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Sample records for human cell survival

  1. Human neural progenitor cells promote photoreceptor survival in retinal explants.

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    Englund-Johansson, Ulrica; Mohlin, Camilla; Liljekvist-Soltic, Ingela; Ekström, Per; Johansson, Kjell

    2010-02-01

    Different types of progenitor and stem cells have been shown to provide neuroprotection in animal models of photoreceptor degeneration. The present study was conducted to investigate whether human neural progenitor cells (HNPCs) have neuroprotective properties on retinal explants models with calpain- and caspase-3-dependent photoreceptor cell death. In the first experiments, HNPCs in a feeder layer were co-cultured for 6 days either with postnatal rd1 mouse or normal rat retinas. Retinal histological sections were used to determine outer nuclear layer (ONL) thickness, and to detect the number of photoreceptors with labeling for calpain activity, cleaved caspase-3 and TUNEL. The ONL thickness of co-cultured rat and rd1 retinas was found to be almost 10% and 40% thicker, respectively, compared to controls. Cell counts of calpain activity, cleaved caspase-3 and TUNEL labeled photoreceptors in both models revealed a 30-50% decrease when co-cultured with HNPCs. The results represent significant increases of photoreceptor survival in the co-cultured retinas. In the second experiments, for an identification of putative survival factors, or a combination of them, a growth factor profile was performed on conditioned medium. The relative levels of various growth factors were analyzed by densitometric measurements of growth factor array membranes. Following growth factors were identified as most potential survival factors; granulocyte colony stimulating factor (G-CSF), granulocyte-macrophage colony stimulating factor (GMCSF), insulin-like growth factor II (IGF-II), neurotrophic factor 3 (NT-3), placental growth factor (PIGF), transforming growth factors (TGF-beta1 and TGF-beta2) and vascular endothelial growth factor (VEGF-D). HNPCs protect both against calpain- and caspase-3-dependent photoreceptor cell death in the rd1 mouse and against caspase-3-dependent photoreceptor cell death in normal rat retinas in vitro. The protective effect is possibly achieved by a variety of

  2. Rhodococcus equi human clinical isolates enter and survive within human alveolar epithelial cells.

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    Ramos-Vivas, J; Pilares-Ortega, L; Remuzgo-Martínez, S; Padilla, D; Gutiérrez-Díaz, J L; Navas-Méndez, J

    2011-05-01

    Rhodococcus equi is an emerging opportunistic human pathogen associated with immunosuppressed people, especially those infected with the human immunodeficiency virus (HIV). This pathogen resides primarily within lung macrophages of infected patients, which may explain in part its ability to escape normal pulmonary defense mechanisms. Despite numerous studies as a pulmonary pathogen in foals, where a plasmid seems to play an important role in virulence, information on the pathogenesis of this pathogen in humans is still scarce. In this study, fluorescence microscopy and vancomycin protection assays were used to investigate the ability of R. equi human isolates to adhere to and to invade the human alveolar epithelial cell line A549. Our findings indicate that some R. equi clinical strains are capable of adhering, entering and surviving within the alveolar cell line, which may contribute to the pathogen persistence in lung tissues. Copyright © 2011 Institut Pasteur. Published by Elsevier SAS. All rights reserved.

  3. Time-course regulation of quercetin on cell survival/proliferation pathways in human hepatoma cells.

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    Granado-Serrano, Ana Belén; Angeles Martín, María; Bravo, Laura; Goya, Luis; Ramos, Sonia

    2008-04-01

    Quercetin, a dietary flavonoid, has been shown to possess anticarcinogenic properties, but the precise molecular mechanisms of action are not thoroughly elucidated. This study was aimed at investigating the time-course regulation effect of quercetin on survival/proliferation pathways in a human hepatoma cell line (HepG2). Quercetin induced a significant time-dependent inactivation of the major survival signaling proteins, i. e., phosphatidylinositol 3-kinase (PI 3-kinase)/protein kinase B (AKT), extracellular regulated kinase (ERK), protein kinase C-alpha (PKC-alpha), in concert with a time-dependent activation of key death-related signals: c-jun amino-terminal kinase (JNK) and PKC-delta. These data suggest that quercetin exerts a tight regulation of survival/proliferation pathways that requires the integration of different signals and persists over time, being the balance of these regulatory signals what determines the fate of HepG2 cells.

  4. Molecular Mechanisms Regulating Human Dendritic Cell Development, Survival and Function

    NARCIS (Netherlands)

    L. van de Laar (Lianne)

    2011-01-01

    textabstractDendritic cells (DC) are professional antigen presenting cells (APC) with a dual function in the immune system. On the one hand, these specialized leukocytes are equipped to alert the immune system to invading pathogens or other danger signals. On the other, DC can promote tolerogenic re

  5. Survival of human embryonic stem cells implanted in the guinea pig auditory epithelium

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    Young Lee, Min; Hackelberg, Sandra; Green, Kari L.; Lunghamer, Kelly G.; Kurioka, Takaomi; Loomis, Benjamin R.; Swiderski, Donald L.; Duncan, R. Keith; Raphael, Yehoash

    2017-01-01

    Hair cells in the mature cochlea cannot spontaneously regenerate. One potential approach for restoring hair cells is stem cell therapy. However, when cells are transplanted into scala media (SM) of the cochlea, they promptly die due to the high potassium concentration. We previously described a method for conditioning the SM to make it more hospitable to implanted cells and showed that HeLa cells could survive for up to a week using this method. Here, we evaluated the survival of human embryonic stem cells (hESC) constitutively expressing GFP (H9 Cre-LoxP) in deaf guinea pig cochleae that were pre-conditioned to reduce potassium levels. GFP-positive cells could be detected in the cochlea for at least 7 days after the injection. The cells appeared spherical or irregularly shaped, and some were aggregated. Flushing SM with sodium caprate prior to transplantation resulted in a lower proportion of stem cells expressing the pluripotency marker Oct3/4 and increased cell survival. The data demonstrate that conditioning procedures aimed at transiently reducing the concentration of potassium in the SM facilitate survival of hESCs for at least one week. During this time window, additional procedures can be applied to initiate the differentiation of the implanted hESCs into new hair cells. PMID:28387239

  6. Human Hematopoietic Stem Cells Can Survive In Vitro for Several Months

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    Taro Ishigaki

    2009-01-01

    Full Text Available We previously reported that long-lasting in vitro hematopoiesis could be achieved using the cells differentiated from primate embryonic stem (ES cells. Thus, we speculated that hematopoietic stem cells differentiated from ES cells could sustain long-lasting in vitro hematopoiesis. To test this hypothesis, we investigated whether human hematopoietic stem cells could similarly sustain long-lasting in vitro hematopoiesis in the same culture system. Although the results varied between experiments, presumably due to differences in the quality of each hematopoietic stem cell sample, long-lasting in vitro hematopoiesis was observed to last up to nine months. Furthermore, an in vivo analysis in which cultured cells were transplanted into immunodeficient mice indicated that even after several months of culture, hematopoietic stem cells were still present in the cultured cells. To the best of our knowledge, this is the first report to show that human hematopoietic stem cells can survive in vitro for several months.

  7. Effect of fatty acids on human bone marrow mesenchymal stem cell energy metabolism and survival.

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    Fillmore, Natasha; Huqi, Alda; Jaswal, Jagdip S; Mori, Jun; Paulin, Roxane; Haromy, Alois; Onay-Besikci, Arzu; Ionescu, Lavinia; Thébaud, Bernard; Michelakis, Evangelos; Lopaschuk, Gary D

    2015-01-01

    Successful stem cell therapy requires the optimal proliferation, engraftment, and differentiation of stem cells into the desired cell lineage of tissues. However, stem cell therapy clinical trials to date have had limited success, suggesting that a better understanding of stem cell biology is needed. This includes a better understanding of stem cell energy metabolism because of the importance of energy metabolism in stem cell proliferation and differentiation. We report here the first direct evidence that human bone marrow mesenchymal stem cell (BMMSC) energy metabolism is highly glycolytic with low rates of mitochondrial oxidative metabolism. The contribution of glycolysis to ATP production is greater than 97% in undifferentiated BMMSCs, while glucose and fatty acid oxidation combined only contribute 3% of ATP production. We also assessed the effect of physiological levels of fatty acids on human BMMSC survival and energy metabolism. We found that the saturated fatty acid palmitate induces BMMSC apoptosis and decreases proliferation, an effect prevented by the unsaturated fatty acid oleate. Interestingly, chronic exposure of human BMMSCs to physiological levels of palmitate (for 24 hr) reduces palmitate oxidation rates. This decrease in palmitate oxidation is prevented by chronic exposure of the BMMSCs to oleate. These results suggest that reducing saturated fatty acid oxidation can decrease human BMMSC proliferation and cause cell death. These results also suggest that saturated fatty acids may be involved in the long-term impairment of BMMSC survival in vivo.

  8. Human mesenchymal stem cells promote survival of T cells in a quiescent state.

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    Benvenuto, Federica; Ferrari, Stefania; Gerdoni, Ezio; Gualandi, Francesca; Frassoni, Francesco; Pistoia, Vito; Mancardi, Gianluigi; Uccelli, Antonio

    2007-07-01

    Mesenchymal stem cells (MSC) are part of the bone marrow that provides signals supporting survival and growth of bystander hematopoietic stem cells (HSC). MSC modulate also the immune response, as they inhibit proliferation of lymphocytes. In order to investigate whether MSC can support survival of T cells, we investigated MSC capacity of rescuing T lymphocytes from cell death induced by different mechanisms. We observed that MSC prolong survival of unstimulated T cells and apoptosis-prone thymocytes cultured under starving conditions. MSC rescued T cells from activation induced cell death (AICD) by downregulation of Fas receptor and Fas ligand on T cell surface and inhibition of endogenous proteases involved in cell death. MSC dampened also Fas receptor mediated apoptosis of CD95 expressing Jurkat leukemic T cells. In contrast, rescue from AICD was not associated with a significant change of Bcl-2, an inhibitor of apoptosis induced by cell stress. Accordingly, MSC exhibited a minimal capacity of rescuing Jurkat cells from chemically induced apoptosis, a process disrupting the mitochondrial membrane potential regulated by Bcl-2. These results suggest that MSC interfere with the Fas receptor regulated process of programmed cell death. Overall, MSC can inhibit proliferation of activated T cells while supporting their survival in a quiescent state, providing a model of their activity inside the HSC niche. Disclosure of potential conflicts of interest is found at the end of this article.

  9. AKT/GSK3β signaling pathway is critically involved in human pluripotent stem cell survival

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    Romorini, Leonardo; Garate, Ximena; Neiman, Gabriel; Luzzani, Carlos; Furmento, Verónica Alejandra; Guberman, Alejandra Sonia; Sevlever, Gustavo Emilio; Scassa, María Elida; Miriuka, Santiago Gabriel

    2016-01-01

    Human embryonic and induced pluripotent stem cells are self-renewing pluripotent stem cells (PSC) that can differentiate into a wide range of specialized cells. Basic fibroblast growth factor is essential for PSC survival, stemness and self-renewal. PI3K/AKT pathway regulates cell viability and apoptosis in many cell types. Although it has been demonstrated that PI3K/AKT activation by bFGF is relevant for PSC stemness maintenance its role on PSC survival remains elusive. In this study we explored the molecular mechanisms involved in the regulation of PSC survival by AKT. We found that inhibition of AKT with three non-structurally related inhibitors (GSK690693, AKT inhibitor VIII and AKT inhibitor IV) decreased cell viability and induced apoptosis. We observed a rapid increase in phosphatidylserine translocation and in the extent of DNA fragmentation after inhibitors addition. Moreover, abrogation of AKT activity led to Caspase-9, Caspase-3, and PARP cleavage. Importantly, we demonstrated by pharmacological inhibition and siRNA knockdown that GSK3β signaling is responsible, at least in part, of the apoptosis triggered by AKT inhibition. Moreover, GSK3β inhibition decreases basal apoptosis rate and promotes PSC proliferation. In conclusion, we demonstrated that AKT activation prevents apoptosis, partly through inhibition of GSK3β, and thus results relevant for PSC survival. PMID:27762303

  10. SIRT1 and c-Myc Promote Liver Tumor Cell Survival and Predict Poor Survival of Human Hepatocellular Carcinomas

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    Jang, Kyu Yun; Noh, Sang Jae; Lehwald, Nadja; Tao, Guo-Zhong; Bellovin, David I.; Park, Ho Sung; Moon, Woo Sung; Felsher, Dean W.; Sylvester, Karl G.

    2012-01-01

    The increased expression of SIRT1 has recently been identified in numerous human tumors and a possible correlation with c-Myc oncogene has been proposed. However, it remains unclear whether SIRT1 functions as an oncogene or tumor suppressor. We sought to elucidate the role of SIRT1 in liver cancer under the influence of c-Myc and to determine the prognostic significance of SIRT1 and c-Myc expression in human hepatocellular carcinoma. The effect of either over-expression or knock down of SIRT1 on cell proliferation and survival was evaluated in both mouse and human liver cancer cells. Nicotinamide, an inhibitor of SIRT1, was also evaluated for its effects on liver tumorigenesis. The prognostic significance of the immunohistochemical detection of SIRT1 and c-Myc was evaluated in 154 hepatocellular carcinoma patients. SIRT1 and c-Myc regulate each other via a positive feedback loop and act synergistically to promote hepatocellular proliferation in both mice and human liver tumor cells. Tumor growth was significantly inhibited by nicotinamide in vivo and in vitro. In human hepatocellular carcinoma, SIRT1 expression positively correlated with c-Myc, Ki67 and p53 expression, as well as high á-fetoprotein level. Moreover, the expression of SIRT1, c-Myc and p53 were independent prognostic indicators of hepatocellular carcinoma. In conclusion, this study demonstrates that SIRT1 expression supports liver tumorigenesis and is closely correlated with oncogenic c-MYC expression. In addition, both SIRT1 and c-Myc may be useful prognostic indicators of hepatocellular carcinoma and SIRT1 targeted therapy may be beneficial in the treatment of hepatocellular carcinoma. PMID:23024800

  11. Intracellular trehalose improves the survival of human red blood cells by freeze-drying

    Institute of Scientific and Technical Information of China (English)

    HE Hui; LIU Baolin; HUA Zezhao; LI Chuan; WU Zhengzheng

    2007-01-01

    Freeze-drying of human red blood cells has a potential important application for blood transfusion.The aim of this study was to investigate the effects ofintracellular trehalose on the survival of red blood cells after freeze-drying and rehydration.Fresh red blood cells were incubated in trehalose solutions of various concentrations at 37℃ for 7 h following freeze-drying.Polyvinylpyrrolidone,Trehalose,sodium citrate,and human serum albumin were used as extracellular protective agents for the freeze-drying of red blood cells.The results indicated that the intracellular trehalose concentration was increased with increasing concentration of extracellular trehalose solution,and the maximum concen tration of intracellular trehalose reached 35 mmol/L.The viability of freeze-dried red blood cells increased with the increment of intracellular trehalose concentration.

  12. Effects of Thapsigargin on the Proliferation and Survival of Human Rheumatoid Arthritis Synovial Cells

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    Hui Wang

    2014-01-01

    Full Text Available A series of experiments have been carried out to investigate the effects of different concentrations of thapsigargin (0, 0.001, 0.1, and 1 μM on the proliferation and survival of human rheumatoid arthritis synovial cells (MH7A. The results showed that thapsigargin can block the cell proliferation in human rheumatoid arthritis synovial cells in a time- and dose-dependent manner. Results of Hoechst staining suggested that thapsigargin may induce cell apoptosis in MH7A cells in a time- and dose-dependent manner, and the percentages of cell death reached 44.6% at thapsigargin concentration of 1 μM treated for 4 days compared to the control. The protein and mRNA levels of cyclin D1 decreased gradually with the increasing of thapsigargin concentration and treatment times. Moreover, the protein levels of mTORC1 downstream indicators pS6K and p4EBP-1 were reduced by thapsigargin treatment at different concentrations and times, which should be responsible for the reduced cyclin D1 expressions. Our results revealed that thapsigargin may effectively impair the cell proliferation and survival of MH7A cells. The present findings will help to understand the molecular mechanism of fibroblast-like synoviocytes proliferations and suggest that thapsigargin is of potential for the clinical treatment of rheumatoid arthritis.

  13. Human ribosomal protein L9 is a Bax suppressor that promotes cell survival in yeast.

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    Eid, Rawan; Sheibani, Sara; Gharib, Nada; Lapointe, Jason F; Horowitz, Avital; Vali, Hojatollah; Mandato, Craig A; Greenwood, Michael T

    2014-05-01

    The identification of a human ribosomal protein L9 (hRPL9) cDNA as a sequence capable of suppressing the lethal effects of heterologously expressed murine Bax in yeast led us to investigate its antiapoptotic potential. Using growth and viability assays, we show that yeast cells heterologously expressing hRPL9 are resistant to the growth inhibitory and lethal effects of exogenously supplied copper, indicating that it has pro-survival properties. To explore potential mechanisms, we used yeast mutants defective in all three types of programmed cell death (apoptosis, necrosis, and autophagy). The ability to retain pro-survival function in all the mutants suggests that hRPL9 may regulate a common pro-death process. In contrast, the yeast RPL9 orthologues, RPL9A and RPL9B, have opposite effects when overexpressed in yeast. In effect, instead of showing resistance to stress, RPL9A and RPL9B overexpressing cells show reduced cell growth. Further analysis indicates that the effects of overexpressed RPL9A and RPL9B are not in themselves lethal, instead, they serve to increase cell doubling time. Thus, yeast RPL9s are more representative of RPs whose extra-ribosomal function is similar to that of tumor suppressors. Taken together, our results demonstrate that RPL9 represents a species- and sequence-specific regulator of cell growth and survival.

  14. Human and Autologous Adipose-derived Stromal Cells Increase Flap Survival in Rats Independently of Host Immune Response

    DEFF Research Database (Denmark)

    Toyserkani, Navid Mohamadpour; Jensen, Charlotte Harken; Andersen, Ditte Caroline

    2017-01-01

    evaluated after 7 days. RESULTS: The mean survival rates for SVF treatment regardless of human or autologous origin were significantly increased as compared with the control group. Adipose stem/stromal cell and SVF lysate injection did not increase flap survival. Vessel density was increased for human...... and rat SVF and human ASC but not for SVF lysate. Human cells were not detected in the flaps after 7 days. CONCLUSIONS: Flap survival increased with SVF treatment regardless of human or autologous origin, suggesting that increased flap survival is independent of the host immune response. All cell...... injections lead to increased vessel density, but it did not necessarily lead to increased flap survival. Further research should elaborate which molecular events make SVF treatment more efficacious than ASC....

  15. Androgen receptor signaling is required for androgen-sensitive human prostate cancer cell proliferation and survival

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    Day Wanda V

    2005-04-01

    Full Text Available Abstract Background Androgens and androgen receptors (AR regulate normal prostate development and growth. They also are involved in pathological development of prostatic diseases, including benign prostatic hyperplasia (BPH and prostate cancer (PCa. Antiandrogen therapy for PCa, in conjunction with chemical or surgical castration, offers initial positive responses and leads to massive prostate cell death. However, cancer cells later appear as androgen-independent PCa. To investigate the role of AR in prostate cell proliferation and survival, we introduced a vector-based small interfering RNA (siRNA. This siRNA targeted 5'-untranslated region of AR mRNA for extended suppression of AR expression in androgen-sensitive human prostate LNCaP cells. Results The siRNA design successfully suppressed endogenous AR expression, as revealed by western blotting and immunofluorescence staining in LNCaP cells. LNCaP cells did not proliferate in the absence of AR and underwent apoptosis, based on elevated phospho-Histone H2B expression and higher number of apoptotic body as compared to control cells. Conclusion We demonstrated that AR is vital for prostate cell proliferation and survival in this androgen-sensitive prostate cell line. These results further strengthen the hypothesis that AR can be a therapeutic target for treating androgen-sensitive stages of PCa. Unlike antiandorgens, however, siRNA targeting AR provides a direct inactivation of AR function through the suppression of AR protein expression.

  16. Survival and differentiation of transplanted neural stem cells derived from human induced pluripotent stem cells in a rat stroke model.

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    Jensen, Matthew B; Yan, Hongmei; Krishnaney-Davison, Rajeev; Al Sawaf, Abdullah; Zhang, Su-Chun

    2013-05-01

    Although administration of various stem cells has shown promise in stroke models, neural stem cells (NSCs) derived from human induced pluripotent stem cells (iPSCs) have advantages over other cell types. We studied whether these cells could survive, differentiate, and improve stroke recovery in an ischemic stroke model. Human iPSCs were induced in vitro to an early NSC stage. One week after focal cerebral ischemia, 20 rats received cells or vehicle by intracerebral injection. Graft cell fate, infarct volume, and behavioral deficits were assessed. Graft cells were found in 8 of the transplanted rats (80%), with estimated mean graft cell numbers nearly double the amount transplanted 1 month later. Graft cells also expressed markers of NSCs in 5 rats (63%), neurons in all 8 rats (100%), rare astrocytes in 4 rats (50%), and signs of proliferation in 4 rats (50%), but no tumor formation was observed. Stroke volume and behavioral recovery were similar between the groups. To our knowledge, this is the first report of transplantation of NSCs derived from human iPSCs in a stroke model. Human iPSC-derived NSCs survived in the postischemic rat brain and appeared to differentiate, primarily into neurons. This cell transplantation approach for stroke appears to be feasible, but further optimization is needed. Copyright © 2013 National Stroke Association. Published by Elsevier Inc. All rights reserved.

  17. Entrance and Survival of Brucella pinnipedialis Hooded Seal Strain in Human Macrophages and Epithelial Cells

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    Briquemont, Benjamin; Sørensen, Karen K.; Godfroid, Jacques

    2013-01-01

    Marine mammal Brucella spp. have been isolated from pinnipeds (B. pinnipedialis) and cetaceans (B. ceti) from around the world. Although the zoonotic potential of marine mammal brucellae is largely unknown, reports of human disease exist. There are few studies of the mechanisms of bacterial intracellular invasion and multiplication involving the marine mammal Brucella spp. We examined the infective capacity of two genetically different B. pinnipedialis strains (reference strain; NTCT 12890 and a hooded seal isolate; B17) by measuring the ability of the bacteria to enter and replicate in cultured phagocytes and epithelial cells. Human macrophage-like cells (THP-1), two murine macrophage cell lines (RAW264.7 and J774A.1), and a human malignant epithelial cell line (HeLa S3) were challenged with bacteria in a gentamicin protection assay. Our results show that B. pinnipedialis is internalized, but is then gradually eliminated during the next 72 – 96 hours. Confocal microscopy revealed that intracellular B. pinnipedialis hooded seal strain colocalized with lysosomal compartments at 1.5 and 24 hours after infection. Intracellular presence of B. pinnipedialis hooded seal strain was verified by transmission electron microscopy. By using a cholesterol-scavenging lipid inhibitor, entrance of B. pinnipedialis hooded seal strain in human macrophages was significantly reduced by 65.8 % (± 17.3), suggesting involvement of lipid-rafts in intracellular entry. Murine macrophages invaded by B. pinnipedialis do not release nitric oxide (NO) and intracellular bacterial presence does not induce cell death. In summary, B. pinnipedialis hooded seal strain can enter human and murine macrophages, as well as human epithelial cells. Intracellular entry of B. pinnipedialis hooded seal strain involves, but seems not to be limited to, lipid-rafts in human macrophages. Brucella pinnipedialis does not multiply or survive for prolonged periods intracellulary. PMID:24376851

  18. Entrance and survival of Brucella pinnipedialis hooded seal strain in human macrophages and epithelial cells.

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    Anett K Larsen

    Full Text Available Marine mammal Brucella spp. have been isolated from pinnipeds (B. pinnipedialis and cetaceans (B. ceti from around the world. Although the zoonotic potential of marine mammal brucellae is largely unknown, reports of human disease exist. There are few studies of the mechanisms of bacterial intracellular invasion and multiplication involving the marine mammal Brucella spp. We examined the infective capacity of two genetically different B. pinnipedialis strains (reference strain; NTCT 12890 and a hooded seal isolate; B17 by measuring the ability of the bacteria to enter and replicate in cultured phagocytes and epithelial cells. Human macrophage-like cells (THP-1, two murine macrophage cell lines (RAW264.7 and J774A.1, and a human malignant epithelial cell line (HeLa S3 were challenged with bacteria in a gentamicin protection assay. Our results show that B. pinnipedialis is internalized, but is then gradually eliminated during the next 72-96 hours. Confocal microscopy revealed that intracellular B. pinnipedialis hooded seal strain colocalized with lysosomal compartments at 1.5 and 24 hours after infection. Intracellular presence of B. pinnipedialis hooded seal strain was verified by transmission electron microscopy. By using a cholesterol-scavenging lipid inhibitor, entrance of B. pinnipedialis hooded seal strain in human macrophages was significantly reduced by 65.8 % (± 17.3, suggesting involvement of lipid-rafts in intracellular entry. Murine macrophages invaded by B. pinnipedialis do not release nitric oxide (NO and intracellular bacterial presence does not induce cell death. In summary, B. pinnipedialis hooded seal strain can enter human and murine macrophages, as well as human epithelial cells. Intracellular entry of B. pinnipedialis hooded seal strain involves, but seems not to be limited to, lipid-rafts in human macrophages. Brucella pinnipedialis does not multiply or survive for prolonged periods intracellulary.

  19. Signal transducer and activator of transcription 3 is involved in cell growth and survival of human rhabdomyosarcoma and osteosarcoma cells

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    Qualman Stephen J

    2007-06-01

    Full Text Available Abstract Background Stat3 has been classified as a proto-oncogene and constitutive Stat3 signaling appears to be involved in oncogenesis of human cancers. However, whether constitutive Stat3 signaling plays a role in the survival and growth of osteosarcomas, rhabdomyosarcomas, and soft-tissue sarcomas is still unclear. Methods To examine whether Stat3 is activated in osteosarcomas, rhabdomyosarcomas and other soft-tissue sarcomas we analyzed sarcoma tissue microarray slides and sarcoma cell lines using immunohistochemistry and Western blot analysis, respectively, with a phospho-specific Stat3 antibody. To examine whether the activated Stat3 pathway is important for sarcoma cell growth and survival, adenovirus-mediated expression of a dominant-negative Stat3 (Y705F and a small molecule inhibitor (termed STA-21 were used to inhibit constitutive Stat3 signaling in human sarcoma cell lines expressing elevated levels of Stat3 phosphorylation. Cell viability was determined by MTT assays and induction of apoptosis was analyzed by western blotting using antibodies that specifically recognize cleaved caspases-3, 8, and 9. Results Stat3 phosphorylation is elevated in 19% (21/113 of osteosarcoma, 27% (17/64 of rhabdomyosarcoma, and 15% (22/151 of other soft-tissue sarcoma tissues as well as in sarcoma cell lines. Expression of the dominant-negative Stat3 and treatment of STA-21 inhibited cell viability and growth and induced apoptosis through caspases 3, 8 and 9 pathways in human sarcoma cell lines expressing elevated levels of phosphorylated Stat3. Conclusion This study demonstrates that Stat3 phosphorylation is elevated in human rhabdomyosarcoma, osteosarcomas and soft-tissue sarcomas. Furthermore, the activated Stat3 pathway is important for cell growth and survival of human sarcoma cells.

  20. Identification of cytoskeleton-associated proteins essential for lysosomal stability and survival of human cancer cells

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    Groth-Pedersen, Line; Aits, Sonja; Corcelle-Termeau, Elisabeth

    2012-01-01

    Microtubule-disturbing drugs inhibit lysosomal trafficking and induce lysosomal membrane permeabilization followed by cathepsin-dependent cell death. To identify specific trafficking-related proteins that control cell survival and lysosomal stability, we screened a molecular motor siRNA library...... in human MCF7 breast cancer cells. SiRNAs targeting four kinesins (KIF11/Eg5, KIF20A, KIF21A, KIF25), myosin 1G (MYO1G), myosin heavy chain 1 (MYH1) and tropomyosin 2 (TPM2) were identified as effective inducers of non-apoptotic cell death. The cell death induced by KIF11, KIF21A, KIF25, MYH1 or TPM2 si......), increased dextran accumulation (KIF20A), or reduced autophagic flux (MYO1G, MYH1). Importantly, all seven siRNAs also killed human cervix cancer (HeLa) and osteosarcoma (U-2-OS) cells and sensitized cancer cells to other lysosome-destabilizing treatments, i.e. photo-oxidation, siramesine, etoposide...

  1. Human hepatocyte growth factor (hHGF-modified hepatic oval cells improve liver transplant survival.

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    Zhu Li

    Full Text Available Despite progress in the field of immunosuppression, acute rejection is still a common postoperative complication following liver transplantation. This study aims to investigate the capacity of the human hepatocyte growth factor (hHGF in modifying hepatic oval cells (HOCs administered simultaneously with orthotopic liver transplantation as a means of improving graft survival. HOCs were activated and isolated using a modified 2-acetylaminofluorene/partial hepatectomy (2-AAF/PH model in male Lewis rats. A HOC line stably expressing the HGF gene was established following stable transfection of the pBLAST2-hHGF plasmid. Our results demonstrated that hHGF-modified HOCs could efficiently differentiate into hepatocytes and bile duct epithelial cells in vitro. Administration of HOCs at the time of liver transplantation induced a wider distribution of SRY-positive donor cells in liver tissues. Administration of hHGF-HOC at the time of transplantation remarkably prolonged the median survival time and improved liver function for recipients compared to these parameters in the other treatment groups (P<0.05. Moreover, hHGF-HOC administration at the time of liver transplantation significantly suppressed elevation of interleukin-2 (IL-2, tumor necrosis factor-α (TNF-α and interferon-γ (IFN-γ levels while increasing the production of IL-10 and TGF-β1 (P<0.05. HOC or hHGF-HOC administration promoted cell proliferation, reduced cell apoptosis, and decreased liver allograft rejection rates. Furthermore, hHGF-modified HOCs more efficiently reduced acute allograft rejection (P<0.05 versus HOC transplantation only. Our results indicate that the combination of hHGF-modified HOCs with liver transplantation decreased host anti-graft immune responses resulting in a reduction of allograft rejection rates and prolonging graft survival in recipient rats. This suggests that HOC-based cell transplantation therapies can be developed as a means of treating severe liver

  2. The pro-adhesive and pro-survival effects of glucocorticoid in human ovarian cancer cells.

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    Yin, Lijuan; Fang, Fang; Song, Xinglei; Wang, Yan; Huang, Gaoxiang; Su, Jie; Hui, Ning; Lu, Jian

    2016-07-01

    Cell adhesion to extracellular matrix (ECM) is controlled by multiple signaling molecules and intracellular pathways, and is pivotal for survival and growth of cells from most solid tumors. Our previous works demonstrated that dexamethasone (DEX) significantly enhances cell adhesion and cell resistance to chemotherapeutics by increasing the levels of integrin β1, α4, and α5 in human ovarian cancer cells. However, it is unclear whether the components of ECM or other membrane molecules are also involved in the pro-adhesive effect of DEX in ovarian cancer cells. In this study, we demonstrated that the treatment of cells with DEX did not change the expression of collagens (I, III, and IV), laminin, CD44, and its principal ligand hyaluronan (HA), but significantly increased the levels of intracellular and secreted fibronectin (FN). Inhibiting the expression of FN with FN1 siRNA or blocking CD44, another FN receptor, with CD44 blocking antibody significantly attenuated the pro-adhesion of DEX, indicating that upregulation of FN mediates the pro-adhesive effect of DEX by its interaction with CD44 besides integrin β1. Moreover, DEX significantly enhanced cell resistance to the chemotherapeutic agent paclitaxel (PTX) by activating PI-3K-Akt pathway. Finally, we found that DEX also significantly upregulated the expression of MUC1, a transmembrane glycoprotein. Inhibiting the expression of MUC1 with MUC1 siRNA significantly attenuated the DEX-induced effects of pro-adhesion, Akt-activation, and pro-survival. In conclusion, these results provide new data that upregulation of FN and MUC1 by DEX contributes to DEX-induced pro-adhesion and protects ovarian cancer cells from chemotherapy.

  3. Human and Autologous Adipose-derived Stromal Cells Increase Flap Survival in Rats Independently of Host Immune Response.

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    Toyserkani, Navid Mohamadpour; Jensen, Charlotte Harken; Andersen, Ditte Caroline; Sheikh, Søren Paludan; Sørensen, Jens Ahm

    2017-07-22

    There is a rising interest in adipose-derived stromal cells for clinical use; however, it is unknown whether freshly isolated stromal cells (SVF) or culture-expanded cells (ASCs) are more efficacious. We therefore aimed to compare the 2 cellular therapies in an in vivo model of angiogenesis, the ischemic flap in rats, which induces acute ischemia. We also aimed to determine the importance of cell presence and the host immune response. A total of 96 rats (n = 12 in each group) were used, and in each rat, a caudally based random flap measuring 2 × 7 cm was made. The study was conducted in 3 phases. First, each rat was treated with human SVF cells, human ASCs, or vehicle. Second, each rat was treated with human SVF, human SVF lysate, or vehicle. Finally, each rat was treated with rat (autologous) SVF cells or vehicle. Flap survival, vessel density, and stromal cell retention were evaluated after 7 days. The mean survival rates for SVF treatment regardless of human or autologous origin were significantly increased as compared with the control group. Adipose stem/stromal cell and SVF lysate injection did not increase flap survival. Vessel density was increased for human and rat SVF and human ASC but not for SVF lysate. Human cells were not detected in the flaps after 7 days. Flap survival increased with SVF treatment regardless of human or autologous origin, suggesting that increased flap survival is independent of the host immune response. All cell injections lead to increased vessel density, but it did not necessarily lead to increased flap survival. Further research should elaborate which molecular events make SVF treatment more efficacious than ASC.

  4. Necdin modulates proliferative cell survival of human cells in response to radiation-induced genotoxic stress

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    Lafontaine Julie

    2012-06-01

    Full Text Available Abstract Background The finite replicative lifespan of cells, termed cellular senescence, has been proposed as a protective mechanism against the proliferation of oncogenically damaged cells, that fuel cancer. This concept is further supported by the induction of premature senescence, a process which is activated when an oncogene is expressed in normal primary cells as well as following intense genotoxic stresses. Thus, deregulation of genes that control this process, like the tumor suppressor p53, may contribute to promoting cancer by allowing cells to bypass senescence. A better understanding of the genes that contribute to the establishment of senescence is therefore warranted. Necdin interacts with p53 and is also a p53 target gene, although the importance of Necdin in the p53 response is not clearly understood. Methods In this study, we first investigated Necdin protein expression during replicative senescence and premature senescence induced by gamma irradiation and by the overexpression of oncogenic RasV12. Gain and loss of function experiments were used to evaluate the contribution of Necdin during the senescence process. Results Necdin expression declined during replicative aging of IMR90 primary human fibroblasts or following induction of premature senescence. Decrease in Necdin expression seemed to be a consequence of the establishment of senescence since the depletion of Necdin in human cells did not induce a senescence-like growth arrest nor a flat morphology or SA-β-galactosidase activity normally associated with senescence. Similarly, overexpression of Necdin did not affect the life span of IMR90 cells. However, we demonstrate that in normal human cells, Necdin expression mimicked the effect of p53 inactivation by increasing radioresistance. Conclusion This result suggests that Necdin potentially attenuate p53 signaling in response to genotoxic stress in human cells and supports similar results describing an inhibitory function

  5. IMMUNE MODULATORY EFFECTS of HUMAN CHORIONIC GONADOTROPIN on DENDRITIC CELLS SUPPORTING FETAL SURVIVAL in MURINE PREGNANCY

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    Dominique Dauven

    2016-11-01

    Full Text Available Dendritic cells (DCs are critically involved in the determination of immunity versus tolerance. Hence, DCs are key regulators of immune responses either favoring or disfavoring fetal survival. Several factors were proposed to modulate DC phenotype and function during preg-nancy. Here, we studied whether the pregnancy hormone human Chorionic Gonadotropin (hCG is involved in DC regulation.In vitro, bone-marrow-derived DCs (BMDCs were stimulated in the presence or absence of urine-purified (uhCG or recombinant hCG (rhCG preparations. Subsequently, BMDC matu-ration was assessed. Cytokine secretion of activated BMDCs and their capability to enforce TH1, TH2, TH17 or Treg cell differentiation was determined after rhCG treatment. Moreover, the in vivo potential of hCG-modulated BMDCs to influence pregnancy outcome, Treg cell number and local cytokine expression was evaluated after adoptive transfer in a murine abor-tion-prone model before and after conception. Both hCG preparations impaired the maturation process of BMDCs. rhCG treatment did nei-ther alter cytokine secretion by BMDCs nor their ability to drive TH1, TH2 or TH17 differen-tiation. rhCG-treated BMDCs augmented the number of Treg cells within the T cell popula-tion. Adoptive transfer of rhCG-treated BMDCs after conception did not influence pregnancy outcome. However, transfer of hCG-treated BMDCs prior to mating had a protective effect on pregnancy. This positive effect was accompanied by increased Treg cell numbers and decidual IL-10 and TGF-β expression. Our results unveil the importance of hCG in retaining DCs in a tolerogenic state, thereby promoting Treg cell increment and supporting fetal survival.

  6. Immune Modulatory Effects of Human Chorionic Gonadotropin on Dendritic Cells Supporting Fetal Survival in Murine Pregnancy

    Science.gov (United States)

    Dauven, Dominique; Ehrentraut, Stefanie; Langwisch, Stefanie; Zenclussen, Ana Claudia; Schumacher, Anne

    2016-01-01

    Dendritic cells (DCs) are critically involved in the determination of immunity vs. tolerance. Hence, DCs are key regulators of immune responses either favoring or disfavoring fetal survival. Several factors were proposed to modulate DC phenotype and function during pregnancy. Here, we studied whether the pregnancy hormone human chorionic gonadotropin (hCG) is involved in DC regulation. In vitro, bone marrow-derived DCs (BMDCs) were stimulated in the presence or absence of urine-purified or recombinant hCG (rhCG) preparations. Subsequently, BMDC maturation was assessed. Cytokine secretion of activated BMDCs and their capability to enforce TH1, TH2, TH17, or Treg cell differentiation was determined after rhCG treatment. Moreover, the in vivo potential of hCG-modulated BMDCs to influence pregnancy outcome, Treg cell number, and local cytokine expression was evaluated after adoptive transfer in a murine abortion-prone model before and after conception. Both hCG preparations impaired the maturation process of BMDCs. rhCG treatment did neither alter cytokine secretion by BMDCs nor their ability to drive TH1, TH2, or TH17 differentiation. rhCG-treated BMDCs augmented the number of Treg cells within the T cell population. Adoptive transfer of rhCG-treated BMDCs after conception did not influence pregnancy outcome. However, transfer of hCG-treated BMDCs prior to mating had a protective effect on pregnancy. This positive effect was accompanied by increased Treg cell numbers and decidual IL-10 and TGF-β expression. Our results unveil the importance of hCG in retaining DCs in a tolerogenic state, thereby promoting Treg cell increment and supporting fetal survival. PMID:27895621

  7. Thermal survival characteristics of cell subpopulations isolated from a heterogeneous human colon tumor.

    Science.gov (United States)

    Leith, J T; Heyman, P; DeWyngaert, J K; Dexter, D L; Calabresi, P; Glicksman, A S

    1983-07-01

    Responses of a heterogeneous human colon adenocarcinoma model tumor system to in vitro hyperthermic treatment at various temperatures have been studied. This model tumor system consists of an original tumor line (DLD-1) obtained from surgical biopsy, and two derivative subpopulations termed clones A and D. These 3 tumor cell populations differ in many properties, including karyotype and DNA content, production of specific antigens, and sensitivities to other cytotoxic agents such as chemotherapeutic drugs and X-irradiation. In these experiments, exponentially growing tumor cells were exposed to hyperthermia (42.2, 42.5, 43.0, 44.0, or 45.0 degrees) for graded time periods. A single-hit, multitarget equation was used to express the dependence of survival on time at a given temperature, and values for extrapolation numbers, quasi-threshold time (min), and T0 (mean lethal time; min) were obtained for the initial regions of survival. At the lower temperatures of 42.2 and 42.5 degrees, biphasic survival curves were obtained for all three tumor lines and, as a consequence, a second mean lethal time (T0,f) was also determined for the final thermal-resistant portion of the survival curves. Using the T0 values as an index of relative resistance, values at 42.2 and 42.5 degrees indicated that, in this temperature region, the parent (DLD-1) line was the most sensitive, the clone A line showed intermediate sensitivity, and the clone D line was the most resistant. In the thermally resistant portion of the survival curve, T0 values indicated that the clone A subpopulation was the most sensitive, the DLD-1 line showed intermediate sensitivity, and the clone D tumor subpopulation remained the most resistant. At the higher temperatures of 43, 44, and 45 degrees, in which thermotolerance is not observed during heat treatment, values for T0 indicated the parent (DLD-1) tumor line was still the most sensitive tumor line, and the clone A and clone D lines showed approximately equal

  8. Identification of cytoskeleton-associated proteins essential for lysosomal stability and survival of human cancer cells.

