SH-SY5Y cells ► CPF-induced cytotoxicity is mediated by

  4. Fibrinogen-Induced Streptococcus mutans Biofilm Formation and Adherence to Endothelial Cells

    Directory of Open Access Journals (Sweden)

    Telma Blanca Lombardo Bedran

    2013-01-01

    Full Text Available Streptococcus mutans, the predominant bacterial species associated with dental caries, can enter the bloodstream and cause infective endocarditis. The aim of this study was to investigate S. mutans biofilm formation and adherence to endothelial cells induced by human fibrinogen. The putative mechanism by which biofilm formation is induced as well as the impact of fibrinogen on S. mutans resistance to penicillin was also evaluated. Bovine plasma dose dependently induced biofilm formation by S. mutans. Of the various plasma proteins tested, only fibrinogen promoted the formation of biofilm in a dose-dependent manner. Scanning electron microscopy observations revealed the presence of complex aggregates of bacterial cells firmly attached to the polystyrene support. S. mutans in biofilms induced by the presence of fibrinogen was markedly resistant to the bactericidal effect of penicillin. Fibrinogen also significantly increased the adherence of S. mutans to endothelial cells. Neither S. mutans cells nor culture supernatants converted fibrinogen into fibrin. However, fibrinogen is specifically bound to the cell surface of S. mutans and may act as a bridging molecule to mediate biofilm formation. In conclusion, our study identified a new mechanism promoting S. mutans biofilm formation and adherence to endothelial cells which may contribute to infective endocarditis.

  5. Theory of activated-rate processes under shear with application to shear-induced aggregation of colloids.

    Science.gov (United States)

    Zaccone, Alessio; Wu, Hua; Gentili, Daniele; Morbidelli, Massimo

    2009-11-01

    Using an approximation scheme within the convective diffusion (two-body Smoluchowski) equation framework, we unveil the shear-driven aggregation mechanism at the origin of structure formation in sheared colloidal systems. The theory, verified against numerics and experiments, explains the induction time followed by explosive (irreversible) rise of viscosity observed in charge-stabilized colloidal and protein systems under steady shear. The Arrhenius-type equation with shear derived here, extending Kramers' theory in the presence of shear, clearly demonstrates the important role of shear drive in activated-rate processes as they are encountered in soft condensed matter.

  6. Interplay Between Side Chain Pattern, Polymer Aggregation, and Charge Carrier Dynamics in PBDTTPD:PCBM Bulk-Heterojunction Solar Cells

    KAUST Repository

    Dyer-Smith, Clare

    2015-05-01

    Poly(benzo[1,2-b:4,5-b′]dithiophene–alt–thieno[3,4-c]pyrrole-4,6-dione) (PBDTTPD) polymer donors with linear side-chains yield bulk-heterojunction (BHJ) solar cell power conversion efficiencies (PCEs) of about 4% with phenyl-C71-butyric acid methyl ester (PC71BM) as the acceptor, while a PBDTTPD polymer with a combination of branched and linear substituents yields a doubling of the PCE to 8%. Using transient optical spectroscopy it is shown that while the exciton dissociation and ultrafast charge generation steps are not strongly affected by the side chain modifications, the polymer with branched side chains exhibits a decreased rate of nongeminate recombination and a lower fraction of sub-nanosecond geminate recombination. In turn the yield of long-lived charge carriers increases, resulting in a 33% increase in short circuit current (J sc). In parallel, the two polymers show distinct grazing incidence X-ray scattering spectra indicative of the presence of stacks with different orientation patterns in optimized thin-film BHJ devices. Independent of the packing pattern the spectroscopic data also reveals the existence of polymer aggregates in the pristine polymer films as well as in both blends which trap excitons and hinder their dissociation.

  7. Abiotic factors in colony formation: effects of nutrition and light on extracellular polysaccharide production and cell aggregates of Microcystis aeruginosa

    Institute of Scientific and Technical Information of China (English)

    YANG Zhen; KONG Fanxiang

    2013-01-01

    Colony morphology is important for Microcystis to sustain a competitive advantage in eutrophic lakes.The mechanism of colony formation in Microcystis is currently unclear.Extracellular polysaccharide (EPS) has been reported to play an important role in cell aggregate formation of some phytoplankton.Microcystis aeruginosa was cultivated under varied abiotic conditions,including different nutrient,light,and temperature conditions,to investigate their effects on EPS production and morphological change.The results show that nutrient concentration and light intensity have great effects on EPS production in M.aeruginosa.There was a considerable increase in EPS production after M.aeruginosa was cultivated in adjusted culture conditions similar to those present in the field (28.9 mg C/L,1.98 mg N/L,0.65 mg P/L,light intensity:100 μmol/(m2·s)).These results indicate that abiotic factors might be one of the triggers for colony formation in Microcystis.

  8. Chemical -induced apoptotic cell death in tomato cells : involvement of caspase-like proteases

    NARCIS (Netherlands)

    Jong, de A.J.; Hoeberichts, F.A.; Yakimova, E.T.; Maximova, E.; Woltering, E.J.