    Science.gov (United States)

    Groth-Pedersen, Line; Aits, Sonja; Corcelle-Termeau, Elisabeth; Petersen, Nikolaj H T; Nylandsted, Jesper; Jäättelä, Marja

    2012-01-01

    Microtubule-disturbing drugs inhibit lysosomal trafficking and induce lysosomal membrane permeabilization followed by cathepsin-dependent cell death. To identify specific trafficking-related proteins that control cell survival and lysosomal stability, we screened a molecular motor siRNA library in human MCF7 breast cancer cells. SiRNAs targeting four kinesins (KIF11/Eg5, KIF20A, KIF21A, KIF25), myosin 1G (MYO1G), myosin heavy chain 1 (MYH1) and tropomyosin 2 (TPM2) were identified as effective inducers of non-apoptotic cell death. The cell death induced by KIF11, KIF21A, KIF25, MYH1 or TPM2 siRNAs was preceded by lysosomal membrane permeabilization, and all identified siRNAs induced several changes in the endo-lysosomal compartment, i.e. increased lysosomal volume (KIF11, KIF20A, KIF25, MYO1G, MYH1), increased cysteine cathepsin activity (KIF20A, KIF25), altered lysosomal localization (KIF25, MYH1, TPM2), increased dextran accumulation (KIF20A), or reduced autophagic flux (MYO1G, MYH1). Importantly, all seven siRNAs also killed human cervix cancer (HeLa) and osteosarcoma (U-2-OS) cells and sensitized cancer cells to other lysosome-destabilizing treatments, i.e. photo-oxidation, siramesine, etoposide or cisplatin. Similarly to KIF11 siRNA, the KIF11 inhibitor monastrol induced lysosomal membrane permeabilization and sensitized several cancer cell lines to siramesine. While KIF11 inhibitors are under clinical development as mitotic blockers, our data reveal a new function for KIF11 in controlling lysosomal stability and introduce six other molecular motors as putative cancer drug targets.

  9. Human primary brain tumor cell growth inhibition in serum-free medium optimized for neuron survival.

    Science.gov (United States)

    Brewer, Gregory J; LeRoux, Peter D

    2007-07-09

    Glioblastoma is the most common primary brain tumor in adults from which about 15,000 patients die each year in the United States. Despite aggressive surgery, radiotherapy and chemotherapy, median survival remains only 1 year. Here we evaluate growth of primary human brain tumor cells in a defined nutrient culture medium (Neuregen) that was optimized for neuron regeneration. We hypothesized that Neuregen would inhibit tumor cell growth because of its ability to inhibit gliosis in rat brain. Tumor tissue was collected from 18 patients including 10 males and 8 females (mean age 60+/-12 years) who underwent craniotomy for newly diagnosed, histologically confirmed brain tumors. The tissue was shipped overnight in Hibernate transport medium. Tumor cells were isolated and plated in Neurobasal/serum or Neuregen on culture plastic. After 1 week, growth in Neuregen was significantly less in 9/10 glioblastoma multiforme cases, 5/5 meningioma cases and 3/3 cases of brain metastasis. Analysis of deficient formulations of Neuregen and formulations to which selected components were added back implicate no single active component. However, individual cases were sensitive to corticosterone, selenium, ethanolamine, fatty acids and/or antioxidants. Therefore, a defined culture medium that promotes neuron regeneration inhibits the growth of human primary glioblastoma, meningioma and metastatic tumor cells in culture. The possible in vivo efficacy of Neuregen for treatment of brain tumor resections remains to be determined.

  10. Anti-apoptotic BFL-1 is the major effector in activation-induced human mast cell survival.

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    Maria Ekoff

    Full Text Available Mast cells are best known for their role in allergic reactions, where aggregation of FcεRI leads to the release of mast cell mediators causing allergic symptoms. The activation also induces a survival program in the cells, i.e., activation-induced mast cell survival. The aim of the present study was to investigate how the activation-induced survival is mediated. Cord blood-derived mast cells and the mast cell line LAD-2 were activated through FcεRI crosslinking, with or without addition of chemicals that inhibit the activity or expression of selected Bcl-2 family members (ABT-737; roscovitine. Cell viability was assessed using staining and flow cytometry. The expression and function of Bcl-2 family members BFL-1 and MCL-1 were investigated using real-time quantitative PCR and siRNA treatment. The mast cell expression of Bfl-1 was investigated in skin biopsies. FcεRI crosslinking promotes activation-induced survival of human mast cells and this is associated with an upregulation of the anti-apoptotic Bcl-2 family member Bfl-1. ABT-737 alone or in combination with roscovitine decreases viability of human mast cells although activation-induced survival is sustained, indicating a minor role for Bcl-X(L, Bcl-2, Bcl-w and Mcl-1. Reducing BFL-1 but not MCL-1 levels by siRNA inhibited activation-induced mast cell survival. We also demonstrate that mast cell expression of Bfl-1 is elevated in birch-pollen-provocated skin and in lesions of atopic dermatitis and psoriasis patients. Taken together, our results highlight Bfl-1 as a major effector in activation-induced human mast cell survival.

  11. Survival of human periodontal ligament cells in media proposed for transport of avulsed teeth.

    Science.gov (United States)

    Sigalas, Emmanouil; Regan, John D; Kramer, Phillip R; Witherspoon, David E; Opperman, Lynne A

    2004-02-01

    Many solutions have been examined as possible storage media for avulsed teeth. In this report, human periodontal ligament (PDL) cells were exposed for 1 h to culture medium, milk, Hanks Balanced Salt Solution (HBSS), Soft Wear, Opti Free, and Solo Care contact lens solutions, Gatorade, and tap water, at room temperature and on ice. The number of viable cells was counted using the trypan blue exclusion technique, immediately after exposure (0 h) and at 24 and 48 h, to test the proliferative capacity of the cells after treatment. The results indicated that a significantly higher number of cells survived and proliferated when the exposures were performed at 0 degrees C. Water had a detrimental effect on the cells, whereas culture medium and HBSS preserved significantly more viable cells than the other experimental solutions. Within the parameters of this study, it appears that HBSS is the optimal storage medium for avulsed teeth. Low-fat milk could serve as an alternative if ice is available. Contact lens solutions or Gatorade on ice could serve as short-term (1 h) storage media if the other solutions are not readily available.

  12. Extracellular sphingosine-1-phosphate: a novel actor in human glioblastoma stem cell survival.

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    Elena Riccitelli

    Full Text Available Glioblastomas are the most frequent and aggressive intracranial neoplasms in humans, and despite advances and the introduction of the alkylating agent temozolomide in therapy have improved patient survival, resistance mechanisms limit benefits. Recent studies support that glioblastoma stem-like cells (GSCs, a cell subpopulation within the tumour, are involved in the aberrant expansion and therapy resistance properties of glioblastomas, through still unclear mechanisms. Emerging evidence suggests that sphingosine-1-phosphate (S1P a potent onco-promoter able to act as extracellular signal, favours malignant and chemoresistance properties in GSCs. Notwithstanding, the origin of S1P in the GSC environment remains unknown. We investigated S1P metabolism, release, and role in cell survival properties of GSCs isolated from either U87-MG cell line or a primary culture of human glioblastoma. We show that both GSC models, grown as neurospheres and expressing GSC markers, are resistant to temozolomide, despite not expressing the DNA repair protein MGMT, a major contributor to temozolomide-resistance. Pulse experiments with labelled sphingosine revealed that both GSC types are able to rapidly phosphorylate the long-chain base, and that the newly produced S1P is efficiently degraded. Of relevance, we found that S1P was present in GSC extracellular medium, its level being significantly higher than in U87-MG cells, and that the extracellular/intracellular ratio of S1P was about ten-fold higher in GSCs. The activity of sphingosine kinases was undetectable in GSC media, suggesting that mechanisms of S1P transport to the extracellular environment are constitutive in GSCs. In addition we found that an inhibitor of S1P biosynthesis made GSCs sensitive to temozolomide (TMZ, and that exogenous S1P reverted this effect, thus involving extracellular S1P as a GSC survival signal in TMZ resistance. Altogether our data implicate for the first time GSCs as a pivotal source

  13. Hydrogen sulfide mediates the anti-survival effect of sulforaphane on human prostate cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Pei, Yanxi [Department of Biology, Lakehead University, Thunder Bay (Canada); College of Life Science, Shanxi University, Taiyuan (China); Wu, Bo [Department of Biology, Lakehead University, Thunder Bay (Canada); Department of Pathophysiology, Harbin Medical University, Harbin (China); Cao, Qiuhui [Department of Biology, Lakehead University, Thunder Bay (Canada); Wu, Lingyun [Department of Pathophysiology, Harbin Medical University, Harbin (China); Department of Pharmacology, University of Saskatchewan, Saskatoon (Canada); Yang, Guangdong, E-mail: gyang@lakeheadu.ca [The School of Kinesiology, Lakehead University, Thunder Bay (Canada)

    2011-12-15

    Hydrogen sulfide (H{sub 2}S) is a novel gasotransmitter that regulates cell proliferation and other cellular functions. Sulforaphane (SFN) is a sulfur-containing compound that exhibits anticancer properties, and young sprouts of broccoli are particularly rich in SFN. There is consistent epidemiological evidence that the consumption of sulfur-containing vegetables, such as garlic and cruciferous vegetables, may help reduce the occurrence of prostate cancer. Here we found that a large amount of H{sub 2}S is released when SFN is added into cell culture medium or mixed with mouse liver homogenates, respectively. Both SFN and NaHS (a H{sub 2}S donor) decreased the viability of PC-3 cells (a human prostate cancer cell line) in a dose-dependent manner, and supplement of methemoglobin or oxidized glutathione (two H{sub 2}S scavengers) reversed SFN-reduced cell viability. We further found both cystathionine gamma-lyase (CSE) and cystathionine beta-synthase are expressed in PC-3 cells and mouse prostate tissues. H{sub 2}S production in prostate tissues from CSE knockout mice was only 20% of that from wild-type mice, suggesting CSE is a major H{sub 2}S-producing enzyme in prostate. CSE overexpression enhanced H{sub 2}S production and inhibited cell viability in PC-3 cells. In addition, both SFN and NaHS activated p38 mitogen-activated protein kinases (MAPK) and c-Jun N-terminal kinase (JNK). Pre-treatment of PC-3 cells with methemoglobin decreased SFN-stimulated MAPK activities. Suppression of both p38 MAPK and JNK reversed H{sub 2}S- or SFN-reduced viability of PC-3 cells. Our results demonstrated that H{sub 2}S mediates the inhibitory effect of SFN on the proliferation of PC-3 cells, which suggests that H{sub 2}S-releasing diet or drug might be beneficial in the treatment of prostate cancer. Highlights: Black-Right-Pointing-Pointer A large amount of H{sub 2}S is released from sulforaphane. Black-Right-Pointing-Pointer H{sub 2}S mediates the anti-survival effect of

  14. Survival, proliferation, and migration of human meningioma stem-like cells in a nanopeptide scaffold

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    Sajad Sahab Negah

    2016-12-01

    Full Text Available Objective(s: In order to grow cells in a three-dimensional (3D microenvironment, self-assembling peptides, such as PuraMatrix, have emerged with potential to mimic the extracellular matrix. The aim of the present study was to investigate the influence of the self-assembling peptide on the morphology, survival, proliferation rate, migration potential, and differentiation of human meningioma stem-like cells (hMgSCs. Materials and Methods: The efficacy of a novel method for placing hMgSCs in PuraMatrix (the injection approach was compared to the encapsulation and surface plating methods. In addition, we designed a new method for measurement of migration distance in 3D cultivation of hMgSCs in PuraMatrix. Results: Our results revealed that hMgSCs have the ability to form spheres in stem cell culture condition. These meningioma cells expressed GFAP, CD133, vimentin, and nestin. Using the injection method, a higher proliferation rate of the hMgSCs was observed after seven days of culture. Furthermore, the novel migration assay was able to measure the migration of a single cell alone in 3D environment. Conclusion: The results indicate the injection method as an efficient technique for culturing hMgSCs in PuraMatrix. Furthermore, the novel migration assay enables us to evaluate the migration of hMgSCs.

  15. Influence of surface-modified maghemite nanoparticles on in vitro survival of human stem cells

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    Michal Babič

    2014-10-01

    Full Text Available Surface-modified maghemite (γ-Fe2O3 nanoparticles were obtained by using a conventional precipitation method and coated with D-mannose and poly(N,N-dimethylacrylamide. Both the initial and the modified particles were characterized by transmission electron microscopy and dynamic light scattering with regard to morphology, particle size and polydispersity. In vitro survival of human stem cells was then investigated by using the methyl thiazolyl tetrazolium (MTT assay, which showed that D-mannose- and poly(N,N-dimethylacrylamide-coated γ-Fe2O3 particles exhibit much lower level of cytotoxicity than the non-coated γ-Fe2O3.

  16. Sertraline increases the survival of retinoic acid induced neuronal cells but not glial cells from human mesenchymal stem cells.

    Science.gov (United States)

    Verdi, Javad; Sharif, Shiva; Banafshe, Hamid Reza; Shoae-Hassani, Alireza

    2014-08-01

    An increase in the number of viable in vitro differentiated neuronal cells is important for their use in clinics. A proportion of differentiated cells lose their viability before being used, and therefore we decided to use a pharmacological agent, sertraline, to increase neural cell differentiation and their survival. Purified endometrial stem cells (EnSCs) were examined for neuronal and glial cell specific markers after retinoic acid (RA) and sertraline treatment via RT-PCR, immunocytochemistry and Western blot analysis. The survival of differentiated cells was measured by MTT assay and the frequency of apoptosis, demonstrated by caspase-3-like activity. EnSCs were differentiated into neuronal cells after RA induction. Sertraline increased neuronal cell differentiation by 1.2-fold and their survival by 1.4-fold, and decreased from glial cell differentiation significantly. The findings indicate that sertraline could be used to improve the in vitro differentiation process of stem cells into neuronal cells, and may be involved in regenerative pharmacology in future. © 2014 International Federation for Cell Biology.

  17. Hydrostatic pressure does not cause detectable changes in survival of human retinal ganglion cells.

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    Andrew Osborne

    Full Text Available PURPOSE: Elevated intraocular pressure (IOP is a major risk factor for glaucoma. One consequence of raised IOP is that ocular tissues are subjected to increased hydrostatic pressure (HP. The effect of raised HP on stress pathway signaling and retinal ganglion cell (RGC survival in the human retina was investigated. METHODS: A chamber was designed to expose cells to increased HP (constant and fluctuating. Accurate pressure control (10-100 mmHg was achieved using mass flow controllers. Human organotypic retinal cultures (HORCs from donor eyes (<24 h post mortem were cultured in serum-free DMEM/HamF12. Increased HP was compared to simulated ischemia (oxygen glucose deprivation, OGD. Cell death and apoptosis were measured by LDH and TUNEL assays, RGC marker expression by qRT-PCR (THY-1 and RGC number by immunohistochemistry (NeuN. Activated p38 and JNK were detected by Western blot. RESULTS: Exposure of HORCs to constant (60 mmHg or fluctuating (10-100 mmHg; 1 cycle/min pressure for 24 or 48 h caused no loss of structural integrity, LDH release, decrease in RGC marker expression (THY-1 or loss of RGCs compared with controls. In addition, there was no increase in TUNEL-positive NeuN-labelled cells at either time-point indicating no increase in apoptosis of RGCs. OGD increased apoptosis, reduced RGC marker expression and RGC number and caused elevated LDH release at 24 h. p38 and JNK phosphorylation remained unchanged in HORCs exposed to fluctuating pressure (10-100 mmHg; 1 cycle/min for 15, 30, 60 and 90 min durations, whereas OGD (3 h increased activation of p38 and JNK, remaining elevated for 90 min post-OGD. CONCLUSIONS: Directly applied HP had no detectable impact on RGC survival and stress-signalling in HORCs. Simulated ischemia, however, activated stress pathways and caused RGC death. These results show that direct HP does not cause degeneration of RGCs in the ex vivo human retina.

  18. Differential depletion of total T cells and regulatory T cells and prolonged allotransplant survival in CD3Ɛ humanized mice treated with polyclonal anti human thymocyte globulin

    Science.gov (United States)

    Buszko, Maja; Cardini, Benno; Oberhuber, Rupert; Oberhuber, Lukas; Jakic, Bojana; Beierfuss, Anja; Wick, Georg; Cappellano, Giuseppe

    2017-01-01

    Thymoglobulin (ATG) is a polyclonal rabbit antibody against human thymocytes used as a T cell-depleting agent to prevent or treat allotransplant rejection. The aim of the present study was to investigate the effect of low dose ATG treatment exclusively on T cells using a humanized BALB/c human CD3Ɛ transgenic mouse model expressing both human and murine T cell receptors (TCR). Mice received a single intravenous (i.v.) injection of ATG. Blood and peripheral lymphoid organs were obtained after different time points. We found a significant T cell depletion in this mouse model. In addition, regulatory T cells (Tregs) proved to be less sensitive to depletion than the rest of T cells and the Treg:non-Treg ratio was therefore increased. Finally, we also investigated the effect of ATG in a heterotopic allogenic murine model of heart transplantation. Survival and transplant function were significantly prolonged in ATG-treated mice. In conclusion, we showed (a) an immunosuppressive effect of ATG in this humanized mouse model which is exclusively mediated by reactivity against human CD3Ɛ; (b) provided evidence for a relative resistance of Tregs against this regimen; and (c) demonstrated the immunomodulatory effect of ATG under these experimental circumstances by prolongation of heart allograft survival. PMID:28257450

  19. Mitochondrial dysfunction in primary human fibroblasts triggers an adaptive cell survival program that requires AMPK-alpha

    NARCIS (Netherlands)

    Distelmaier, F.; Valsecchi, F.; Liemburg-Apers, D.; Lebiedzinska, M.; Rodenburg, R.; Heil, S.; Keijer, J.; Fransen, J.; Imamura, H.; Danhauser, K.; Seibt, A.; Viollet, B.; Gellerich, F.; Smeitink, J.; Wieckowski, M.; Willems, P.; Koopman, W.J.H.

    2015-01-01

    Dysfunction of complex I (CI) of the mitochondrial electron transport chain (ETC) features prominently in human pathology. Cell models of ETC dysfunction display adaptive survival responses that still are poorly understood but of relevance for therapy development. Here we comprehensively examined ho

  20. Mitochondrial dysfunction in primary human fibroblasts triggers an adaptive cell survival program that requires AMPK-alpha

    NARCIS (Netherlands)

    Distelmaier, F.; Valsecchi, F.; Liemburg-Apers, D.; Lebiedzinska, M.; Rodenburg, R.; Heil, S.; Keijer, J.; Fransen, J.; Imamura, H.; Danhauser, K.; Seibt, A.; Viollet, B.; Gellerich, F.; Smeitink, J.; Wieckowski, M.; Willems, P.; Koopman, W.J.H.

    2015-01-01

    Dysfunction of complex I (CI) of the mitochondrial electron transport chain (ETC) features prominently in human pathology. Cell models of ETC dysfunction display adaptive survival responses that still are poorly understood but of relevance for therapy development. Here we comprehensively examined

  1. Mitochondrial dysfunction in primary human fibroblasts triggers an adaptive cell survival program that requires AMPK-alpha

    NARCIS (Netherlands)

    Distelmaier, F.; Valsecchi, F.; Liemburg-Apers, D.C.; Lebiedzinska, M.; Rodenburg, R.J.T.; Heil, S.; Keijer, J.; Fransen, J.A.; Imamura, H.; Danhauser, K.; Seibt, A.; Viollet, B.; Gellerich, F.N.; Smeitink, J.; Wieckowski, M.R.; Willems, P.H.G.M.; Koopman, W.J.H.

    2015-01-01

    Dysfunction of complex I (CI) of the mitochondrial electron transport chain (ETC) features prominently in human pathology. Cell models of ETC dysfunction display adaptive survival responses that still are poorly understood but of relevance for therapy development. Here we comprehensively examined ho

  2. Mitochondrial dysfunction in primary human fibroblasts triggers an adaptive cell survival program that requires AMPK-α

    NARCIS (Netherlands)

    F. Distelmaier (Felix); F. Valsecchi (Federica); D.C. Liemburg-Apers (Dania C.); M. Lebiedzinska (Magdalena); R.J.T. Rodenburg (Richard); S.G. Heil (Sandra); J. Keijer (Jaap); J.A.M. Fransen (Jack); H. Imamura (Hiromi); K. Danhauser (Katharina); A. Seibt (Annette); B. Viollet (Benoit); F.N. Gellerich (Frank); J.A.M. Smeitink (Jan); M.R. Wieckowski (Mariusz R.); P.H.G.M. Willems (Peter H.G.M.); W.J.H. Koopman (W. J H)

    2015-01-01

    textabstractDysfunction of complex I (CI) of the mitochondrial electron transport chain (ETC) features prominently in human pathology. Cell models of ETC dysfunction display adaptive survival responses that still are poorly understood but of relevance for therapy development. Here we comprehensively

  3. Active Stat3 is required for survival of human squamous cell carcinoma cells in serum-free conditions

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    DiGiovanni John

    2006-04-01

    Full Text Available Abstract Background Squamous cell carcinoma (SCC of the skin is the most aggressive form of non-melanoma skin cancer (NMSC, and is the single most commonly diagnosed cancer in the U.S., with over one million new cases reported each year. Recent studies have revealed an oncogenic role of activated signal transducer and activator of transcription 3 (Stat3 in many human tumors, especially in those of epithelial origin, including skin SCC. Stat3 is a mediator of numerous growth factor and cytokine signaling pathways, all of which activate it through phosphorylation of tyrosine 705. Results To further address the role of Stat3 in skin SCC tumorigenesis, we have analyzed a panel of human skin-derived cell lines ranging from normal human epidermal keratinocytes (NHEK, to non-tumorigenic transformed skin cells (HaCaT, to highly tumorigenic cells (SRB1-m7 and SRB12-p9 and observed a positive correlation between Stat3 phosphorylation and SCC malignancy. We next determined the role of Stat3 activity in cell proliferation and viability under serum-free culture conditions. This was accomplished by suppressing Stat3 activity in the SRB12-p9 cells through stable expression of a dominant negative acting form of Stat3β, which contains a tyrosine 705 to phenylalanine mutation (S3DN. The S3DN cells behaved similar to parental SRB12-p9 cells when cultured in optimal growth conditions, in the presence of 10% fetal calf serum. However, unlike the SRB12-p9 cells, S3DN cells underwent apoptotic cell death when cultured in serum-free medium (SFM. This was evidenced by multiple criteria, including accumulation of sub-G1 particles, induced PARP cleavage, and acquisition of the characteristic morphological changes associated with apoptosis. Conclusion This study provides direct evidence for a role for Stat3 in maintaining cell survival in the conditions of exogenous growth factor deprivation produced by culture in SFM. We also propose that delivery of the S3DN gene or

  4. Substitued (E-b-(benzoylacrylic acids suppressed survival of neoplastic human HeLa cells

    Directory of Open Access Journals (Sweden)

    I. JURANIC

    1999-09-01

    Full Text Available The bacteriostatic activity of some of alkyl substituted (E-b-(benzoylacrylic acids was shown earlier. The aim of this study was to investigate the antiproliferative action of 19 alkyl-, or halogeno-, or methoxy-, or acetamido- substituted (E-b-(benzoylacrylic acids, against human cervix carcinoma, HeLa, cells. Target HeLa cells were continuously treated with increasing concentrations of substituted (E-b-(benzoylacrylic acids during two days. The MTT test was used for assessment of the antiproliferative action of this group of compounds. Treatment of HeLa cells with 4-methyl-, 4-fluoro-, 4-chloro-, 4-bromo- and 4-methoxy- derivatives of (E-b-(benzoyl acrylic acid leads to the expression of cytostatic activity against HeLa cells (IC50 were in the range from 31-40 µM. Their antiproliferative action was less than that of the basic compound (E-b-(benzoylacrylic acid whose IC50 was 28.5 µM. The 3,4-dimethyl-, 2,4-dimethyl- and 2,5-dimethyl- derivatives as well as the 4-ethyl- and 3,4-dichloro- and 2,4-dichloro-derivatives, have stronger cytostatic activity than the correspoding monosubstituted and parent compound. Their IC50 were 18.5 µM; 17.5 µM; 17.0 mM; 17.5 µM; 22.0 µM and 18 µM, respectively. The 4-iso-propyl- and 4-n-butyl-derivatives exerted higher cytostatic activity than the compounds with a lower number of methylene -CH2- groups in the substitutent. Their IC50 were 14.5 µM and 6.5 µM respectively. The 2,5-di-iso-propyl- and 4-tert-butyl-derivatives expressed the most strong antiproliferative action against the investigated HeLa cells, IC50 being 4.5 µM and 5.5 µM, respectively. The investigated compounds affected the survival of HeLa cells, expressing a strong structure-activity relationship of the Hansch type.

  5. Mast cell reactivity at the margin of human skin wounds: an early cell marker of wound survival?

    Science.gov (United States)

    Oehmichen, M; Gronki, T; Meissner, C; Anlauf, M; Schwark, T

    2009-10-30

    Detecting the vitality of mechanical skin wounds (antemortem versus postmortem injury) in human cadavers remains a specifically forensic problem. To determine whether skin mast cells (MCs) are activated during the very early phase of human wound healing we performed a histomorphometric evaluation of the extent of MC enzyme loss as an indication of MC degranulation at the wound margins of skin wounds in 64 human cadavers. We compared the number of tryptase-reactive MCs, which are said not to loose all of their enzyme activity during degranulation process, with the number of naphthol AS-D chloroacetate esterase (NAS-DClAE)-positive MCs, which loose their complete enzyme activity in the form of enzyme-positive granula after activation. The enzyme activity was evaluated on sequential histological sections after autopsy as an indirect quantification of the number of degranulated MCs. Most of the victims had died within 10-60 min after injury (n=50), 12 survived between 60 min and 24h, and only 2 victims survived more than 24h (12 days each). The number of enzyme-positive MCs were counted in six successive visual fields (0.785 mm(2)) on the one hand located parallel to and--on the other hand--at increasing distances outward from the wound margins. In victims surviving the injury less than 60 min the average number of NAS-DClAE-reactive MCs next to the wound margin was significantly lower than the number of tryptase-reactive MCs. The extent of the reduction in NAS-DClAE-reactive MC counts correlated inversely with the distance from the wound edges. Our findings show that MCs undergo very early loss of NAS-DClAE activity at wound margins, and thus appear to be an early cell marker of wound survival. However, definitive evidence that the enzyme loss (degranulation) represents a vital process can only be obtained by comparing MC enzyme loss induced by injury during intact circulation with the MC reaction to injury inflicted very shortly after cardiac arrest, a question that

  6. Cultured human embryonic neocortical cells survive and grow in infarcted cavities of adult rat brains and interconnect with host brain

    Institute of Scientific and Technical Information of China (English)

    ZENG Jin-sheng; YU Jian; CUI Chun-mei; ZHAO Zhan; HONG Hua; SHENG Wen-li; TAO Yu-qian; LI Ling; HUANG Ru-xun

    2005-01-01

    Background There are no reports on exnografting cultured human fetal neocortical cells in this infracted cavities of adult rat brains. This study was undertaken to observe whether cultured human cortical neurons and astrocytes can survive and grow in the infarcted cavities of adult rat brains and whether they interconnect with host brains.Methods The right middle cerebral artery was ligated distal to the striatal branches in 16 adult stroke-prone renovascular hypertensive rats. One week later, cultured cells from human embryonic cerebral cortexes were stereotaxically transferred to the infarcted cavity of 11 rats. The other 5 rats receiving sham transplants served as controls. For immunosuppression, all transplanted rats received intraperitoneal injection of cyclosporine A daily starting on the day of grafting. Immunohistochemistry for glial fibrillary acidic protein (GFAP), synaptophysin, neurofilament, and microtubule associated protein-2 (MAP-2) was performed on brain sections perfused in situ 8 weeks after transplantation.Results Grafts in the infarcted cavities of 6 of 10 surviving rats consisted of bands of neurons with an immature appearance, bundles of fibers, and GFAP-immunopositive astrocytes, which were unevenly distributed. The grafts were rich in synaptophysin, neurofilament, and MAP2-positive neurons with long processes. The graft/host border was diffuse with dendrites apparently bridging over to the host brain, into which neurofilament immunopositive fibers protruded. Conclusion Cultured human fetal brain cells can survive and grow in the infarcted cavities of immunodepressed rats and integrate with the host brain.

  7. Survival of human mesenchymal stromal cells from bone marrow and adipose tissue after xenogenic transplantation in immunocompetent mice

    DEFF Research Database (Denmark)

    Niemeyer, P; Vohrer, J; Schmal, H

    2008-01-01

    of the current paper was to evaluate the survival of undifferentiated and osteogenically induced human MSC from different origins after transplantation in immunocompetent mice. METHODS: Human MSC were isolated from bone marrow (BMSC) and adipose tissue (ASC). After cultivation on mineralized collagen, MSC were...... was performed as a measure of immunologic rejection. Unloaded scaffolds served as controls (group C). Specimens were harvested at 4 and 8 weeks. RESULTS: Undifferentiated BMSC and ASC were detected in the majority of cases after xenogenic transplantation (group A, a total of 22 out of 24 cases), while......INTRODUCTION: Mesenchymal stromal cells (MSC) represent an attractive cell population for tissue engineering purposes. As MSC are described as immunoprivileged, non-autologous applications seem possible. A basic requirement is the survival of MSC after transplantation in the host. The purpose...

  8. Errantum: Treatment of human astrocytoma U87 cells with silicon dioxide nanoparticles lowers their survival and alters their expression of mitochondrial and cell signaling proteins

    Directory of Open Access Journals (Sweden)

    Lai JCK

    2010-12-01

    Full Text Available Lai JCK, Ananthakrishnan G, Jandhyam S, et al. Treatment of human astrocytoma U87 cells with silicon dioxide nanoparticles lowers their survival and alters their expression of mitochondrial and cell signaling proteins. Int J Nanomedicine. 2010;5:715–723.The wrong image was used in Figure 5 on page 719.

  9. Radiation-induced PGE2 sustains human glioma cells growth and survival through EGF signaling.

    Science.gov (United States)

    Brocard, Emeline; Oizel, Kristell; Lalier, Lisenn; Pecqueur, Claire; Paris, François; Vallette, François M; Oliver, Lisa

    2015-03-30

    Glioblastoma Multiforme (GBM) is the most common brain cancer in adults. Radiotherapy (RT) is the most effective post-operative treatment for the patients even though GBM is one of the most radio-resistant tumors. Dead or dying cells within the tumor are thought to promote resistance to treatment through mechanisms that are very poorly understood. We have evaluated the role of Prostaglandin E2 (PGE2), a versatile bioactive lipid, in GBM radio-resistance. We used an in vitro approach using 3D primary cultures derived from representative GBM patients. We show that irradiated glioma cells produced and released PGE2 in important quantities independently of the induction of cell death. We demonstrate that the addition of PGE2 enhances cell survival and proliferation though its ability to trans-activate the Epithelial Growth Factor receptor (EGFR) and to activate β-catenin. Indeed, PGE2 can substitute for EGF to promote primary cultures survival and growth in vitro and the effect is likely to occur though the Prostaglandin E2 receptor EP2.

  10. Radiation-induced PGE2 sustains human glioma cell growth and survival through EGF signaling

    Science.gov (United States)

    Lalier, Lisenn; Pecqueur, Claire; Paris, François; Vallette, François M.; Oliver, Lisa

    2015-01-01

    Glioblastoma Multiforme (GBM) is the most common brain cancer in adults. Radiotherapy (RT) is the most effective post-operative treatment for the patients even though GBM is one of the most radio-resistant tumors. Dead or dying cells within the tumor are thought to promote resistance to treatment through mechanisms that are very poorly understood. We have evaluated the role of Prostaglandin E2 (PGE2), a versatile bioactive lipid, in GBM radio-resistance. We used an in vitro approach using 3D primary cultures derived from representative GBM patients. We show that irradiated glioma cells produced and released PGE2 in important quantities independently of the induction of cell death. We demonstrate that the addition of PGE2 enhances cell survival and proliferation though its ability to trans-activate the Epithelial Growth Factor receptor (EGFR) and to activate β-catenin. Indeed, PGE2 can substitute for EGF to promote primary cultures survival and growth in vitro and the effect is likely to occur though the Prostaglandin E2 receptor EP2. PMID:25749386

  11. Taurine transport in human placental trophoblast is important for regulation of cell differentiation and survival.

    Science.gov (United States)

    Desforges, M; Parsons, L; Westwood, M; Sibley, C P; Greenwood, S L

    2013-03-21

    The outer epithelial cell layer of human placenta, the syncytiotrophoblast, is a specialised terminally differentiated multinucleate tissue. It is generated and renewed from underlying cytotrophoblast cells that undergo proliferation, differentiation and fusion with syncytiotrophoblast. Acquisition of fresh cellular components is thought to be balanced by apoptosis and shedding of aged nuclei. This process of trophoblast cell turnover maintains a functional syncytiotrophoblast, capable of sufficient nutrient transfer from mother to foetus. Foetal growth restriction (FGR) is a pregnancy complication associated with aberrant trophoblast turnover and reduced activity of certain amino acid transporters, including the taurine transporter (TauT). Taurine is the most abundant amino acid in human placenta implying an important physiological role within this tissue. Unlike other amino acids, taurine is not incorporated into proteins and in non-placental cell types represents an important osmolyte involved in cell volume regulation, and is also cytoprotective. Here, we investigated the role of taurine in trophoblast turnover using RNA interference to deplete primary human trophoblast cells of TauT and reduce intracellular taurine content. Trophoblast differentiation was compromised in TauT-deficient cells, and susceptibility of these cells to an inflammatory cytokine that is elevated in FGR was increased, evidenced by elevated levels of apoptosis. These data suggest an important role for taurine in trophoblast turnover and cytoprotection.

  12. Mirk/Dyrk1B mediates G0/G1 to S phase cell cycle progression and cell survival involving MAPK/ERK signaling in human cancer cells

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    Gao Jingchun

    2013-01-01

    Full Text Available Abstract Background Mirk/Dyrk1B contributes to G0 arrest by destabilization of cyclin D1 and stabilization of p27kip1 to maintain the viability of quiescent human cancer cells, and it could be negatively regulated by mitogenic-activated protein kinase (MAPK/extracellular signal-regulated kinase (ERK signaling. This study was performed to investigate the effect of Mirk/Dyrk1B on cell cycle and survival of human cancer cells involving MAPK/ERK signaling. Methods The correlations between Mirk/Dyrk1B expression and active ERK1/2 detected by western blot in both ovarian cancer and non-small cell lung cancer (NSCLC cells were analyzed by simple regression. Mirk/Dyrk1B unique phosphopeptides with sites associated with Mirk/Dyrk1B protein were isolated and quantitated by liquid chromatography coupled to tandem mass/mass spectrometry (LC-MS/MS proteomics analysis. The human cancer cells were treated with small interfering RNAs (siRNAs and/or U0126, an inhibitor of MEK for indicated duration, followed by investigating the alterations of cell cycle and apoptosis as well as related proteins examined by flow cytometry and Western blot, respectively. Results Our study demonstrated the widely expressed Mirk/Dyrk1B proteins in the human cancer cells were positively correlated with the levels of activated ERK1/2. Moreover, Mirk/Dyrk1B protein expressions consistent with the tyrosine autophosphorylated levels in the human cancer cells were increased by U0126 or growth factor-depleted culture. Conversely, knockdown of Mirk/Dyrk1B by siRNA led to up-regulated activation of c-Raf-MEK-ERK1/2 pathway and subsequent changes in cell cycle proteins (cyclin D1, p27kip1, accompanied by increased growth rate and cells from G0/G1 into S of cell cycle which could be blocked by U0126 in a dose-dependent manner, indicating Mirk/Dyrk1B may sequester MAPK/ERK pathway, and vice versa. Whereas, combined Mirk siRNA and U0126 induced cell apoptosis in the human cancer cells

  13. Overexpression of FABP3 inhibits human bone marrow derived mesenchymal stem cell proliferation but enhances their survival in hypoxia

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Suna, E-mail: wangs3@mail.nih.gov; Zhou, Yifu; Andreyev, Oleg; Hoyt, Robert F.; Singh, Avneesh; Hunt, Timothy; Horvath, Keith A.

    2014-04-15

    Studying the proliferative ability of human bone marrow derived mesenchymal stem cells in hypoxic conditions can help us achieve the effective regeneration of ischemic injured myocardium. Cardiac-type fatty acid binding protein (FABP3) is a specific biomarker of muscle and heart tissue injury. This protein is purported to be involved in early myocardial development, adult myocardial tissue repair and responsible for the modulation of cell growth and proliferation. We have investigated the role of FABP3 in human bone marrow derived mesenchymal stem cells under ischemic conditions. MSCs from 12 donors were cultured either in standard normoxic or modified hypoxic conditions, and the differential expression of FABP3 was tested by quantitative {sup RT}PCR and western blot. We also established stable FABP3 expression in MSCs and searched for variation in cellular proliferation and differentiation bioprocesses affected by hypoxic conditions. We identified: (1) the FABP3 differential expression pattern in the MSCs under hypoxic conditions; (2) over-expression of FABP3 inhibited the growth and proliferation of the MSCs; however, improved their survival in low oxygen environments; (3) the cell growth factors and positive cell cycle regulation genes, such as PCNA, APC, CCNB1, CCNB2 and CDC6 were all down-regulated; while the key negative cell cycle regulation genes TP53, BRCA1, CASP3 and CDKN1A were significantly up-regulated in the cells with FABP3 overexpression. Our data suggested that FABP3 was up-regulated under hypoxia; also negatively regulated the cell metabolic process and the mitotic cell cycle. Overexpression of FABP3 inhibited cell growth and proliferation via negative regulation of the cell cycle and down-regulation of cell growth factors, but enhances cell survival in hypoxic or ischemic conditions. - Highlights: • FABP3 expression pattern was studied in 12 human hypoxic-MSCs. • FABP3 mRNA and proteins are upregulated in the MSCs under hypoxic conditions.