    2000-01-01

    A new system to study programmed cell death in plants is described. Tomato (Lycopersicon esculentum Mill.) suspension cells were induced to undergo programmed cell death by treatment with known inducers of apoptosis in mammalian cells. This chemical-induced cell death was accompanied by the characte

  9. Uranium induces apoptosis in lung epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Periyakaruppan, Adaikkappan; Sarkar, Shubhashish; Sadanandan, Bindu; Thomas, Renard; Wilson, Bobby L. [Texas Southern University, Environmental Toxicology Program, Department of Chemistry, Houston, TX (United States); Ravichandran, Prabakaran; Sharma, Chidananda S.; Ramesh, Vani; Hall, Joseph C.; Ramesh, Govindarajan T. [Norfolk State University, Molecular Toxicology Laboratory, Department of Biology, Center for Biotechnology and Biomedical Sciences, Norfolk, VA (United States)

    2009-06-15

    Uranium is a naturally occurring radioactive material present everywhere in the environment. It is toxic because of its chemical or radioactive properties. Uranium enters environment mainly from mines and industry and cause threat to human health by accumulating in lungs as a result of inhalation. In our previous study, we have shown the effectiveness of antioxidant system response to the oxidative stress induced by uranyl acetate (UA) in rat lung epithelial (LE) cells. As part of our continuing studies; here, we investigated the mechanism underlying when LE cells are exposed to different concentration of UA. Oxidative stress may lead to apoptotic signaling pathways. LE cells treated with 0.25, 0.5 and 1 mM of UA results in dose and time-dependent increase in activity of both caspases-3 and -8. Increase in the concentration of cytochrome-c oxidase in cytosol was seen in LE cells treated with 1 mM UA as a result of mitochondria membrane permeability. The cytochrome-c leakage may trigger the apoptotic pathway. TUNEL assay performed in LE cells treated with 1 mM of UA showed significant incorporation of dNTPs in the nucleus after 24 h. In the presence of the caspase inhibitors, we observed the significant decrease in the activity of caspases-8 and -3 in 0.5 and 1 mM UA-treated LE cells. (orig.)

  10. Activation-Induced Cell Death in T Cells and Autoimmunity

    Institute of Scientific and Technical Information of China (English)

    Jian Zhang; Xuemei Xu; Yong Liu

    2004-01-01

    Activation-induced cell death (AICD), which results from the interaction between Fas and Fas ligand, is responsible for maintaining tolerance to self-antigen. A defect in AICD may lead to development of autoimmunity. During the last several years, much progress has been made in understanding the mechanism(s) of AICD and its potential role in the pathogenesis of autoimmune diseases. In this review, we summarize the most recent progress on the regulation of the susceptibility of T cells to AICD and its possible involvement in autoimmune diseases.

  11. Cell signalling pathways underlying induced pluripotent stem cell reprogramming

    Institute of Scientific and Technical Information of China (English)

    Kate; Hawkins; Shona; Joy; Tristan; Mc; Kay

    2014-01-01

    Induced pluripotent stem(i PS) cells, somatic cells reprogrammed to the pluripotent state by forced expression of defined factors, represent a uniquely valuable resource for research and regenerative medicine. However, this methodology remains inefficient due to incomplete mechanistic understanding of the reprogramming process. In recent years, various groups have endeavoured to interrogate the cell signalling that governs the reprogramming process, including LIF/STAT3, BMP, PI3 K, FGF2, Wnt, TGFβ and MAPK pathways, with the aim of increasing our understanding and identifying new mechanisms of improving safety, reproducibility and efficiency. This has led to a unified model of reprogramming that consists of 3 stages: initiation, maturation and stabilisation. Initiation of reprogramming occurs in almost all cells that receive the reprogramming transgenes; most commonly Oct4, Sox2, Klf4 and c Myc, and involves a phenotypic mesenchymal-to-epithelial transition. The initiation stage is also characterised by increased proliferation and a metabolic switch from oxidative phosphorylation to glycolysis. The maturation stage is considered the major bottleneck within the process, resulting in very few "stabilisation competent" cells progressing to the final stabilisation phase. To reach this stage in both mouse and human cells, pre-i PS cells must activate endogenous expression of the core circuitry of pluripotency, comprising Oct4, Sox2, and Nanog, and thus reach a state of transgene independence. By the stabilisation stage, i PS cells generally use the same signalling networks that govern pluripotency in embryonic stem cells. These pathways differ between mouse and human cells although recent work has demonstrated that this is context dependent. As i PS cell generation technologies move forward, tools are being developed to interrogate the process in more detail, thus allowing a greater understanding of this intriguing biological phenomenon.

  12. Management of trauma-induced inflammatory root resorption using mineral trioxide aggregate obturation: two-year follow up.

    Science.gov (United States)

    Güzeler, Irem; Uysal, Serdar; Cehreli, Zafer C

    2010-12-01

    Inflammatory root resorption is a serious complication of dental trauma, which leads to progressive loss of the root structure. This report describes the treatment a previously traumatized young maxillary lateral incisor, severely affected by inflammatory root resorption. An 11-year-old boy presented with pain and mobility in his maxillary incisors which experienced fall trauma 2 years earlier. Radiographic examination revealed incomplete root development of the right central incisor, associated with advanced inflammatory root resorption and a periapical lesion. Following removal of a prior long-term calcium hydroxide dressing, the root canal was submitted to a 2-week irrigation regimen involving 1.25% sodium hypochlorite and 2% chlorhexidine gluconate. Thereafter, the entire root was filled with mineral trioxide aggregate. The radiographic follow up at 6 months showed arrest of root resorption and initiation of periapical healing in the absence of clinical symptoms and mobility. This was followed by advanced osseous regeneration and re-establishment of the periodontal space at 12 and 24 months. From the present case, it can be concluded that mineral trioxide aggregate obturation can be a viable option that can improve the healing outcomes in cases of severe inflammatory root resorption in young permanent teeth.

  13. Langhans giant cells from M. tuberculosis-induced human granulomas cannot mediate mycobacterial uptake.

    Science.gov (United States)

    Lay, G; Poquet, Y; Salek-Peyron, P; Puissegur, M-P; Botanch, C; Bon, H; Levillain, F; Duteyrat, J-L; Emile, J-F; Altare, F

    2007-01-01

    Tuberculosis is characterized by a tight interplay between Mycobacterium tuberculosis (M. tb) and host cells within granulomas. These cellular aggregates restrain M. tb spreading but do not kill all bacilli, which persist for years. A more detailed investigation of the interaction between M. tb and granuloma cells is needed to improve our understanding of this persistence and to explain the physiopathology of tuberculosis. In the present study, a recently developed in vitro human model of tuberculous granulomas has been used to analyse the modulation of granuloma cell differentiation by M. tb, in comparison to poorly virulent mycobacteria, which do not persist. It is reported that whilst all mycobacteria species induce granuloma formation, only M. tb triggers the differentiation of granuloma macrophages into very large multinucleated giant cells (MGCs) that are unable to mediate any bacterial uptake. This loss of function is not due to cell quiescence, as MGCs still display NADPH oxidase activity, but it correlates with decreased expression of phagocytosis receptors. This phenomenon is specific for the virulent species of M. tuberculosis complex, as poorly virulent species only induce the formation of small multinucleated cells (MCs) with conserved mycobacterial uptake ability, which never reach the MGC differentiation stage. The phenotype of MGCs thus strongly resembles mature dendritic cells with a loss of microbial uptake ability, despite conserved antigen presentation. In M. tb-induced granulomas, MGCs thus seem to be devoted to the destruction of bacilli that have been ingested in previous differentiation stages, ie in macrophages and MCs.

  14. Derivation of induced pluripotent stem cells from pig somatic cells.

    Science.gov (United States)

    Ezashi, Toshihiko; Telugu, Bhanu Prakash V L; Alexenko, Andrei P; Sachdev, Shrikesh; Sinha, Sunilima; Roberts, R Michael

    2009-07-07

    For reasons that are unclear the production of embryonic stem cells from ungulates has proved elusive. Here, we describe induced pluripotent stem cells (iPSC) derived from porcine fetal fibroblasts by lentiviral transduction of 4 human (h) genes, hOCT4, hSOX2, hKLF4, and hc-MYC, the combination commonly used to create iPSC in mouse and human. Cells were cultured on irradiated mouse embryonic fibroblasts (MEF) and in medium supplemented with knockout serum replacement and FGF2. Compact colonies of alkaline phosphatase-positive cells emerged after approximately 22 days, providing an overall reprogramming efficiency of approximately 0.1%. The cells expressed porcine OCT4, NANOG, and SOX2 and had high telomerase activity, but also continued to express the 4 human transgenes. Unlike human ESC, the porcine iPSC (piPSC) were positive for SSEA-1, but negative for SSEA-3 and -4. Transcriptional profiling on Affymetrix (porcine) microarrays and real time RT-PCR supported the conclusion that reprogramming to pluripotency was complete. One cell line, ID6, had a normal karyotype, a cell doubling time of approximately 17 h, and has been maintained through >220 doublings. The ID6 line formed embryoid bodies, expressing genes representing all 3 germ layers when cultured under differentiating conditions, and teratomas containing tissues of ectoderm, mesoderm, and endoderm origin in nude mice. We conclude that porcine somatic cells can be reprogrammed to form piPSC. Such cell lines derived from individual animals could provide a means for testing the safety and efficacy of stem cell-derived tissue grafts when returned to the same pigs at a later age.

  15. [Lysophosphatidic acid and human erythrocyte aggregation].

    Science.gov (United States)

    Sheremet'ev, Iu A; Popovicheva, A N; Levin, G Ia

    2014-01-01

    The effects of lysophosphatidic acid on the morphology and aggregation of human erythrocytes has been studied. Morphology of erythrocytes and their aggregates were studied by light microscopy. It has been shown that lysophosphatidic acid changes the shape of red blood cells: diskocyte become echinocytes. Aggregation of red blood cells (rouleaux) was significantly reduced in autoplasma. At the same time there is a strong aggregation of echinocytes. This was accompanied by the formation of microvesicles. Adding normal plasma to echinocytes restores shape and aggregation of red blood cells consisting of "rouleaux". A possible mechanism of action of lysophosphatidic acid on erythrocytes is discussed.

  16. The dynamic relationship between low birthweight and induced abortion in New York City. An aggregate time-series analysis.

    Science.gov (United States)

    Joyce, T; Grossman, M

    1990-11-01

    We use a vector autoregression to examine the dynamic relationship between the race-specific percentage of pregnancies terminated by induced abortion and the race-specific percentage of low-birthweight births in New York City. With monthly data beginning in 1972, we find that induced abortion explains low birthweight for blacks, but not for whites. There is no evidence of feedback from low birthweight to induced abortion. The findings suggest that unanticipated decreases in the percentage of pregnancies terminated by induced abortion would worsen birth outcomes among blacks in New York City.

  17. Decreased platelet aggregation by shear stress-stimulated endothelial cells in vitro: description of a method and first results in diabetes.