  14. Impact of alemtuzumab treatment on the survival and function of human regulatory T cells in vitro.

    Science.gov (United States)

    Havari, Evis; Turner, Michael J; Campos-Rivera, Juanita; Shankara, Srinivas; Nguyen, Tri-Hung; Roberts, Bruce; Siders, William; Kaplan, Johanne M

    2014-01-01

    Alemtuzumab is a humanized monoclonal antibody specific for the CD52 protein present at high levels on the surface of B and T lymphocytes. In clinical trials, alemtuzumab has shown a clinical benefit superior to that of interferon-β in relapsing-remitting multiple sclerosis patients. Treatment with alemtuzumab leads to the depletion of circulating lymphocytes followed by a repopulation process characterized by alterations in the number, proportions and properties of lymphocyte subsets. Of particular interest, an increase in the percentage of T cells with a regulatory phenotype (Treg cells) has been observed in multiple sclerosis patients after alemtuzumab. Since Treg cells play an important role in the control of autoimmune responses, the effect of alemtuzumab on Treg cells was further studied in vitro. Alemtuzumab effectively mediated complement-dependent cytolysis of human T lymphocytes and the remaining population was enriched in T cells with a regulatory phenotype. The alemtuzumab-exposed T cells displayed functional regulatory characteristics including anergy to stimulation with allogeneic dendritic cells and ability to suppress the allogeneic response of autologous T cells. Consistent with the observed increase in Treg cell frequency, the CD25(hi) T-cell population was necessary for the suppressive activity of alemtuzumab-exposed T cells. The mechanism of this suppression was found to be dependent on both cell-cell contact and interleukin-2 consumption. These findings suggest that an alemtuzumab-mediated increase in the proportion of Treg cells may play a role in promoting the long-term efficacy of alemtuzumab in patients with multiple sclerosis.

  15. Mesenchymal stem cells from the human umbilical cord ameliorate fulminant hepatic failure and increase survival in mice

    Institute of Scientific and Technical Information of China (English)

    Jin-Feng Yang; Hong-Cui Cao; Qiao-Ling Pan; Jiong Yu; Jun Li; Lan-Juan Li

    2015-01-01

    BACKGROUND:Cell therapy has been promising for various diseases. We investigated whether transplantation of human umbilical cord mesenchymal stem cells (hUCMSCs) has any therapeutic effects on D-galactosamine/lipopolysaccharide (GalN/LPS)-induced fulminant hepatic failure in mice. METHODS:hUCMSCs isolated from human umbilical cord were cultured and transplanted via the tail vein into severe combined immune deifciency mice with GalN/LPS-induced fulminant hepatic failure. After transplantation, the localiza-tion and differentiation of hUCMSCs in the injured livers were investigated by immunohistochemical and genetic analy-ses. The recovery of the injured livers was evaluated histologi-cally. The survival rate of experimental animals was analyzed by the Kaplan-Meier method and log-rank test. RESULTS:hUCMSCs expressed high levels of CD29, CD73, CD13, CD105 and CD90, but did not express CD31, CD79b, CD133, CD34, and CD45. Cultured hUCMSCs displayed adip-ogenic and osteogenic differentiation potential. Hematoxylin and eosin staining revealed that transplantation of hUCMSCs reduced hepatic necrosis and promoted liver regeneration. Transplantation of hUCMSCs prolonged the survival rate of mice with fulminant hepatic failure. Polymerase chain reaction for humanalu sequences showed the presence of human cells in mouse livers. Positive staining for human albumin, human alpha-fetoprotein and human cytokeratin 18 suggested the for-mation of hUCMSCs-derived hepatocyte-like cellsin vivo. CONCLUSIONS:hUCMSC was a potential candidate for stem cell based therapies. After transplantation, hUCMSCs partially repaired hepatic damage induced by GalN/LPS in mice. hUC-MSCs engrafted into the injured liver and differentiated into hepatocyte-like cells.

  16. Artichoke compound cynarin differentially affects the survival, growth and stress response of normal, immortalized and cancerous human cells

    DEFF Research Database (Denmark)

    Gezer, Ceren; Yücecan, Sevinç; Rattan, Suresh Inder Singh

    2015-01-01

    Cynarin (CYN) is the main derivative of caffeoylquinic acid, found in leaves and heads of artichoke. Potential health-beneficial effects of CYN include as being choloretic-cholesterol lowering, hepatoprotective, anti-atherosclerotic, and antioxidative. We have tested the effects of various doses...... of CYN on the proliferative potential, survival, morphology, and stress response (SR) markers haemoxygenase-1 (HO-1) and heat shock protein-70 (HSP70) in normal human skin fibroblasts (FSF-1), telomerase-immortalized mesenchymal stem cells (hTERT-MSC) and cervical cancer cells, HeLa. Effects of CYN...

  17. Artichoke compound cynarin differentially affects the survival, growth and stress response of normal, immortalized and cancerous human cells

    DEFF Research Database (Denmark)

    Gezer, Ceren; Yücecan, Sevinç; Rattan, Suresh Inder Singh

    2015-01-01

    Cynarin (CYN) is the main derivative of caffeoylquinic acid, found in leaves and heads of artichoke. Potential health-beneficial effects of CYN include as being choloretic-cholesterol lowering, hepatoprotective, anti-atherosclerotic, and antioxidative. We have tested the effects of various doses...... of CYN on the proliferative potential, survival, morphology, and stress response (SR) markers haemoxygenase-1 (HO-1) and heat shock protein-70 (HSP70) in normal human skin fibroblasts (FSF-1), telomerase-immortalized mesenchymal stem cells (hTERT-MSC) and cervical cancer cells, HeLa. Effects of CYN...

  18. Dopaminergic neurons derived from human induced pluripotent stem cells survive and integrate into 6-OHDA-lesioned rats.

    Science.gov (United States)

    Cai, Jingli; Yang, Ming; Poremsky, Elizabeth; Kidd, Sarah; Schneider, Jay S; Iacovitti, Lorraine

    2010-07-01

    Cell replacement therapy could be an important treatment strategy for Parkinson's disease (PD), which is caused by the degeneration of dopamine neurons in the midbrain (mDA). The success of this approach greatly relies on the discovery of an abundant source of cells capable of mDAergic function in the brain. With the paucity of available human fetal tissue, efforts have increasingly focused on renewable stem cells. Human induced pluripotent stem (hiPS) cells offer great promise in this regard. If hiPS cells can be differentiated into authentic mDA neuron, hiPS could provide a potential autologous source of transplant tissue when generated from PD patients, a clear advantage over human embryonic stem (hES) cells. Here, we report that mDA neurons can be derived from a commercially available hiPS cell line, IMR90 clone 4, using a modified hES differentiation protocol established in our lab. These cells express all the markers (Lmx1a, Aldh1a1, TH, TrkB), follow the same mDA lineage pathway as H9 hES cells, and have similar expression levels of DA and DOPAC. Moreover, when hiPS mDA progenitor cells are transplanted into 6-OHDA-lesioned PD rats, they survive long term and many develop into bona fide mDA neurons. Despite their differentiation and integration into the brain, many Nestin+ tumor-like cells remain at the site of the graft. Our data suggest that as with hES cells, selecting the appropriate population of mDA lineage cells and eliminating actively dividing hiPS cells before transplantation will be critical for the future success of hiPS cell replacement therapy in PD patients.

  19. The suppression of manganese superoxide dismutase decreased the survival of human glioblastoma multiforme T98G cells

    Directory of Open Access Journals (Sweden)

    Novi S. Hardiany

    2017-05-01

    Full Text Available Background: Glioblastoma multiforme (GBM is a primary malignant brain tumor which has poor prognosis. High incidence of oxidative stress-based therapy resistance could be related to the high antioxidant status of GBM cells. Our previous study has reported that manganese superoxide dismutase (MnSOD antioxidant expression was significantly higher in high grade glioma than in low grade. The aim of this study was to analyze the impact of MnSOD suppression toward GBM cell survival.Methods: This study is an experimental study using human glioblastoma multiforme T98G cell line. Suppression of MnSOD expression was performed using in vitro transfection MnSOD-siRNA. The MnSOD expression was analyzed by measuring the mRNA using real time RT-PCR, protein using ELISA technique, and specific activity of enzyme using inhibition of xantine oxidase. Concentration of reactive oxygen species (ROS intracellular was determined by measuring superoxide radical and hydrogen peroxide. Cell survival was analyzed by measuring viability, proliferation, and cell apoptosis.Results: In vitro transfection of MnSOD-siRNA suppressed the mRNA, protein, and specific activity of MnSOD. This treatment significantly increased the concentration of superoxide radical; however, it did not influence the concentration of hydrogen peroxide. Moreover, viability MnSOD-suppressing cell significantly decreased, accompanied by increase of cell apoptosis without affecting cell proliferation.Conclusion: The suppression of MnSOD expression leads to decrease glioblastoma multiforme cell survival, which was associated to the increase of cell apoptotic.

  20. Cyclooxygenase 2-dependent and independent activation of Akt through casein kinase 2α contributes to human bladder cancer cell survival

    Directory of Open Access Journals (Sweden)

    Fujimoto Kiyohide

    2011-05-01

    Full Text Available Abstract Background Survival rate for patients presenting muscle invasive bladder cancer is very low, and useful therapeutic target has not been identified yet. In the present study, new COX2 downstream signals involved in urothelial carcinoma cell survival were investigated in vitro and in vivo. Methods COX2 gene was silenced by siRNA transfection. Orthotopic implantation animal model and transurethral instillation of siRNA with atelocollagen was constructed to examine the effects of COX2 knockdown in vivo. Cell cycle was examined by flowcytoketry. Surgical specimens derived from patients with urinary bladder cancer (all were initially diagnosed cases were used for immunohistochemical analysis of the indicated protein expression in urothelial carcinoma cells. Results Treatment with the COX2 inhibitor or knockdown of COX2 reduced expression of casein kinase (CK 2 α, a phophorylated Akt and urokinase type plasminogen activator (uPA, resulting in p27 induction, cell cycle arrest at G1 phase and cell growth suppression in human urothelial carcinoma cell lines expressing COX2. Silencing of CK2α exhibited the similar effects. Even in UMUC3 cells lacking the COX2 gene, COX2 inhibition also inhibited cell growth through down-regulation of the CK2α-Akt/uPA axis. The mouse orthotropic bladder cancer model demonstrated that the COX2 inhibitor, meloxicam significantly reduced CK2α, phosphorylated Akt and uPA expression, whereas induced p27 by which growth and invasiveness of bladder cancer cells were strongly inhibited. Immunohistochemically, high expression of COX2, CK2α and phosphorylated form of Akt was found in high-grade, invasive carcinomas as well as carcinoma in situ, but not in low-grade and noninvasive phenotypes. Conclusions COX2-dependent and independent activation of CK2α-Akt/uPA signal is mainly involved in urothelial carcinoma cell survival, moreover, not only COX2 but also CK2α could be direct targets of COX2 inhibitors.

  1. Mastic oil from Pistacia lentiscus var. chia inhibits growth and survival of human K562 leukemia cells and attenuates angiogenesis.

    Science.gov (United States)

    Loutrari, Heleni; Magkouta, Sophia; Pyriochou, Anastasia; Koika, Vasiliki; Kolisis, Fragiskos N; Papapetropoulos, Andreas; Roussos, Charis

    2006-01-01

    Mastic oil from Pistacia lentiscus var. chia, a natural plant extract traditionally used as a food additive, has been extensively studied for its antimicrobial activity attributed to the combination of its bioactive components. One of them, perillyl alcohol (POH), displays tumor chemopreventive, chemotherapeutic, and antiangiogenic properties. We investigated whether mastic oil would also suppress tumor cell growth and angiogenesis. We observed that mastic oil concentration and time dependently exerted an antiproliferative and proapoptotic effect on K562 human leukemia cells and inhibited the release of vascular endothelial growth factor (VEGF) from K562 and B16 mouse melanoma cells. Moreover, mastic oil caused a concentration-dependent inhibition of endothelial cell (EC) proliferation without affecting cell survival and a significant decrease of microvessel formation both in vitro and in vivo. Investigation of underlying mechanism(s) demonstrated that mastic oil reduced 1) in K562 cells the activation of extracellular signal-regulated kinases 1/2 (Erk1/2) known to control leukemia cell proliferation, survival, and VEGF secretion and 2) in EC the activation of RhoA, an essential regulator of neovessel organization. Overall, our results underscore that mastic oil, through its multiple effects on malignant cells and ECs, may be a useful natural dietary supplement for cancer prevention.

  2. Galectin-9 enhances cytokine secretion, but suppresses survival and degranulation, in human mast cell line.

    Directory of Open Access Journals (Sweden)

    Reiji Kojima

    Full Text Available Galectin-9 (Gal-9, a lectin having a β-galactoside-binding domain, can induce apoptosis of Th1 cells by binding to TIM-3. In addition, Gal-9 inhibits IgE/Ag-mediated degranulation of mast cell/basophilic cell lines by binding to IgE, thus blocking IgE/Ag complex formation. However, the role of Gal-9 in mast cell function in the absence of IgE is not fully understood. Here, we found that recombinant Gal-9 directly induced phosphorylation of Erk1/2 but not p38 MAPK in a human mast cell line, HMC-1, which does not express FcεRI. Gal-9 induced apoptosis and inhibited PMA/ionomycin-mediated degranulation of HMC-1 cells. On the other hand, Gal-9 induced cytokine and/or chemokine production by HMC-1 cells, dependent on activation of ERK1/2 but not p38 MAPK. In addition, the lectin activity of Gal-9 was required for Gal-9-mediated cytokine secretion by HMC-1 cells. These observations suggest that Gal-9 has dual properties as both a regulator and an activator of mast cells.

  3. Survival of primary human hepatocytes and death of induced pluripotent stem cells in media lacking glucose and arginine.

    Directory of Open Access Journals (Sweden)

    Minoru Tomizawa

    Full Text Available BACKGROUND: Tumorigenicity is an associated risk for transplantation of hepatocytes differentiated from human induced pluripotent stem (hiPS cells. Hepatocytes express the enzymes galactokinase and ornithine transcarbamylase (OTC to aid in their own survival. However, hiPS cells do not express these enzymes, and therefore, are not be expected to survive in a medium containing galactose and ornithine and lacking glucose and arginine. MATERIALS AND METHODS: Real-time quantitative polymerase chain reaction (PCR was performed to analyze the expression of galactokinase 1 (GALK11 and GALK2, ornithine carbamyltransferase, and phenylalanine hydroxylase (PAH. The hiPS cell line 201B7 was cultured in hepatocyte selection medium (HSM, which lacks glucose and arginine but contains galactose and ornithine. Furthermore, microscopic analysis of the cultured cells was performed after hematoxylin and eosin (H&E staining, terminal deoxynucleotidyl transferase (TdT-mediated dUTP nick end labeling (TUNEL. The hiPS cells were immunostained to assess their pluripotency in HSM. In addition, the primary human hepatocytes were cultured with or without hiPS cells in HSM. RESULTS: The expression levels of GALK1, GALK2, OTC, and PAH in 201B7 were 22.2±5.0 (average ± standard deviation, 14.2% ±1.1%, 1.2% ±0.2%, and 8.4% ±0.7% respectively, compared with those in the adult liver. The hiPS cell population diminished when cultured in HSM and completely disappeared after 3 days. The cultured cells showed condensation or fragmentation of their nuclei, thereby suggesting apoptosis. TUNEL staining confirmed that the cells had undergone apoptosis. The 201B7 cells were positive for Nanog, SSEA-4, and TRA-1-60. The primary human hepatocytes survived when cultured alone in HSM and when co-cultured with hiPS cells. CONCLUSION: Therefore, HSM is and ideal medium for eliminating hiPS cells and purifying hepatocytes without inducing any damage.

  4. Neuropilin-2 mediated β-catenin signaling and survival in human gastro-intestinal cancer cell lines.

    Directory of Open Access Journals (Sweden)

    Shaija Samuel

    Full Text Available NRP-2 is a high-affinity kinase-deficient receptor for ligands belonging to the class 3 semaphorin and vascular endothelial growth factor families. NRP-2 has been detected on the surface of several types of human cancer cells, but its expression and function in gastrointestinal (GI cancer cells remains to be determined. We sought to determine the function of NRP-2 in mediating downstream signals regulating the growth and survival of human gastrointestinal cancer cells. In human gastric cancer specimens, NRP-2 expression was detected in tumor tissues but not in adjacent normal mucosa. In CNDT 2.5 cells, shRNA mediated knockdown NRP-2 expression led to decreased migration and invasion in vitro (p<0.01. Focused gene-array analysis demonstrated that loss of NRP-2 reduced the expression of a critical metastasis mediator gene, S100A4. Steady-state levels and function of β-catenin, a known regulator of S100A4, were also decreased in the shNRP-2 clones. Furthermore, knockdown of NRP-2 sensitized CNDT 2.5 cells in vitro to 5FU toxicity. This effect was associated with activation of caspases 3 and 7, cleavage of PARP, and downregulation of Bcl-2. In vivo growth of CNDT 2.5 cells in the livers of nude mice was significantly decreased in the shNRP-2 group (p<0.05. Intraperitoneal administration of NRP-2 siRNA-DOPC decreased the tumor burden in mice (p = 0.01. Collectively, our results demonstrate that tumor cell-derived NRP-2 mediates critical survival signaling in gastrointestinal cancer cells.

  5. Transplantation of human oligodendrocyte progenitor cells in an animal model of diffuse traumatic axonal injury: survival and differentiation.

    Science.gov (United States)

    Xu, Leyan; Ryu, Jiwon; Hiel, Hakim; Menon, Adarsh; Aggarwal, Ayushi; Rha, Elizabeth; Mahairaki, Vasiliki; Cummings, Brian J; Koliatsos, Vassilis E

    2015-05-14

    Diffuse axonal injury is an extremely common type of traumatic brain injury encountered in motor vehicle crashes, sports injuries, and in combat. Although many cases of diffuse axonal injury result in chronic disability, there are no current treatments for this condition. Its basic lesion, traumatic axonal injury, has been aggressively modeled in primate and rodent animal models. The inexorable axonal and perikaryal degeneration and dysmyelination often encountered in traumatic axonal injury calls for regenerative therapies, including therapies based on stem cells and precursors. Here we explore the proof of concept that treatments based on transplants of human oligodendrocyte progenitor cells can replace or remodel myelin and, eventually, contribute to axonal regeneration in traumatic axonal injury. We derived human oligodendrocyte progenitor cells from the human embryonic stem cell line H9, purified and characterized them. We then transplanted these human oligodendrocyte progenitor cells into the deep sensorimotor cortex next to the corpus callosum of nude rats subjected to traumatic axonal injury based on the impact acceleration model of Marmarou. We explored the time course and spatial distribution of differentiation and structural integration of these cells in rat forebrain. At the time of transplantation, over 90 % of human oligodendrocyte progenitor cells expressed A2B5, PDGFR, NG2, O4, Olig2 and Sox10, a profile consistent with their progenitor or early oligodendrocyte status. After transplantation, these cells survived well and migrated massively via the corpus callosum in both injured and uninjured brains. Human oligodendrocyte progenitor cells displayed a striking preference for white matter tracts and were contained almost exclusively in the corpus callosum and external capsule, the striatopallidal striae, and cortical layer 6. Over 3 months, human oligodendrocyte progenitor cells progressively matured into myelin basic protein(+) and adenomatous

  6. PEA-15 Induces Autophagy in Human Ovarian Cancer Cells and is Associated with Prolonged Overall Survival

    OpenAIRE

    Bartholomeusz, Chandra; Rosen, Daniel; Wei, Caimiao; Kazansky, Anna; Yamasaki, Fumiyuki; Takahashi, Takeshi; Itamochi, Hiroaki; KONDO, Seiji; Liu, Jinsong; Ueno, Naoto T

    2008-01-01

    Phospho-enriched protein in astrocytes (PEA-15) is a 15-kDa phosphoprotein that slows cell proliferation by binding to and sequestering extracellular signal-regulated kinase (ERK) in the cytoplasm, thereby inhibiting ERK-dependent transcription and proliferation. In previous studies of E1A human gene therapy for ovarian cancer, we discovered that PEA-15 induced the antitumor effect of E1A by sequestering activated ERK in the cytoplasm of cancer cells. Here, we investigated the role of PEA-15 ...

  7. Adenoviral E4 Gene Stimulates Secretion of Pigmental Epithelium Derived Factor (PEDF) that Maintains Long-term Survival of Human Glomerulus-derived Endothelial Cells*

    Science.gov (United States)

    Jerebtsova, Marina; Kumari, Namita; Obuhkov, Yuri; Nekhai, Sergei

    2012-01-01

    Renal glomerular endothelial cells are specialized cells with an important role in physiological filtration and glomerular disease. However, maintenance of human primary endothelial cells requires stimulation with serum and growth factors that often results in modification of the cells properties. Previously, expression of early adenovirus region E4 was shown to help maintaining long-term survival of human endothelial cells in serum free media without addition of growth factors. In the current study, we showed that media conditioned with human epithelial cells stably transfected with Ad E4 region also supported survival of human glomerulus-derived endothelial cells in serum-free media. Mass-spectrometry analysis of the conditioned media identified pigmental epithelium derived factor (PEDF) as a major component of the conditioned media. PEDF expression in 293-E4 cells was validated by RT-PCR, Western blot and ELISA analysis. PEDF expression was detected in mouse glomeruli. Supplementation with recombinant PEDF supported survival of primary endothelial cells and the cells transformed with SV40 large T antigen in serum-free media, and extended the life-span of both cell cultures. PEDF did not inhibit FGF-2 stimulated growth and tubulogenesis of endothelial cells. Thus we demonstrated that adenoviral E4 region stimulated expression and secretion of PEDF by human renal epithelial cells that acted as a survival factor for glomerulus-derived endothelial cells. PMID:22915824

  8. Human alpha-lactalbumin made lethal to tumor cells (HAMLET) kills human glioblastoma cells in brain xenografts by an apoptosis-like mechanism and prolongs survival.

    Science.gov (United States)

    Fischer, Walter; Gustafsson, Lotta; Mossberg, Ann-Kristin; Gronli, Janne; Mork, Sverre; Bjerkvig, Rolf; Svanborg, Catharina

    2004-03-15

    Malignant brain tumors present a major therapeutic challenge because no selective or efficient treatment is available. Here, we demonstrate that intratumoral administration of human alpha-lactalbumin made lethal to tumor cells (HAMLET) prolongs survival in a human glioblastoma (GBM) xenograft model, by selective induction of tumor cell apoptosis. HAMLET is a protein-lipid complex that is formed from alpha-lactalbumin when the protein changes its tertiary conformation and binds oleic acid as a cofactor. HAMLET induces apoptosis in a wide range of tumor cells in vitro, but the therapeutic effect in vivo has not been examined. In this study, invasively growing human GBM tumors were established in nude rats (Han:rnu/rnu Rowett, n = 20) by transplantation of human GBM biopsy spheroids. After 7 days, HAMLET was administered by intracerebral convection-enhanced delivery for 24 h into the tumor area; and alpha-lactalbumin, the native, folded variant of the same protein, was used as a control. HAMLET reduced the intracranial tumor volume and delayed the onset of pressure symptoms in the tumor-bearing rats. After 8 weeks, all alpha-lactalbumin-treated rats had developed pressure symptoms, but the HAMLET-treated rats remained asymptomatic. Magnetic resonance imaging scans revealed large differences in tumor volume (456 versus 63 mm(3)). HAMLET caused apoptosis in vivo in the tumor but not in adjacent intact brain tissue or in nontransformed human astrocytes, and no toxic side effects were observed. The results identify HAMLET as a new candidate in cancer therapy and suggest that HAMLET should be additionally explored as a novel approach to controlling GBM progression.

  9. Transmission of clonal chromosomal abnormalities in human hematopoietic stem and progenitor cells surviving radiation exposure

    Energy Technology Data Exchange (ETDEWEB)

    Kraft, Daniela, E-mail: d.kraft@gsi.de [GSI Helmholtz Center for Heavy Ion Research, Department of Biophysics, Planckstr. 1, 64291 Darmstadt (Germany); Institute for Transfusion Medicine und Immunohematology, DRK-Blutspendedienst Baden-Wuerttemberg—Hessen, Johann Wolfgang Goethe-University Hospital, Sandhofstrasse 1, 60528 Frankfurt (Germany); Ritter, Sylvia, E-mail: s.ritter@gsi.de [GSI Helmholtz Center for Heavy Ion Research, Department of Biophysics, Planckstr. 1, 64291 Darmstadt (Germany); Durante, Marco, E-mail: m.durante@gsi.de [GSI Helmholtz Center for Heavy Ion Research, Department of Biophysics, Planckstr. 1, 64291 Darmstadt (Germany); Institute for Condensed Matter Physics, Physics Department, Technical University Darmstadt, Hochschulstraße 6-8, 64289 Darmstadt (Germany); Seifried, Erhard, E-mail: e.seifried@blutspende.de [Institute for Transfusion Medicine und Immunohematology, DRK-Blutspendedienst Baden-Wuerttemberg—Hessen, Johann Wolfgang Goethe-University Hospital, Sandhofstrasse 1, 60528 Frankfurt (Germany); Fournier, Claudia, E-mail: c.fournier@gsi.de [GSI Helmholtz Center for Heavy Ion Research, Department of Biophysics, Planckstr. 1, 64291 Darmstadt (Germany); Tonn, Torsten, E-mail: t.tonn@blutspende.de [Institute for Transfusion Medicine und Immunohematology, DRK-Blutspendedienst Baden-Wuerttemberg—Hessen, Johann Wolfgang Goethe-University Hospital, Sandhofstrasse 1, 60528 Frankfurt (Germany); Technische Universität Dresden, Med. Fakultät Carl Gustav Carus, Institute for Transfusion Medicine Dresden, German Red Cross Blood Donation Service North-East, Blasewitzer Straße 68/70, 01307 Dresden (Germany)

    2015-07-15

    Highlights: • Radiation induced formation and transmission of chromosomal aberrations were assessed. • Cytogenetic analysis was performed in human CD34+ HSPC by mFISH. • We report transmission of stable aberrations in irradiated, clonally expanded HSPC. • Unstable aberrations in clonally expanded HSPC occur independently of irradiation. • Carbon ions and X-rays bear a similar risk for propagation of cytogenetic changes. - Abstract: In radiation-induced acute myeloid leukemia (rAML), clonal chromosomal abnormalities are often observed in bone marrow cells of patients, suggesting that their formation is crucial in the development of the disease. Since rAML is considered to originate from hematopoietic stem and progenitor cells (HSPC), we investigated the frequency and spectrum of radiation-induced chromosomal abnormalities in human CD34{sup +} cells. We then measured stable chromosomal abnormalities, a possible biomarker of leukemia risk, in clonally expanded cell populations which were grown for 14 days in a 3D-matrix (CFU-assay). We compared two radiation qualities used in radiotherapy, sparsely ionizing X-rays and densely ionizing carbon ions (29 and 60–85 keV/μm, doses between 0.5 and 4 Gy). Only a negligible number of de novo arising, unstable aberrations (≤0.05 aberrations/cell, 97% breaks) were measured in the descendants of irradiated HSPC. However, stable aberrations were detected in colonies formed by irradiated HSPC. All cells of the affected colonies exhibited one or more identical aberrations, indicating their clonal origin. The majority of the clonal rearrangements (92%) were simple exchanges such as translocations (77%) and pericentric inversions (15%), which are known to contribute to the development of rAML. Carbon ions were more efficient in inducing cell killing (maximum of ∼30–35% apoptotic cells for 2 Gy carbon ions compared to ∼25% for X-rays) and chromosomal aberrations in the first cell-cycle after exposure (∼70% and

  10. Silibinin inhibits fibronectin induced motility, invasiveness and survival in human prostate carcinoma PC3 cells via targeting integrin signaling

    Energy Technology Data Exchange (ETDEWEB)

    Deep, Gagan [Department of Pharmaceutical Sciences, Skaggs School of Pharmacy and Pharmaceutical Sciences, University of Colorado Denver, Aurora, CO (United States); University of Colorado Cancer Center, University of Colorado Denver, Aurora, CO (United States); Kumar, Rahul; Jain, Anil K. [Department of Pharmaceutical Sciences, Skaggs School of Pharmacy and Pharmaceutical Sciences, University of Colorado Denver, Aurora, CO (United States); Agarwal, Chapla [Department of Pharmaceutical Sciences, Skaggs School of Pharmacy and Pharmaceutical Sciences, University of Colorado Denver, Aurora, CO (United States); University of Colorado Cancer Center, University of Colorado Denver, Aurora, CO (United States); Agarwal, Rajesh, E-mail: Rajesh.agarwal@ucdenver.edu [Department of Pharmaceutical Sciences, Skaggs School of Pharmacy and Pharmaceutical Sciences, University of Colorado Denver, Aurora, CO (United States); University of Colorado Cancer Center, University of Colorado Denver, Aurora, CO (United States)

    2014-10-15

    Highlights: • Silibinin inhibits fibronectin-induce motile morphology in PC3 cells. • Silibinin inhibits fibronectin-induced migration and invasion in PC3 cells. • Silibinin targets fibronectin-induced integrins and downstream signaling molecule. - Abstract: Prostate cancer (PCA) is the 2nd leading cause of cancer-related deaths among men in the United States. Preventing or inhibiting metastasis-related events through non-toxic agents could be a useful approach for lowering high mortality among PCA patients. We have earlier reported that natural flavonoid silibinin possesses strong anti-metastatic efficacy against PCA however, mechanism/s of its action still remains largely unknown. One of the major events during metastasis is the replacement of cell–cell interaction with integrins-based cell–matrix interaction that controls motility, invasiveness and survival of cancer cells. Accordingly, here we examined silibinin effect on advanced human PCA PC3 cells’ interaction with extracellular matrix component fibronectin. Silibinin (50–200 μM) treatment significantly decreased the fibronectin (5 μg/ml)-induced motile morphology via targeting actin cytoskeleton organization in PC3 cells. Silibinin also decreased the fibronectin-induced cell proliferation and motility but significantly increased cell death in PC3 cells. Silibinin also inhibited the PC3 cells invasiveness in Transwell invasion assays with fibronectin or cancer associated fibroblasts (CAFs) serving as chemoattractant. Importantly, PC3-luc cells cultured on fibronectin showed rapid dissemination and localized in lungs following tail vein injection in athymic male nude mice; however, in silibinin-treated PC3-luc cells, dissemination and lung localization was largely compromised. Molecular analyses revealed that silibinin treatment modulated the fibronectin-induced expression of integrins (α5, αV, β1 and β3), actin-remodeling (FAK, Src, GTPases, ARP2 and cortactin), apoptosis (cPARP and

  11. Endocannabinoids regulate growth and survival of human eccrine sweat gland-derived epithelial cells.

    Science.gov (United States)

    Czifra, Gabriella; Szöllősi, Attila G; Tóth, Balázs I; Demaude, Julien; Bouez, Charbel; Breton, Lionel; Bíró, Tamás

    2012-08-01

    The functional existence of the emerging endocannabinoid system (ECS), one of the new neuroendocrine players in cutaneous biology, is recently described in the human skin. In this study, using human eccrine sweat gland-derived immortalized NCL-SG3 model cells and a wide array of cellular and molecular assays, we investigated the effects of prototypic endocannabinoids (anandamide, 2-arachidonoylglycerol) on cellular functions. We show here that both endocannabinoids dose-dependently suppressed proliferation, induced apoptosis, altered expressions of various cytoskeleton proteins (e.g., cytokeratins), and upregulated lipid synthesis. Interestingly, as revealed by specific agonists and antagonists as well as by RNA interference, neither the metabotropic cannabinoid receptors (CB) nor the "ionotropic" CB transient receptor potential ion channels, expressed by these cells, mediated the cellular actions of the endocannabinoids. However, the endocannabinoids selectively activated the mitogen-activated protein kinase signaling pathway. Finally, other elements of the ECS (i.e., enzymes involved in the synthesis and degradation of endocannabinoids) were also identified on NCL-SG3 cells. These results collectively suggest that cannabinoids exert a profound regulatory role in the biology of the appendage. Therefore, from a therapeutic point of view, upregulation of endocannabinoid levels might help to manage certain sweat gland-derived disorders (e.g., tumors) characterized by unwanted growth.

  12. Regulation of HBEGF by Micro-RNA for Survival of Developing Human Trophoblast Cells.

    Science.gov (United States)

    Jain, Chandni V; Jessmon, Philip; Kilburn, Brian A; Jodar, Meritxell; Sendler, Edward; Krawetz, Stephen A; Armant, D Randall

    2016-01-01

    The growth factor HBEGF is upregulated post-transcriptionally in the low O2 environment of the human placenta during the first 10 weeks of pregnancy. We have examined the possible roles of HBEGF turnover and micro-RNA (miRNA) in its regulation by O2 in human first trimester trophoblast. HTR-8/SVneo trophoblast cells were cultured at 2% or 20% O2. The cells were transfected with a dual luciferase reporter construct (psiCHECK-2) containing no insert (control), the HBEGF 3' untranslated region (3'UTR), or sub-regions of the 3'UTR, as well as with siRNA for DGCR8. RNA was extracted from trophoblast cells cultured at 2% O2 for 0-4 h for next-generation sequencing. HBEGF was quantified by ELISA. HBEGF, DGCR8, and β-actin were examined by western blotting. Protein turnover studies, using 10 μg/ml cyclohexamide, 1 μg/ml lactocystin, or 100 μg/ml MG132, demonstrated faster HBEGF degradation at 20% O2 than 2% O2, mediated by the proteasome. However, proteasome inhibition failed to initiate HBEGF accumulation at 20% O2. Reporter assays, comparing to empty vector, demonstrated that the intact HBEGF 3' UTR inhibited expression (0.26), while fragments containing only its flanking regions increased reporter activity (3.15; 3.43). No differential expression of miRNAs was found in trophoblast cells cultured at 2% and 20% O2. Nevertheless, HBEGF upregulation at 2% O2 was blocked when the miRNA-processing protein DGCR8 was silenced, suggesting a role for miRNA. Our findings suggest involvement of flanking regions of the 3'UTR in activating HBEGF protein synthesis in response to 2% O2, possibly through a miRNA-mediated mechanism.

  13. CD9 may contribute to the survival of human germinal center B cells by facilitating the interaction with follicular dendritic cells

    Directory of Open Access Journals (Sweden)

    Sun-Ok Yoon

    2014-01-01

    Full Text Available The germinal center (GC is a dynamic microenvironment where antigen (Ag-activated B cells rapidly expand and differentiate, generating plasma cells (PC that produce high-affinity antibodies. Precise regulation of survival and proliferation of Ag-activated B cells within the GC is crucial for humoral immune responses. The follicular dendritic cells (FDC are the specialized stromal cells in the GC that prevent apoptosis of GC-B cells. Recently, we reported that human GC-B cells consist of CD9+ and CD9− populations and that it is the CD9+ cells that are committed to the PC lineage. In this study, we investigated the functional role of CD9 on GC-B cells. Tonsillar tissue section staining revealed that in vivo CD9+ GC-B cells localized in the light zone FDC area. Consistent this, in vitro CD9+ GC-B cells survived better than CD9− GC-B cells in the presence of HK cells, an FDC line, in a cell–cell contact-dependent manner. The frozen tonsillar tissue section binding assay showed that CD9+ GC-B cells bound to the GC area of tonsillar tissues significantly more than the CD9− GC-B cells did and that the binding was significantly inhibited by neutralizing anti-integrin β1 antibody. Furthermore, CD9+ cells bound to soluble VCAM-1 more than CD9− cells did, resulting in activation and stabilization of the active epitope of integrin β1. All together, our data suggest that CD9 on GC-B cells contributes to survival by strengthening their binding to FDC through the VLA4/VCAM-1 axis.

  14. Human mesenchymal stem cells prolong survival and ameliorate motor deficit through trophic support in Huntington's disease mouse models.

    Directory of Open Access Journals (Sweden)

    Yuan-Ta Lin

    Full Text Available We investigated the therapeutic potential of human bone marrow-derived mesenchymal stem cells (hBM-MSCs in Huntington's disease (HD mouse models. Ten weeks after intrastriatal injection of quinolinic acid (QA, mice that received hBM-MSC transplantation showed a significant reduction in motor function impairment and increased survival rate. Transplanted hBM-MSCs were capable of survival, and inducing neural proliferation and differentiation in the QA-lesioned striatum. In addition, the transplanted hBM-MSCs induced microglia, neuroblasts and bone marrow-derived cells to migrate into the QA-lesioned region. Similar results were obtained in R6/2-J2, a genetically-modified animal model of HD, except for the improvement of motor function. After hBM-MSC transplantation, the transplanted hBM-MSCs may integrate with the host cells and increase the levels of laminin, Von Willebrand Factor (VWF, stromal cell-derived factor-1 (SDF-1, and the SDF-1 receptor Cxcr4. The p-Erk1/2 expression was increased while Bax and caspase-3 levels were decreased after hBM-MSC transplantation suggesting that the reduced level of apoptosis after hBM-MSC transplantation was of benefit to the QA-lesioned mice. Our data suggest that hBM-MSCs have neural differentiation improvement potential, neurotrophic support capability and an anti-apoptotic effect, and may be a feasible candidate for HD therapy.

  15. B-cell receptor-associated protein 31 regulates human embryonic stem cell adhesion, stemness, and survival via control of epithelial cell adhesion molecule.

    Science.gov (United States)

    Kim, Won-Tae; Seo Choi, Hong; Min Lee, Hyun; Jang, Young-Joo; Ryu, Chun Jeih

    2014-10-01

    B-Cell receptor-associated protein 31 (BAP31) regulates the export of secreted membrane proteins from the endoplasmic reticulum (ER) to the downstream secretory pathway. Previously, we generated a monoclonal antibody 297-D4 against the surface molecule on undifferentiated human embryonic stem cells (hESCs). Here, we found that 297-D4 antigen was localized to pluripotent hESCs and downregulated during early differentiation of hESCs and identified that the antigen target of 297-D4 was BAP31 on the hESC-surface. To investigate the functional role of BAP31 in hESCs, BAP31 expression was knocked down by small interfering RNA. BAP31 depletion impaired hESC self-renewal and pluripotency and drove hESC differentiation into multicell lineages. BAP31 depletion hindered hESC proliferation by arresting cell cycle at G0/G1 phase and inducing caspase-independent cell death. Interestingly, BAP31 depletion reduced hESC adhesion to extracellular matrix (ECM). Analysis of cell surface molecules showed decreased expression of epithelial cell adhesion molecule (EpCAM) in BAP31-depleted hESCs, while ectopic expression of BAP31 elevated the expression of EpCAM. EpCAM depletion also reduced hESC adhesion to ECM, arrested cell cycle at G0/G1 phase and induced cell death, producing similar effects to those of BAP31 depletion. BAP31 and EpCAM were physically associated and colocalized at the ER and cell surface. Both BAP31 and EpCAM depletion decreased cyclin D1 and E expression and suppressed PI3K/Akt signaling, suggesting that BAP31 regulates hESC stemness and survival via control of EpCAM expression. These findings provide, for the first time, mechanistic insights into how BAP31 regulates hESC stemness and survival via control of EpCAM expression.

  16. PPARγ activation inhibits growth and survival of human endometriotic cells by suppressing estrogen biosynthesis and PGE2 signaling.

    Science.gov (United States)

    Lebovic, Dan I; Kavoussi, Shahryar K; Lee, JeHoon; Banu, Sakhila K; Arosh, Joe A

    2013-12-01

    Endometriosis is a chronic inflammatory disease of reproductive age women leading to chronic pelvic pain and infertility. Current antiestrogen therapies are temporizing measures, and endometriosis often recurs. Potential nonestrogenic or nonsteroidal targets are needed for treating endometriosis. Peroxisome proliferator-activated receptor (PPAR)γ, a nuclear receptor, is activated by thiazolidinediones (TZDs). In experimental endometriosis, TZDs inhibit growth of endometriosis. Clinical data suggest potential use of TZDs for treating pain and fertility concurrently in endometriosis patients. Study objectives were to 1) determine the effects of PPARγ action on growth and survival of human endometriotic epithelial and stromal cells and 2) identify the underlying molecular links between PPARγ activation and cell cycle regulation, apoptosis, estrogen biosynthesis, and prostaglandin E2 biosynthesis and signaling in human endometriotic epithelial and stromal cells. Results indicate that activation of PPARγ by TZD ciglitazone 1) inhibits growth of endometriotic epithelial cells 12Z up to 35% and growth of endometriotic stromal cells 22B up to 70% through altered cell cycle regulation and intrinsic apoptosis, 2) decreases expression of PGE2 receptors (EP)2 and EP4 mRNAs in 12Z and 22B cells, and 3) inhibits expression and function of P450 aromatase mRNA and protein and estrone production in 12Z and 22B cells through EP2 and EP4 in a stromal-epithelial cell-specific manner. Collectively, these results indicate that PGE2 receptors EP2 and EP4 mediate actions of PPARγ by incorporating multiple cell signaling pathways. Activation of PPARγ combined with inhibition of EP2 and EP4 may emerge as novel nonsteroidal therapeutic targets for endometriosis-associated pain and infertility, if clinically proven safe and efficacious.

  17. In vivo tracking of T cells in humans unveils decade-long survival and activity of genetically modified T memory stem cells.

    Science.gov (United States)

    Biasco, Luca; Scala, Serena; Basso Ricci, Luca; Dionisio, Francesca; Baricordi, Cristina; Calabria, Andrea; Giannelli, Stefania; Cieri, Nicoletta; Barzaghi, Federica; Pajno, Roberta; Al-Mousa, Hamoud; Scarselli, Alessia; Cancrini, Caterina; Bordignon, Claudio; Roncarolo, Maria Grazia; Montini, Eugenio; Bonini, Chiara; Aiuti, Alessandro

    2015-02-04

    A definitive understanding of survival and differentiation potential in humans of T cell subpopulations is of paramount importance for the development of effective T cell therapies. In particular, uncovering the dynamics in vivo in humans of the recently described T memory stem cells (TSCM) would be crucial for therapeutic approaches that aim at taking advantage of a stable cellular vehicle with precursor potential. We exploited data derived from two gene therapy clinical trials for an inherited immunodeficiency, using either retrovirally engineered hematopoietic stem cells or mature lymphocytes to trace individual T cell clones directly in vivo in humans. We compared healthy donors and bone marrow-transplanted patients, studied long-term in vivo T cell composition under different clinical conditions, and specifically examined TSCM contribution according to age, conditioning regimen, disease background, cell source, long-term reconstitution, and ex vivo gene correction processing. High-throughput sequencing of retroviral vector integration sites (ISs) allowed tracing the fate of more than 1700 individual T cell clones in gene therapy patients after infusion of gene-corrected hematopoietic stem cells or mature lymphocytes. We shed light on long-term in vivo clonal relationships among different T cell subtypes, and we unveiled that TSCM are able to persist and to preserve their precursor potential in humans for up to 12 years after infusion of gene-corrected lymphocytes. Overall, this work provides high-resolution tracking of T cell fate and activity and validates, in humans, the safe and functional decade-long survival of engineered TSCM, paving the way for their future application in clinical settings.

  18. Mesenchymal Stem Cells Support Survival and Proliferation of Primary Human Acute Myeloid Leukemia Cells through Heterogeneous Molecular Mechanisms

    Science.gov (United States)

    Brenner, Annette K.; Nepstad, Ina; Bruserud, Øystein

    2017-01-01

    Acute myeloid leukemia (AML) is a bone marrow malignancy, and various bone marrow stromal cells seem to support leukemogenesis, including osteoblasts and endothelial cells. We have investigated how normal bone marrow mesenchymal stem cells (MSCs) support the in vitro proliferation of primary human AML cells. Both MSCs and primary AML cells show constitutive release of several soluble mediators, and the mediator repertoires of the two cell types are partly overlapping. The two cell populations were cocultured on transwell plates, and MSC effects on AML cells mediated through the local cytokine/soluble mediator network could thus be evaluated. The presence of normal MSCs had an antiapoptotic and growth-enhancing effect on primary human AML cells when investigating a group of 51 unselected AML patients; this was associated with increased phosphorylation of mTOR and its downstream targets, and the effect was independent of cytogenetic or molecular-genetic abnormalities. The MSCs also supported the long-term proliferation of the AML cells. A subset of the patients also showed an altered cytokine network with supra-additive levels for several cytokines. The presence of cytokine-neutralizing antibodies or receptor inhibitors demonstrated that AML cells derived from different patients were heterogeneous with regard to effects of various cytokines on AML cell proliferation or regulation of apoptosis. We conclude that even though the effects of single cytokines derived from bone marrow MSCs on human AML cells differ among patients, the final cytokine-mediated effects of the MSCs during coculture is growth enhancement and inhibition of apoptosis.

  19. Cell division cycle-associated 7-like gene: A novel biomarker for adverse survival in human high-grade gliomas

    Directory of Open Access Journals (Sweden)

    Chia-Kuang Tsai

    2016-01-01

    Full Text Available Background: High-grade primary gliomas are aggressively growing and have an unfavorable prognosis. The utility of prognostic biomarkers of outcome in glioma patients is important for medical practice. Cell division cycle-associated 7-like (CDCA7L protein modifies cancer progression and metastasis. Nevertheless, its character in defining the clinical prognosis of human gliomas has not been illuminated. Subjects and Methods: The hypothesis of this study was that CDCA7L is upregulated in human gliomas. We studied two de-linked data from Gene Expression Omnibus (GEO profile. The first dataset (GDS1816/225081_s_at/CDCA7L in primary high-grade glioma included age, gender, and survival time. Another dataset (GDS1962/225081_s_at/CDCA7L was also encompassed to estimate CDCA7L gene expression in each pathological grading. Search Tool for the Retrieval of Interacting Genes/Proteins (STRING was used to survey the protein-protein interaction (PPI network of CDCA7L-regulated oncogenesis. Results: Statistical analysis of the GEO profile revealed that the World Health Organization (WHO Grade IV (n = 81 gliomas had higher CDCA7L mRNA expression level than in Grade II (n = 7, P = 2.15 × 10 −14 gliomas and nontumor controls (n = 23, P = 2.87 × 10 − 18. Kaplan-Meier analysis reported that patients with high CDCA7L mRNA levels (n = 49 had adverse survival than those with low CDCA7L expression (n = 28. The PPI analysis of CDCA7L-regulated oncogenesis showed CDCA7L as a potential hub protein. Conclusions: The expression of CDCA7L has a positive correlation with the WHO pathological grading and shorter survival. This finding suggests that CDCA7L may be a potential biomarker of prognosis in human gliomas.

  20. HIV-1 and recombinant gp120 affect the survival and differentiation of human vessel wall-derived mesenchymal stem cells

    Directory of Open Access Journals (Sweden)

    Pasquinelli Gianandrea

    2011-05-01

    Full Text Available Abstract Background HIV infection elicits the onset of a progressive immunodeficiency and also damages several other organs and tissues such as the CNS, kidney, heart, blood vessels, adipose tissue and bone. In particular, HIV infection has been related to an increased incidence of cardiovascular diseases and derangement in the structure of blood vessels in the absence of classical risk factors. The recent characterization of multipotent mesenchymal cells in the vascular wall, involved in regulating cellular homeostasis, suggests that these cells may be considered a target of HIV pathogenesis. This paper investigated the interaction between HIV-1 and vascular wall resident human mesenchymal stem cells (MSCs. Results MSCs were challenged with classical R5 and X4 HIV-1 laboratory strains demonstrating that these strains are able to enter and integrate their retro-transcribed proviral DNA in the host cell genome. Subsequent experiments indicated that HIV-1 strains and recombinant gp120 elicited a reliable increase in apoptosis in sub-confluent MSCs. Since vascular wall MSCs are multipotent cells that may be differentiated towards several cell lineages, we challenged HIV-1 strains and gp120 on MSCs differentiated to adipogenesis and endotheliogenesis. Our experiments showed that the adipogenesis is increased especially by upregulated PPARγ activity whereas the endothelial differentiation induced by VEGF treatment was impaired with a downregulation of endothelial markers such as vWF, Flt-1 and KDR expression. These viral effects in MSC survival and adipogenic or endothelial differentiation were tackled by CD4 blockade suggesting an important role of CD4/gp120 interaction in this context. Conclusions The HIV-related derangement of MSC survival and differentiation may suggest a direct role of HIV infection and gp120 in impaired vessel homeostasis and in genesis of vessel damage observed in HIV-infected patients.

  1. EBIO Does Not Induce Cardiomyogenesis in Human Pluripotent Stem Cells but Modulates Cardiac Subtype Enrichment by Lineage-Selective Survival

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    Monica Jara-Avaca

    2017-02-01

    Full Text Available Subtype-specific human cardiomyocytes (CMs are valuable for basic and applied research. Induction of cardiomyogenesis and enrichment of nodal-like CMs was described for mouse pluripotent stem cells (mPSCs in response to 1-ethyl-2-benzimidazolinone (EBIO, a chemical modulator of small-/intermediate-conductance Ca2+-activated potassium channels (SKs 1–4. Investigating EBIO in human pluripotent stem cells (PSCs, we have applied three independent differentiation protocols of low to high cardiomyogenic efficiency. Equivalent to mPSCs, timed EBIO supplementation during hPSC differentiation resulted in dose-dependent enrichment of up to 80% CMs, including an increase in nodal- and atrial-like phenotypes. However, our study revealed extensive EBIO-triggered cell loss favoring cardiac progenitor preservation and, subsequently, CMs with shortened action potentials. Proliferative cells were generally more sensitive to EBIO, presumably via an SK-independent mechanism. Together, EBIO did not promote cardiogenic differentiation of PSCs, opposing previous findings, but triggered lineage-selective survival at a cardiac progenitor stage, which we propose as a pharmacological strategy to modulate CM subtype composition.

  2. Cisplatin-induced apoptosis inhibits autophagy, which acts as a pro-survival mechanism in human melanoma cells.

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    Barbara Del Bello

    Full Text Available The interplay between a non-lethal autophagic response and apoptotic cell death is still a matter of debate in cancer cell biology. In the present study performed on human melanoma cells, we investigate the role of basal or stimulated autophagy in cisplatin-induced cytotoxicity, as well as the contribution of cisplatin-induced activation of caspases 3/7 and conventional calpains. The results show that, while down-regulating Beclin-1, Atg14 and LC3-II, cisplatin treatment inhibits the basal autophagic response, impairing a physiological pro-survival response. Consistently, exogenously stimulated autophagy, obtained with trehalose or calpains inhibitors (MDL-28170 and calpeptin, protects from cisplatin-induced apoptosis, and such a protection is reverted by inhibiting autophagy with 3-methyladenine or ATG5 silencing. In addition, during trehalose-stimulated autophagy, the cisplatin-induced activation of calpains is abrogated, suggesting the existence of a feedback loop between the autophagic process and calpains. On the whole, our results demonstrate that in human melanoma cells autophagy may function as a beneficial stress response, hindered by cisplatin-induced death mechanisms. In a therapeutic perspective, these findings suggest that the efficacy of cisplatin-based polychemotherapies for melanoma could be potentiated by inhibitors of autophagy.

  3. Effects of resolvin D1 on cell survival and cytokine expression of human gingival fibroblasts.

    Science.gov (United States)

    Khaled, Mohamed; Shibani, Nouf-Al; Labban, Nawaf; Batarseh, Ghada; Song, Fengyu; Ruby, John; Windsor, L Jack

    2013-12-01

    Tissue breakdown in periodontitis is initiated by bacteria, such as Porphyromonas gingivalis, and is caused largely by host responses. Resolvins protect the host against acute inflammation by blocking the migration of polymorphonuclear neutrophils to initiate resolution. The effects of resolvins on human gingival fibroblasts (HGFs) are unknown. This study examines the effects of resolvin D1 on HGF survival and cytokine expression when treated with or without P. gingivalis supernatant. Cytotoxicity of resolvin D1 on HGFs with or without a toxic level of P. gingivalis supernatant was measured with lactate dehydrogenase assays. Cytokine arrays were performed on HGF-conditioned media treated with or without resolvin D1 and with or without P. gingivalis supernatant. Resolvin D1 had no cytotoxic effects on HGFs at concentrations between 1 and 1,000 nM (all P > 0.05). Resolvin D1 (1,000 nM) significantly inhibited the toxic effects of 13.5% (v/v) P. gingivalis supernatant on HGFs (P = 0.002). Resolvin D1 significantly reduced the expression of interleukin (IL)-6 (P = 0.010) and monocyte chemoattractant protein (MCP)-1 (P = 0.04) in untreated fibroblasts. P. gingivalis (10%) supernatant significantly increased the expression levels of granulocyte-macrophage colony-stimulating factor (CSF), granulocyte CSF, growth-regulated oncogene (GRO), IL-5, IL-6, IL-7, IL-8, IL-10, MCP-1, MCP-2, MCP-3, and monokine induced by γ-interferon. Resolvin D1 significantly reduced the expression of GRO (P = 0.04), marginally reduced the levels of MCP-1 (P = 0.10), and marginally increased the levels of transforming growth factor (TGF)-β1 (P = 0.07) from HGFs treated with P. gingivalis supernatant. Resolvin D1 altered the cytotoxicity of P. gingivalis supernatant on HGFs. Resolvin D1 significantly reduced GRO, marginally reduced MCP-1, and marginally increased TGF-β1 from P. gingivalis-treated HGFs, which could alter the ability of P. gingivalis to induce inflammation.

  4. STAT3 signaling pathway is necessary for cell survival and tumorsphere forming capacity in ALDH{sup +}/CD133{sup +} stem cell-like human colon cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Lin, Li, E-mail: lin.796@osu.edu [Center for Childhood Cancer, The Research Institute at Nationwide Children' s Hospital, Department of Pediatrics, Internal Medicine, College of Medicine, The Ohio State University, Columbus, OH 43205 (United States); Division of Cardiology, Department of Internal Medicine, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030 (China); Fuchs, James; Li, Chenglong [Division of Medicinal Chemistry and Pharmacognosy, College of Pharmacy, The Ohio State University, Columbus, OH 43210 (United States); Olson, Veronica [Center for Childhood Cancer, The Research Institute at Nationwide Children' s Hospital, Department of Pediatrics, Internal Medicine, College of Medicine, The Ohio State University, Columbus, OH 43205 (United States); Bekaii-Saab, Tanios [Internal Medicine, College of Medicine, The Ohio State University, Columbus, OH 43210 (United States); Lin, Jiayuh, E-mail: lin.674@osu.edu [Center for Childhood Cancer, The Research Institute at Nationwide Children' s Hospital, Department of Pediatrics, Internal Medicine, College of Medicine, The Ohio State University, Columbus, OH 43205 (United States)

    2011-12-16

    Highlights: Black-Right-Pointing-Pointer The phosphorylated or activated form of STAT3 was expressed in colon cancer stem-like cells. Black-Right-Pointing-Pointer STAT3 inhibitor, FLLL32 inhibits P-STAT3 and STAT3 target genes in colon cancer stem-like cells. Black-Right-Pointing-Pointer Inhibition of STAT3 resulted in decreased cell viability and reduced numbers of tumorspheres. Black-Right-Pointing-Pointer STAT3 is required for survival and tumorsphere forming capacity in colon cancer stem-like cells. Black-Right-Pointing-Pointer Targeting STAT3 in cancer stem-like cells may offer a novel treatment approach for colon cancer. -- Abstract: Persistent activation of Signal Transducers and Activators of Transcription 3 (STAT3) is frequently detected in colon cancer. Increasing evidence suggests the existence of a small population of colon cancer stem or cancer-initiating cells may be responsible for tumor initiation, metastasis, and resistance to chemotherapy and radiation. Whether STAT3 plays a role in colon cancer-initiating cells and the effect of STAT3 inhibition is still unknown. Flow cytometry was used to isolate colon cancer stem-like cells from three independent human colon cancer cell lines characterized by both aldehyde dehydrogenase (ALDH)-positive and CD133-positive subpopulation (ALDH{sup +}/CD133{sup +}). The effects of STAT3 inhibition in colon cancer stem-like cells were examined. The phosphorylated or activated form of STAT3 was expressed in colon cancer stem-like cells and was reduced by a STAT3-selective small molecular inhibitor, FLLL32. FLLL32 also inhibited the expression of potential STAT3 downstream target genes in colon cancer stem-like cells including survivin, Bcl-XL, as well as Notch-1, -3, and -4, which may be involved in stem cell function. Furthermore, FLLL32 inhibited cell viability and tumorsphere formation as well as induced cleaved caspase-3 in colon cancer stem-like cells. FLLL32 is more potent than curcumin as evidenced with lower

  5. Cytokine profile of conditioned medium from human tumor cell lines after acute and fractionated doses of gamma radiation and its effect on survival of bystander tumor cells.

    Science.gov (United States)

    Desai, Sejal; Kumar, Amit; Laskar, S; Pandey, B N

    2013-01-01

    Cytokines are known to play pivotal roles in cancer initiation, progression and pathogenesis. Accumulating evidences suggest differences in basal and stress-induced cytokine profiles of cancers with diverse origin. However, a comprehensive investigation characterising the cytokine profile of various tumor types after acute and fractionated doses of gamma-irradiation, and its effect on survival of bystander cells is not well known in literature. In the present study, we have evaluated the cytokine secretion profile of human tumor cell lines (HT1080, U373MG, HT29, A549 and MCF-7) either before (basal) or after acute (2, 6 Gy) and fractionated doses (3×2 Gy) of gamma-irradiation in culture medium obtained from these cells by multiplex bead array/ELISA. Moreover, clonogenic assays were performed to evaluate the effect of conditioned medium (CM) on the survival and growth of respective cells. Based on the screening of 28 analytes, our results showed that the basal profiles of these cell lines varied considerably in terms of the number and magnitude of secreted factors, which was minimum in MCF-7. Interestingly, TNF-α, IL-1β, PDGF-AA, TGF-β1, fractalkine, IL-8, VEGF and GCSF were found in CM of all the cell lines. However, secretion of certain cytokines was cell line-specific. Moreover, CM caused increase in clonogenic survival of respective tumor cells (in the order HT1080>U373MG>HT29>A549>MCF-7), which was correlated with the levels of IL-1β, IL-6, IL-8, GMCSF and VEGF in their CM. After irradiation, the levels of most of the cytokines increased markedly in a dose dependent manner. The fold change in cytokine levels was lower in irradiated conditioned medium (ICM) of tumor cells collected after fractionated than respective acute dose, except in MCF-7. Interestingly, amongst these cell lines, the radiation-induced fold increase in cytokine levels was maximum in ICM of A549 cells. Moreover, bystander A549 cells treated with respective ICM showed dose dependent

  6. Influence of inorganic and organic iron compounds on parameters of cell growth and survival in human colon cells.

    Science.gov (United States)

    Klenow, Stefanie; Pool-Zobel, Beatrice L; Glei, Michael

    2009-04-01

    Increased risk for development of colon cancer is associated with red meat intake and iron toxicity is discussed for one underlying mechanism. Anyhow, for iron itself only limited evidence is found. In this study, effects of different iron compounds on proliferation of HT29 carcinoma and LT97 adenoma human colon cells were investigated. After treatment of cells with inorganic (ferrous sulfate: FeSO4 and ferric nitrilotriacetate: FeNTA) and organic (hemoglobin and hemin) iron sources (24-72 h), number of cells and metabolic activity were measured. Under normal cell culture conditions, neither iron compound elevated cell growth in either cell line with the exception of FeNTA which induced LT97 cell growth significantly. Distinct inhibition of cell proliferation was measured for organic iron. Serum-free incubation of HT29 cells revealed growth promoting properties of iron under deficiency. Even though organic iron, especially hemin, was a potent growth factor, both substances showed also dose-dependent cytotoxic effects. In conclusion, these data emphasize that not iron itself, but merely organic iron may promote carcinogenic events. Since promotion of proliferation was only detectable under deficiency, cytotoxic properties of organic iron may be of more importance in colon carcinogenesis.

  7. Inositol hexaphosphate downregulates both constitutive and ligand-induced mitogenic and cell survival signaling, and causes caspase-mediated apoptotic death of human prostate carcinoma PC-3 cells.

    Science.gov (United States)

    Gu, Mallikarjuna; Raina, Komal; Agarwal, Chapla; Agarwal, Rajesh

    2010-01-01

    Constitutively active mitogenic and prosurvival signaling cascades due to aberrant expression and interaction of growth factors and their receptors are well documented in human prostate cancer (PCa). Epidermal growth factor (EGF) and insulin-like growth factor-1 (IGF-1) are potent mitogens that regulate proliferation and survival of PCa cells via autocrine and paracrine loops involving both mitogen-activated protein kinase (MAPK)- and Akt-mediated signaling. Accordingly, here we assessed the effect of inositol hexaphosphate (IP6) on constitutive and ligand (EGF and IGF-1)-induced biological responses and associated signaling cascades in advanced and androgen-independent human PCa PC-3 cells. Treatment of PC-3 cells with 2 mM IP6 strongly inhibited both growth and proliferation and decreased cell viability; similar effects were also observed in other human PCa DU145 and LNCaP cells. IP6 also caused a strong apoptotic death of PC-3 cells together with caspase 3 and PARP cleavage. Mechanistic studies showed that biological effects of IP6 were associated with inhibition of both constitutive and ligand-induced Akt phosphorylation together with a decrease in total Akt levels, but a differential inhibitory effect on MAPKs extra cellular signal-regulated kinase 1/2 (ERK1/2), c-Jun N-terminal protein kinase (JNK1/2), and p38 under constitutive and ligand-activated conditions. Under similar condition, IP6 also inhibited AP-1 DNA-binding activity and decreased nuclear levels of both phospho and total c-Fos and c-Jun. Together, these findings for the first time establish IP6 efficacy in inhibiting aberrant EGF receptor (EGFR) or IGF-1 receptor (IGF-1R) pathway-mediated sustained growth promoting and survival signaling cascades in advanced and androgen-independent human PCa PC-3 cells, which might have translational implications in advanced human PCa control and management.

  8. Functional switching of ATM: sensor of DNA damage in proliferating cells and mediator of Akt survival signal in post-mitotic human neuron-like cells

    Institute of Scientific and Technical Information of China (English)

    Yan Li; Hua Xiong; Da-Qing Yang

    2012-01-01

    Ataxia-telangiectasia (A-T) is an autosomal recessive disorder characterized by cerebellar ataxia and oculocutaneous telangiectasias.The gene mutated in this disease,ATM (A-T,mutated),encodes a 370-kDa Ser/Thr protein kinase.ATM not only mediates cellular response to DNA damage but also acts as an activator of Akt in response to insulin.However,despite intensive studies,the mechanism underlying the neuronal degeneration symptoms of human A-T is still poorly understood.We found that the topoisomerase inhibitors etoposide and camptothecin readily induced apoptosis in undifferentiated proliferating SH-SY5Y cells but could not induce apoptosis in neuronally differentiated SH-SY5Y cells.In addition,etoposide induced p53 phosphorylation and H2AX foci formation in proliferating SH-SY5Y cells but failed to do so in differentiated SH-SY5Y cells.Moreover,while inhibition of ATM in undifferentiated SH-SY5Y cells partially protected them from etoposide-induced apoptosis,the same treatment had no effect on cell viability in differentiated SH-SY5Y cells.These results suggest that DNA damage or defective response to DNA damage is not the cause of neuronal cell death in human A-T.In contrast,we discovered that Akt phosphorylation was inhibited when ATM activity was suppressed in differentiated SH-SY5Y cells.Furthermore,inhibition of ATM induced apoptosis following serum starvation in neuronally differentiated SH-SY5Y cells but could not trigger apoptosis under the same conditions in undifferentiated proliferating SH-SY5Y cells.These results demonstrate that ATM mediates the Akt signaling and promotes cell survival in neuron-like human SH-SY5Y cells,suggesting that impaired activation of Akt is the reason for neuronal degeneration in human A-T.

  9. RIPK1 regulates survival of human melanoma cells upon endoplasmic reticulum stress through autophagy.

    Science.gov (United States)

    Luan, Qi; Jin, Lei; Jiang, Chen Chen; Tay, Kwang Hong; Lai, Fritz; Liu, Xiao Ying; Liu, Yi Lun; Guo, Su Tang; Li, Chun Ying; Yan, Xu Guang; Tseng, Hsin-Yi; Zhang, Xu Dong

    2015-01-01

    Although RIPK1 (receptor [TNFRSF]-interacting protein kinase 1) is emerging as a critical determinant of cell fate in response to cellular stress resulting from activation of death receptors and DNA damage, its potential role in cell response to endoplasmic reticulum (ER) stress remains undefined. Here we report that RIPK1 functions as an important prosurvival mechanism in melanoma cells undergoing pharmacological ER stress induced by tunicamycin (TM) or thapsigargin (TG) through activation of autophagy. While treatment with TM or TG upregulated RIPK1 and triggered autophagy in melanoma cells, knockdown of RIPK1 inhibited autophagy and rendered the cells sensitive to killing by TM or TG, recapitulating the effect of inhibition of autophagy. Consistently, overexpression of RIPK1 enhanced induction of autophagy and conferred resistance of melanoma cells to TM- or TG-induced cell death. Activation of MAPK8/JNK1 or MAPK9/JNK2, which phosphorylated BCL2L11/BIM leading to its dissociation from BECN1/Beclin 1, was involved in TM- or TG-induced, RIPK1-mediated activation of autophagy; whereas, activation of the transcription factor HSF1 (heat shock factor protein 1) downstream of the ERN1/IRE1-XBP1 axis of the unfolded protein response was responsible for the increase in RIPK1 in melanoma cells undergoing pharmacological ER stress. Collectively, these results identify upregulation of RIPK1 as an important resistance mechanism of melanoma cells to TM- or TG-induced ER stress by protecting against cell death through activation of autophagy, and suggest that targeting the autophagy-activating mechanism of RIPK1 may be a useful strategy to enhance sensitivity of melanoma cells to therapeutic agents that induce ER stress.

  10. Long-term survival of human neural stem cells in the ischemic rat brain upon transient immunosuppression.

    Directory of Open Access Journals (Sweden)

    Laura Rota Nodari

    Full Text Available Understanding the physiology of human neural stem cells (hNSCs in the context of cell therapy for neurodegenerative disorders is of paramount importance, yet large-scale studies are hampered by the slow-expansion rate of these cells. To overcome this issue, we previously established immortal, non-transformed, telencephalic-diencephalic hNSCs (IhNSCs from the fetal brain. Here, we investigated the fate of these IhNSC's immediate progeny (i.e. neural progenitors; IhNSC-Ps upon unilateral implantation into the corpus callosum or the hippocampal fissure of adult rat brain, 3 days after global ischemic injury. One month after grafting, approximately one fifth of the IhNSC-Ps had survived and migrated through the corpus callosum, into the cortex or throughout the dentate gyrus of the hippocampus. By the fourth month, they had reached the ipsilateral subventricular zone, CA1-3 hippocampal layers and the controlateral hemisphere. Notably, these results could be accomplished using transient immunosuppression, i.e administering cyclosporine for 15 days following the ischemic event. Furthermore, a concomitant reduction of reactive microglia (Iba1+ cells and of glial, GFAP+ cells was also observed in the ipsilateral hemisphere as compared to the controlateral one. IhNSC-Ps were not tumorigenic and, upon in vivo engraftment, underwent differentiation into GFAP+ astrocytes, and β-tubulinIII+ or MAP2+ neurons, which displayed GABAergic and GLUTAmatergic markers. Electron microscopy analysis pointed to the formation of mature synaptic contacts between host and donor-derived neurons, showing the full maturation of the IhNSC-P-derived neurons and their likely functional integration into the host tissue. Thus, IhNSC-Ps possess long-term survival and engraftment capacity upon transplantation into the globally injured ischemic brain, into which they can integrate and mature into neurons, even under mild, transient immunosuppressive conditions. Most notably

  11. BIRC6 protein, an inhibitor of apoptosis: role in survival of human prostate cancer cells.

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    Christopher G Low

    Full Text Available BACKGROUND: BIRC6 is a member of the Inhibitors of Apoptosis Protein (IAP family which is thought to protect a variety of cancer cells from apoptosis. The main objective of the present study was to investigate whether BIRC6 plays a role in prostate cancer and could be useful as a novel therapeutic target. METHODS: BIRC6 expression in cell lines was assessed using Western blot analysis and in clinical samples using immunohistochemistry of tissue microarrays. The biological significance of BIRC6 was determined by siRNA-induced reduction of BIRC6 expression in LNCaP cells followed by functional assays. RESULTS: Elevated BIRC6 protein expression was found in prostate cancer cell lines and clinical specimens as distinct from their benign counterparts. Increased BIRC6 expression was associated with Gleason 6-8 cancers and castration resistance. Reduction of BIRC6 expression in LNCaP cells led to a marked reduction in cell proliferation which was associated with an increase in apoptosis and a decrease in autophagosome formation. Doxorubicin-induced apoptosis was found to be coupled to a reduction in BIRC6 protein expression. CONCLUSION: The data suggest a role for BIRC6 in prostate cancer progression and treatment resistance, and indicate for the first time that the BIRC6 gene and its product are potentially valuable targets for treatment of prostate cancers.

  12. Curcumin sensitizes human gastric cancer cells to 5-fluorouracil through inhibition of the NFκB survival-signaling pathway

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    Kang Y

    2016-12-01

    Full Text Available Yanting Kang,1,2,* Wanle Hu,3,* Encheng Bai,1,2 Hailun Zheng,1 Zhiguo Liu,1 Jianzhang Wu,1 Rong Jin,2 Chengguang Zhao,1 Guang Liang1 1Chemical Biology Research Center, School of Pharmaceutical Sciences, 2Department of Epidemiology, First Affiliated Hospital, 3Department of Coloproctology, Second Affiliated Hospital, Wenzhou Medical University, Wenzhou, Zhejiang, China *These authors contributed equally to this work Abstract: Fluorouracil (5-FU is the most commonly used chemotherapeutic agent for gastric cancer (GC. However, the occurrence of resistance to 5-FU treatment poses a major problem for its clinical efficacy. In this study, we found that the NFκB-signaling pathway can mediate 5-FU resistance in GC cells. We developed a 5-FU-resistant GC cell line named SGCR/5-FU and found that the 5-FU-induced resistance increased cytosolic IκBα degradation and promoted NFκB nuclear translocation in GC cells. These findings were further confirmed by the activation of the NFκB survival-signaling pathway in clinical specimens. Curcumin, a natural compound, can reverse 5-FU resistance and inhibits proliferation in GC cells by downregulating the NFκB-signaling pathway. Moreover, it can also decrease the expression level of TNFα messenger RNA. Flow cytometry and Western blot analysis results showed that the combination of curcumin and 5-FU caused synergistic inhibition of growth and induction of potent apoptosis in the resistant cancer cell lines in vitro. In conclusion, our results demonstrate that the combination of 5-FU and curcumin could be further developed as a potential therapy for human GC. Keywords: 5-FU, curcumin, NFκB, drug resistance, gastric cancer

  13. Neonatal Desensitization Supports Long-Term Survival and Functional Integration of Human Embryonic Stem Cell-Derived Mesenchymal Stem Cells in Rat Joint Cartilage Without Immunosuppression

    Science.gov (United States)

    Zhang, Shufang; Jiang, Yang Zi; Zhang, Wei; Chen, Longkun; Tong, Tong; Liu, Wanlu; Mu, Qin; Liu, Hua; Ji, Junfeng; Ouyang, Hong Wei

    2013-01-01

    Immunological response hampers the investigation of human embryonic stem cells (hESCs) or their derivates for tissue regeneration in vivo. Immunosuppression is often used after surgery, but exhibits side effects of significant weight loss and allows only short-term observation. The purpose of this study was to investigate whether neonatal desensitization supports relative long-term survival of hESC-derived mesenchymal stem cells (hESC-MSCs) and promotes cartilage regeneration. hESC-MSCs were injected on the day of birth in rats. Six weeks after neonatal injection, a full-thickness cylindrical cartilage defect was created and transplanted with a hESC-MSC-seeded collagen bilayer scaffold (group d+s+c) or a collagen bilayer scaffold (group d+s). Rats without neonatal injection were transplanted with the hESC-MSC-seeded collagen bilayer scaffold to serve as controls (group s+c). Cartilage regeneration was evaluated by histological analysis, immunohistochemical staining, and biomechanical test. The role of hESC-MSCs in cartilage regeneration was analyzed by CD4 immunostaining, cell death detection, and visualization of human cells in regenerated tissues. hESC-MSCs expressed CD105, CD73, CD90, CD29, and CD44, but not CD45 and CD34, and possessed trilineage differentiation potential. Group d+s+c exhibited greater International Cartilage Repair Society (ICRS) scores than group d+s or group s+c. Abundant collagen type II and improved mechanical properties were detected in group d+s+c. There were less CD4+ inflammatory cell infiltration and cell death at week 1, and hESC-MSCs were found to survive as long as 8 weeks after transplantation in group d+s+c. Our study suggests that neonatal desensitization before transplantation may be an efficient way to develop a powerful tool for preclinical study of human cell-based therapies in animal models. PMID:22788986

  14. Clinical significance of SOX9 in human non-small cell lung cancer progression and overall patient survival

    Directory of Open Access Journals (Sweden)

    Zhou Chun-Hui

    2012-03-01

    Full Text Available Abstract Background Sex determining region Y (SRY-related high mobility groupbox 9 (SOX9 is an important transcription factor required for development, which regulates the expression of target genes in the associated pathway. The aim of this study was to describe the expression of SOX9 in human non-small cell lung cancer (NSCLC and to investigate the association between SOX9 expression and progression of NSCLC. Methods SOX9 protein and mRNA expression in normal human pneumonocytes, lung cancer cell lines, and eight pairs of matched lung cancer tissues and their adjacent normal lung tissues were detected by Western blotting and real-time reverse transcription-polymerase chain reaction (RT-PCR. Immunohistochemistry was used to determine SOX9 protein expression in 142 cases of histologically characterized NSCLC. Statistical analyses were applied to test for prognostic and diagnostic associations. Results SOX9 in lung cancer cell lines was upregulated at both mRNA and protein levels, and SOX9 mRNA and protein were also elevated in NSCLC tissues compared with levels in corresponding adjacent non-cancerous lung tissues. Immunohistochemical analysis demonstrated a high expression of SOX9 in 74/142 (52.1% paraffin-embedded archival lung cancer biopsies. Statistical analysis indicated that upregulation of SOX9 was significantly correlated with the histological stage of NSCLC (P = 0.017 and that patients with a high SOX9 level exhibited a shorter survival time (P Conclusions This work shows that SOX9 may serve as a novel and prognostic marker for NSCLC, and play a role during the development and progression of the disease.

  15. Temporal gene expression profile of human precursor B leukemia cells induced by adhesion receptor: identification of pathways regulating B-cell survival.

    Science.gov (United States)

    Astier, Anne Laurence; Xu, Ronghui; Svoboda, Marek; Hinds, Esther; Munoz, Olivier; de Beaumont, Rosalie; Crean, Colin Daniel; Gabig, Theodore; Freedman, Arnold Stephen

    2003-02-01

    The physical interactions between B cells and stromal cells from the lymphoid tissue microenvironment are critical to the survival of normal and malignant B cells. They are principally mediated by integrins expressed on B cells and counterreceptors on stromal cells. Specifically, alpha4beta1 integrin engagement rescues B cells from physiological or drug-induced apoptosis. Therefore, in order to understand the mechanisms by which integrins prevent apoptosis in leukemia B cells, we compared the temporal gene expression profiles induced by beta1-integrin ligation with fibronectin (Fn) or adhesion by poly-L-Lysine in serum-starved precursor B leukemia cells. Among the 38 selected differentially expressed genes, 6 genes involved in adhesion (VAV2, EPB41L1, CORO1A), proliferation (FRAP1, CCT4), and intercellular communication (GJB3) were validated by real-time quantitative polymerase chain reaction (RT-Q-PCR). Gene expression modulation could also be validated at the protein level for 5 other genes. We show that integrin stimulation up-regulated FBI-1 expression but inhibited CD79a, Requiem, c-Fos, and caspase 7 induction when the cells underwent apoptosis. We further demonstrate that Fn stimulation also inhibits caspase 3 activation but increases XIAP and survivin expression. Moreover, integrin stimulation also prevents caspase activation induced by doxorubicin. Therefore, we identified genes modulated by adhesion of human precursor B leukemia cells that regulate proliferation and apoptosis, highlighting new pathways that might provide insights into future therapy aiming at targeting apoptosis of leukemia cells.

  16. Fludarabine and cladribine induce changes in surface proteins on human B-lymphoid cell lines involved with apoptosis, cell survival, and antitumor immunity.

    Science.gov (United States)

    Kohnke, Philippa L; Mactier, Swetlana; Almazi, Juhura G; Crossett, Ben; Christopherson, Richard I

    2012-09-07

    Fludarabine and cladribine are purine analogues used to treat hematological malignancies. Alone or in combination with therapeutic antibodies, they are effective in treating patients with chronic lymphocytic leukemia and non-Hodgkin's lymphoma. However, the mechanisms of action of these drugs are not well understood. Plasma membrane proteins perform a variety of essential functions that can be affected by malignancy and perturbed by chemotherapy. Analysis of surface proteins may contribute to an understanding of the mechanisms of action of purine analogues and identify biomarkers for targeted therapy. The surface of human cells is rich in N-linked glycoproteins, enabling use of a hydrazide-coupling technique to enrich for glycoproteins, with iTRAQ labeling for quantitative comparison. A number of plasma membrane proteins on human leukemia and lymphoma cells were affected by treatment with a purine analogue, including decreases in CD22 (an adhesion and signaling molecule) and increases in CD205 (a "damaged cell marker") and CD80 and CD50 (T-cell interaction molecules). Purine analogues may affect B-cell receptor (BCR) signaling and costimulatory molecules, leading to multiple signals for apoptosis and cell clearance. Fludarabine and cladribine induce differential effects, with some cell survival proteins (ECE-1 and CD100) more abundant after fludarabine treatment. Cell surface proteins induced by fludarabine and cladribine may be targets for therapeutic antibodies.

  17. IL-17A promotes immune cell recruitment in human esophageal cancers and the infiltrating dendritic cells represent a positive prognostic marker for patient survival.