    Science.gov (United States)

    De Franceschi, Maria S; Palange, Anna L; Mancuso, Anna; Grande, Laura; Muccari, Domenico; Scavelli, Faustina B; Irace, Concetta; Gnasso, Agostino; Carallo, Claudio

    2015-01-01

    The interaction between platelets and endothelium in vivo is a complex phenomenon. Our aim was to develop an in vitro system that mimics the in vivo environment and investigate platelet function in a common pathological condition. Human umbilical vein endothelial cells were used and platelets from 28 type 2 diabetes patients were studied under shear stress conditions. Mean coefficient of variation of platelet aggregation was 10% in dynamic conditions in the presence of endothelium. Endothelial cells increased the concentration of inductor needed to achieve 50% platelet aggregation to adenosine diphosphate from 2.6 ± 1.3 in static conditions to 3.7 ± 1.3 µM in dynamic conditions. A similar pattern was observed when collagen was used for platelet activation. Incubation of endothelium with a nitric oxide inhibitor abolished this effect, indicating platelet inhibitory effect of endothelial cells is nitric oxide mediated. Platelet reactivity of healthy controls was less influenced by the presence of endothelial cells and displayed reduced basal platelet reactivity compared with platelets from diabetes patients. We show that platelet aggregation in diabetes as commonly reported in vitro may not fully reflect the in vivo pathophysiological process. Future studies are warranted to investigate other pathological conditions and analyse the effects of antiplatelet agents using this system.

  18. Sequence-dependent internalization of aggregating peptides.

    Science.gov (United States)

    Couceiro, José R; Gallardo, Rodrigo; De Smet, Frederik; De Baets, Greet; Baatsen, Pieter; Annaert, Wim; Roose, Kenny; Saelens, Xavier; Schymkowitz, Joost; Rousseau, Frederic

    2015-01-02

    Recently, a number of aggregation disease polypeptides have been shown to spread from cell to cell, thereby displaying prionoid behavior. Studying aggregate internalization, however, is often hampered by the complex kinetics of the aggregation process, resulting in the concomitant uptake of aggregates of different sizes by competing mechanisms, which makes it difficult to isolate pathway-specific responses to aggregates. We designed synthetic aggregating peptides bearing different aggregation propensities with the aim of producing modes of uptake that are sufficiently distinct to differentially analyze the cellular response to internalization. We found that small acidic aggregates (≤500 nm in diameter) were taken up by nonspecific endocytosis as part of the fluid phase and traveled through the endosomal compartment to lysosomes. By contrast, bigger basic aggregates (>1 μm) were taken up through a mechanism dependent on cytoskeletal reorganization and membrane remodeling with the morphological hallmarks of phagocytosis. Importantly, the properties of these aggregates determined not only the mechanism of internalization but also the involvement of the proteostatic machinery (the assembly of interconnected networks that control the biogenesis, folding, trafficking, and degradation of proteins) in the process; whereas the internalization of small acidic aggregates is HSF1-independent, the uptake of larger basic aggregates was HSF1-dependent, requiring Hsp70. Our results show that the biophysical properties of aggregates determine both their mechanism of internalization and proteostatic response. It remains to be seen whether these differences in cellular response contribute to the particular role of specific aggregated proteins in disease.

  19. Exquisite 1D Assemblies Arising from Rationally Designed Asymmetric Donor-Acceptor Architectures Exhibiting Aggregation-Induced Emission as a Function of Auxiliary Acceptor Strength.

    Science.gov (United States)

    Singh, Roop Shikha; Mukhopadhyay, Sujay; Biswas, Arnab; Pandey, Daya Shankar

    2016-01-11

    One-dimensional nanostructures with aggregation-induced emission (AIE) properties have been fabricated to keep the pace with growing demand from optoelectronics applications. The compounds 2-[4-(4-methylpiperazin-1-yl)benzylidene]malononitrile (PM1), 2-{4-[4-(pyridin-2-yl)piperazin-1-yl]-benzylidene}malononitrile (PM2), and 2-{4-[4-(pyrimidin-2-yl)piperazin-1-yl]benzylidene}malononitrile (PM3) have been designed and synthesized by melding piperazine and dicyanovinylene to investigate AIE in an asymmetric donor-acceptor (D-A) construct of A'-D-π-A- topology. The synthetic route has been simplified by using phenylpiperazine as a weak donor (D), dicyanovinylene as an acceptor (A), and pyridyl/pyrimidyl groups (PM2/PM3) as auxiliary acceptors (A'). It has been established that A' plays a vital role in triggering AIE in these compounds because the same D-A construct led to aggregation-caused quenching upon replacing A' with an electron-donating ethyl group (PM1). Moreover, the effect of restricted intramolecular rotation and twisted intramolecular charge transfer on the mechanism of AIE has also been investigated. Furthermore, it has been clearly shown that the optical disparities of these A'-D-π-A architectures are a direct consequence of comparative A' strength. Single-crystal X-ray analyses provided justification for role of intermolecular interactions in aggregate morphology. Electrochemical and theoretical studies affirmed the effect of the A' strength on the overall properties of the A'-D-π-A system.

  20. Exosomes of BV-2 cells induced by alpha-synuclein: important mediator of neurodegeneration in PD.

    Science.gov (United States)

    Chang, Chongwang; Lang, Hongjuan; Geng, Ning; Wang, Jing; Li, Nan; Wang, Xuelian

    2013-08-26

    Parkinson's disease (PD) is a progressive neurodegenerative disease. Alpha-synuclein aggregation, which can activate microglia to enhance its dopaminergic neurotoxicity, plays a central role in the progression of PD. However the mechanism is still unclear. To investigate how alpha-synuclein affects the neuron, exosomes were derived from alpha-synuclein treated mouse microglia cell line BV-2 cells by differential centrifugation and ultracentrifugation. We found that alpha-synuclein can induce an increase of exosomal secretion by microglia. These activated exosomes expressed a high level of MHC class II molecules and membrane TNF-α. In addition, the activated exosomes cause increased apoptosis. Exosomes secreted from activated microglias might be important mediator of alpha-synuclein-induced neurodegeneration in PD.