    Science.gov (United States)

    Lu, Lin; Pan, Ke; Zheng, Hai-Xia; Li, Jian-Jun; Qiu, Hui-Juan; Zhao, Jing-Jing; Weng, De-Sheng; Pan, Qiu-Zhong; Wang, Dan-Dan; Jiang, Shan-Shan; Chang, Alfred E; Li, Qiao; Xia, Jian-Chuan

    2013-10-01

    We previously reported that tumor-infiltrating interleukin (IL)-17A-producing cells play a protective role in human esophageal squamous cell carcinoma (ESCC). However, the potential mechanisms involved remain unclear. In the present study, we investigated the effects of IL-17A on immune cell recruitment and function in ESCC. In vitro chemotaxis assays using the ESCC cell lines EC109 and KYSE30 demonstrated that although IL-17A showed no significant direct effects on the migration of T cells, natural killer (NK) cells as well as dendritic cells (DCs), it could induce ESCC tumor cells to produce inflammatory chemokines, for example, CXCL9, CXCL10 and CCL2, CCL20, which are associated with the migration of T cells, NK cells, and DCs, respectively. In addition, IL-17A enhanced the cytotoxic effects of NK cells against tumor cells by augmenting the expression of cytotoxic molecules, for example, tumor necrosis factor-α, interferon-γ, Perforin, and Granzyme B and activation receptors, for example, NKp46, NKp44, NTB-A, and NKG2D on NK cells. Furthermore, immunohistochemical analysis revealed that the density of IL-17A-producing cells was positively and significantly associated with the density of CD1a DCs in tumor tissues. With the analyses of 181 ESCC patients, we found a correlation of higher number of tumor-infiltrating CD1a DCs with significantly improved overall survival of patients with ESCC. This study provides further understanding of the roles of Th17 cells in ESCC, which may contribute to the development of novel cancer immunotherapy strategies.

  18. Cell culture condition-dependent impact of AGE-rich food extracts on kinase activation and cell survival on human fibroblasts.

    Science.gov (United States)

    Nass, Norbert; Weissenberg, Kristian; Somoza, Veronika; Ruhs, Stefanie; Silber, Rolf-Edgar; Simm, Andreas

    2014-03-01

    Advanced glycation end products (AGEs) are stable end products of the Maillard reaction. Effects of food extracts are often initially analysed in cellular test systems and it is not clear how different cell culture conditions might influence the results. Therefore, we compared the effects of two models for AGE-rich food, bread crust and coffee extract (CE) on WI-38 human lung fibroblasts under different cell culture conditions (sub-confluent versus confluent cells, with and without serum). WI-38 cells responded to coffee and bread crust extract (BCE) with a rapid phosphorylation of PKB (AKT), p42/44 MAPK (ERK 1/2) and p38 MAPK, strongly depending on culture conditions. BCE resulted in increased cell numbers, whereas CE appeared to be cytotoxic. When cell numbers under all culture conditions and treatments were correlated with kinase phosphorylation, the relation between phospho-p38 MAPK and phospho-AKT represented a good, cell culture condition-independent predictor of cell survival.

  19. Human T-lymphotropic virus type-1 p30 alters cell cycle G2 regulation of T lymphocytes to enhance cell survival

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    Silverman Lee

    2007-07-01

    Full Text Available Abstract Background Human T-lymphotropic virus type-1 (HTLV-1 causes adult T-cell leukemia/lymphoma and is linked to a number of lymphocyte-mediated disorders. HTLV-1 contains both regulatory and accessory genes in four pX open reading frames. pX ORF-II encodes two proteins, p13 and p30, whose roles are still being defined in the virus life cycle and in HTLV-1 virus-host cell interactions. Proviral clones of HTLV-1 with pX ORF-II mutations diminish the ability of the virus to maintain viral loads in vivo. p30 expressed exogenously differentially modulates CREB and Tax-responsive element-mediated transcription through its interaction with CREB-binding protein/p300 and while acting as a repressor of many genes including Tax, in part by blocking tax/rex RNA nuclear export, selectively enhances key gene pathways involved in T-cell signaling/activation. Results Herein, we analyzed the role of p30 in cell cycle regulation. Jurkat T-cells transduced with a p30 expressing lentivirus vector accumulated in the G2-M phase of cell cycle. We then analyzed key proteins involved in G2-M checkpoint activation. p30 expression in Jurkat T-cells resulted in an increase in phosphorylation at serine 216 of nuclear cell division cycle 25C (Cdc25C, had enhanced checkpoint kinase 1 (Chk1 serine 345 phosphorylation, reduced expression of polo-like kinase 1 (PLK1, diminished phosphorylation of PLK1 at tyrosine 210 and reduced phosphorylation of Cdc25C at serine 198. Finally, primary human lymphocyte derived cell lines immortalized by a HTLV-1 proviral clone defective in p30 expression were more susceptible to camptothecin induced apoptosis. Collectively these data are consistent with a cell survival role of p30 against genotoxic insults to HTLV-1 infected lymphocytes. Conclusion Collectively, our data are the first to indicate that HTLV-1 p30 expression results in activation of the G2-M cell cycle checkpoint, events that would promote early viral spread and T-cell

  20. Human umbilical cord matrix-derived stem cells exert trophic effects on β-cell survival in diabetic rats and isolated islets

    Directory of Open Access Journals (Sweden)

    Yunting Zhou

    2015-12-01

    Full Text Available Human umbilical cord matrix-derived stem cells (uMSCs, owing to their cellular and procurement advantages compared with mesenchymal stem cells derived from other tissue sources, are in clinical trials to treat type 1 (T1D and type 2 diabetes (T2D. However, the therapeutic basis remains to be fully understood. The immunomodulatory property of uMSCs could explain the use in treating T1D; however, the mere immune modulation might not be sufficient to support the use in T2D. We thus tested whether uMSCs could exert direct trophic effects on β-cells. Infusion of uMSCs into chemically induced diabetic rats prevented hyperglycemic progression with a parallel preservation of islet size and cellularity, demonstrating the protective effect of uMSCs on β-cells. Mechanistic analyses revealed that uMSCs engrafted long-term in the injured pancreas and the engraftment markedly activated the pancreatic PI3K pathway and its downstream anti-apoptotic machinery. The pro-survival pathway activation was associated with the expression and secretion of β-cell growth factors by uMSCs, among which insulin-like growth factor 1 (IGF1 was highly abundant. To establish the causal relationship between the uMSC-secreted factors and β-cell survival, isolated rat islets were co-cultured with uMSCs in the transwell system. Co-culturing improved the islet viability and insulin secretion. Furthermore, reduction of uMSC-secreted IGF1 via siRNA knockdown diminished the protective effects on islets in the co-culture. Thus, our data support a model whereby uMSCs exert trophic effects on islets by secreting β-cell growth factors such as IGF1. The study reveals a novel therapeutic role of uMSCs and suggests that multiple mechanisms are employed by uMSCs to treat diabetes.

  1. A novel MEK-ERK-AMPK signaling axis controls chemokine receptor CCR7-dependent survival in human mature dendritic cells.

    Science.gov (United States)

    López-Cotarelo, Pilar; Escribano-Díaz, Cristina; González-Bethencourt, Ivan Luis; Gómez-Moreira, Carolina; Deguiz, María Laura; Torres-Bacete, Jesús; Gómez-Cabañas, Laura; Fernández-Barrera, Jaime; Delgado-Martín, Cristina; Mellado, Mario; Regueiro, José Ramón; Miranda-Carús, María Eugenia; Rodríguez-Fernández, José Luis

    2015-01-09

    Chemokine receptor CCR7 directs mature dendritic cells (mDCs) to secondary lymph nodes where these cells regulate the activation of T cells. CCR7 also promotes survival in mDCs, which is believed to take place largely through Akt-dependent signaling mechanisms. We have analyzed the involvement of the AMP-dependent kinase (AMPK) in the control of CCR7-dependent survival. A pro-apoptotic role for AMPK is suggested by the finding that pharmacological activators induce apoptosis, whereas knocking down of AMPK with siRNA extends mDC survival. Pharmacological activation of AMPK also induces apoptosis of mDCs in the lymph nodes. Stimulation of CCR7 leads to inhibition of AMPK, through phosphorylation of Ser-485, which was mediated by G(i)/Gβγ, but not by Akt or S6K, two kinases that control the phosphorylation of AMPK on Ser-485 in other settings. Using selective pharmacological inhibitors, we show that CCR7-induced phosphorylation of AMPK on Ser-485 is mediated by MEK and ERK. Coimmunoprecipitation analysis and proximity ligation assays indicate that AMPK associates with ERK, but not with MEK. These results suggest that in addition to Akt-dependent signaling mechanisms, CCR7 can also promote survival of mDCs through a novel MEK1/2-ERK1/2-AMPK signaling axis. The data also suggest that AMPK may be a potential target to modulate mDC lifespan and the immune response.

  2. The effects of altered fractionation schedules on the survival of human cell lines differing in their proliferative activity and repair capacity

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    Konefal, J.B.; Taylor, Y.C. (Washington Univ. School of Medicine, St. Louis, MO (USA))

    1989-11-01

    Plateau phase cultures of human cell lines were used as model systems to study the relative influences of proliferation and repair on the effectiveness of altered fractionation schedules. A human normal diploid fibroblast cell line (AG1522) which has a high capacity to repair potentially lethal radiation damage (PLD) and very little proliferative activity when grown to confluence was compared to human tumor cell lines which maintain significant cell-cycle activity in plateau phase. The human fibrosarcoma cell line, HT1080, used in the present study exhibited a much greater rate of turnover than the normal fibroblasts as determined from tritiated thymidine incorporation (5 day labeling index of 66% vs 20%) and no PLD repair, as determined by delayed plating experiments, in plateau phase. Twenty Gy were delivered to both cell lines over 5 days in 3 regimens: one 4 Gy fraction/day, two 2 Gy fractions/day with a 2 hr interval between doses, and two 2 Gy fractions/day with a 6 hr interval between doses. Although the normal fibroblasts demonstrated the greatest sparing between acute single doses and one 4 Gy fraction/day, there was little additional benefit (increased survival) from the increased dose fractionation. In contrast, the twice daily fractionation schedules resulted in significant differential sparing of the fibrosarcoma cells compared to the normal fibroblasts. With the 6 hr interval between doses, the survival advantages of the cell line with the slow turnover rate and high PLD repair capacity were completely lost. Split-dose experiments indicated slightly less sublethal damage repair in the fibrosarcoma cell line, but for both cell lines recovery was complete in 2 hr. DNA distributions were measured by flow cytometry and long term labeling index measurements performed in parallel with the multifraction radiation survival studies.

  3. Protein kinase A inhibitor, H89, enhances survival and clonogenicity of dissociated human embryonic stem cells through Rho-associated coiled-coil containing protein kinase (ROCK) inhibition.

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    Zhang, Liang; Xu, Yanqing; Xu, Jiandong; Wei, Yuping; Xu, Xia

    2016-04-01

    Can cell survival of dissociated human embryonic stem cells (hESCs) be increased during culture? A protein kinase A (PKA) inhibitor, H89, can significantly enhance survival and clonogenicity of dissociated hESCs without affecting their pluripotency. hESCs are vulnerable to massive cell death upon cellular detachment and dissociation. hESCs were dissociated into single cells and then cultured in feeder-dependent and -independent manners. H89 was added to the culture medium at different concentrations for 1 day. The statistical results were obtained from at least three independent experiments (n ≥ 4). The group without treatment was used as the negative control. 4 µM H89 was added in the culture medium to promote cell survival and colony formation of dissociated hESCs. MTT method and propidium iodide (PI) staining were used to determine cell proliferation, cell death and cell cycle, respectively. To count colony formation, alkaline phosphatase (AP) staining was carried out. Western blot was performed to determine protein expression. Except AP staining, immunofluorescence, RT-PCR and karyotype analysis were used to confirm the pluripotent state of H89 treated hESCs. H89 inhibits the dissociation-induced phosphorylation of PKA and two substrates of Rho-associated coiled-coil containing protein kinase (ROCK), myosin light chain (MLC2) and myosin phosphatase target subunit 1 (MYPT1), significantly increases cell survival and colony formation, and strongly depresses dissociation-induced cell death and cell blebbing without affecting the pluripotency of hESCs and their differentiation in vitro. Appropriate H89 concentration should be used and 1 day of H89 treatment is sufficient for promoting survival and colony formation of dissociated hESCs. These results provide an alternative for human pluripotent stem cell (hPSC) culture, broaden the scope of participants in the cell death of single hES cells after dissociation and further enlighten clues to understand the

  4. Combination of immortalization and inducible death strategies to generate a human mesenchymal stromal cell line with controlled survival

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    Paul Bourgine

    2014-03-01

    By combining the opposite concepts of ‘induced-life’ and ‘inducible-death’, we generated a hMSCs line with defined properties and allowing for temporally controlled survival. The cell line represents a relevant tool for medical discovery in regenerative medicine and a potential means to address availability, standardization and safety requirements in cell & gene therapy. The concept of a hTERT-iCasp9 combination, here explored in the context of hMSCs, could be extended to other types of progenitor/stem cells.

  5. Human Schwann cells exhibit long-term cell survival, are not tumorigenic and promote repair when transplanted into the contused spinal cord.

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    Bastidas, Johana; Athauda, Gagani; De La Cruz, Gabriela; Chan, Wai-Man; Golshani, Roozbeh; Berrocal, Yerko; Henao, Martha; Lalwani, Anil; Mannoji, Chikato; Assi, Mazen; Otero, P Anthony; Khan, Aisha; Marcillo, Alexander E; Norenberg, Michael; Levi, Allan D; Wood, Patrick M; Guest, James D; Dietrich, W Dalton; Bartlett Bunge, Mary; Pearse, Damien D

    2017-08-01

    The transplantation of rodent Schwann cells (SCs) provides anatomical and functional restitution in a variety of spinal cord injury (SCI) models, supporting the recent translation of SCs to phase 1 clinical trials for human SCI. Whereas human (Hu)SCs have been examined experimentally in a complete SCI transection paradigm, to date the reported behavior of SCs when transplanted after a clinically relevant contusive SCI has been restricted to the use of rodent SCs. Here, in a xenotransplant, contusive SCI paradigm, the survival, biodistribution, proliferation and tumorgenicity as well as host responses to HuSCs, cultured according to a protocol analogous to that developed for clinical application, were investigated. HuSCs persisted within the contused nude rat spinal cord through 6 months after transplantation (longest time examined), exhibited low cell proliferation, displayed no evidence of tumorigenicity and showed a restricted biodistribution to the lesion. Neuropathological examination of the CNS revealed no adverse effects of HuSCs. Animals exhibiting higher numbers of surviving HuSCs within the lesion showed greater volumes of preserved white matter and host rat SC and astrocyte ingress as well as axon ingrowth and myelination. These results demonstrate the safety of HuSCs when employed in a clinically relevant experimental SCI paradigm. Further, signs of a potentially positive influence of HuSC transplants on host tissue pathology were observed. These findings show that HuSCs exhibit a favorable toxicity profile for up to 6 months after transplantation into the contused rat spinal cord, an important outcome for FDA consideration of their use in human clinical trials. © 2017 Wiley Periodicals, Inc.

  6. Human Chorionic Gonadotropin Protects Vascular Endothelial Cells from Oxidative Stress by Apoptosis Inhibition, Cell Survival Signalling Activation and Mitochondrial Function Protection

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    Daniela Surico

    2015-07-01

    Full Text Available Background/Aim: Previous reports have made it hypothetically possible that human chorionic gonadotropin (hCG could protect against the onset of pregnancy-related pathological conditions by acting as an antioxidant. In the present study we planned to examine the effects of hCG against oxidative stress in human umbilical vein endothelial cells (HUVEC. Methods: HUVEC were subjected to peroxidation by hydrogen peroxide. The modulation of nitric oxide (NO release by hCG and its effects on cell viability, glutathione (GSH levels, mitochondrial membrane potential and mitochondrial transition pore opening (MPTP were examined by specific dyes. Endothelial and inducible NO synthase (eNOS and iNOS, Akt and extracellular -signal-regulated kinases 1/2 (ERK1/2 activation and markers of apoptosis were analyzed by Western Blot. Results: In HUVEC, hCG reduced NO release by modulating eNOS and iNOS. Moreover, hCG protected HUVEC against oxidative stress by preventing GSH reduction and apoptosis, by maintaining Akt and ERK1/2 activation and by keeping mitochondrial function. Conclusion: The present results have for the first time shown protective effects exerted by hCG on vascular endothelial function, which would be achieved by modulation of NO release, antioxidant and antiapoptotic actions and activation of cell survival signalling. These findings could have clinical implications in the management of pregnancy-related disorders.

  7. New blocking antibodies impede adhesion, migration and survival of ovarian cancer cells, highlighting MFGE8 as a potential therapeutic target of human ovarian carcinoma.

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    Lorenzo Tibaldi

    Full Text Available Milk Fat Globule--EGF--factor VIII (MFGE8, also called lactadherin, is a secreted protein, which binds extracellularly to phosphatidylserine and to αvβ3 and αvβ5 integrins. On human and mouse cells expressing these integrins, such as endothelial cells, phagocytes and some tumors, MFGE8/lactadherin has been shown to promote survival, epithelial to mesenchymal transition and phagocytosis. A protumoral function of MFGE8 has consequently been documented for a few types of human cancers, including melanoma, a subtype of breast cancers, and bladder carcinoma. Inhibiting the functions of MFGE8 could thus represent a new type of therapy for human cancers. Here, we show by immunohistochemistry on a collection of human ovarian cancers that MFGE8 is overexpressed in 45% of these tumors, and we confirm that it is specifically overexpressed in the triple-negative subtype of human breast cancers. We have established new in vitro assays to measure the effect of MFGE8 on survival, adhesion and migration of human ovarian and triple-negative breast cancer cell lines. Using these assays, we could identify new MFGE8-specific monoclonal antibodies, which efficiently blocked these three tumor-promoting effects of MFGE8. Our results suggest future use of MFGE8-blocking antibodies as new anti-cancer therapeutics in subgroups of ovarian carcinoma, and triple-negative breast carcinoma patients.

  8. Survival responses of cell subpopulations isolated from a heterogeneous human colon tumour after combinations of hyperthermia and X-irradiation

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    Leith, J.T.; Heyman, P.; Dewyngaert, J.K.; Glicksman, A.S. (Brown Univ., Providence, RI (USA). Div. of Biological and Medical Sciences; Rhode Island Hospital, Providence (USA)); Dexter, D.L.; Calabresi, P. (Roger Williams General Hospital, Providence, RI (USA))

    1983-03-01

    This research has investigated the effects of combined modality treatment on the survival responses of two tumour subpopulations obtained from a heterogeneous human colon adenocarcinoma. An isobologram analysis of the clonogenic survival responses of the two tumour subpopulations showed that the clone A responses were within the envelope of additivity for either sequence of application. In contrast, the responses of the clone D tumour subpopulation exhibited a supra-additive response to the combined treatments with the sequence of heat followed by X-irradiation being somewhat more effective than the sequence of X-irradiation followed by heat. These data indicate that the responses of tumour subpopulations obtained from heterogeneous solid tumours to combined modality treatments may vary in an, at present, unpredictable manner.

  9. Interleukin 10 promotes immune response by increasing the survival of activated CD8(+) T cells in human papillomavirus 16-infected cervical cancer.

    Science.gov (United States)

    Li, Li; Ma, Yan; Liu, Shuang; Zhang, Jin; Xu, Xin-Yan

    2016-10-11

    Human papillomavirus (HPV)-specific CD8(+) T cells are present in HPV-infected cervical cancer patients and have demonstrated potent antitumor properties. However, these cells cannot control tumor progression in most patients. To investigate the underlying mechanisms involved in suppressing or promoting CD8(+) T cell functions, we focused on interleukin 10 (IL-10), a pleiotropic cytokine with controversial roles in antitumor immunity. We found that compared to healthy controls, circulating CD8(+) T cells in HPV 16-infected cervical cancer patients expressed significantly higher levels of IL-10. Interestingly, these CD8(+) T cells from cervical cancer patients, but not those from healthy controls, responded to HPV 16 E6/E7 peptide stimulation by increasing IL-10 expression, demonstrating an antigen-specific IL-10 release. Addition of exogenous IL-10 improved the survival, but did not increase the proliferation, of peptide-stimulated CD8(+) T cells. CD8(+) T cells cultured in the presence of IL-10 also resulted in significantly higher interferon gamma (IFN-gamma) and granzyme B concentration, primarily due to improved cell survival. In resected cervical tumors, the frequency of tumor-infiltrating IL-10(+) CD8(+) T cells was positively correlated with the frequency of tumor-infiltrating IFN-gamma(+) and granzyme B(+) CD8(+) T cells. Tumor-associated macrophages were more potent than peripheral blood monocyte-derived macrophages at inducing IL-10 expression in CD8(+) T cells, possibly explaining the elevated IL-10(+) CD8(+) T cell frequency in cervical cancer patients. Together, these results are consistent with an immunostimulatory role of IL-10, which promoted CD8(+) T cell response by increasing the survival of activated CD8(+) T cells.

  10. Influence of Five Potential Anticancer Drugs on Wnt Pathway and Cell Survival in Human Biliary Tract Cancer Cells

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    Julia WACHTER, Daniel NEUREITER, Beate ALINGER, Martin PICHLER, Julia FUEREDER, Christian OBERDANNER, Pietro Di FAZIO, Matthias OCKER, Frieder BERR, Tobias KIESSLICH

    2012-01-01

    Full Text Available Background: The role of Wnt signalling in carcinogenesis suggests compounds targeting this pathway as potential anti-cancer drugs. Several studies report activation of Wnt signalling in biliary tract cancer (BTC thus rendering Wnt inhibitory drugs as potential candidates for targeted therapy of this highly chemoresistant disease.Methods: In this study we analysed five compounds with suggested inhibitory effects on Wnt signalling (DMAT, FH535, myricetin, quercetin, and TBB for their cytotoxic efficiency, mode of cell death, time- and cell line-dependent characteristics as well as their effects on Wnt pathway activity in nine different BTC cell lines.Results: Exposure of cancer cells to different concentrations of the compounds results in a clear dose-dependent reduction of viability for all drugs in the order FH535 > DMAT > TBB > myricetin > quercetin. The first three substances show high cytotoxicity in all tested cell lines, cause a direct cytotoxic effect by induction of apoptosis and inhibit pathway-specific signal transduction in a Wnt transcription factor reporter activity assay. Selected target genes such as growth-promoting cyclin D1 and the cell cycle progression inhibitor p27 are down- and up-regulated after treatment, respectively.Conclusions: Taken together, these data demonstrate that the small molecular weight inhibitors DMAT, F535 and TBB have a considerable cytotoxic and possibly Wnt-specific effect on BTC cell lines in vitro. Further in vivo investigation of these drugs as well as of new Wnt inhibitors may provide a promising approach for targeted therapy of this difficult-to-treat tumour.

  11. Caffeine Induces Cell Death via Activation of Apoptotic Signal and Inactivation of Survival Signal in Human Osteoblasts

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    Wen-Hsiung Chan

    2008-05-01

    Full Text Available Caffeine consumption is a risk factor for osteoporosis, but the precise regulatory mechanisms are currently unknown. Here, we show that cell viability decreases in osteoblasts treated with caffeine in a dose-dependent manner. This cell death is attributed primarily to apoptosis and to a smaller extent, necrosis. Moreover, caffeine directly stimulates intracellular oxidative stress. Our data support caffeine-induced apoptosis in osteoblasts via a mitochondria-dependent pathway. The apoptotic biochemical changes were effectively prevented upon pretreatment with ROS scavengers, indicating that ROS plays a critical role as an upstream controller in the caffeine-induced apoptotic cascade. Additionally, p21-activated protein kinase 2 (PAK2 and c-Jun N-terminal kinase (JNK were activated in caffeine-treated osteoblasts. Experiments further found that PAK2 activity is required for caffeine-induced JNK activation and apoptosis. Importantly, our data also show that caffeine triggers cell death via inactivation of the survival signal, including the ERK- and Akt-mediated anti-apoptotic pathways. Finally, exposure of rats to dietary water containing 10~20 μM caffeine led to bone mineral density loss. These results demonstrate for the first time that caffeine triggers apoptosis in osteoblasts via activation of mitochondria-dependent cell death signaling and inactivation of the survival signal, and causes bone mineral density loss in vivo.

  12. Progranulin Deficiency Reduces CDK4/6/pRb Activation and Survival of Human Neuroblastoma SH-SY5Y Cells.

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    de la Encarnación, Ana; Alquézar, Carolina; Esteras, Noemí; Martín-Requero, Ángeles

    2015-12-01

    Null mutations in GRN are associated with frontotemporal lobar degeneration with TDP-43 inclusions (FTLD-TDP). However, the influence of progranulin (PGRN) deficiency in neurodegeneration is largely unknown. In neuroblastoma cells, silencing of GRN gene causes significantly reduced cell survival after serum withdrawal. The following observations suggest that alterations of the CDK4/6/retinoblastoma protein (pRb) pathway, secondary to changes in PI3K/Akt and ERK1/2 activation induced by PGRN deficiency, are involved in the control of serum deprivation-induced apoptosis: (i) inhibiting CDK4/6 levels or their associated kinase activity by sodium butyrate or PD332991 sensitized control SH-SY5Y cells to serum deprivation-induced apoptosis without affecting survival of PGRN-deficient cells; (ii) CDK4/6/pRb seems to be downstream of the PI3K/Akt and ERK1/2 signaling pathways since their specific inhibitors, LY294002 and PD98059, were able to decrease CDK6-associated kinase activity and induce death of control SH-SY5Y cells; (iii) PGRN-deficient cells show reduced stimulation of PI3K/Akt, ERK1/2, and CDK4/6 activities compared with control cells in the absence of serum; and (iv) supplementation of recombinant human PGRN was able to rescue survival of PGRN-deficient cells. These observations highlight the important role of PGRN-mediated stimulation of the PI3K/Akt-ERK1/2/CDK4/6/pRb pathway in determining the cell fate survival/death under serum deprivation.

  13. Balance between apoptosis or survival induced by changes in extracellular-matrix composition in human mesangial cells: a key role for ILK-NFκB pathway.

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    del Nogal, María; Luengo, Alicia; Olmos, Gemma; Lasa, Marina; Rodriguez-Puyol, Diego; Rodriguez-Puyol, Manuel; Calleros, Laura

    2012-12-01

    Renal fibrosis is the final outcome of many clinical conditions that lead to chronic renal failure, characterized by a progressive substitution of cellular elements by extracellular-matrix proteins, in particular collagen type I. The aim of this study was to identify the mechanisms responsible for human mesangial cell survival, conditioned by changes in extracellular-matrix composition. Our results indicate that collagen I induces apoptosis in cells but only after inactivation of the pro-survival factor NFκB by either the super-repressor IκBα or the PDTC inhibitor. Collagen I activates a death pathway, through ILK/GSK-3β-dependent Bim expression. Moreover, collagen I significantly increases NFκB-dependent transcription, IκBα degradation and p65/NFκB translocation to the nucleus; it activates β1 integrin and this is accompanied by increased activity of ILK which leads to AKT activation. Knockdown of ILK or AKT with small interfering RNA suppresses the increase in NFκB activity. NFκB mediates cell survival through the antiapoptotic protein Bcl-xL. Our data suggest that human mesangial cells exposed to abnormal collagen I are protected against apoptosis by a complex mechanism involving integrin β1/ILK/AKT-dependent NFκB activation with consequent Bcl-xL overexpression, that opposes a simultaneously activated ILK/GSK-3β-dependent Bim expression and this dual mechanism may play a role in the progression of glomerular dysfunction.

  14. The novel mTORC1/2 dual inhibitor INK-128 suppresses survival and proliferation of primary and transformed human pancreatic cancer cells

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    Lou, Hai-zhou [Department of Medical Oncology, Sir Run Run Shaw Hospital, College of Medicine, Zhejiang University, Hangzhou 310016 (China); Weng, Xiao-chuan [Department of Anesthesiology, Hangzhou Xia-sha Hospital, Hangzhou 310018 (China); Pan, Hong-ming; Pan, Qin [Department of Medical Oncology, Sir Run Run Shaw Hospital, College of Medicine, Zhejiang University, Hangzhou 310016 (China); Sun, Peng [Department of Medical Oncology, Sun Yat-Sen University Cancer Center, Guangzhou 510060 (China); Liu, Li-li [Department of Medical Oncology, Sir Run Run Shaw Hospital, College of Medicine, Zhejiang University, Hangzhou 310016 (China); Chen, Bin, E-mail: chenbinhangzhou126@126.com [Department of Hepatopancreatobiliary Surgery, First People’s Hospital of Hangzhou, Hangzhou 310006 (China)

    2014-07-25

    Highlights: • INK-128 inhibits the survival and growth of human pancreatic cancer cells. • INK-128 induced pancreatic cancer cell apoptosis and necrosis simultaneously. • INK-128 blocks mTORC1/2 activation simultaneously in pancreatic cancer cells. • INK-128 down-regulates cyclin D1 and causes pancreatic cancer cell cycle arrest. • INK-128 significantly increases sensitivity of pancreatic cancer cells to gemcitabine. - Abstract: Pancreatic cancer has one of worst prognosis among all human malignancies around the world, the development of novel and more efficient anti-cancer agents against this disease is urgent. In the current study, we tested the potential effect of INK-128, a novel mammalian target of rapamycin (mTOR) complex 1 and 2 (mTORC1/2) dual inhibitor, against pancreatic cancer cells in vitro. Our results demonstrated that INK-128 concentration- and time-dependently inhibited the survival and growth of pancreatic cancer cells (both primary cells and transformed cells). INK-128 induced pancreatic cancer cell apoptosis and necrosis simultaneously. Further, INK-128 dramatically inhibited phosphorylation of 4E-binding protein 1 (4E-BP1), ribosomal S6 kinase 1 (S6K1) and Akt at Ser 473 in pancreatic cancer cells. Meanwhile, it downregulated cyclin D1 expression and caused cell cycle arrest. Finally, we found that a low concentration of INK-128 significantly increased the sensitivity of pancreatic cancer cells to gemcitabine. Together, our in vitro results suggest that INK-128 might be further investigated as a novel anti-cancer agent or chemo-adjuvant for pancreatic cancer treatment.

  15. Inhibition of connective tissue growth factor (CTGF/CCN2) expression decreases the survival and myogenic differentiation of human rhabdomyosarcoma cells.

    Science.gov (United States)

    Croci, Stefania; Landuzzi, Lorena; Astolfi, Annalisa; Nicoletti, Giordano; Rosolen, Angelo; Sartori, Francesca; Follo, Matilde Y; Oliver, Noelynn; De Giovanni, Carla; Nanni, Patrizia; Lollini, Pier-Luigi

    2004-03-01

    Connective tissue growth factor (CTGF/CCN2), a cysteine-rich protein of the CCN (Cyr61, CTGF, Nov) family of genes, emerged from a microarray screen of genes expressed by human rhabdomyosarcoma cells. Rhabdomyosarcoma is a soft tissue sarcoma of childhood deriving from skeletal muscle cells. In this study, we investigated the role of CTGF in rhabdomyosarcoma. Human rhabdomyosarcoma cells of the embryonal (RD/12, RD/18, CCA) and the alveolar histotype (RMZ-RC2, SJ-RH4, SJ-RH30), rhabdomyosarcoma tumor specimens, and normal skeletal muscle cells expressed CTGF. To determine the function of CTGF, we treated rhabdomyosarcoma cells with a CTGF antisense oligonucleotide or with a CTGF small interfering RNA (siRNA). Both treatments inhibited rhabdomyosarcoma cell growth, suggesting the existence of a new autocrine loop based on CTGF. CTGF antisense oligonucleotide-mediated growth inhibition was specifically due to a significant increase in apoptosis, whereas cell proliferation was unchanged. CTGF antisense oligonucleotide induced a strong decrease in the level of myogenic differentiation of rhabdomyosarcoma cells, whereas the addition of recombinant CTGF significantly increased the proportion of myosin-positive cells. CTGF emerges as a survival and differentiation factor and could be a new therapeutic target in human rhabdomyosarcoma.

  16. BDNF Increases Survival and Neuronal Differentiation of Human Neural Precursor Cells Cotransplanted with a Nanofiber Gel to the Auditory Nerve in a Rat Model of Neuronal Damage

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    Yu Jiao

    2014-01-01

    Full Text Available Objectives. To study possible nerve regeneration of a damaged auditory nerve by the use of stem cell transplantation. Methods. We transplanted HNPCs to the rat AN trunk by the internal auditory meatus (IAM. Furthermore, we studied if addition of BDNF affects survival and phenotypic differentiation of the grafted HNPCs. A bioactive nanofiber gel (PA gel, in selected groups mixed with BDNF, was applied close to the implanted cells. Before transplantation, all rats had been deafened by a round window niche application of β-bungarotoxin. This neurotoxin causes a selective toxic destruction of the AN while keeping the hair cells intact. Results. Overall, HNPCs survived well for up to six weeks in all groups. However, transplants receiving the BDNF-containing PA gel demonstrated significantly higher numbers of HNPCs and neuronal differentiation. At six weeks, a majority of the HNPCs had migrated into the brain stem and differentiated. Differentiated human cells as well as neurites were observed in the vicinity of the cochlear nucleus. Conclusion. Our results indicate that human neural precursor cells (HNPC integration with host tissue benefits from additional brain derived neurotrophic factor (BDNF treatment and that these cells appear to be good candidates for further regenerative studies on the auditory nerve (AN.

  17. Molecular mechanisms of (-)-epicatechin and chlorogenic acid on the regulation of the apoptotic and survival/proliferation pathways in a human hepatoma cell line.

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    Granado-Serrano, Ana Belén; Martín, María Angeles; Izquierdo-Pulido, María; Goya, Luis; Bravo, Laura; Ramos, Sonia

    2007-03-07

    Dietary polyphenols have been associated with reduced risk of chronic diseases, but the precise molecular mechanisms of protection remain unclear. This work was aimed at studying the effect of (-)-epicatechin (EC) and chlorogenic acid (CGA) on the regulation of apoptotic and survival/proliferation pathways in a human hepatoma cell line (HepG2). EC or CGA treatment for 18 h had a slight effect on cell viability and decreased reactive oxygen species formation, and EC alone promoted cell proliferation, whereas CGA increased glutathione levels. Phenols neither induced the caspase cascade for apoptosis nor affected expression levels of Bcl-xL or Bax. A sustained activation of the major survival signals AKT/PI-3-kinase and ERK was shown in EC-treated cells, rather than in CGA-exposed cells. These data suggest that EC and CGA have no effect on apoptosis and enhance the intrinsic cellular tolerance against oxidative insults either by activating survival/proliferation pathways or by increasing antioxidant potential in HepG2.

  18. Depression of p53-independent Akt survival signals in human oral cancer cells bearing mutated p53 gene after exposure to high-LET radiation

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    Nakagawa, Yosuke [Department of Oral and Maxillofacial Surgery, School of Medicine, Nara Medical University, 840 Shijo-cho, Kashihara, Nara 634-8521 (Japan); Takahashi, Akihisa [Advanced Scientific Research Leader Development Unit, Gunma University, 3-39-22 Showa-machi, Maebashi, Gunma 371-8511 (Japan); Kajihara, Atsuhisa; Yamakawa, Nobuhiro; Imai, Yuichiro [Department of Oral and Maxillofacial Surgery, School of Medicine, Nara Medical University, 840 Shijo-cho, Kashihara, Nara 634-8521 (Japan); Ota, Ichiro; Okamoto, Noritomo [Department of Otorhinolaryngology, School of Medicine, Nara Medical University, 840 Shijo-cho, Kashihara, Nara 634-8521 (Japan); Mori, Eiichiro [Department of Radiation Oncology, School of Medicine, Nara Medical University, 840 Shijo-cho, Kashihara, Nara 634-8521 (Japan); Noda, Taichi [Department of Dermatology, School of Medicine, Nara Medical University, 840 Shijo-cho, Kashihara, Nara 634-8521 (Japan); Furusawa, Yoshiya [Heavy-ion Radiobiology Research Group, Research Center for Charged Particle Therapy, National Institute of Radiological Sciences, 4-9-1 Anagawa, Inage-ku, Chiba 263-8555 (Japan); Kirita, Tadaaki [Department of Oral and Maxillofacial Surgery, School of Medicine, Nara Medical University, 840 Shijo-cho, Kashihara, Nara 634-8521 (Japan); Ohnishi, Takeo, E-mail: tohnishi@naramed-u.ac.jp [Department of Radiation Oncology, School of Medicine, Nara Medical University, 840 Shijo-cho, Kashihara, Nara 634-8521 (Japan)

    2012-07-13

    Highlights: Black-Right-Pointing-Pointer High-LET radiation induces efficiently apoptosis regardless of p53 gene status. Black-Right-Pointing-Pointer We examined whether high-LET radiation depresses the Akt-survival signals. Black-Right-Pointing-Pointer High-LET radiation depresses of survival signals even in the mp53 cancer cells. Black-Right-Pointing-Pointer High-LET radiation activates Caspase-9 through depression of survival signals. Black-Right-Pointing-Pointer High-LET radiation suppresses cell growth through depression of survival signals. -- Abstract: Although mutations and deletions in the p53 tumor suppressor gene lead to resistance to low linear energy transfer (LET) radiation, high-LET radiation efficiently induces cell lethality and apoptosis regardless of the p53 gene status in cancer cells. Recently, it has been suggested that the induction of p53-independent apoptosis takes place through the activation of Caspase-9 which results in the cleavage of Caspase-3 and poly (ADP-ribose) polymerase (PARP). This study was designed to examine if high-LET radiation depresses serine/threonine protein kinase B (PKB, also known as Akt) and Akt-related proteins. Human gingival cancer cells (Ca9-22 cells) harboring a mutated p53 (mp53) gene were irradiated with 2 Gy of X-rays or Fe-ion beams. The cellular contents of Akt-related proteins participating in cell survival signaling were analyzed with Western Blotting 1, 2, 3 and 6 h after irradiation. Cell cycle distributions after irradiation were assayed with flow cytometric analysis. Akt-related protein levels decreased when cells were irradiated with high-LET radiation. High-LET radiation increased G{sub 2}/M phase arrests and suppressed the progression of the cell cycle much more efficiently when compared to low-LET radiation. These results suggest that high-LET radiation enhances apoptosis through the activation of Caspase-3 and Caspase-9, and suppresses cell growth by suppressing Akt-related signaling, even in mp

  19. BDNF Pretreatment of Human Embryonic-Derived Neural Stem Cells Improves Cell Survival and Functional Recovery After Transplantation in Hypoxic-Ischemic Stroke.

    Science.gov (United States)

    Rosenblum, Sahar; Smith, Tenille N; Wang, Nancy; Chua, Joshua Y; Westbroek, Erick; Wang, Kendrick; Guzman, Raphael

    2015-01-01

    Intra-arterial neural stem cell (NSC) therapy has the potential to improve long-term outcomes after stroke. Here we evaluate if pretreatment of NSCs with brain-derived neurotrophic factor (BDNF) prior to transplantation improves cell engraftment and functional recovery following hypoxic-ischemic (HI) stroke. Human embryonic-derived NSCs with or without BDNF pretreatment (1 h, 100 ng/ml) were transplanted 3 days after HI stroke. Functional recovery was assessed using the horizontal ladder test. Cell engraftment was evaluated using bioluminescence imaging (BLI) and histological counts of SC121(+) cells. Fluoro-Jade C (FJC) and NeuN stains were used to evaluate neuroprotection. The effect of BDNF on NSCs was analyzed using a migration assay, immunocytochemistry, Luminex proteomic assay, and RT-qPCR.BLI analysis demonstrated significantly higher photon flux in the BDNF-treated NSC group compared to untreated NSC (p = 0.049) and control groups (p = 0.0021) at 1 week after transplantation. Immunohistochemistry confirmed increased transplanted cell survival in the cortex (p = 0.0126) and hippocampus (p = 0.0098) of animals injected with BDNF-treated NSCs compared to untreated NSCs. Behavioral testing revealed that the BDNF-treated NSC group demonstrated increased sensorimotor recovery compared to the untreated NSC and control groups (p < 0.001) over the 1-month period (p < 0.001) following transplantation. A significant improvement in performance was found in the BDNF-treated NSC group compared to the control group at 14, 21, and 28 (p < 0.05) days after transplantation. The cortex and hippocampus of the BDNF-treated NSC group had significantly more SC121(+) NSCs (p = 0.0125, p = 0.0098), fewer FJC(+) neurons (p = 0.0370, p = 0.0285), and a higher percentage of NeuN(+) expression (p = 0.0354) in the cortex compared to the untreated NSC group. BDNF treatment of NSCs resulted in significantly greater migration to SDF-1, secretion of M-CSF, VEGF, and expression of CXCR4

  20. Survivin as a potential mediator to support autoreactive cell survival in myasthenia gravis: a human and animal model study.

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    Linda L Kusner

    Full Text Available The mechanisms that underlie the development and maintenance of autoimmunity in myasthenia gravis are poorly understood. In this investigation, we evaluate the role of survivin, a member of the inhibitor of apoptosis protein family, in humans and in two animal models. We identified survivin expression in cells with B lymphocyte and plasma cells markers, and in the thymuses of patients with myasthenia gravis. A portion of survivin-expressing cells specifically bound a peptide derived from the alpha subunit of acetylcholine receptor indicating that they recognize the peptide. Thymuses of patients with myasthenia gravis had large numbers of survivin-positive cells with fewer cells in the thymuses of corticosteroid-treated patients. Application of a survivin vaccination strategy in mouse and rat models of myasthenia gravis demonstrated improved motor assessment, a reduction in acetylcholine receptor specific autoantibodies, and a retention of acetylcholine receptor at the neuromuscular junction, associated with marked reduction of survivin-expressing circulating CD20+ cells. These data strongly suggest that survivin expression in cells with lymphocyte and plasma cell markers occurs in patients with myasthenia gravis and in two animal models of myasthenia gravis. Survivin expression may be part of a mechanism that inhibits the apoptosis of autoreactive B cells in myasthenia gravis and other autoimmune disorders.

  1. Survivin as a potential mediator to support autoreactive cell survival in myasthenia gravis: a human and animal model study.

    Science.gov (United States)

    Kusner, Linda L; Ciesielski, Michael J; Marx, Alexander; Kaminski, Henry J; Fenstermaker, Robert A

    2014-01-01

    The mechanisms that underlie the development and maintenance of autoimmunity in myasthenia gravis are poorly understood. In this investigation, we evaluate the role of survivin, a member of the inhibitor of apoptosis protein family, in humans and in two animal models. We identified survivin expression in cells with B lymphocyte and plasma cells markers, and in the thymuses of patients with myasthenia gravis. A portion of survivin-expressing cells specifically bound a peptide derived from the alpha subunit of acetylcholine receptor indicating that they recognize the peptide. Thymuses of patients with myasthenia gravis had large numbers of survivin-positive cells with fewer cells in the thymuses of corticosteroid-treated patients. Application of a survivin vaccination strategy in mouse and rat models of myasthenia gravis demonstrated improved motor assessment, a reduction in acetylcholine receptor specific autoantibodies, and a retention of acetylcholine receptor at the neuromuscular junction, associated with marked reduction of survivin-expressing circulating CD20+ cells. These data strongly suggest that survivin expression in cells with lymphocyte and plasma cell markers occurs in patients with myasthenia gravis and in two animal models of myasthenia gravis. Survivin expression may be part of a mechanism that inhibits the apoptosis of autoreactive B cells in myasthenia gravis and other autoimmune disorders.

  2. Survival responses of cell subpopulations isolated from a heterogeneous human colon tumour after combinations of hyperthermia and X-irradiation.

    Science.gov (United States)

    Leith, J T; Heyman, P; Dewyngaert, J K; Glicksman, A S; Dexter, D L; Calabresi, P

    1983-03-01

    In summary, this research has investigated the effects of combined modality treatment (i.e., low linear energy transfer ionizing radiation and hyperthermia at 42.5 degrees C) on the survival responses of two tumour subpopulations (designated clones A and D) obtained from a heterogeneous human colon adenocarcinoma. A constant hyperthermic exposure (2 hours at 42.5 degrees C) was given either 3 min before or 3 min after graded exposure to X-rays. An isobologram analysis (Steel and Peckham 1979) of the clonogenic survival responses of the two tumour subpopulations showed that the clone A responses were within the envelope of additivity for either sequence of application. In contrast, the responses of the clone D tumour subpopulation exhibited a supra-additive response to the combined treatments with the sequence of heat followed by X-irradiation being somewhat more effective than the sequence of X-irradiation followed by heat. These data indicate that the responses of tumour subpopulations obtained from heterogeneous solid tumours to combined modality treatments may vary in an, at present, unpredictable manner.

  3. IL-26 promotes the proliferation and survival of human gastric cancer cells by regulating the balance of STAT1 and STAT3 activation.

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    Wei You

    Full Text Available Interleukin-26 (IL-26 is one of the cytokines secreted by Th17 cells whose role in human tumors remains unknown. Here, we investigated the expression and potential role of IL-26 in human gastric cancer (GC. The expression of IL-26 and related molecules such as IL-20R1, STAT1 and STAT3 was examined by real-time PCR and immunohistochemisty. The effects of IL-26 on cell proliferation and cisplatin-induced apoptosis were analyzed by BrdU cooperation assay and PI-Annexin V co-staining, respectively. Lentiviral mediated siRNA was used to explore its mechanism of action, and IL-26 related signaling was analyzed by western blotting. Human GC tissues showed increased levels of IL-26 and its related molecules and activation of STAT3 signaling, whereas STAT1 activation did not differ significantly between GC and normal gastric tissues. Moreover, IL-26 was primarily produced by Th17 and NK cells. IL-26 promoted the proliferation and survival of MKN45 and SGC-7901 gastric cancer cells in a dose-dependent manner. Furthermore, IL-20R2 and IL-10R1, which are two essential receptors for IL-26 signaling, were expressed in both cell lines. IL-26 activated STAT1 and STAT3 signaling; however, the upregulation of the expression of Bcl-2, Bcl-xl and c-myc indicated that the effect of IL-26 is mediated by STAT3 activation. Knockdown of STAT1 and STAT3 expression suggested that the proliferative and anti-apoptotic effects of IL-26 are mediated by the modulation of STAT1/STAT3 activation. In summary, elevated levels of IL-26 in human GC promote proliferation and survival by modulating STAT1/STAT3 signaling.

  4. Dormant origins licensed by excess Mcm2-7 are required for human cells to survive replicative stress.

    Science.gov (United States)

    Ge, Xin Quan; Jackson, Dean A; Blow, J Julian

    2007-12-15

    In late mitosis and early G1, Mcm2-7 complexes are loaded onto DNA to license replication origins for use in the upcoming S phase. However, the amount of Mcm2-7 loaded is in significant excess over the number of origins normally used. We show here that in human cells, excess chromatin-bound Mcm2-7 license dormant replication origins that do not fire during normal DNA replication, in part due to checkpoint activity. Dormant origins were activated within active replicon clusters if replication fork progression was inhibited, despite the activation of S-phase checkpoints. After lowering levels of chromatin-bound Mcm2-7 in human cells by RNA interference (RNAi), the use of dormant origins was suppressed in response to replicative stress. Although cells with lowered chromatin-bound Mcm2-7 replicated at normal rates, when challenged with replication inhibitors they had dramatically reduced rates of DNA synthesis and reduced viability. These results suggest that the use of dormant origins licensed by excess Mcm2-7 is a new and physiologically important mechanism that cells utilize to maintain DNA replication rates under conditions of replicative stress. We propose that checkpoint kinase activity can preferentially suppress initiation within inactive replicon clusters, thereby directing new initiation events toward active clusters that are experiencing replication problems.

  5. Association between FAT1 mutation and overall survival in patients with human papillomavirus–negative head and neck squamous cell carcinoma

    Science.gov (United States)

    Kim, Ki Tae; Kim, Bo‐Sung

    2016-01-01

    Abstract Background The purpose of this study was to characterize the mutation profile of FAT atypical cadherin 1 (FAT1) and determine the prognostic significance of FAT1 mutation for overall survival in patients with human papillomavirus (HPV)‐negative head and neck squamous cell carcinoma (HNSCC). Methods Data were downloaded from The Cancer Genome Atlas (TCGA) and International Cancer Genome Consortium (ICGC) data portals and used as discovery and validation sets. FAT1 mutational status was determined in 234 and 37 patients with HPV‐negative HNSCC, respectively, and overall survival analysis was performed. For comparison, HPV‐positive patients were also analyzed for overall survival. Results Most of the identified nonsynonymous somatic FAT1 mutations were loss‐of‐function mutations. FAT1 mutation was significantly associated with better overall survival in HPV‐negative patients from both the TCGA cohort (p = .026) and the ICGC cohort (p = .047), but not in HPV‐positive patients. Conclusion FAT1 mutational status is a strong independent prognostic factor in patients with HPV‐negative HNSCC. © 2016 The Authors Head & Neck Published by Wiley Periodicals, Inc. Head Neck 38: E2021–E2029, 2016 PMID:26876381

  6. p16 expression independent of human papillomavirus is associated with lower stage and longer disease-free survival in oral cavity squamous cell carcinoma.

    Science.gov (United States)

    Satgunaseelan, Laveniya; Virk, Sohaib A; Lum, Trina; Gao, Kan; Clark, Jonathan R; Gupta, Ruta

    2016-08-01

    There is limited information regarding the incidence of p16 expression, its association with human papillomavirus (HPV) and prognosis in oral cavity squamous cell carcinoma (OSCC). The role of p16 in OSCC is evaluated in 215 cases using tissue microarrays (TMAs). p16 immunohistochemistry and HPV in situ hybridisation were performed on TMAs following histopathology review of 215 patients with OSCC in the Sydney Head and Neck Cancer Institute database. Thirty-seven (17.2%) cases showed p16 expression without association with HPV. p16 expression significantly decreased with increasing pT category (p=0.002). p16 expression was associated with longer disease-specific survival on univariable analysis (p=0.044) but not on multivariable analysis adjusting for depth of invasion. Amongst patients receiving adjuvant radiotherapy, patients with p16 expression had significantly longer disease-free and overall survival. p16 expression was seen in early stage OSCCs and was associated with better survival following surgery and radiotherapy. While not an independent predictor of survival, p16 may mediate its effects by contributing to reduced proliferative capacity, leading to smaller tumour size and lower invasive potential.

  7. Celecoxib and LLW-3-6 Reduce Survival of Human Glioma Cells Independently and Synergistically with Sulfasalazine.

    Science.gov (United States)

    Yerokun, Tokunbo; Winfield, Leyte L

    2015-12-01

    Gliomas are among the most commonly diagnosed central nervous system tumors. Celecoxib has been utilized with success in the treatment of several types of cancer, including gliomas. The present study examined the antiproliferative effects of celecoxib and its benzimidazole-based analog, LLW-3-6, when used as co-treatment with sulfasalazine against human glioma LN18 cells. At 48-h treatment, the glioma cells maintained 60% viability in the presence of celecoxib or LLW-3-6 at the maximum concentration tested (40 μM). Co-treatment of glioma cells under a non-lethal dose (50 μM) of sulfasalazine and either celecoxib or LLW-3-6 (administered at different concentrations) resulted in improved inhibition of cell viability. The concentration of the molecules required to reduce cell growth in the combined treatment was significantly less than that needed when either molecule was administered independently. Based on computational values, LLW-3-6 has physiochemical characteristics that should allow for improved bioavailability in comparison to that of celecoxib.

  8. A novel angiogenesis inhibitor impairs lovo cell survival via targeting against human VEGFR and its signaling pathway of phosphorylation.

    Science.gov (United States)

    Zhang, Y M; Dai, B L; Zheng, L; Zhan, Y Z; Zhang, J; Smith, W W; Wang, X L; Chen, Y N; He, L C

    2012-10-11

    Colorectal cancer represents the fourth commonest malignancy, and constitutes a major cause of significant morbidity and mortality among other diseases. However, the chemical therapy is still under development. Angiogenesis plays an important role in colon cancer development. We developed HMQ18-22 (a novel analog of taspine) with the aim to target angiogenesis. We found that HMQ18-22 significantly reduced angiogenesis of chicken chorioallantoic membrane (CAM) and mouse colon tissue, and inhibited cell migration and tube formation as well. Then, we verified the interaction between HMQ18-22 and VEGFR2 by AlphaScreen P-VEGFR assay, screened the targets on angiogenesis by VEGF Phospho Antibody Array, validated the target by western blot and RNAi in lovo cells. We found HMQ18-22 could decrease phosphorylation of VEGFR2(Tyr(1214)), VEGFR1(Tyr(1333)), Akt(Tyr(326)), protein kinase Cα (PKCα) (Tyr(657)) and phospholipase-Cγ-1 (PLCγ-1) (Tyr(771)). Most importantly, HMQ18-22 inhibited proliferation of lovo cell and tumor growth in a human colon tumor xenografted model of athymic mice. Compared with normal lovo cells proliferation, the inhibition on proliferation of knockdown cells (VEGFR2, VEGFR1, Akt, PKCα and PLCγ-1) by HMQ18-22 decreased. These results suggested that HMQ18-22 is a novel angiogenesis inhibitor and can be a useful therapeutic candidate for colon cancer intervention.

  9. Transposon mutagenesis of Salmonella enterica serovar Enteritidis identifies genes that contribute to invasiveness in human and chicken cells and survival in egg albumen.

    Science.gov (United States)

    Shah, Devendra H; Zhou, Xiaohui; Kim, Hye-Young; Call, Douglas R; Guard, Jean

    2012-12-01

    Salmonella enterica serovar Enteritidis is an important food-borne pathogen, and chickens are a primary reservoir of human infection. While most knowledge about Salmonella pathogenesis is based on research conducted on Salmonella enterica serovar Typhimurium, S. Enteritidis is known to have pathobiology specific to chickens that impacts epidemiology in humans. Therefore, more information is needed about S. Enteritidis pathobiology in comparison to that of S. Typhimurium. We used transposon mutagenesis to identify S. Enteritidis virulence genes by assay of invasiveness in human intestinal epithelial (Caco-2) cells and chicken liver (LMH) cells and survival within chicken (HD-11) macrophages as a surrogate marker for virulence. A total of 4,330 transposon insertion mutants of an invasive G1 Nal(r) strain were screened using Caco-2 cells. This led to the identification of attenuating mutations in a total of 33 different loci, many of which include genes previously known to contribute to enteric infection (e.g., Salmonella pathogenicity island 1 [SPI-1], SPI-4, SPI-5, CS54, fliH, fljB, csgB, spvR, and rfbMN) in S. Enteritidis and other Salmonella serovars. Several genes or genomic islands that have not been reported previously (e.g., SPI-14, ksgA, SEN0034, SEN2278, and SEN3503) or that are absent in S. Typhimurium or in most other Salmonella serovars (e.g., pegD, SEN1152, SEN1393, and SEN1966) were also identified. Most mutants with reduced Caco-2 cell invasiveness also showed significantly reduced invasiveness in chicken liver cells and impaired survival in chicken macrophages and in egg albumen. Consequently, these genes may play an important role during infection of the chicken host and also contribute to successful egg contamination by S. Enteritidis.

  10. Lixisenatide accelerates restoration of normoglycemia and improves human beta-cell function and survival in diabetic immunodeficient NOD–scid IL-2rgnull RIP-DTR mice engrafted with human islets

    Directory of Open Access Journals (Sweden)

    Yang C

    2015-08-01

    Full Text Available Chaoxing Yang,1 Matthias Loehn,2 Agata Jurczyk,1 Natalia Przewozniak,1 Linda Leehy,1 Pedro L Herrera,3 Leonard D Shultz,4 Dale L Greiner,1 David M Harlan,5 Rita Bortell1 1Program in Molecular Medicine, Diabetes Center of Excellence, University of Massachusetts Medical School, Worcester, MA, USA; 2Sanofi-Aventis, Diabetes Division, Frankfurt, Germany; 3University of Geneva, Geneva, Switzerland; 4The Jackson Laboratory, Bar Harbor, ME, USA; 5Department of Medicine, Diabetes Center of Excellence, University of Massachusetts Medical School, Worcester, MA, USA Objective: Glucagon-like peptide-1 induces glucose-dependent insulin secretion and, in rodents, increases proliferation and survival of pancreatic beta cells. To investigate the effects on human beta cells, we used immunodeficient mice transplanted with human islets. The goal was to determine whether lixisenatide, a glucagon-like peptide-1 receptor agonist, improves human islet function and survival in vivo. Methods: Five independent transplant studies were conducted with human islets from five individual donors. Diabetic human islet-engrafted immunodeficient mice were treated with lixisenatide (50, 150, and 500 µg/kg or vehicle. Islet function was determined by blood glucose, plasma human insulin/C-peptide, and glucose tolerance tests. Grafts were analyzed for total beta- and alpha-cell number, percent proliferation, and levels of apoptosis. Results: Diabetic mice transplanted with marginal human islet mass and treated with lixisenatide were restored to euglycemia more rapidly than vehicle-treated mice. Glucose tolerance tests, human plasma insulin, and glucose-stimulation indices of lixisenatide-treated mice were significantly improved compared to vehicle-treated mice. The percentages of proliferating or apoptotic beta cells at graft recovery were not different between lixisenatide-treated and vehicle-treated mice. Nevertheless, in one experiment we found a significant twofold to threefold

  11. Two types of human malignant melanoma cell lines revealed by expression patterns of mitochondrial and survival-apoptosis genes: implications for malignant melanoma therapy.

    Science.gov (United States)

    Su, David M; Zhang, Qiuyang; Wang, Xuexi; He, Ping; Zhu, Yuelin Jack; Zhao, Jianxiong; Rennert, Owen M; Su, Yan A

    2009-05-01

    Human malignant melanoma has poor prognosis because of resistance to apoptosis and therapy. We describe identification of the expression profile of 1,037 mitochondria-focused genes and 84 survival-apoptosis genes in 21 malignant melanoma cell lines and 3 normal melanocyte controls using recently developed hMitChip3 cDNA microarrays. Unsupervised hierarchical clustering analysis of 1,037 informative genes, and 84 survival-apoptosis genes, classified these malignant melanoma cell lines into type A (n = 12) and type B (n = 9). Three hundred fifty-five of 1,037 (34.2%) genes displayed significant (P ≤ 0.030; false discovery rate ≤ 3.68%) differences (± ≥ 2.0-fold) in average expression, with 197 genes higher and 158 genes lower in type A than in type B. Of 84 genes with known survival-apoptosis functions, 38 (45.2%) displayed the significant (P genes expressed at higher levels in type A than in type B, whereas the different set of antiapoptotic (PSEN1, PPP2CA, API5, PPP2R1B, PPP2R1A, and FIS1), antioxidant (HSPD1, GSS, SOD1, ATOX1, and CAT), and proapoptotic (ENDOG, BAK1, CASP2, CASP4, PDCD5, HTRA2, SEPT4, TNFSF10, and PRODH) genes expressed at lower levels in type A than in type B. Microarray data were validated by quantitative reverse transcription-PCR. These results showed the presence of two types of malignant melanoma, each with a specific set of dysregulated survival-apoptosis genes, which may prove useful for development of new molecular targets for therapeutic intervention and novel diagnostic biomarkers for treatment and prognosis of malignant melanoma.

  12. A Neisseria meningitidis NMB1966 mutant is impaired for invasion of respiratory epithelial cells, survival in human blood and for virulence in vivo.

    Science.gov (United States)

    Li, Ming-Shi; Chow, Noel Y S; Sinha, Sunita; Halliwell, Denise; Finney, Michelle; Gorringe, Andrew R; Watson, Mark W; Kroll, J Simon; Langford, Paul R; Webb, Steven A R

    2009-02-01

    We sought to determine whether NMB1966, encoding a putative ABC transporter, has a role in pathogenesis. Compared to its isogenic wild-type parent strain Neisseria meningitidis MC58, the NMB1966 knockout mutant was less adhesive and invasive for human bronchial epithelial cells, had reduced survival in human blood and was attenuated in a systemic mouse model of infection. The transcriptome of the wild-type and the NMB1966 mutant was compared. The data are consistent with a previous functional assignment of NMB1966 being the ABC transporter component of a glutamate transporter operon. Forty-seven percent of all the differentially regulated genes encoded known outer membrane proteins or pathways generating complex surface structures such as adhesins, peptidoglycan and capsule. The data show that NMB1966 has a role in virulence and that remodelling of the outer membrane and surface/structures is associated with attenuation of the NMB1966 mutant.

  13. Human papillomavirus in head and neck squamous cell carcinoma of unknown primary is a common event and a strong predictor of survival.

    Directory of Open Access Journals (Sweden)

    David Hebbelstrup Jensen

    Full Text Available BACKGROUND: The purpose of this study was to examine the prevalence of human papillomavirus (HPV in patients with head and neck squamous cell carcinoma of unknown primary (CUP. METHODS: All patients diagnosed with and treated for CUP between January 1, 2000, and June 1, 2011, at two Danish medical centers were included. All patients received a thorough diagnostic work-up, including FDG-PET, before being diagnosed as CUP. We determined the HPV status in all patients using a combination of HPV DNA PCR and p16 stain. In addition, clinical information on the study patients was retrieved from clinical records. RESULTS: Of the identified 60 patients with CUP, 13 were shown to be positive for HPV DNA, amounting to 22% of the study population. In addition, we were able to show a clear disease-free and overall-survival benefit in the HPV-positive group, with a hazard ratio of 0.16 (95% CI: 0.038-0.67 for over-all survival. This survival benefit was also apparent when adjusted for advanced age in a multivariate Cox regression analysis. CONCLUSION: A fairly large percentage of CUP cases are HPV-related, and because this is related to both the location and prognosis, we recommend HPV testing as part of the diagnostic work-up.

  14. Effects of platelet-rich plasma, adipose-derived stem cells, and stromal vascular fraction on the survival of human transplanted adipose tissue.

    Science.gov (United States)

    Kim, Deok-Yeol; Ji, Yi-Hwa; Kim, Deok-Woo; Dhong, Eun-Sang; Yoon, Eul-Sik

    2014-11-01

    Traditional adipose tissue transplantation has unpredictable viability and poor absorption rates. Recent studies have reported that treatment with platelet-rich plasma (PRP), adipose-derived stem cells (ASCs), and stromal vascular fraction (SVF) are related to increased survival of grafted adipose tissue. This study was the first simultaneous comparison of graft survival in combination with PRP, ASCs, and SVF. Adipose tissues were mixed with each other, injected subcutaneously into the back of nude mice, and evaluated at 4, 8, and 12 weeks. Human adipocytes were grossly maintained in the ASCs and SVF mixtures. Survival of the adipose tissues with PRP was observed at 4 weeks and with SVF at 8 and 12 weeks. At 12 weeks, volume reduction in the ASCs and SVF mixtures were 36.9% and 32.1%, respectively, which were significantly different from that of the control group without adjuvant treatment, 51.0%. Neovascular structures were rarely observed in any of the groups. Our results suggest that the technique of adding ASCs or SVF to transplanted adipose tissue might be more effective than the conventional grafting method. An autologous adipose tissue graft in combination with ASCs or SVF may potentially contribute to stabilization of engraftment.

  15. Resveratrol triggers the pro-apoptotic endoplasmic reticulum stress response and represses the pro-survival XBP1 signaling in human multiple myeloma cells

    Science.gov (United States)

    Wang, Feng-Ming; Galson, Deborah L.; Roodman, G. David; Ouyang, Hongjiao

    2012-01-01

    Objective Resveratrol, trans-3, 4’, 5,-trihydroxystilbene, suppresses multiple myeloma (MM). The endoplasmic reticulum (ER) stress response component IRE1α/XBP1 axis is essential for MM pathogenesis. We investigated the molecular action of resveratrol on IRE1α/XBP1 axis in human MM cells. Methods Human MM cell lines ANBL-6, OPM2, and MM.1S were utilized to determine the molecular signaling events following the treatment with resveratrol. The stimulation of IRE1α/XBP1 axis was analyzed by Western blot and reverse transcription polymerase chain reaction. The effect of resveratrol on the transcriptional activity of spliced XBP1 was assessed by luciferase assays. Chromatin immunoprecipitation (ChIP) was performed to determine the effects of resveratrol on the DNA binding activity of XBP1 in MM cells. Results Resveratrol activated IRE1α as evidenced by XBP1 mRNA splicing and the phosphorylation of both IRE1α and its downstream kinase JNK in MM cells. These responses were associated with resveratrol-induced cytotoxicity of MM cells. Resveratrol selectively suppressed the transcriptional activity of XBP1s while it stimulated gene expression of the molecules that are regulated by non-IRE1/XBP1 axis of the ER stress response. Luciferase assays indicated that resveratrol suppressed the transcriptional activity of XBP1s through sirtuin 1 (SIRT1), a downstream molecular target of resveratrol. ChIP studies revealed that resveratrol decreased the DNA binding capacity of XBP1 and increased the enrichment of SIRT1 at the XBP1 binding region in the XBP1 promoter. Conclusion Resveratrol exerts its chemotherapeutic effect on human MM cells through mechanisms involving the impairment of the pro-survival XBP1 signaling and the activation of pro-apoptotic ER stress response. PMID:21723843

  16. In situ normoxia enhances survival and proliferation rate of human adipose tissue-derived stromal cells without increasing the risk of tumourigenesis.

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    Jane Ru Choi

    Full Text Available Adipose tissue-derived stromal cells (ASCs natively reside in a relatively low-oxygen tension (i.e., hypoxic microenvironment in human body. Low oxygen tension (i.e., in situ normoxia, has been known to enhance the growth and survival rate of ASCs, which, however, may lead to the risk of tumourigenesis. Here, we investigated the tumourigenic potential of ASCs under their physiological condition to ensure their safe use in regenerative therapy. Human ASCs isolated from subcutaneous fat were cultured in atmospheric O2 concentration (21% O2 or in situ normoxia (2% O2. We found that ASCs retained their surface markers, tri-lineage differentiation potential, and self-renewal properties under in situ normoxia without altering their morphology. In situ normoxia displayed a higher proliferation and viability of ASCs with less DNA damage as compared to atmospheric O2 concentration. Moreover, low oxygen tension significantly up-regulated VEGF and bFGF mRNA expression and protein secretion while reducing the expression level of tumour suppressor genes p16, p21, p53, and pRb. However, there were no significant differences in ASCs telomere length and their relative telomerase activity when cultured at different oxygen concentrations. Collectively, even with high proliferation and survival rate, ASCs have a low tendency of developing tumour under in situ normoxia. These results suggest 2% O2 as an ideal culture condition for expanding ASCs efficiently while maintaining their characteristics.

  17. Genome-wide studies reveal that Lin28 enhances the translation of genes important for growth and survival of human embryonic stem cells.

    Science.gov (United States)

    Peng, Shuping; Chen, Ling-Ling; Lei, Xin-Xiang; Yang, Li; Lin, Haifan; Carmichael, Gordon G; Huang, Yingqun

    2011-03-01

    Lin28 inhibits the expression of let-7 microRNAs but also exhibits let-7-independent functions. Using immunoprecipitation and deep sequencing, we show here that Lin28 preferentially associates with a small subset of cellular mRNAs. Of particular interest are those for ribosomal proteins and metabolic enzymes, the expression levels of which are known to be coupled to cell growth and survival. Polysome profiling and reporter analyses suggest that Lin28 stimulates the translation of many or most of these targets. Moreover, Lin28-responsive elements were found within the coding regions of all target genes tested. Finally, a mutant Lin28 that still binds RNA but fails to interact with RNA helicase A (RHA), acts as a dominant-negative inhibitor of Lin28-dependent stimulation of translation. We suggest that Lin28, working in concert with RHA, enhances the translation of genes important for the growth and survival of human embryonic stem cells. Copyright © 2011 AlphaMed Press.

  18. Inositol Hexaphosphate Down-regulates both Constitutive and Ligand-Induced Mitogenic and Cell Survival Signaling, and Causes Caspase-Mediated Apoptotic Death of Human Prostate Carcinoma PC-3 cells

    Science.gov (United States)

    Gu, Mallikarjuna; Raina, Komal; Agarwal, Chapla; Agarwal, Rajesh

    2009-01-01

    Constitutively active mitogenic and pro-survival signaling cascades due to aberrant expression and interaction of growth factors and their receptors are well documented in human prostate cancer (PCa). EGF and IGF-1 are potent mitogens that regulate proliferation and survival of PCa cells via autocrine and paracrine loops involving both MAPK- and Akt-mediated signaling. Accordingly, here we assessed the effect of inositol hexaphosphate (IP6) on constitutive and ligand (EGF and IGF-1)-induced biological responses and associated signaling cascades in advanced and androgen-independent human PCa PC-3 cells. Treatment of PC-3 cells with 2 mM IP6 strongly inhibited both growth and proliferation and decreased cell viability; similar effects were also observed in other human PCa DU145 and LNCaP cells. IP6 also caused a strong apoptotic death of PC-3 cells together with caspase 3 and PARP cleavage. Mechanistic studies showed that biological effects of IP6 were associated with inhibition of both constitutive and ligand-induced Akt phosphorylation together with a decrease in total Akt levels, but a differential inhibitory effect on MAPKs ERK1/2, JNK1/2 and p38 under constitutive and ligand-activated conditions. Under similar condition, IP6 also inhibited AP-1 DNA binding activity and decreased nuclear levels of both phospho and total c-Fos and c-Jun. Together, these findings for the first time establish IP6 efficacy in inhibiting aberrant EGFR or IGF-1R pathway-mediated sustained growth promoting and survival signaling cascades in advanced and androgen-independent human PCa PC-3 cells, which might have translational implications in advanced human PCa control and management. PMID:19544333

  19. EphrinA4 plays a critical role in α4 and αL mediated survival of human CLL cells during extravasation

    Science.gov (United States)

    Trinidad, Eva M.; García, Dolores; Soler, Gloria; Ortuño, Francisco J.; Zapata, Agustín G.; Alonso, Luis M.

    2016-01-01

    A role of endothelial cells in the survival of CLL cells during extravasation is presently unknown. Herein we show that CLL cells but not normal B cells can receive apoptotic signals through physical contact with TNF-α activated endothelium impairing survival in transendothelial migration (TEM) assays. In addition, the CLL cells of patients having lymphadenopathy (LApos) show a survival advantage during TEM that can be linked to increased expression of α4 and αL integrin chains. Within this context, ephrinA4 expressed on the surface of CLL cells sequestrates integrins and inactivates them resulting in reduced adhesion and inhibition of apoptotic/survival signals through them. In agreement, ephrinA4 silencing resulted in increased survival of CLL cells of LApos patients but not LA neg patients. Similarly was observed when a soluble ephrinA4 isoform was added to TEM assays strongly suggesting that accumulation of this isoform in the serum of LApos patients could contribute to CLL cells dissemination and survival in vivo. In supporting, CLL lymphadenopathies showed a preferential accumulation of apoptotic CLL cells around high endothelial venules lacking ephrinA4. Moreover, soluble ephrinA4 isolated from sera of patients increased the number and viability of CLL cells recovered from the lymph nodes of adoptively transferred mice. Finally, we present evidence suggesting that soluble ephrinA4 mediated survival during TEM could enhance a transcellular TEM route of the CLL cells. Together these findings point to an important role of ephrinA4 in the nodal dissemination of CLL cells governing extravasation and survival. PMID:27374180

  20. Triiodothyronine regulates cell growth and survival in renal cell cancer.

    Science.gov (United States)

    Czarnecka, Anna M; Matak, Damian; Szymanski, Lukasz; Czarnecka, Karolina H; Lewicki, Slawomir; Zdanowski, Robert; Brzezianska-Lasota, Ewa; Szczylik, Cezary

    2016-10-01

    Triiodothyronine plays an important role in the regulation of kidney cell growth, differentiation and metabolism. Patients with renal cell cancer who develop hypothyreosis during tyrosine kinase inhibitor (TKI) treatment have statistically longer survival. In this study, we developed cell based model of triiodothyronine (T3) analysis in RCC and we show the different effects of T3 on renal cell cancer (RCC) cell growth response and expression of the thyroid hormone receptor in human renal cell cancer cell lines from primary and metastatic tumors along with human kidney cancer stem cells. Wild-type thyroid hormone receptor is ubiquitously expressed in human renal cancer cell lines, but normalized against healthy renal proximal tube cell expression its level is upregulated in Caki-2, RCC6, SKRC-42, SKRC-45 cell lines. On the contrary the mRNA level in the 769-P, ACHN, HKCSC, and HEK293 cells is significantly decreased. The TRβ protein was abundant in the cytoplasm of the 786-O, Caki-2, RCC6, and SKRC-45 cells and in the nucleus of SKRC-42, ACHN, 769-P and cancer stem cells. T3 has promoting effect on the cell proliferation of HKCSC, Caki-2, ASE, ACHN, SK-RC-42, SMKT-R2, Caki-1, 786-0, and SK-RC-45 cells. Tyrosine kinase inhibitor, sunitinib, directly inhibits proliferation of RCC cells, while thyroid hormone receptor antagonist 1-850 (CAS 251310‑57-3) has less significant inhibitory impact. T3 stimulation does not abrogate inhibitory effect of sunitinib. Renal cancer tumor cells hypostimulated with T3 may be more responsive to tyrosine kinase inhibition. Moreover, some tumors may be considered as T3-independent and present aggressive phenotype with thyroid hormone receptor activated independently from the ligand. On the contrary proliferation induced by deregulated VHL and or c-Met pathways may transgress normal T3 mediated regulation of the cell cycle.

  1. R132H mutation in IDH1 gene reduces proliferation, cell survival and invasion of human glioma by downregulating Wnt/β-catenin signaling.

    Science.gov (United States)

    Cui, Daming; Ren, Jie; Shi, Jinlong; Feng, Lijing; Wang, Ke; Zeng, Tao; Jin, Yi; Gao, Liang

    2016-04-01

    Mutations in the isocitrate dehydrogenase 1 (IDH1) gene commonly occur in gliomas. Remarkably, the R132H mutation in IDH1 (IDH1-R132H) is associated with better prognosis and increased survival than patients lacking this mutation. The molecular mechanism underlying this phenomenon is largely unknown. In this study, we investigated potential cross-talk between IDH1-R132H and Wnt/β-catenin signaling in regulating the cellular properties of human glioma. Although aberrant nuclear accumulation of β-catenin is linked to the malignant progression of gliomas, its association with IDH1 remains unknown. We identified an inverse correlation between IDH1-R132H and the expression and activity of β-catenin in human gliomas. In addition, overexpression of IDH1-R132H in glioblastoma cell lines U87 and U251 led to reduced cell proliferation, migration and invasion, accompanied by increased apoptosis. At the molecular level, we detected a significant reduction in the expression, nuclear accumulation and activity of β-catenin following overexpression of IDH1-R132H. A microarray-based comparison of gene expression indicated that several mediators, effectors and targets of Wnt/β-catenin signaling are downregulated, while negative regulators are upregulated in IDH1-R132H gliomas. Further, overexpression of β-catenin in IDH1-R132H glioma cells restored the cellular phenotype induced by this mutation. Specifically, β-catenin abrogated the decrease in proliferation, invasion and migration, and the increase in apoptosis, triggered by overexpression of IDH1-R132H. Finally, we demonstrate that xenografts of IDH1-R132H overexpressing U87 cells can significantly decrease the growth of tumors in vivo. Altogether, our results strongly suggest that the R132H mutation in IDH1 serves a tumor suppressor function in human glioma by negatively regulating Wnt/β-catenin signaling.

  2. Survival and Functionality of Human Induced Pluripotent Stem Cell-Derived Oligodendrocytes in a Nonhuman Primate Model for Multiple Sclerosis

    NARCIS (Netherlands)

    Thiruvalluvan, Arun; Czepiel, Marcin; Kap, Yolanda A; Mantingh-Otter, Ietje; Vainchtein, Ilia; Kuipers, Jeroen; Bijlard, Marjolein; Baron, Wia; Giepmans, Ben; Brück, Wolfgang; 't Hart, Bert A; Boddeke, Erik; Copray, Sjef

    2016-01-01

    : Fast remyelination by endogenous oligodendrocyte precursor cells (OPCs) is essential to prevent axonal and subsequent retrograde neuronal degeneration in demyelinating lesions in multiple sclerosis (MS). In chronic lesions, however, the remyelination capacity of OPCs becomes insufficient. Cell the

  3. Inhibitory Effects of Salinomycin on Cell Survival, Colony Growth, Migration, and Invasion of Human Non-Small Cell Lung Cancer A549 and LNM35: Involvement of NAG-1.

    Science.gov (United States)

    Arafat, Kholoud; Iratni, Rabah; Takahashi, Takashi; Parekh, Khatija; Al Dhaheri, Yusra; Adrian, Thomas E; Attoub, Samir

    2013-01-01

    A major challenge for oncologists and pharmacologists is to develop more potent and less toxic drugs that will decrease the tumor growth and improve the survival of lung cancer patients. Salinomycin is a polyether antibiotic used to kill gram-positive bacteria including mycobacteria, protozoans such as plasmodium falciparum, and the parasites responsible for the poultry disease coccidiosis. This old agent is now a serious anti-cancer drug candidate that selectively inhibits the growth of cancer stem cells. We investigated the impact of salinomycin on survival, colony growth, migration and invasion of the differentiated human non-small cell lung cancer lines LNM35 and A549. Salinomycin caused concentration- and time-dependent reduction in viability of LNM35 and A549 cells through a caspase 3/7-associated cell death pathway. Similarly, salinomycin (2.5-5 µM for 7 days) significantly decreased the growth of LNM35 and A549 colonies in soft agar. Metastasis is the main cause of death related to lung cancer. In this context, salinomycin induced a time- and concentration-dependent inhibition of cell migration and invasion. We also demonstrated for the first time that salinomycin induced a marked increase in the expression of the pro-apoptotic protein NAG-1 leading to the inhibition of lung cancer cell invasion but not cell survival. These findings identify salinomycin as a promising novel therapeutic agent for lung cancer.

  4. Inhibitory Effects of Salinomycin on Cell Survival, Colony Growth, Migration, and Invasion of Human Non-Small Cell Lung Cancer A549 and LNM35: Involvement of NAG-1.

    Directory of Open Access Journals (Sweden)

    Kholoud Arafat

    Full Text Available A major challenge for oncologists and pharmacologists is to develop more potent and less toxic drugs that will decrease the tumor growth and improve the survival of lung cancer patients. Salinomycin is a polyether antibiotic used to kill gram-positive bacteria including mycobacteria, protozoans such as plasmodium falciparum, and the parasites responsible for the poultry disease coccidiosis. This old agent is now a serious anti-cancer drug candidate that selectively inhibits the growth of cancer stem cells. We investigated the impact of salinomycin on survival, colony growth, migration and invasion of the differentiated human non-small cell lung cancer lines LNM35 and A549. Salinomycin caused concentration- and time-dependent reduction in viability of LNM35 and A549 cells through a caspase 3/7-associated cell death pathway. Similarly, salinomycin (2.5-5 µM for 7 days significantly decreased the growth of LNM35 and A549 colonies in soft agar. Metastasis is the main cause of death related to lung cancer. In this context, salinomycin induced a time- and concentration-dependent inhibition of cell migration and invasion. We also demonstrated for the first time that salinomycin induced a marked increase in the expression of the pro-apoptotic protein NAG-1 leading to the inhibition of lung cancer cell invasion but not cell survival. These findings identify salinomycin as a promising novel therapeutic agent for lung cancer.

  5. Down-regulation of c9orf86 in human breast cancer cells inhibits cell proliferation, invasion and tumor growth and correlates with survival of breast cancer patients.