  1. Aggregation of the rhizospheric bacterium Azospirillum brasilense in response to oxygen

    Science.gov (United States)

    Abdoun, Hamid; McMillan, Mary; Pereg, Lily

    2016-04-01

    Azospirillum brasilense spp. have ecological, scientific and agricultural importance. As model plant growth promoting rhizobacteria they interact with a large variety of plants, including important food and cash crops. Azospirillum strains are known for their production of plant growth hormones that enhance root systems and for their ability to fix nitrogen. Azospirillum cells transform in response to environmental cues. The production of exopolysaccharides and cell aggregation during cellular transformation are important steps in the attachment of Azospirillum to roots. We investigate signals that induce cellular transformation and aggregation in the Azospirillum and report on the importance of oxygen to the process of aggregation in this rhizospheric bacterium.

  2. Calcitonin gene-related peptide induced migraine attacks in patients with and without familial aggregation of migraine

    DEFF Research Database (Denmark)

    Guo, Song; Christensen, Anne Francke; Liu, Marie Louise

    2017-01-01

    BACKGROUND: Calcitonin gene-related peptide provokes migraine attacks in 65% of patients with migraine without aura. Whether aggregation of migraine in first-degree relatives (family load) or a high number of risk-conferring single nucleotide polymorphisms contributes to migraine susceptibility...... to calcitonin gene-related peptide infusion in migraine patients is unknown. We hypothesized that genetic enrichment plays a role in triggering of migraine and, therefore, migraine without aura patients with high family load would report more migraine attacks after calcitonin gene-related peptide infusion than...... patients with low family load. METHODS: We allocated 40 previously genotyped migraine without aura patients to receive intravenous infusion of 1.5 µg/min calcitonin gene-related peptide and recorded migraine attacks including headache characteristics and associated symptoms. Information of familial...

  3. Ultrafast fluorescent decay induced by metal-mediated dipole-dipole interaction in two-dimensional molecular aggregates

    CERN Document Server

    Hu, Qing; Nam, Sang Hoon; Xiao, Jun; Liu, Yongmin; Zhang, Xiang; Fang, Nicholas X

    2016-01-01

    Two-dimensional molecular aggregate (2DMA), a thin sheet of strongly interacting dipole molecules self-assembled at close distance on an ordered lattice, is a fascinating fluorescent material. It is distinctively different from the single or colloidal dye molecules or quantum dots in most previous research. In this paper, we verify for the first time that when a 2DMA is placed at a nanometric distance from a metallic substrate, the strong and coherent interaction between the dipoles inside the 2DMA dominates its fluorescent decay at picosecond timescale. Our streak-camera lifetime measurement and interacting lattice-dipole calculation reveal that the metal-mediated dipole-dipole interaction shortens the fluorescent lifetime to about one half and increases the energy dissipation rate by ten times than expected from the noninteracting single-dipole picture. Our finding can enrich our understanding of nanoscale energy transfer in molecular excitonic systems and may designate a new direction for developing fast a...

  4. DRAG ON SUBMICRON NANOPARTICLE AGGREGATES

    Institute of Scientific and Technical Information of China (English)

    F.; Einar; Kruis

    2005-01-01

    A new procedure was developed for estimating the effective collision diameter of an aggregate composed of primary particles of any size. The coagulation coefficient of two oppositely charged particles was measured experimentally and compared with classic Fuchs theory, including a new method to account for particle non-sphericity. A second set of experiments were performed on well-defined nanoparticle aggregates at different stages of sintering, i.e. from the aggregate to the fully sintered stage. Here, electrical mobility was used to characterize the particle drag. The aggregates are being built from two different size-fractionated nanoparticle aerosols, the non-aggregated particles are discarded by an electrofilter and then they are passed through a furnace at concentrations low enough not to induce coagulation.

  5. Reprogramming T cell Lymphocytes to Induced Pluripotent Stem Cells

    Science.gov (United States)

    Bared, Kalia

    The discovery of induced pluripotent stem cells (iPSC) provided a novel technology for the study of development and pharmacology and complement embryonic stem cells (ES) for cell therapy applications. Though iPSC are derived from adult tissue they are comparable to ES cells in their behavior; multi-lineage differentiation and self-renewal. This makes iPSC research appealing because they can be studied in great detail and expanded in culture broadly. Fibroblasts were the first cell type reprogrammed to an iPSC using a retrovirus vector, since then alternative cell types including lymphocytes have been used to generate iPSC. Different types of vectors have also been developed to enhance iPSC formation and quality. However, specific T lymphocyte subsets have not been shown to reprogram to a pluripotent state to date. Here, we proposed to derive iPSC from peripheral blood effector and central memory T cells, reasoning that the resultant iPSC will maintain the epigenetic memory of a T lymphocyte, including the T cell receptor (TCR) gene rearrangement. This epigenetic memory will enable the differentiation and expansion of T cell iPSC into professional T cells containing a specific TCR. These could then be used for cell therapy to target specific antigens, as well as to improve culture techniques to expand T cells in vitro. We studied different gene delivery methods to derive iPSC from different types of T lymphocytes. We assessed the viability of viral transduction using flow cytometry to detect green fluorescent marker contained in the viral construct and quantitative real time polymerase chain reaction (qRT-PCR) to detect Oct4, Klf4, Sox2, and c-Myc gene expression. Our results demonstrate that the Sendai virus construct is the most feasible platform to reprogram T lymphocytes. We anticipate that this platform will provide an efficient and safe approach to derive iPSC from different T cell subsets, including memory T cells.