    Directory of Open Access Journals (Sweden)

    Yang-Yang Li

    Full Text Available C9orf86 which is a novel subfamily within the Ras superfamily of GTPases, is overexpressed in the majority of primary breast tumors. Few functional studies have focused on the C9orf86 protein; therefore, in this study, we explored the role of C9orf86 in breast carcinogenesis. In our study, we found that silencing of C9orf86 by siRNA in MCF-7 and SK-BR-3 cells resulted in suppressed cell proliferation as well as in vitro cell invasion capabilities. Moreover, knockdown of C9orf86 inhibited tumor growth in nude mice. Cell cycle and apoptotic assays showed that the anti-proliferative effect of C9orf86-siRNA was mediated by arresting cells in the G1 phase and promoting apoptosis. In addition, we found that patients with high levels of C9orf86 expression showed a significant trend towards worse survival compared to patients with low C9orf86 expression (P = 0.002. These results provide evidence that C9orf86 represents a novel and clinically useful biomarker for BC patients and plays an important role during the progression of BC.

  6. Computational Modeling of Cell Survival Using VHDL

    Directory of Open Access Journals (Sweden)

    Shruti Jain1,

    2010-01-01

    Full Text Available The model for cell survival has been implemented using VeryHigh Speed Integrated Circuit Hardware DescriptionLanguage (VHDL (Xilinx Tool taking three input signals:Tumor necrosis factor-α (TNF, Epidermal growth factor(EGF and Insulin. Cell survival has been regulated by theinteraction of five proteins viz P13K, TNFR1, EGFR, IRS andIKK in a network. In the absence of any one, in protein networkleads to cell death. For the EGF input signal the proteins likeMEK, ERK, AkT, Rac & JNK have been important forregulation of cell survival. Similarly for TNF and Insulin inputsignal proteins like NFκB, AkT, XIAP, JNK, MAP3K & MK2and MEK, ERK, AkT, Rac, mTOR & JNK respectively havebeen important for regulation of cell survival.

  7. Human Müller stem cell (MIO-M1) transplantation in a rat model of glaucoma: survival, differentiation, and integration.

    Science.gov (United States)

    Bull, Natalie D; Limb, G Astrid; Martin, Keith R

    2008-08-01

    Stem cell transplantation is a potential treatment strategy for neurodegenerative diseases such as glaucoma. The Müller stem cell line MIO-M1 can be differentiated to produce retinal neurons and glia. The survival, migration, differentiation, and integration of MIO-M1 cells were investigated in a rat model of glaucoma. The effect of modulating the retinal environment with either chondroitinase ABC or erythropoietin was also studied. Intraocular pressure was chronically increased unilaterally by using a laser glaucoma model in adult rats. EGFP-transduced MIO-M1 cells were transplanted into the vitreous or subretinal space of glaucomatous or untreated eyes. Oral immune suppressants were administered to reduce xenograft rejection. Survival, migration, differentiation, and integration of grafted cells were assessed by immunohistochemistry. Transplanted cells survived for 2 to 3 weeks in vivo, although microglia/macrophage infiltration and a reduction in graft survival were seen by 4 weeks. Grafted cells displayed a migratory phenotype with an elongated bipolar shape often oriented toward the retina. Transplanted cells expressed markers such as PSA-NCAM, GFAP, and beta-III-tubulin. The host retina was resistant to MIO-M1 migration, but modification of the local environment with erythropoietin or chondroitinase ABC facilitated retinal infiltration by MIO-M1 cells. The results demonstrate that differentiating MIO-M1 cells within the glaucomatous eye produced cells that expressed neuronal and glial cell markers. The retina was relatively resistant to transplant integration, and long-term xenograft survival was limited. However, local modulation of the retinal environment enhanced the integration of MIO-M1 cells into the glaucomatous retina.

  8. Small ubiquitin-like modifier 1-3 conjugation [corrected] is activated in human astrocytic brain tumors and is required for glioblastoma cell survival.

    Science.gov (United States)

    Yang, Wei; Wang, Liangli; Roehn, Gabriele; Pearlstein, Robert D; Ali-Osman, Francis; Pan, Hongjie; Goldbrunner, Roland; Krantz, Matthew; Harms, Christoph; Paschen, Wulf

    2013-01-01

    Small ubiquitin-like modifier (SUMO1-3) constitutes a group of proteins that conjugate to lysine residues of target proteins thereby modifying their activity, stability, and subcellular localization. A large number of SUMO target proteins are transcription factors and other nuclear proteins involved in gene expression. Furthermore, SUMO conjugation plays key roles in genome stability, quality control of newly synthesized proteins, proteasomal degradation of proteins, and DNA damage repair. Any marked increase in levels of SUMO-conjugated proteins is therefore expected to have a major impact on the fate of cells. We show here that SUMO conjugation is activated in human astrocytic brain tumors. Levels of both SUMO1- and SUMO2/3-conjugated proteins were markedly increased in tumor samples. The effect was least pronounced in low-grade astrocytoma (WHO Grade II) and most pronounced in glioblastoma multiforme (WHO Grade IV). We also found a marked rise in levels of Ubc9, the only SUMO conjugation enzyme identified so far. Blocking SUMO1-3 conjugation in glioblastoma cells by silencing their expression blocked DNA synthesis, cell growth, and clonogenic survival of cells. It also resulted in DNA-dependent protein kinase-induced phosphorylation of H2AX, indicative of DNA double-strand damage, and G(2) /M cell cycle arrest. Collectively, these findings highlight the pivotal role of SUMO conjugation in DNA damage repair processes and imply that the SUMO conjugation pathway could be a new target of therapeutic intervention aimed at increasing the sensitivity of glioblastomas to radiotherapy and chemotherapy.

  9. PATZ1 expression correlates positively with BAX and negatively with BCL6 and survival in human diffuse large B cell lymphomas

    Science.gov (United States)

    Valentino, Elena; Vitiello, Michela; Luciano, Antonio; Palma, Giuseppe; Arra, Claudio; Mantia, Elvira La; Panico, Luigi; Tenneriello, Valentina; Pinto, Antonello; Frigeri, Ferdinando; Capobianco, Gaetana; Botti, Gerardo; Cerchia, Laura; De Chiara, Annarosaria; Fedele, Monica

    2016-01-01

    Non-Hodgkin lymphomas (NHLs) include a heterogeneous group of diseases, which differ in both cellular origin and clinical behavior. Among the aggressive malignancies of this group, the diffuse large B-cell lymphomas (DLBCLs) are the most frequently observed. They are themselves clinically and molecularly heterogeneous and have been further sub-divided in three sub-types according to different cell of origin, mechanisms of oncogenesis and clinical outcome. Among them, the germinal center B-cell-like (GCB) derives from the germinal center and expresses the BCL6 oncogene. We have previously shown that Patz1-knockout mice develop B-cell neoplasias, suggesting a tumor suppressor role for PATZ1 in human NHLs. Here, by immunohistochemical analysis of a tissue-microarray including 170 NHLs, we found that PATZ1 nuclear expression is down-regulated in follicular lymphomas and DLBCLs. Moreover, consistent with our previous results showing a PATZ1-dependent regulation of BCL6 and BAX transcription, we show that low PATZ1 nuclear expression significantly correlates with high BCL6 expression, mainly in DLBCLs, and with low BAX expression, also considering separately follicular lymphomas and DLBCLs. Finally, by analyzing overall and progression-free survival in DLBCL patients that underwent rituximab plus cyclophosphamide, doxorubicin, vincristine, and prednisone chemotherapy, low levels of PATZ1 were significantly associated to a worst outcome and demonstrated an independent prognostic factor in multivariate analysis, including known prognostic factors of DLBCL, IPI score and cell of origin (GCB/non-GCB). Therefore, we propose PATZ1 as a new prognostic marker of DLBCLs, which may act as a tumor suppressor by enhancing apoptosis through inhibiting and enhancing transcription of BCL6 and BAX, respectively. PMID:27494852

  10. Gene Network Analysis of Glucose Linked Signaling Pathways and Their Role in Human Hepatocellular Carcinoma Cell Growth and Survival in HuH7 and HepG2 Cell Lines

    Science.gov (United States)

    Berger, Emmanuelle; Vega, Nathalie; Weiss-Gayet, Michèle; Géloën, Alain

    2015-01-01

    Cancer progression may be affected by metabolism. In this study, we aimed to analyze the effect of glucose on the proliferation and/or survival of human hepatocellular carcinoma (HCC) cells. Human gene datasets regulated by glucose were compared to gene datasets either dysregulated in HCC or regulated by other signaling pathways. Significant numbers of common genes suggested putative involvement in transcriptional regulations by glucose. Real-time proliferation assays using high (4.5 g/L) versus low (1 g/L) glucose on two human HCC cell lines and specific inhibitors of selected pathways were used for experimental validations. High glucose promoted HuH7 cell proliferation but not that of HepG2 cell line. Gene network analyses suggest that gene transcription by glucose could be mediated at 92% through ChREBP in HepG2 cells, compared to 40% in either other human cells or rodent healthy liver, with alteration of LKB1 (serine/threonine kinase 11) and NOX (NADPH oxidases) signaling pathways and loss of transcriptional regulation of PPARGC1A (peroxisome-proliferator activated receptors gamma coactivator 1) target genes by high glucose. Both PPARA and PPARGC1A regulate transcription of genes commonly regulated by glycolysis, by the antidiabetic agent metformin and by NOX, suggesting their major interplay in the control of HCC progression. PMID:26380295

  11. Survival of transplanted neurotrophin-3 expressing human neural stem cells and motor function in a rat model of spinal cord injury

    Institute of Scientific and Technical Information of China (English)

    Peiqiang Cai; Guangyun Sun; Peishu Cai; Martin Oudega; Rui Xiao; Xuewen Wang; Wei Li; Yunbing Shu; Cheng Cai; Haihao Yang; Xuebing Shan; Wuhua Luo

    2009-01-01

    BACKGROUND: Many methods have been attempted to repair nerves following spinal cord injury,including peripheral nerve transplantation, Schwann cell transplantation, olfactory ensheathing cell transplantation, and embryonic neural tissue transplantation. However, there is a need for improved outcomes.OBJECTIVE: To investigate the repair feasibility for rat spinal cord injury using human neural stem cells (hNSCs) genetically modified by lentivirus to express neurotrophin-3.DESIGN, TIME AND SETTING: In vitro cell biological experiment and in vivo randomized, controlled,genetic engineering experiment were performed at the Third Military Medical University of Chinese PLA and First People's Hospital of Yibin, China from March 2006 to December 2007.MATERIALS: A total of 64 adult, female, Wistar rats were used for the in vivo study. Of them, 48 rats were used to establish models of spinal cord hemisection, and were subsequently equally and randomly assigned to model, genetically modified hNSC, and normal hNSC groups. The remaining 16 rats served as normal controls.METHODS: hNSCs were in vitro genetically modified by lentivirus to secrete both green fluorescence protein and neurotrophin-3. Neurotrophin-3 expression was measured by Westem blot.Genetically modified hNSC or normal hNSC suspension (5×105) was injected into the rat spinal cord following T10 spinal cord hemisection. A total of 5μL Dulbecco's-modified Eagle's medium was infused into the rat spinal cord in the model group. Transgene expression and survival of transplanted hNSCs were determined by immunohistochemistry. Motor function was evaluatedusing the Basso, Beattie, and Bresnahan (BBB) scale.MAIN OUTCOME MEASURES: The following parameters were measured: expression ofneurotrophin-3 produced by genetically modified hNSCs, transgene expression and survival ofhNSCs in rats, motor function in rats.RESULTS: hNSCs were successfully genetically modified by lentivirus to stably express neurotrophin-3. The

  12. Human Leukocyte Antigen Class I and II Alleles and Overall Survival in Diffuse Large B-Cell Lymphoma and Follicular Lymphoma

    Directory of Open Access Journals (Sweden)

    Yani Lu

    2011-01-01

    Full Text Available Genetic variation in the 6p21 chromosomal region, including human leukocyte antigen (HLA genes and tumor necrosis factor (TNF, has been linked to both etiology and clinical outcomes of lymphomas. We estimated the effects of HLA class I (A, B, and C, class II DRB1 alleles, and the ancestral haplotype (AH 8.1 (HLAA*01-B*08-DRB1*03-TNF-308A on overall survival (OS among patients with diffuse large B-cell lymphoma (DLBCL and follicular lymphoma (FL in a population-based study of non-Hodgkin lymphoma. During a median followup of 89 months, 31% (52 of 166 DLBCL and 28% (46 of 165 FL patients died. Using multivariate Cox regression models, we observed statistically significant associations between genetic variants and survival: HLA-Cw*07:01 was associated with poorer OS among DLBCL patients (Hazard ratio [HR] = 1.76, 95% confidence interval [CI] = 1.01–3.05; HLA-A*01:01 was associated with poorer OS (HR = 2.23, 95% CI = 1.24–4.01, and HLA-DRB1*13 (HR = 0.12, 95% CI = 0.02–0.90 and HLA-B Bw4 (HR = 0.36, 95% CI = 0.20–0.63 with better OS among FL patients. These results support a role for HLA in the prognosis of DLBCL and FL and represent a promising class of prognostic factors that warrants further evaluation.

  13. Staphylococcus aureus survival in human blood.

    Science.gov (United States)

    Malachowa, Natalia; DeLeo, Frank R

    2011-01-01

    Methicillin-resistant Staphylococcus aureus (MRSA) is abundant in hospitals and in the United States is a leading cause of mortality due to infectious agents. Community-associated MRSA (CA-MRSA) strains such as USA300, which typically cause disease outside of healthcare settings, are also prevalent in the United States. Although most CA-MRSA infections affect skin and soft tissue, the pathogen can enter the bloodstream and ultimately cause severe disease. In a recent paper, we used USA300-specific microarrays to generate a comprehensive view of the molecules that facilitate S. aureus immune evasion and survival in human blood. Notably, genes encoding proteins involved in iron-uptake and utilization and gamma-hemolysin (hlgABC) are highly up-regulated by USA300 during culture in human blood. Here we discuss the potential implication of these findings and the possible role of gamma-hemolysin in the success of S. aureus as a human pathogen.

  14. In vitro cytotoxicity of {sup 211}At-labeled trastuzumab in human breast cancer cell lines: effect of specific activity and HER2 receptor heterogeneity on survival fraction

    Energy Technology Data Exchange (ETDEWEB)

    Akabani, Gamal [Department of Radiology, Duke University Medical Center, P.O. Box 3808, Durham, NC 27710 (United States); Carlin, Sean [Department of Radiology, Duke University Medical Center, P.O. Box 3808, Durham, NC 27710 (United States); Welsh, Phil [Department of Radiology, Duke University Medical Center, P.O. Box 3808, Durham, NC 27710 (United States); Zalutsky, Michael R. [Department of Radiology, Duke University Medical Center, P.O. Box 3808, Durham, NC 27710 (United States)]. E-mail: zalut001@mc.duke.edu

    2006-04-15

    Introduction: Radioimmunotherapy with anti-HER2 monoclonal antibodies (mAbs) such as trastuzumab is a promising strategy for treating HER2-positive breast and ovarian carcinoma patients. The objective of this study was to determine the cytotoxic effectiveness of trastuzumab labeled with the 7.2-h half-life {alpha}-particle emitter {sup 211}At. Methods: Experiments were performed on SKBr-3, BT-474 and the transfected MCF7/HER2-18 human breast carcinoma cell lines. Intrinsic radiosensitivity was determined after exposure to external beam irradiation. The cytotoxicity of {sup 211}At-labeled trastuzumab was measured by clonogenic assays. The distribution of HER2 receptor expression on the cell lines was measured using fluorescence-activated cell sorting. A pharmacokinetic (PK)/microdosimetric model was established to assess the effects of specific activity (SA), HER2 receptor expression and absorbed dose on survival fraction (SF). Results: With external beam irradiation, the 2-Gy SF for BT-474, SKBr-3 and MCF7/HER2-18 cells was 0.78, 0.53 and 0.64 Gy, respectively. Heterogeneous HER2 expression was observed, with a subpopulation of cells lacking measurable receptor (14.5%, SKBr-3; 0.34%, MCF-7/HER2; 1.73%, BT-474). When plotted as a function of activity concentration, SF curves were biphasic and inversely proportional to SA; however, when the model was applied and absorbed doses calculated, the SF curve was monoexponential independent of SA. Thus, the PK model was able to demonstrate the effects of competition between cold and labeled mAb. These studies showed that the relative biological effectiveness of {sup 211}At-labeled trastuzaumab was about 10 times higher than that of external beam therapy. Conclusion: These in vitro studies showed that {sup 211}At-labeled trastuzumab mAb is an effective cytotoxic agent for the treatment of HER2-positive tumor cells. The SA of the labeled mAb and the homogeneity of HER2 receptor expression are important variables influencing

  15. Corynebacterium diphtheriae putative tellurite-resistance protein (CDCE8392_0813 contributes to the intracellular survival in human epithelial cells and lethality of Caenorhabditis elegans

    Directory of Open Access Journals (Sweden)

    Louisy Sanches dos Santos

    2015-01-01

    Full Text Available Corynebacterium diphtheriae, the aetiologic agent of diphtheria, also represents a global medical challenge because of the existence of invasive strains as causative agents of systemic infections. Although tellurite (TeO32- is toxic to most microorganisms, TeO32--resistant bacteria, including C. diphtheriae, exist in nature. The presence of TeO32--resistance (TeR determinants in pathogenic bacteria might provide selective advantages in the natural environment. In the present study, we investigated the role of the putative TeR determinant (CDCE8392_813 gene in the virulence attributes of diphtheria bacilli. The disruption of CDCE8392_0813 gene expression in the LDCIC-L1 mutant increased susceptibility to TeO32- and reactive oxygen species (hydrogen peroxide, but not to other antimicrobial agents. The LDCIC-L1 mutant also showed a decrease in both the lethality of Caenorhabditis elegans and the survival inside of human epithelial cells compared to wild-type strain. Conversely, the haemagglutinating activity and adherence to and formation of biofilms on different abiotic surfaces were not regulated through the CDCE8392_0813 gene. In conclusion, the CDCE8392_813 gene contributes to the TeR and pathogenic potential of C. diphtheriae.

  16. Corynebacterium diphtheriae putative tellurite-resistance protein (CDCE8392_0813) contributes to the intracellular survival in human epithelial cells and lethality of Caenorhabditis elegans.

    Science.gov (United States)

    Santos, Louisy Sanches Dos; Antunes, Camila Azevedo; Santos, Cintia Silva Dos; Pereira, José Augusto Adler; Sabbadini, Priscila Soares; Luna, Maria das Graças de; Azevedo, Vasco; Hirata Júnior, Raphael; Burkovski, Andreas; Asad, Lídia Maria Buarque de Oliveira; Mattos-Guaraldi, Ana Luíza

    2015-08-01

    Corynebacterium diphtheriae, the aetiologic agent of diphtheria, also represents a global medical challenge because of the existence of invasive strains as causative agents of systemic infections. Although tellurite (TeO32-) is toxic to most microorganisms, TeO32--resistant bacteria, including C. diphtheriae, exist in nature. The presence of TeO32--resistance (TeR) determinants in pathogenic bacteria might provide selective advantages in the natural environment. In the present study, we investigated the role of the putative TeR determinant (CDCE8392_813gene) in the virulence attributes of diphtheria bacilli. The disruption of CDCE8392_0813 gene expression in the LDCIC-L1 mutant increased susceptibility to TeO32- and reactive oxygen species (hydrogen peroxide), but not to other antimicrobial agents. The LDCIC-L1 mutant also showed a decrease in both the lethality of Caenorhabditis elegans and the survival inside of human epithelial cells compared to wild-type strain. Conversely, the haemagglutinating activity and adherence to and formation of biofilms on different abiotic surfaces were not regulated through the CDCE8392_0813 gene. In conclusion, the CDCE8392_813 gene contributes to the TeR and pathogenic potential of C. diphtheriae.

  17. Corynebacterium diphtheriae putative tellurite-resistance protein (CDCE8392_0813 contributes to the intracellular survival in human epithelial cells and lethality of Caenorhabditis elegans

    Directory of Open Access Journals (Sweden)

    Louisy Sanches dos Santos

    2015-08-01

    Full Text Available Corynebacterium diphtheriae, the aetiologic agent of diphtheria, also represents a global medical challenge because of the existence of invasive strains as causative agents of systemic infections. Although tellurite (TeO32- is toxic to most microorganisms, TeO32--resistant bacteria, including C. diphtheriae, exist in nature. The presence of TeO32--resistance (TeR determinants in pathogenic bacteria might provide selective advantages in the natural environment. In the present study, we investigated the role of the putative TeR determinant (CDCE8392_813gene in the virulence attributes of diphtheria bacilli. The disruption of CDCE8392_0813 gene expression in the LDCIC-L1 mutant increased susceptibility to TeO32- and reactive oxygen species (hydrogen peroxide, but not to other antimicrobial agents. The LDCIC-L1 mutant also showed a decrease in both the lethality of Caenorhabditis elegansand the survival inside of human epithelial cells compared to wild-type strain. Conversely, the haemagglutinating activity and adherence to and formation of biofilms on different abiotic surfaces were not regulated through the CDCE8392_0813 gene. In conclusion, the CDCE8392_813 gene contributes to the TeR and pathogenic potential of C. diphtheriae.

  18. Enhanced survival effect of pyruvate correlates MAPK and NF-kappaB activation in hydrogen peroxide-treated human endothelial cells.

    Science.gov (United States)

    Lee, Yong-Jin; Kang, Il-Jun; Bünger, Rolf; Kang, Young-Hee

    2004-02-01

    We recently reported that pyruvate inhibited translocation and activation of p53 caused by DNA damage due to oxidant injury (Lee YJ, Kang IJ, Bünger R, and Kang YH. Microvasc Res 66: 91-101, 2003); this was associated with increased expression of apoptosis-related bcl-2 and decreased expression of bax gene. This study attempted to delineate possible regulatory sites and mechanisms of antiapoptotic pyruvate, focusing on reactive oxygen species-mediated signaling in a human umbilical vein endothelial cell model. We compared the effects of the cytosolic reductant l-lactate and malate-aspartate shuttle blocker aminooxyacetate, both of which increase cytosolic NADH, on the downstream signaling pathway. Hydrogen peroxide (0.5 mM H2O2) depleted intracellular total glutathione that was prevented by pyruvate but not by l-lactate or aminooxyacetate. Activation of caspase-3 and the cleavage of procaspase-6 and procaspase-7 were strongly inhibited by pyruvate but markedly enhanced by l-lactate and aminooxyacetate, implicating redox-related antiapoptotic mechanisms of pyruvate. Western blot analysis and immunochemical data revealed that H2O2-induced transactivation of nuclear factor-kappaB (NF-kappaB) was also inhibited by pyruvate but not by l-lactate or aminooxyacetate. In addition, H2O2 downregulated extracellular signal-regulated kinase (ERK1/2) and phosphorylated p38 mitogen-activated protein kinase (MAPK), effects that were fully reversed by pyruvate within 2 h. Collectively, these findings indicate that pyruvate can protect cellular glutathione, thus enhancing cellular antioxidant potential, and that enhanced antioxidant potential can desensitize NF-kappaB transactivation due to reactive oxygen species, suggesting possible metabolic redox relations to NF-kappaB. Furthermore, pyruvate blocked the p38 MAPK pathway and activated the ERK pathway in an apparently redox-sensitive manner, which may regulate expression of genes believed to prevent apoptosis and promote cell

  19. Citicoline induces angiogenesis improving survival of vascular/human brain microvessel endothelial cells through pathways involving ERK1/2 and insulin receptor substrate-1

    Directory of Open Access Journals (Sweden)

    Krupinski Jerzy

    2012-12-01

    Full Text Available Abstract Background Citicoline is one of the neuroprotective agents that have been used as a therapy in stroke patients. There is limited published data describing the mechanisms through which it acts. Methods We used in vitro angiogenesis assays: migration, proliferation, differentiation into tube-like structures in Matrigel™ and spheroid development assays in human brain microvessel endothelial cells (hCMEC/D3. Western blotting was performed on protein extraction from hCMEC/D3 stimulated with citicoline. An analysis of citicoline signalling pathways was previously studied using a Kinexus phospho-protein screening array. A staurosporin/calcium ionophore-induced apoptosis assay was performed by seeding hCMEC/D3 on to glass coverslips in serum poor medium. In a pilot in vivo study, transient MCAO in rats was carried out with and without citicoline treatment (1000 mg/Kg applied at the time of occlusion and subsequently every 3 days until euthanasia (21 days. Vascularity of the stroke-affected regions was examined by immunohistochemistry. Results Citicoline presented no mitogenic and chemotactic effects on hCMEC/D3; however, it significantly increased wound recovery, the formation of tube-like structures in Matrigel™ and enhanced spheroid development and sprouting. Citicoline induced the expression of phospho-extracellular-signal regulated kinase (ERK-1/2. Kinexus assays showed an over-expression of insulin receptor substrate-1 (IRS-1. Knock-down of IRS-1 with targeted siRNA in our hCMEC/D3 inhibited the pro-angiogenic effects of citicoline. The percentage of surviving cells was higher in the presence of citicoline. Citicoline treatment significantly increased the numbers of new, active CD105-positive microvessels following MCAO. Conclusions The findings demonstrate both a pro-angiogenic and protective effect of citicoline on hCMEC/D3 in vitro and following middle cerebral artery occlusion (MCAO in vivo.

  20. Citicoline induces angiogenesis improving survival of vascular/human brain microvessel endothelial cells through pathways involving ERK1/2 and insulin receptor substrate-1.

    Science.gov (United States)

    Krupinski, Jerzy; Abudawood, Manal; Matou-Nasri, Sabine; Al-Baradie, Raid; Petcu, Eugen Bogdan; Justicia, Carlos; Planas, Anna; Liu, Donghui; Rovira, Norma; Grau-Slevin, Marta; Secades, Julio; Slevin, Mark

    2012-12-10

    Citicoline is one of the neuroprotective agents that have been used as a therapy in stroke patients. There is limited published data describing the mechanisms through which it acts. We used in vitro angiogenesis assays: migration, proliferation, differentiation into tube-like structures in Matrigel™ and spheroid development assays in human brain microvessel endothelial cells (hCMEC/D3). Western blotting was performed on protein extraction from hCMEC/D3 stimulated with citicoline. An analysis of citicoline signalling pathways was previously studied using a Kinexus phospho-protein screening array. A staurosporin/calcium ionophore-induced apoptosis assay was performed by seeding hCMEC/D3 on to glass coverslips in serum poor medium. In a pilot in vivo study, transient MCAO in rats was carried out with and without citicoline treatment (1000 mg/Kg) applied at the time of occlusion and subsequently every 3 days until euthanasia (21 days). Vascularity of the stroke-affected regions was examined by immunohistochemistry. Citicoline presented no mitogenic and chemotactic effects on hCMEC/D3; however, it significantly increased wound recovery, the formation of tube-like structures in Matrigel™ and enhanced spheroid development and sprouting. Citicoline induced the expression of phospho-extracellular-signal regulated kinase (ERK)-1/2. Kinexus assays showed an over-expression of insulin receptor substrate-1 (IRS-1). Knock-down of IRS-1 with targeted siRNA in our hCMEC/D3 inhibited the pro-angiogenic effects of citicoline. The percentage of surviving cells was higher in the presence of citicoline. Citicoline treatment significantly increased the numbers of new, active CD105-positive microvessels following MCAO. The findings demonstrate both a pro-angiogenic and protective effect of citicoline on hCMEC/D3 in vitro and following middle cerebral artery occlusion (MCAO) in vivo.

  1. Anemia and survival in human immunodeficiency virus

    DEFF Research Database (Denmark)

    Lundgren, Jens Dilling; Mocroft, Amanda

    2003-01-01

    The prospective, multicenter cohort study EuroSIDA has previously reported on predictors and outcomes of anemia in patients infected with human immunodeficiency virus. In a Cox proportional-hazards model with serial measures of CD4+ cell count, plasma viral load, and degrees of anemia fitted...... marker and whether correction of anemia itself results in a better prognosis remain to be determined....

  2. Intracoronary artery transplantation of cardiomyoblast-like cells from human adipose tissue-derived multi-lineage progenitor cells improve left ventricular dysfunction and survival in a swine model of chronic myocardial infarction

    Energy Technology Data Exchange (ETDEWEB)

    Okura, Hanayuki [The Center for Medical Engineering and Informatics, Osaka University, 2-2 Yamada-oka, Suita, Osaka 565-0879 (Japan); Department of Somatic Stem Cell Therapy and Health Policy, Institute of Biomedical Research and Innovation, Foundation for Biomedical Research and Innovation, 2-2 Minatojima-minamimachi, Chuo-ku, Kobe, Hyogo 650-0047 (Japan); Saga, Ayami; Soeda, Mayumi [Department of Somatic Stem Cell Therapy and Health Policy, Institute of Biomedical Research and Innovation, Foundation for Biomedical Research and Innovation, 2-2 Minatojima-minamimachi, Chuo-ku, Kobe, Hyogo 650-0047 (Japan); Miyagawa, Shigeru; Sawa, Yoshiki [Department of Surgery, Osaka University Graduate School of Medicine, 2-2 Yamada-oka, Suita, Osaka 565-0879 (Japan); Daimon, Takashi [Division of Biostatistics, Hyogo College of Medicine, 1-1 Mukogawa-cho, Nishinomiya, Hyogo 663-8501 (Japan); Ichinose, Akihiro [Department of Plastic Surgery, Kobe University Hospital, 7-5-2 Kusunoki-cho, Chuo-ku, Kobe, Hyogo (Japan); Matsuyama, Akifumi, E-mail: akifumi-matsuyama@umin.ac.jp [The Center for Medical Engineering and Informatics, Osaka University, 2-2 Yamada-oka, Suita, Osaka 565-0879 (Japan); Department of Plastic Surgery, Kobe University Hospital, 7-5-2 Kusunoki-cho, Chuo-ku, Kobe, Hyogo (Japan); RIKEN Program for Drug Discovery and Medical Technology Platforms, 1-7-22 Suehiro-cho, Tsurumi-ku, Yokohama, Kanagawa 230-0045 (Japan)

    2012-09-07

    Highlights: Black-Right-Pointing-Pointer We administered human CLCs in a swine model of MI via intracoronary artery. Black-Right-Pointing-Pointer Histological studies demonstrated engraftment of hCLCs into the scarred myocardium. Black-Right-Pointing-Pointer Echocardiography showed rescue of cardiac function in the hCLCs transplanted swine. Black-Right-Pointing-Pointer Transplantation of hCLCs is an effective therapeutics for cardiac regeneration. -- Abstract: Transplantation of human cardiomyoblast-like cells (hCLCs) from human adipose tissue-derived multi-lineage progenitor cells improved left ventricular function and survival of rats with myocardial infarction. Here we examined the effect of intracoronary artery transplantation of human CLCs in a swine model of chronic heart failure. Twenty-four pigs underwent balloon-occlusion of the first diagonal branch followed by reperfusion, with a second balloon-occlusion of the left ascending coronary artery 1 week later followed by reperfusion. Four weeks after the second occlusion/reperfusion, 17 of the 18 surviving animals with severe chronic MI (ejection fraction <35% by echocardiography) were immunosuppressed then randomly assigned to receive either intracoronary artery transplantation of hCLCs hADMPCs or placebo lactic Ringer's solution with heparin. Intracoronary artery transplantation was followed by the distribution of DiI-stained hCLCs into the scarred myocardial milieu. Echocardiography at post-transplant days 4 and 8 weeks showed rescue and maintenance of cardiac function in the hCLCs transplanted group, but not in the control animals, indicating myocardial functional recovery by hCLCs intracoronary transplantation. At 8 week post-transplantation, 7 of 8 hCLCs transplanted animals were still alive compared with only 1 of the 5 control (p = 0.0147). Histological studies at week 12 post-transplantation demonstrated engraftment of the pre DiI-stained hCLCs into the scarred myocardium and their expression of

  3. Irisin ameliorates hepatic glucose/lipid metabolism and enhances cell survival in insulin-resistant human HepG2 cells through adenosine monophosphate-activated protein kinase signaling.

    Science.gov (United States)

    So, Wing Yan; Leung, Po Sing

    2016-09-01

    Irisin is a newly identified myokine that promotes the browning of white adipose tissue, enhances glucose uptake in skeletal muscle and modulates hepatic metabolism. However, the signaling pathways involved in the effects on hepatic glucose and lipid metabolism have not been resolved. This study aimed to examine the role of irisin in the regulation of hepatic glucose/lipid metabolism and cell survival, and whether adenosine monophosphate-activated protein kinase (AMPK), a master metabolic regulator in the liver, is involved in irisin's actions. Human liver-derived HepG2 cells were cultured in normal glucose-normal insulin (NGNI) or high glucose-high insulin (HGHI/insulin-resistant) condition. Hepatic glucose and lipid metabolism was evaluated by glucose output and glycogen content or triglyceride accumulation assays, respectively. Our results showed that irisin stimulated phosphorylation of AMPK and acetyl-CoA-carboxylase (ACC) via liver kinase B1 (LKB1) rather than Ca(2+)/calmodulin-dependent protein kinase kinase β (CaMKKβ) in HepG2 cells. Irisin ameliorated hepatic insulin resistance induced by HGHI condition. Irisin reduced hepatic triglyceride content and glucose output, but increased glycogen content, with those effects reversed by dorsomorphin, an AMPK inhibitor. Furthermore, irisin also stimulated extracellular signal-regulated kinase (ERK) 1/2 phosphorylation and promoted cell survival in an AMPK-dependent manner. In conclusion, our data indicate that irisin ameliorates dysregulation of hepatic glucose/lipid metabolism and cell death in insulin-resistant states via AMPK activation. These findings reveal a novel irisin-mediated protective mechanism in hepatic metabolism which provides a scientific basis for irisin as a potential therapeutic target for the treatment of insulin resistance and type 2 diabetes mellitus. Copyright © 2016 Elsevier Ltd. All rights reserved.

  4. CD84 is a survival receptor for CLL cells.

    Science.gov (United States)

    Binsky-Ehrenreich, I; Marom, A; Sobotta, M C; Shvidel, L; Berrebi, A; Hazan-Halevy, I; Kay, S; Aloshin, A; Sagi, I; Goldenberg, D M; Leng, L; Bucala, R; Herishanu, Y; Haran, M; Shachar, I

    2014-02-20

    Chronic lymphocytic leukemia (CLL) is characterized by the accumulation of CD5+ B lymphocytes in peripheral blood, lymphoid organs and bone marrow. The main feature of the disease is accumulation of the malignant cells due to decreased apoptosis. CD84 belongs to the signaling lymphocyte activating molecule family of immunoreceptors, and has an unknown function in CLL cells. Here, we show that the expression of CD84 is significantly elevated from the early stages of the disease, and is regulated by macrophage migration inhibitory factor and its receptor, CD74. Activation of cell surface CD84 initiates a signaling cascade that enhances CLL cell survival. Both downmodulation of CD84 expression and its immune-mediated blockade induce cell death in vitro and in vivo. In addition, analysis of samples derived from an on-going clinical trial, in which human subjects were treated with humanized anti-CD74 (milatuzumab), shows a decrease in CD84 messenger RNA and protein levels in milatuzumab-treated cells. This downregulation was correlated with reduction of Bcl-2 and Mcl-1 expression. Thus, our data show that overexpression of CD84 in CLL is an important survival mechanism that appears to be an early event in the pathogenesis of the disease. These findings suggest novel therapeutic strategies based on the blockade of this CD84-dependent survival pathway.

  5. The role of caspase 3 and BclxL in the action of interleukin 7 (IL-7): a survival factor in activated human T cells

    DEFF Research Database (Denmark)

    Amos, C L; Woetmann, A; Nielsen, M;

    1998-01-01

    by the synthetic glucocorticoid, dexamethasone. Bcl-2 protein expression was uupregulated by IL-7 with or without dexamethasone, but Bc1-2 was expressed at a much lower level than BclxL in these cells. Levels of Bax did not markedly change on either cytokine stimulation or dexamethasone treatment. An unidentified...... cells. Both cytokines abrogated the dexamethasone-induced stimulation of Caspase 3 and prevented the cleavage of poly (ADP-ribose) polymerase (PARP), a substrate for the Caspase 3. IL-7 upregulated the expression of Bc1xL and counteracted the downregulation of this anti-apoptotic protein...... of apoptosis. A clear role for IL-7 as a survival factor for cytokine withdrawal and glucocorticoid induced apoptosis in activated primary hT cells is implicated. In addition, regulation of BclxL and downstream inhibition of Caspase 3 activity may mediate this rescue signal....

  6. Anemia and survival in human immunodeficiency virus

    DEFF Research Database (Denmark)

    Lundgren, Jens Dilling; Mocroft, Amanda

    2003-01-01

    The prospective, multicenter cohort study EuroSIDA has previously reported on predictors and outcomes of anemia in patients infected with human immunodeficiency virus. In a Cox proportional-hazards model with serial measures of CD4+ cell count, plasma viral load, and degrees of anemia fitted...... as time-dependent variables, the relative hazard of death increased markedly for patients with anemia versus no anemia. A clinical scoring system was developed and validated for patients receiving highly active antiretroviral therapy using the most recent laboratory measures. Mild and severe anemia were...... independently (Panemia. The mechanisms underlying why hemoglobin is such a strong prognostic...

  7. Anemia and survival in human immunodeficiency virus

    DEFF Research Database (Denmark)

    Lundgren, Jens Dilling; Mocroft, Amanda

    2003-01-01

    The prospective, multicenter cohort study EuroSIDA has previously reported on predictors and outcomes of anemia in patients infected with human immunodeficiency virus. In a Cox proportional-hazards model with serial measures of CD4+ cell count, plasma viral load, and degrees of anemia fitted...... as time-dependent variables, the relative hazard of death increased markedly for patients with anemia versus no anemia. A clinical scoring system was developed and validated for patients receiving highly active antiretroviral therapy using the most recent laboratory measures. Mild and severe anemia were...... independently (Panemia. The mechanisms underlying why hemoglobin is such a strong prognostic...

  8. Cigarette smoke extract promotes human vascular smooth muscle cell proliferation and survival through ERK1/2- and NF-κB-dependent pathways

    DEFF Research Database (Denmark)

    Chen, Qing-Wen; Edvinsson, Lars; Xu, Cang-Bao

    2010-01-01

    Tobacco use is one of the major risk factors of cardiovascular disease. The underlying molecular mechanisms that link cigarette smoke to cardiovascular disease remain unclear. The present study was designed to examine the effects of dimethyl sulfoxide (DMSO)-soluble smoke particles (DSPs) on human...... and necrosis were found in serum-starved HASMCs. DSPs decreased cell death and increased B-cell leukemia/lymphoma 2 expression. Blocking phosphorylation of ERK1/2 or NF-κB attenuated DSP-induced cell death inhibition. Cigarette smoke particles stimulate HASMC proliferation and inhibit cell death...

  9. Cigarette smoke extract promotes human vascular smooth muscle cell proliferation and survival through ERK1/2- and NF-κB-dependent pathways

    DEFF Research Database (Denmark)

    Chen, Qing-Wen; Edvinsson, Lars; Xu, Cang-Bao

    2010-01-01

    Tobacco use is one of the major risk factors of cardiovascular disease. The underlying molecular mechanisms that link cigarette smoke to cardiovascular disease remain unclear. The present study was designed to examine the effects of dimethyl sulfoxide (DMSO)-soluble smoke particles (DSPs) on human...... and necrosis were found in serum-starved HASMCs. DSPs decreased cell death and increased B-cell leukemia/lymphoma 2 expression. Blocking phosphorylation of ERK1/2 or NF-¿B attenuated DSP-induced cell death inhibition. Cigarette smoke particles stimulate HASMC proliferation and inhibit cell death...

  10. Reduced expression of DNA repair and redox signaling protein APE1/Ref-1 impairs human pancreatic cancer cell survival, proliferation, and cell cycle progression.