  6. Ectopic expression of cyclase associated protein CAP restores the streaming and aggregation defects of adenylyl cyclase a deficient Dictyostelium discoideum cells

    Directory of Open Access Journals (Sweden)

    Sultana Hameeda

    2012-01-01

    Full Text Available Abstract Background Cell adhesion, an integral part of D. discoideum development, is important for morphogenesis and regulated gene expression in the multicellular context and is required to trigger cell-differentiation. G-protein linked adenylyl cyclase pathways are crucially involved and a mutant lacking the aggregation specific adenylyl cyclase ACA does not undergo multicellular development. Results Here, we have investigated the role of cyclase-associated protein (CAP, an important regulator of cell polarity and F-actin/G-actin ratio in the aca- mutant. We show that ectopic expression of GFP-CAP improves cell polarization, streaming and aggregation in aca- cells, but it fails to completely restore development. Our studies indicate a requirement of CAP in the ACA dependent signal transduction for progression of the development of unicellular amoebae into multicellular structures. The reduced expression of the cell adhesion molecule DdCAD1 together with csA is responsible for the defects in aca- cells to initiate multicellular development. Early development was restored by the expression of GFP-CAP that enhanced the DdCAD1 transcript levels and to a lesser extent the csA mRNA levels. Conclusions Collectively, our data shows a novel role of CAP in regulating cell adhesion mechanisms during development that might be envisioned to unravel the functions of mammalian CAP during animal embryogenesis.

  7. Binase induces apoptosis of transformed myeloid cells and does not induce T-cell immune response.

    Science.gov (United States)

    Ilinskaya, Olga N; Zelenikhin, Pavel V; Petrushanko, Irina Yu; Mitkevich, Vladimir A; Prassolov, Vladimir S; Makarov, Alexander A

    2007-10-01

    Microbial RNases along with such animal RNases as onconase and BS-RNase are a promising basis for developing new antitumor drugs. We have shown that the Bacillus intermedius RNase (binase) induces selective apoptosis of transformed myeloid cells. It attacks artificially expressing activated c-Kit myeloid progenitor FDC cells and chronic myelogenous leukemia cells K562. Binase did not induce apoptosis in leukocytes of healthy donors and in normal myeloid progenitor cells. The inability of binase to initiate expression of activation markers CD69 and IFN-gamma in CD4+ and CD8+ T-lymphocytes testifies that enzyme is devoid of superantigenic properties. Altogether, these results demonstrate that binase possesses therapeutic opportunities for treatment of genotyped human neoplasms expressing activated kit.

  8. Human retinal pigment epithelial cell-induced apoptosis in activated T cells

    DEFF Research Database (Denmark)

    Jørgensen, A; Wiencke, A K; la Cour, M;

    1998-01-01

    induced apoptosis in several activated T-cell populations and T-cell lines, including T-cell antigen receptor (TCR)-CD3-negative T-cell lines. In contrast, RPE cells induced little or no apoptosis in resting peripheral T cells. Major histocompatibility complex (MHC) class II monoclonal antibodies, which...

  9. Erythroid differentiation of human induced pluripotent stem cells is independent of donor cell type of origin

    OpenAIRE

    2015-01-01

    Epigenetic memory in induced pluripotent stem cells, which is related to the somatic cell type of origin of the stem cells, might lead to variations in the differentiation capacities of the pluripotent stem cells. In this context, induced pluripotent stem cells from human CD34+ hematopoietic stem cells might be more suitable for hematopoietic differentiation than the commonly used fibroblast-derived induced pluripotent stem cells. To investigate the influence of an epigenetic memory on the ex...

  10. A label-free colorimetric aptasensor for simple, sensitive and selective detection of Pt (II) based on platinum (II)-oligonucleotide coordination induced gold nanoparticles aggregation.

    Science.gov (United States)

    Fan, Daoqing; Zhai, Qingfeng; Zhou, Weijun; Zhu, Xiaoqing; Wang, Erkang; Dong, Shaojun

    2016-11-15

    Herein, a gold nanoparticles (AuNPs) based label-free colorimetric aptasensor for simple, sensitive and selective detection of Pt (II) was constructed for the first time. Four bases (G-G mismatch) mismatched streptavidin aptamer (MSAA) was used to protect AuNPs from salt-induced aggregation and recognize Pt (II) specifically. Only in the presence of Pt (II), coordination occurs between G-G bases and Pt (II), leading to the activation of streptavidin aptamer. Streptavidin coated magnetic beads (MBs) were used as separation agent to separate Pt (II)-coordinated MSAA. The residual less amount of MSAA could not efficiently protect AuNPs anymore and aggregation of AuNPs will produce a colorimetric product. With the addition of Pt (II), a pale purple-to-blue color variation could be observed by the naked eye. A detection limit of 150nM and a linear range from 0.6μM to 12.5μM for Pt (II) could be achieved without any amplification.

  11. Aggregation of MBP in chronic demyelination

    Science.gov (United States)

    Frid, Kati; Einstein, Ofira; Friedman-Levi, Yael; Binyamin, Orli; Ben-Hur, Tamir; Gabizon, Ruth

    2015-01-01

    Objectives Misfolding of key disease proteins to an insoluble state is associated with most neurodegenerative conditions, such as prion, Parkinson, and Alzheimer’s diseases. In this work, and by studying animal models of multiple sclerosis, we asked whether this is also the case for myelin basic protein (MBP) in the late and neurodegenerative phases of demyelinating diseases. Methods To this effect, we tested whether MBP, an essential myelin component, present prion-like properties in animal models of MS, as is the case for Cuprizone-induced chronic demyelination or chronic phases of Experimental Autoimmune Encephalomyelitis (EAE). Results We show here that while total levels of MBP were not reduced following extensive demyelination, part of these molecules accumulated thereafter as aggregates inside oligodendrocytes or around neuronal cells. In chronic EAE, MBP precipitated concomitantly with Tau, a marker of diverse neurodegenerative conditions, including MS. Most important, analysis of fractions from Triton X-100 floatation gradients suggest that the lipid composition of brain membranes in chronic EAE differs significantly from that of naïve mice, an effect which may relate to oxidative insults and subsequently prevent the appropriate insertion and compaction of new MBP in the myelin sheath, thereby causing its misfolding and aggregation. Interpretation Prion-like aggregation of MBP following chronic demyelination may result from an aberrant lipid composition accompanying this pathological status. Such aggregation of MBP may contribute to neuronal damage that occurs in the progressive phase of MS. PMID:26273684

  12. Marmoset induced pluripotent stem cells: Robust neural differentiation following pretreatment with dimethyl sulfoxide

    Directory of Open Access Journals (Sweden)

    Zhifang Qiu

    2015-07-01

    Full Text Available The marmoset is an important nonhuman primate model for regenerative medicine. For experimental autologous cell therapy based on induced pluripotent (iPS cells in the marmoset, cells must be able to undergo robust and reliable directed differentiation that will not require customization for each specific iPS cell clone. When marmoset iPS cells were aggregated in a hanging drop format for 3 days, followed by exposure to dual SMAD inhibitors and retinoic acid in monolayer culture for 3 days, we found substantial variability in the response of different iPS cell clones. However, when clones were pretreated with 0.05–2% dimethyl sulfoxide (DMSO for 24 hours, all clones showed a very similar maximal response to the directed differentiation scheme. Peak responses were observed at 0.5% DMSO in two clones and at 1% DMSO in a third clone. When patterns of gene expression were examined by microarray analysis, hierarchical clustering showed very similar responses in all 3 clones when they were pretreated with optimal DMSO concentrations. The change in phenotype following exposure to DMSO and the 6 day hanging drop/monolayer treatment was confirmed by immunocytochemistry. Analysis of DNA content in DMSO-exposed cells indicated that it is unlikely that DMSO acts by causing cells to exit from the cell cycle. This approach should be generally valuable in the directed neural differentiation of pluripotent cells for experimental cell therapy.

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

    Science.gov (United States)

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

    2014-07-01

    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.

  14. Detection of the critical micelle concentration of cationic and anionic surfactants based on aggregation-induced emission property of hexaphenylsilole derivatives

    Institute of Scientific and Technical Information of China (English)

    2009-01-01

    We report a fluorescence "turn-on" method to detect the critical micelle concentration (CMC) of surfactants. This method works well for both cationic and anionic surfactants. It employs an unprecedented mechanism (aggregation-induced emission, or AIE) to determine the CMC values, and the results are consistent with the data obtained by the classical techniques. In addition, this method renders the convenient detection of the CMC values. Any large and professional instruments are unnecessary, instead, a portable UV lamp and an ultrasonic generator are enough to carry out the detection in an ordinary laboratory. Considering that micelles are interesting entities and have found applications in many important fields such as emulsion polymerization, template of nanosized materials synthesis, controllable drug delivery and macromolecular self-assembling. Our experimental results may offer a facile, sensitive and promising method to detect the formation of micelles constructed by the new amphiphilic molecules and macromolecules.

  15. Detection of the critical micelle concentration of cationic and anionic surfactants based on aggregation-induced emission property of hexaphenylsilole derivatives

    Institute of Scientific and Technical Information of China (English)

    TANG Li; JIN JiaKe; ZHANG Shuang; MAO Yu; SUN JingZhi; YUAN WangZhang; ZHAO Hui; XU HaiPeng; QIN AnJun; TANG Ben Zhong

    2009-01-01

    We report a fluorescence "turn-on" method to detect the critical micelle concentration (CMC) of sur-factants. This method works well for both cationic and anionic surfactants. It employs an unprece-dented mechanism (aggregation-induced emission, or AIE) to determine the CMC values, and the re-suits are consistent with the data obtained by the classical techniques. In addition, this method renders the convenient detection of the CMC values. Any large and professional instruments are unnecessary, instead, a portable UV lamp and an ultrasonic generator are enough to carry out the detection in an ordinary laboratory. Considering that micelles are interesting entities and have found applications in many important fields such as emulsion polymerization, template of nanoeized materials synthesis, controllable drug delivery and macromolecular self-assembling. Our experimental results may offer a facile, sensitive and promising method to detect the formation of micelles constructed by the new amphiphilic molecules and macromolecules.

  16. BMP-2 induces versican and hyaluronan that contribute to post-EMT AV cushion cell migration.

    Directory of Open Access Journals (Sweden)

    Kei Inai

    Full Text Available Distal outgrowth and maturation of mesenchymalized endocardial cushions are critical morphogenetic events during post-EMT atrioventricular (AV valvuloseptal morphogenesis. We explored the role of BMP-2 in the regulation of valvulogenic extracellular matrix (ECM components, versican and hyaluronan (HA, and cell migration during post-EMT AV cushion distal outgrowth/expansion. We observed intense staining of versican and HA in AV cushion mesenchyme from the early cushion expansion stage, Hamburger and Hamilton (HH stage-17 to the cushion maturation stage, HH stage-29 in the chick. Based on this expression pattern we examined the role of BMP-2 in regulating versican and HA using 3D AV cushion mesenchymal cell (CMC aggregate cultures on hydrated collagen gels. BMP-2 induced versican expression and HA deposition as well as mRNA expression of versican and Has2 by CMCs in a dose dependent manner. Noggin, an antagonist of BMP, abolished BMP-2-induced versican and HA as well as mRNA expression of versican and Has2. We further examined whether BMP-2-promoted cell migration was associated with expression of versican and HA. BMP-2- promoted cell migration was significantly impaired by treatments with versican siRNA and HA oligomer. In conclusion, we provide evidence that BMP-2 induces expression of versican and HA by AV CMCs and that these ECM components contribute to BMP-2-induced CMC migration, indicating critical roles for BMP-2 in distal outgrowth/expansion of mesenchymalized AV cushions.

  17. Arginine-glycine-aspartic acid-polyethylene glycol-polyamidoamine dendrimer conjugate improves liver-cell aggregation and function in 3-D spheroid culture.

    Science.gov (United States)

    Chen, Zhanfei; Lian, Fen; Wang, Xiaoqian; Chen, Yanling; Tang, Nanhong

    The polyamidoamine (PAMAM) dendrimer, a type of macromolecule material, has been used in spheroidal cell culture and drug delivery in recent years. However, PAMAM is not involved in the study of hepatic cell-spheroid culture or its biological activity, particularly in detoxification function. Here, we constructed a PAMAM-dendrimer conjugate decorated by an integrin ligand: arginine-glycine-aspartic acid (RGD) peptide. Our studies demonstrate that RGD-polyethylene glycol (PEG)-PAMAM conjugates can promote singly floating hepatic cells to aggregate together in a sphere-like growth with a weak reactive oxygen species. Moreover, RGD-PEG-PAMAM conjugates can activate the AKT-MAPK pathway in hepatic cells to promote cell proliferation and improve basic function and ammonia metabolism. Together, our data support the hepatocyte sphere treated by RGD-PEG-PAMAM conjugates as a potential source of hepatic cells for a biological artificial liver system.

  18. Generation of healthy mice from gene-corrected disease-specific induced pluripotent stem cells.

    Directory of Open Access Journals (Sweden)

    Guangming Wu

    2011-07-01

    Full Text Available Using the murine model of tyrosinemia type 1 (fumarylacetoacetate hydrolase [FAH] deficiency; FAH⁻/⁻ mice as a paradigm for orphan disorders, such as hereditary metabolic liver diseases, we evaluated fibroblast-derived FAH⁻/⁻-induced pluripotent stem cells (iPS cells as targets for gene correction in combination with the tetraploid embryo complementation method. First, after characterizing the FAH⁻/⁻ iPS cell lines, we aggregated FAH⁻/⁻-iPS cells with tetraploid embryos and obtained entirely FAH⁻/⁻-iPS cell-derived mice that were viable and exhibited the phenotype of the founding FAH⁻/⁻ mice. Then, we transduced FAH cDNA into the FAH⁻/⁻-iPS cells using a third-generation lentiviral vector to generate gene-corrected iPS cells. We could not detect any chromosomal alterations in these cells by high-resolution array CGH analysis, and after their aggregation with tetraploid embryos, we obtained fully iPS cell-derived healthy mice with an astonishing high efficiency for full-term development of up to 63.3%. The gene correction was validated functionally by the long-term survival and expansion of FAH-positive cells of these mice after withdrawal of the rescuing drug NTBC (2-(2-nitro-4-fluoromethylbenzoyl-1,3-cyclohexanedione. Furthermore, our results demonstrate that both a liver-specific promoter (transthyretin, TTR-driven FAH transgene and a strong viral promoter (from spleen focus-forming virus, SFFV-driven FAH transgene rescued the FAH-deficiency phenotypes in the mice derived from the respective gene-corrected iPS cells. In conclusion, our data demonstrate that a lentiviral gene repair strategy does not abrogate the full pluripotent potential of fibroblast-derived iPS cells, and genetic manipulation of iPS cells in combination with tetraploid embryo aggregation provides a practical and rapid approach to evaluate the efficacy of gene correction of human diseases in mouse models.

  19. The Preliminary Experimental Study of Induced Differentiation of Embryonic Stem Cells into Corneal Epithelial Cells

    Institute of Scientific and Technical Information of China (English)

    Ling Yu; Jian Ge; Zhichong Wang; Bing Huang; Keming Yu; Chongde Long; Xigu Chen

    2001-01-01

    Purpose:To study preliminarily induced differentiation of embryonic stem cells intocorneal epithelial cells in vitro.Methods: Murine embryonic stem cells were co-cultured with Rabbit limbal cornealepithelial cells in Transwell system to induce differentiation. Mophological andimmunohistochemical examination were implemented.Results: The induced cells from embryonic stem cells have an epithelial appearance.The cells formed a network and were confluent into film gradually after beingco-cultured with rabbit limbal corneal epithelial cells for 24 ~ 96 hours. The cells rangedmosaic structure and localized together with clear rim. Most of the cells showedpolygonal appearance. Transmission electron microscope showed lots of microvilli on thesurface of induced cells and tight junctions between them. These epithelial-like cellsexpressed the corneal epithelial cell specific marker cytokeratin3/cytokeratinl2.Conclusion: The potential mechanism of the differentiation of murine embryonic stemcells into corneal epithelia