    Science.gov (United States)

    Jiang, Yanlin; Zhou, Shaoyu; Sandusky, George E; Kelley, Mark R; Fishel, Melissa L

    2010-11-01

    Pancreatic cancer is a deadly disease that is virtually never cured. Understanding the chemoresistance intrinsic to this cancer will aid in developing new regimens. High expression of APE1/Ref-1, a DNA repair and redox signaling protein, is associated with resistance, poor outcome, and angiogenesis; little is known in pancreatic cancer. Immunostaining of adenocarcinoma shows greater APE1/Ref-1 expression than in normal pancreas tissue. A decrease in APE1/Ref-1 protein levels results in pancreatic cancer cell growth inhibition, increased apoptosis, and altered cell cycle progression. Endogenous cell cycle inhibitors increase when APE1/ Ref-1 is reduced, demonstrating its importance to proliferation and growth of pancreatic cancer.

  11. Distribution and survival of Borrelia miyamotoi in human blood components.

    Science.gov (United States)

    Thorp, Aaron M; Tonnetti, Laura

    2016-03-01

    Borrelia miyamotoi, the agent of relapsing fever, is a tick-borne spirochete first isolated in Japan in 1994. Since then, the spirochete has been detected in ticks globally, generally in the same vectors as the Lyme disease agent. Human infection has been reported in Russia, Europe, Japan, and the United States, as influenza-like febrile illness. In addition, two cases of meningoencephalitis caused by B. miyamotoi have also been reported in immunocompromised patients. Here we evaluate the ability of the spirochete to survive in human blood components stored under standard blood bank conditions. Freshly collected human whole blood was spiked with in vitro cultured B. miyamotoi or B. miyamotoi-infected mouse plasma and separated into red blood cells (RBCs), plasma, and platelets. Components were either injected into immunocompromised (SCID) or wild-type immunocompetent mice or cultured in vitro, right after separation and after storage at the appropriate conditions. Infection was monitored by microscopic observation, blood smears, and polymerase chain reaction. In vivo, all the SCID mice challenged with the components before storage and the RBCs stored for up to 42 days developed the infection. Wild-type mice also developed the infection when injected with prestorage samples from all components, while a lower number of mice were infected by RBCs stored for 42 days. In vitro, spirochetes grew in all samples but frozen plasma. This study demonstrated that B. miyamotoi can survive standard storage conditions of most human blood components, suggesting the possibility of transmission by blood transfusion. © 2015 AABB.

  12. Novel anti-apoptotic microRNAs 582-5p and 363 promote human glioblastoma stem cell survival via direct inhibition of caspase 3, caspase 9, and Bim.

    Directory of Open Access Journals (Sweden)

    Desiree Hunt Floyd

    Full Text Available Glioblastoma is the most common and lethal primary brain tumor. Tumor initiation and recurrence are likely caused by a sub-population of glioblastoma stem cells, which may derive from mutated neural stem and precursor cells. Since CD133 is a stem cell marker for both normal brain and glioblastoma, and to better understand glioblastoma formation and recurrence, we looked for dys-regulated microRNAs in human CD133+ glioblastoma stem cells as opposed to CD133+ neural stem cells isolated from normal human brain. Using FACS sorting of low-passage cell samples followed by microRNA microarray analysis, we found 43 microRNAs that were dys-regulated in common in three separate CD133+ human glioblastomas compared to CD133+ normal neural stem cells. Among these were several microRNAs not previously associated with cancer. We then verified the microRNAs dys-regulated in glioblastoma using quantitative real time PCR and Taqman analysis of the original samples, as well as human GBM stem cell and established cell lines and many human specimens. We show that two candidate oncogenic microRNAs, miR-363 and miR-582-5p, can positively influence glioblastoma survival, as shown by forced expression of the microRNAs and their inhibitors followed by cell number assay, Caspase 3/7 assay, Annexin V apoptosis/fluorescence activated cell sorting, siRNA rescue of microRNA inhibitor treatment, as well as 3'UTR mutagenesis to show luciferase reporter rescue of the most successful targets. miR-582-5p and miR-363 are shown to directly target Caspase 3, Caspase 9, and Bim.

  13. Intracerebroventricular injection of encapsulated human mesenchymal cells producing glucagon-like peptide 1 prolongs survival in a mouse model of ALS.

    Directory of Open Access Journals (Sweden)

    Sarah Knippenberg

    Full Text Available BACKGROUND: As pharmacological therapies have largely failed so far, stem cell therapy has recently come into the focus of ALS research. Neuroprotective potential was shown for several types of stem and progenitor cells, mainly due to release of trophic factors. In the present study, we assessed the effects of intracerebroventricular injection of glucagon-like peptide 1 (GLP-1 releasing mesenchymal stromal cells (MSC in mutant SOD1 (G93A transgenic mice. METHODOLOGY/PRINCIPAL FINDINGS: To improve the neuroprotective effects of native MSC, they had been transfected with a plasmid vector encoding a GLP-1 fusion gene prior to the injection, as GLP-1 was shown to exhibit neuroprotective properties before. Cells were encapsulated and therefore protected against rejection. After intracerebroventricular injection of these GLP-1 MSC capsules in presymptomatic SOD1 (G93A mice, we assessed possible protective effects by survival analysis, measurement of body weight, daily monitoring and evaluation of motor performance by rotarod and footprint analyses. Motor neuron numbers in the spinal cord as well as the amount of astrocytosis, microglial activation, heat shock response and neuronal nitric oxide synthase (nNOS expression were analyzed by immunohistological methods. Treatment with GLP-1 producing MSC capsules significantly prolonged survival by 13 days, delayed symptom onset by 15 days and weight loss by 14 days and led to significant improvements in motor performance tests compared to vehicle treated controls. Histological data are mainly in favour of anti-inflammatory effects of GLP-1 producing MSC capsules with reduced detection of inflammatory markers and a significant heat shock protein increase. CONCLUSION/SIGNIFICANCE: Intracerebroventricular injection of GLP-1 MSC capsules shows neuroprotective potential in the SOD1 (G93A mouse model.

  14. Cell survival in a simulated Mars environment

    Science.gov (United States)

    Todd, Paul; Kurk, Michael Andy; Boland, Eugene; Thomas, David

    2016-07-01

    The most ancient life forms on earth date back comfortably to the time when liquid water was believed to be abundant on Mars. These ancient life forms include cyanobacteria, contemporary autotrophic earth organisms believed to have descended from ancestors present as long as 3.5 billion years ago. Contemporary cyanobacteria have adapted to the earth environment's harshest conditions (long-term drying, high and low temperature), and, being autotrophic, they are among the most likely life forms to withstand space travel and the Mars environment. However, it is unlikely that humans would unwittingly contaminate a planetary spacecraft with these microbes. One the other hand, heterotrophic microbes that co-habit with humans are more likely spacecraft contaminants, as history attests. Indeed, soil samples from the Atacama desert have yielded colony-forming organisms resembling enteric bacteria. There is a need to understand the survivability of cyanobacteria (likely survivors, unlikely contaminants) and heterotrophic eubacteria (unlikely survivors, likely contaminants) under simulated planetary conditions. A 35-day test was performed in a commercial planetary simulation system (Techshot, Inc., Greenville, IN) in which the minimum night-time temperature was -80 C, the maximum daytime temperature was +26 C, the simulated day-night light cycle in earth hours was 12-on and 12-off, and the total pressure of the pure CO _{2} atmosphere was maintained below 11 mbar. Any water present was allowed to equilibrate with the changing temperature and pressure. The gas phase was sampled into a CR1-A low-pressure hygrometer (Buck Technologies, Boulder, CO), and dew/frost point was measured once every hour and recorded on a data logger, along with the varying temperature in the chamber, from which the partial pressure of water was calculated. According to measurements there was no liquid water present throughout the test except during the initial pump-down period when aqueous specimens

  15. Human embryonic stem cell-derived cardiomyocytes survive and mature in the mouse heart and transiently improve function after myocardial infarction

    NARCIS (Netherlands)

    van Laake, Linda W.; Passier, Robert; Monshouwer-Kloots, Jantine; Verkleij, Arie J.; Lips, Daniel J.; Freund, Christian; den Ouden, Krista; Ward-van Oostwaard, Dorien; Korving, Jeroen; Tertoolen, Leon G.; van Echteld, Cees J.; Doevendans, Pieter A.; Mummery, Christine L.

    2007-01-01

    Regeneration of the myocardium by transplantation of cardiomyocytes is an emerging therapeutic strategy. Human embryonic stem cells (HESC) form cardiomyocytes readily but until recently at low efficiency, so that preclinical studies on transplantation in animals are only just beginning. Here, we sho

  16. Combinatorial and sequential delivery of gemcitabine and oseltamivir phosphate from implantable poly(D,L-lactic-co-glycolic acid cylinders disables human pancreatic cancer cell survival

    Directory of Open Access Journals (Sweden)

    Allison Logan S

    2017-07-01

    Full Text Available Stephanie Allison Logan,1 Amanda J Brissenden,1 Myron R Szewczuk,2 Ronald J Neufeld1 1Department of Chemical Engineering, 2Department of Biomedical and Molecular Sciences, Queen’s University, Kingston, ON, Canada Abstract: Combination therapies against multiple targets are currently being developed to prevent resistance to a single chemotherapeutic agent and to extirpate pre-existing resistance in heterogeneous cancer cells in tumors due to selective pressure from the single agent. Gemcitabine (GEM, a chemotherapeutic agent, is the current standard of care for patients with pancreatic cancer. Patients with pancreatic cancer receiving GEM have a low progression-free survival. Given the poor response rate to GEM, cancer cells are known to develop rapid resistance to this drug. Metronomic chemotherapy using combinatorial and sequential delivery systems are novel developmental approaches to disrupt tumor neovascularization, reduce systemic drug toxicity, and increase the sensitivity of chemotherapeutics in cancer. Here, implantable double-layered poly(D,L-lactic-co-glycolic acid (PLGA cylinders were engineered to sequentially release GEM in combination with oseltamivir phosphate (OP over an extended time. Double-layered PLGA cylindrical implants loaded with these active hydrophilic drugs were fabricated with minimal loss of drugs during the formulation, enabling extensive control of drug loading and establishing uniform drug distribution throughout the polymer matrix. OP is used in the formulation because of its anticancer drug properties targeting mammalian neuraminidase 1 (Neu1 involved in multistage tumorigenesis. OP and GEM encapsulated in inner/outer GEMin/OPout or OPin/GEMout implantable PLGA double-layered cylinders displayed sustained near linear release over 30 days. OP and GEM released from the double-layered cylinders effectively reduced cell viability in pancreatic cancer cell line PANC1 and its GEM-resistant variant for up to 15

  17. Pharmacologically active microcarriers for endothelial progenitor cell support and survival.

    Science.gov (United States)

    Musilli, Claudia; Karam, Jean-Pierre; Paccosi, Sara; Muscari, Claudio; Mugelli, Alessandro; Montero-Menei, Claudia N; Parenti, Astrid

    2012-08-01

    The regenerative potential of endothelial progenitor cell (EPC)-based therapies is limited due to poor cell viability and minimal retention following application. Neovascularization can be improved by means of scaffolds supporting EPCs. The aim of the present study was to investigate whether human early EPCs (eEPCs) could be efficiently cultured on pharmacologically active microcarriers (PAMs), made with poly(d,l-lactic-coglycolic acid) and coated with adhesion/extracellular matrix molecules. They may serve as a support for stem cells and may be used as cell carriers providing a controlled delivery of active protein such as the angiogenic factor, vascular endothelial growth factor-A (VEGF-A). eEPC adhesion to fibronectin-coated PAMs (FN-PAMs) was assessed by means of microscopic evaluation and by means of Alamar blue assay. Phospho ERK(1/2) and PARP-1 expression was measured by means of Western blot to assess the survival effects of FN-PAMs releasing VEGF-A (FN-VEGF-PAMs). The Alamar blue assay or a modified Boyden chamber assay was employed to assess proliferative or migratory capacity, respectively. Our data indicate that eEPCs were able to adhere to empty FN-PAMs within a few hours. FN-VEGF-PAMs increased the ability of eEPCs to adhere to them and strongly supported endothelial-like phenotype and cell survival. Moreover, the release of VEGF-A by FN-PAMs stimulated in vitro HUVEC migration and proliferation. These data strongly support the use of PAMs for supporting eEPC growth and survival and for stimulating resident mature human endothelial cells.

  18. A human iPSC model of Ligase IV deficiency reveals an important role for NHEJ-mediated-DSB repair in the survival and genomic stability of induced pluripotent stem cells and emerging haematopoietic progenitors.

    Science.gov (United States)

    Tilgner, K; Neganova, I; Moreno-Gimeno, I; Al-Aama, J Y; Burks, D; Yung, S; Singhapol, C; Saretzki, G; Evans, J; Gorbunova, V; Gennery, A; Przyborski, S; Stojkovic, M; Armstrong, L; Jeggo, P; Lako, M

    2013-08-01

    DNA double strand breaks (DSBs) are the most common form of DNA damage and are repaired by non-homologous-end-joining (NHEJ) or homologous recombination (HR). Several protein components function in NHEJ, and of these, DNA Ligase IV is essential for performing the final 'end-joining' step. Mutations in DNA Ligase IV result in LIG4 syndrome, which is characterised by growth defects, microcephaly, reduced number of blood cells, increased predisposition to leukaemia and variable degrees of immunodeficiency. In this manuscript, we report the creation of a human induced pluripotent stem cell (iPSC) model of LIG4 deficiency, which accurately replicates the DSB repair phenotype of LIG4 patients. Our findings demonstrate that impairment of NHEJ-mediated-DSB repair in human iPSC results in accumulation of DSBs and enhanced apoptosis, thus providing new insights into likely mechanisms used by pluripotent stem cells to maintain their genomic integrity. Defects in NHEJ-mediated-DSB repair also led to a significant decrease in reprogramming efficiency of human cells and accumulation of chromosomal abnormalities, suggesting a key role for NHEJ in somatic cell reprogramming and providing insights for future cell based therapies for applications of LIG4-iPSCs. Although haematopoietic specification of LIG4-iPSC is not affected per se, the emerging haematopoietic progenitors show a high accumulation of DSBs and enhanced apoptosis, resulting in reduced numbers of mature haematopoietic cells. Together our findings provide new insights into the role of NHEJ-mediated-DSB repair in the survival and differentiation of progenitor cells, which likely underlies the developmental abnormalities observed in many DNA damage disorders. In addition, our findings are important for understanding how genomic instability arises in pluripotent stem cells and for defining appropriate culture conditions that restrict DNA damage and result in ex vivo expansion of stem cells with intact genomes.

  19. Anti-PD-L1 prolongs survival and triggers T cell but not humoral anti-tumor immune responses in a human MUC1-expressing preclinical ovarian cancer model.

    Science.gov (United States)

    Mony, Jyothi Thyagabhavan; Zhang, Lixin; Ma, Tianzhou; Grabosch, Shannon; Tirodkar, Tejas S; Brozick, Joan; Tseng, George; Elishaev, Esther; Edwards, Robert P; Huang, Xin; Vlad, Anda M

    2015-09-01

    Monoclonal antibodies that block inhibitory immune checkpoint molecules and enhance anti-tumor responses show clinical promise in advanced solid tumors. Most of the preliminary evidence on therapeutic efficacy of immune checkpoint blockers comes from studies in melanoma, lung and renal cancer. To test the in vivo potential of programmed death-ligand 1 (PD-L1) blockade in ovarian cancer, we recently generated a new transplantable tumor model using human mucin 1 (MUC1)-expressing 2F8 cells. The MUC1 transgenic (MUC1.Tg) mice develop large number of intraperitoneal (IP) tumors following IP injection of 8 × 10(5) syngeneic 2F8 cells. The tumors are aggressive and display little T cell infiltration. Anti-PD-L1 antibody was administered IP every 2 weeks (200 μg/dose) for a total of three doses. Treatment was started 21 days post-tumor challenge, a time point which corresponds to late tumor stage. The anti-PD-L1 treatment led to substantial T cell infiltration within the tumor and significantly increased survival (p = 0.001) compared to isotype control-treated mice. When the same therapy was administered to wild-type mice challenged with 2F8 tumors, no survival benefit was observed, despite the presence of high titer anti-MUC1 antibodies. However, earlier treatment (day 11) and higher frequency of IP injections restored the T cell responses and led to prolonged survival. Splenocyte profiling via Nanostring using probes for 511 immune genes revealed a treatment-induced immune gene signature consistent with increased T cell-mediated immunity. These findings strongly support further preclinical and clinical strategies exploring PD-L1 blockade in ovarian cancer.

  20. Preserved seminiferous tubule integrity with spermatogonial survival and induction of Sertoli and Leydig cell maturation after long-term organotypic culture of prepubertal human testicular tissue.

    Science.gov (United States)

    de Michele, F; Poels, J; Weerens, L; Petit, C; Evrard, Z; Ambroise, J; Gruson, D; Wyns, C

    2017-01-01

    Is an organotypic culture system able to provide the appropriate testicular microenvironment for in-vitro maturation of human immature testicular tissue (ITT)? Our organotypic culture system provided a microenvironment capable of preserving seminiferous tubule (ST) integrity and Leydig cell (LC) functionality and inducing Sertoli cell (SC) maturation. Cryopreservation of human ITT is a well-established strategy to preserve fertility in prepubertal boys affected by cancer, with a view for obtaining sperm. While spermatogenesis in mice has been replicated in organotypic culture, yielding reproductively efficient spermatozoa, this process has not yet been achieved in humans. The aim of this study was to in vitro mature frozen-thawed ITT. To this end, 1 mm(3) tissue fragments from three prepubertal patients aged 2 (P1), 11 (P2) and 12 (P3) years were placed in organotypic culture for 139 days. Culture media, supplemented with either testosterone or hCG, were compared. ST integrity and tissue viability were assessed by histological score and lactate dehydrogenase (LDH) levels in supernatants. Spermatogonia (SG), proliferating cells and proliferating SG were identified by the use of MAGE-A4 and Ki67 immunohistochemical markers. Glial cell line-derived neurotrophic factor (GDNF) was used as a marker of SC functionality, while SC maturation was evaluated by androgen receptor (AR), anti-Müllerian hormone (AMH) immunohistochemistry (IHC) and AMH immunoenzymatic assay. LC functionality was determined by testosterone levels in supernatants and by 3β-hydroxysteroid dehydrogenase (3β-HSD) IHC. Apoptosis was studied by IHC with active caspases 3 and 8 and by TUNEL (terminal deoxynubocleotidyl transferase-mediated dUTP nick end labeling) analysis. Tissue viability was preserved, as demonstrated by the decrease in and stabilization of LDH release, and evolution of ST scoring, with the percentage of well-preserved STs showing no statistical differences during culture in either

  1. PDK2-mediated alternative splicing switches Bnip3 from cell death to cell survival.

    Science.gov (United States)

    Gang, Hongying; Dhingra, Rimpy; Lin, Junjun; Hai, Yan; Aviv, Yaron; Margulets, Victoria; Hamedani, Mohammad; Thanasupawat, Thatchawan; Leygue, Etienne; Klonisch, Thomas; Davie, James R; Kirshenbaum, Lorrie A

    2015-09-28

    Herein we describe a novel survival pathway that operationally links alternative pre-mRNA splicing of the hypoxia-inducible death protein Bcl-2 19-kD interacting protein 3 (Bnip3) to the unique glycolytic phenotype in cancer cells. While a full-length Bnip3 protein (Bnip3FL) encoded by exons 1-6 was expressed as an isoform in normal cells and promoted cell death, a truncated spliced variant of Bnip3 mRNA deleted for exon 3 (Bnip3Δex3) was preferentially expressed in several human adenocarcinomas and promoted survival. Reciprocal inhibition of the Bnip3Δex3/Bnip3FL isoform ratio by inhibiting pyruvate dehydrogenase kinase isoform 2 (PDK2) in Panc-1 cells rapidly induced mitochondrial perturbations and cell death. The findings of the present study reveal a novel survival pathway that functionally couples the unique glycolytic phenotype in cancer cells to hypoxia resistance via a PDK2-dependent mechanism that switches Bnip3 from cell death to survival. Discovery of the survival Bnip3Δex3 isoform may fundamentally explain how certain cells resist Bnip3 and avert death during hypoxia.

  2. Radiation survival parameters of antineoplastic drug-sensitive and -resistant human ovarian cancer cell lines and their modification by buthionine sulfoximine

    Energy Technology Data Exchange (ETDEWEB)

    Louie, K.G.; Behrens, B.C.; Kinsella, T.J.; Hamilton, T.C.; Grotzinger, K.R.; McKoy, W.M.; Winker, M.A.; Ozols, R.F.

    1985-05-01

    The optimum integration of chemotherapy and irradiation is of potential clinical significance in the treatment of ovarian cancer. A series of human ovarian cancer cell lines have been developed in which dose-response relationships to standard anticancer drugs have been determined, and the patterns of cross-resistance between these drugs and irradiation have been established. By stepwise incubation with drugs, sublines of A2780, a drug-sensitive cell line, have been made 100-fold, 10-fold, and 10-fold more resistant to Adriamycin (2780AD), melphalan (2780ME), and cisplatin (2780CP). Two additional cell lines, NIH:OVCAR-3nu(Ag+) and NIH:OVCAR-4(Ag+), were established from drug-refractory patients. 2780ME, 2780CP, OVCAR-3nu(Ag+), and OVCAR-4(Ag+) are all cross-resistant to irradiation, with DOS of 146, 187, 143, and 203, respectively. However, 2780AD remains sensitive to radiation, with a DO of 111, which is similar to that of A2780 (101). Glutathione (GSH) levels are elevated in 2780ME, 2780CP, OVCAR-3nu(Ag+), and OVCAR-4(Ag+) to 4.58, 6.13, 12.10, and 15.14 nmol/10(6) cells as compared to A2780, with 1.89 nmol/10(6) cells. However, the GSH level in 2780AD is only minimally higher than that in A2780 (2.94 nmol/10(6) cells). Buthionine sulfoximine, a specific inhibitor of GSH synthesis, significantly increases the radiation sensitivity of 2780ME (changing the DO from 143 to 95) and 2780CP to a lesser extent, suggesting that intracellular GSH levels may play an important role in the radiation response of certain neoplastic cells.

  3. Starved epithelial cells uptake extracellular matrix for survival

    Science.gov (United States)

    Muranen, Taru; Iwanicki, Marcin P.; Curry, Natasha L.; Hwang, Julie; DuBois, Cory D.; Coloff, Jonathan L.; Hitchcock, Daniel S.; Clish, Clary B.; Brugge, Joan S.; Kalaany, Nada Y.

    2017-01-01

    Extracellular matrix adhesion is required for normal epithelial cell survival, nutrient uptake and metabolism. This requirement can be overcome by oncogene activation. Interestingly, inhibition of PI3K/mTOR leads to apoptosis of matrix-detached, but not matrix-attached cancer cells, suggesting that matrix-attached cells use alternate mechanisms to maintain nutrient supplies. Here we demonstrate that under conditions of dietary restriction or growth factor starvation, where PI3K/mTOR signalling is decreased, matrix-attached human mammary epithelial cells upregulate and internalize β4-integrin along with its matrix substrate, laminin. Endocytosed laminin localizes to lysosomes, results in increased intracellular levels of essential amino acids and enhanced mTORC1 signalling, preventing cell death. Moreover, we show that starved human fibroblasts secrete matrix proteins that maintain the growth of starved mammary epithelial cells contingent upon epithelial cell β4-integrin expression. Our study identifies a crosstalk between stromal fibroblasts and epithelial cells under starvation that could be exploited therapeutically to target tumours resistant to PI3K/mTOR inhibition. PMID:28071763

  4. Human neural stem cells survive long term in the midbrain of dopamine-depleted monkeys after GDNF overexpression and project neurites toward an appropriate target.

    Science.gov (United States)

    Wakeman, Dustin R; Redmond, D Eugene; Dodiya, Hemraj B; Sladek, John R; Leranth, Csaba; Teng, Yang D; Samulski, R Jude; Snyder, Evan Y

    2014-06-01

    Transplanted multipotent human fetal neural stem cells (hfNSCs) significantly improved the function of parkinsonian monkeys in a prior study primarily by neuroprotection, with only 3%-5% of cells expressing a dopamine (DA) phenotype. In this paper, we sought to determine whether further manipulation of the neural microenvironment by overexpression of a developmentally critical molecule, glial cell-derived neurotrophic factor (GDNF), in the host striatum could enhance DA differentiation of hfNSCs injected into the substantia nigra and elicit growth of their axons to the GDNF-expressing target. hfNSCs were transplanted into the midbrain of 10 green monkeys exposed to 1-methyl-4-phenyl-1,2,3,6-tetrahydro-pyridine. GDNF was delivered concomitantly to the striatum via an adeno-associated virus serotype 5 vector, and the fate of grafted cells was assessed after 11 months. Donor cells remained predominantly within the midbrain at the injection site and sprouted numerous neurofilament-immunoreactive fibers that appeared to course rostrally toward the striatum in parallel with tyrosine hydroxylase-immunoreactive fibers from the host substantia nigra but did not mature into DA neurons. This work suggests that hfNSCs can generate neurons that project long fibers in the adult primate brain. However, in the absence of region-specific signals and despite GDNF overexpression, hfNSCs did not differentiate into mature DA neurons in large numbers. It is encouraging, however, that the adult primate brain appeared to retain axonal guidance cues. We believe that transplantation of stem cells, specifically instructed ex vivo to yield DA neurons, could lead to reconstruction of some portion of the nigrostriatal pathway and prove beneficial for the parkinsonian condition. ©AlphaMed Press.

  5. Human immune system development and survival of non-obese diabetic (NOD)-scid IL2rγ(null) (NSG) mice engrafted with human thymus and autologous haematopoietic stem cells.

    Science.gov (United States)

    Covassin, L; Jangalwe, S; Jouvet, N; Laning, J; Burzenski, L; Shultz, L D; Brehm, M A

    2013-12-01

    Immunodeficient mice bearing targeted mutations in the IL2rg gene and engrafted with human immune systems are effective tools for the study of human haematopoiesis, immunity, infectious disease and transplantation biology. The most robust human immune model is generated by implantation of human fetal thymic and liver tissues in irradiated recipients followed by intravenous injection of autologous fetal liver haematopoietic stem cells [often referred to as the BLT (bone marrow, liver, thymus) model]. To evaluate the non-obese diabetic (NOD)-scid IL2rγ(null) (NSG)-BLT model, we have assessed various engraftment parameters and how these parameters influence the longevity of NSG-BLT mice. We observed that irradiation and subrenal capsule implantation of thymus/liver fragments was optimal for generating human immune systems. However, after 4 months, a high number of NSG-BLT mice develop a fatal graft-versus-host disease (GVHD)-like syndrome, which correlates with the activation of human T cells and increased levels of human immunoglobulin (Ig). Onset of GVHD was not delayed in NSG mice lacking murine major histocompatibility complex (MHC) classes I or II and was not associated with a loss of human regulatory T cells or absence of intrathymic cells of mouse origin (mouse CD45(+) ). Our findings demonstrate that NSG-BLT mice develop robust human immune systems, but that the experimental window for these mice may be limited by the development of GVHD-like pathological changes.

  6. Human survival: evolution, religion and the irrational

    Directory of Open Access Journals (Sweden)

    Milton H. Saier, jr

    2010-06-01

    Full Text Available Is there a possible biological explanation for religion? That is, is there a genetic basis for believing in mystical, supernatural beings when there is no scientifi c evidence for their existence? Can we explain why some people prefer to accept myth over science? Why do so many people still accept creation and refuse to embrace evolution? Is there an evolutionary basis for religious beliefs? It is certainly true that religions have been part of human civilization throughout most of its recent history, at least for the last 5,000 years, and probably for much longer. Even great nonmystical philosophers such as Confucius, Buddha, and Lao Tzu have had their teachings evolve into mystical religions with spiritual ancestors, gods and reincarnation. On the other hand, religion is largely absent in modern Chinese culture, and of diminishing importance in Japanese and European cultures. In all cultures, the degrees of education gained by individuals correlate inversely with attachments to mystical deities. Atheists abound although they may be reluctant to come out of the closet and affi rm their rational convictions. In this article, we seek explanations for human irrationality.

  7. Human papillomavirus in head and neck squamous cell carcinoma of unknown primary is a common event and a strong predictor of survival

    DEFF Research Database (Denmark)

    Jensen, David Hebbelstrup; Hedback, Nora; Specht, Lena

    2014-01-01

    clinical records. RESULTS: Of the identified 60 patients with CUP, 13 were shown to be positive for HPV DNA, amounting to 22% of the study population. In addition, we were able to show a clear disease-free and overall-survival benefit in the HPV-positive group, with a hazard ratio of 0.16 (95% CI: 0.......038-0.67) for over-all survival. This survival benefit was also apparent when adjusted for advanced age in a multivariate Cox regression analysis. CONCLUSION: A fairly large percentage of CUP cases are HPV-related, and because this is related to both the location and prognosis, we recommend HPV testing as part...

  8. Conditional survival predictions after nephrectomy for renal cell carcinoma.

    NARCIS (Netherlands)

    Karakiewicz, P.I.; Suardi, N.; Capitanio, U.; Isbarn, H.; Jeldres, C.; Perrotte, P.; Sun, M.; Ficarra, V.; Zigeuner, R.; Tostain, J.; Mejean, A.; Cindolo, L.; Pantuck, A.J.; Belldegrun, A.S.; Zini, L.; Taille, A. De La; Chautard, D.; Descotes, J.L.; Shariat, S.F.; Valeri, A.; Mulders, P.F.A.; Lang, H.; Lechevallier, E.; Patard, J.J.

    2009-01-01

    PURPOSE: Conditional survival implies that on average long-term cancer survivors have a better prognosis than do newly diagnosed individuals. We explored the effect of conditional survival in renal cell carcinoma. MATERIALS AND METHODS: We studied 3,560 patients with renal cell carcinoma of all stag

  9. Mathematical model of naive T cell division and survival IL-7 thresholds

    Directory of Open Access Journals (Sweden)

    Carmen eMolina-Paris

    2013-12-01

    Full Text Available We develop a mathematical model to describe the homeostasis of naive T~cells in the peripheral immune system of ageing humans, incorporating thymic output and the availability of interleukin-7 (IL-7. The model is formulated as three ordinary differential equations: two describe T~cell numbers, in a resting state and progressing through the cell cycle. The third is introduced to describe changes in IL-7 availability. Thymic output is a decreasing function of time, representative of thymic atrophy found in ageing humans. Each T~cell is assumed to possess two interleukin-7 receptor (IL-7R signalling thresholds: a survival threshold and a second, higher, proliferation threshold. If the IL-7R signalling strength is below its survival threshold, a cell may undergo apoptosis. When the signalling strength is above the survival threshold, but below the proliferation threshold, the cell survives but does not divide. Signalling strength above the proliferation threshold enables entry into cell cycle. Assuming that individual cell thresholds are log-normally distributed, we derive population-average rates for apoptosis and entry into cell cycle. We have analysed the adiabatic change in homeostasis as thymic output decreases. With a parameter set representative of a healthy individual, the model predicts a unique equilibrium number of T~cells. In a parameter range representative of persistent virus or bacterial infection, where naive T~cell survival is impaired, a decrease in thymic output may result in the collapse of the naive T~cell repertoire.

  10. Common raven juvenile survival in a human-augmented landscape

    Science.gov (United States)

    Webb, William C.; Boarman, William I.; Rotenberry, John T.

    2004-01-01

    Anthropogenic resource subsidies have contributed to the dramatic increase in the abundance of Common Ravens (Corvus corax) in the western Mojave Desert, California, during the past 30 years. To better understand the effects of these subsidies on raven demography, we examined whether survival to juvenile departure from the natal territory could be predicted by a set of environmental and morphological variables, such as nest proximity to anthropogenic resources and juvenile condition. We captured 240 juvenile ravens over 2 years and marked them prior to fledging. Nest proximity to anthropogenic resources and earlier fledging dates significantly predicted raven juvenile survival to departure from the natal territory. The best-fitting mark-recapture models predicted postdeparture survival as a function of time since fledging, nest proximity to anthropogenic resources, and year hatched. The positive effect of nest proximity to anthropogenic resources influenced postdeparture survival for at least 9 months after fledging, as revealed by the mark-recapture analysis. Annual survival was 47% for first-year, 81% for second-year, and 83% for third-year birds. Our results support the hypothesis that anthropogenic resources contribute to increasing raven numbers via increased juvenile survival to departure as well as increased postdeparture survival. We expect raven numbers to grow in concert with the growing human presence in the Mojave Desert unless raven access to anthropogenic resources is diminished.

  11. 人胚神经干细胞植入脑损伤大鼠的存活和分化状态%Survival and differentiation of human embryonic neural stem cells in rats with brain injury

    Institute of Scientific and Technical Information of China (English)

    张泽舜; 万虹; 历俊华; 翟晶; 王忠诚

    2007-01-01

    .结论:人胚神经干细胞能够存活于脑损伤区域,移植后逐渐分化为星形胶质细胞,且易被内皮吞噬细胞所消化.提示免疫排斥反应可能影响人胚神经干细胞的存活.%BACKGROUND:Finding of neural stem cells(NSCs)brings new hope for repairing central nervous system(CNS)injury.However,the influence of internal environment after brain injury on the survival and differentiation of NSCs is a complexand variable process.OBJECTIVE:To observe the survival and differentiation of human embryonic NSCs following implantation into rats with fluid percussion brain injury.DESIGN:Open experiment.SETTING:Department of Neurosurgery,Guangdong Provincial Hospital of Traditional Chinese Medicine;Beijing Institute of Neurosurgery.MATERIALS:This experiment was carried out In the Laboratory of Neural Stem Cells,Beijing Institute of Neurosurgery from September 2002 to March 2003.Twenty-four female SD rats,aged 7 weeks,with body mass of(250±10)g,were provided by the Experimental Animal lnstitute, Chinese Academy of Medical Sciences[License Nd. SCXK(Jing)2002-2003]. Cerebrum of 8-week aborted fetus was obtained (Informed consents were obtained from parturients and their relatives). Fetal survival was monitored by B ultrasonic wave during abortion. BrdU monoclonal antibody(Sigma Company),rabbit anti-nidogen polyclonal antibody(Chemicon Company),mouse anti-microtubule-associated protein 2 (MAP-2)monoclonal antibody(Neomarkers Company),rabbit anti-gliaI fibrillary acidic protein(GFAP)polyclonal antibody (Biogenex Company).METHODS:① CerebraI cortex cells of 8-week aborted human fetus was harvested and cultured in vitro for obtaining human embryronic NSCs.②Rat models of hydraulic impact injury were developed.Bone window of motor sensory area of cerebral cortex was set at 2.5mm posterior to bregma which was zero point and 3.0 mm lateral to midline.Hydraulic impact injury parameters were set as impact pressure 0.3 MPa. impact time 25 ms and impact time

  12. A track-event theory of cell survival

    Energy Technology Data Exchange (ETDEWEB)

    Besserer, Juergen; Schneider, Uwe [Zuerich Univ. (Switzerland). Inst. of Physics; Radiotherapy Hirslanden, Zuerich (Switzerland)

    2015-09-01

    When fractionation schemes for hypofractionation and stereotactic body radiotherapy are considered, a reliable cell survival model at high dose is needed for calculating doses of similar biological effectiveness. In this work a simple model for cell survival which is valid also at high dose is developed from Poisson statistics. An event is defined by two double strand breaks (DSB) on the same or different chromosomes. An event is always lethal due to direct lethal damage or lethal binary misrepair by the formation of chromosome aberrations. Two different mechanisms can produce events: one-track events (OTE) or two-track-events (TTE). The target for an OTE is always a lethal event, the target for an TTE is one DSB. At least two TTEs on the same or different chromosomes are necessary to produce an event. Both, the OTE and the TTE are statistically independent. From the stochastic nature of cell kill which is described by the Poisson distribution the cell survival probability was derived. It was shown that a solution based on Poisson statistics exists for cell survival. It exhibits exponential cell survival at high dose and a finite gradient of cell survival at vanishing dose, which is in agreement with experimental cell studies. The model fits the experimental data nearly as well as the three-parameter formula of Hug-Kellerer and is only based on two free parameters. It is shown that the LQ formalism is an approximation of the model derived in this work. It could be also shown that the derived model predicts a fractionated cell survival experiment better than the LQ-model. It was shown that cell survival can be described with a simple analytical formula on the basis of Poisson statistics. This solution represents in the limit of large dose the typical exponential behavior and predicts cell survival after fractionated dose application better than the LQ-model.

  13. The role of estrogen in the survival of ovarian tumors—A study of the human ovarian adenocarcinoma cell lines OC-117-VGH and OVCAR3

    Directory of Open Access Journals (Sweden)

    Kuan-Chong Chao

    2013-02-01

    Conclusion: Although the ER-positive and ER-negative ovarian cancer cell lines were inhibited by estrogen, the influence of cell-cycle regulatory proteins was different between the two, suggesting that the inhibitory effect of estrogen on ovarian cancer cell lines might be mediated through different pathways.

  14. The Mass Media of Entertainment and Human Survival.

    Science.gov (United States)

    Gorney, Roderic; Steele, Gary

    Urgently needed for human survival is a means of influencing large numbers of people to put into rapid action measures which could neutralize such menances as pollution, overpopulation, and violence. Though the cumulative effect of the mass media is not fully established, media entertainment may be the most influential institution in our society.…

  15. The Mass Media of Entertainment and Human Survival.

    Science.gov (United States)

    Gorney, Roderic; Steele, Gary

    Urgently needed for human survival is a means of influencing large numbers of people to put into rapid action measures which could neutralize such menances as pollution, overpopulation, and violence. Though the cumulative effect of the mass media is not fully established, media entertainment may be the most influential institution in our society.…

  16. Restoring KLF5 in esophageal squamous cell cancer cells activates the JNK pathway leading to apoptosis and reduced cell survival.

    Science.gov (United States)

    Tarapore, Rohinton S; Yang, Yizeng; Katz, Jonathan P

    2013-05-01

    Esophageal cancer is the eighth most common cancer in the world and has an extremely dismal prognosis, with a 5-year survival of less than 20%. Current treatment options are limited, and thus identifying new molecular targets and pathways is critical to derive novel therapies. Worldwide, more than 90% of esophageal cancers are esophageal squamous cell cancer (ESCC). Previously, we identified that Krüppel-like factor 5 (KLF5), a key transcriptional regulator normally expressed in esophageal squamous epithelial cells, is lost in human ESCC. To examine the effects of restoring KLF5 in ESCC, we transduced the human ESCC cell lines TE7 and TE15, both of which lack KLF5 expression, with retrovirus to express KLF5 upon doxycycline induction. When KLF5 was induced, ESCC cells demonstrated increased apoptosis and decreased viability, with up-regulation of the proapoptotic factor BAX. Interestingly, c-Jun N-terminal kinase (JNK) signaling, an important upstream mediator of proapoptotic pathways including BAX, was also activated following KLF5 induction. KLF5 activation of JNK signaling was mediated by KLF5 transactivation of two key upstream regulators of the JNK pathway, ASK1 and MKK4, and inhibition of JNK blocked apoptosis and normalized cell survival following KLF5 induction. Thus, restoring KLF5 in ESCC cells promotes apoptosis and decreases cell survival in a JNK-dependent manner, providing a potential therapeutic target for human ESCC.

  17. Lose to win: marT pseudogenization in Salmonella enterica serovar Typhi contributed to the surV-dependent survival to H2O2, and inside human macrophage-like cells.

    Science.gov (United States)

    Ortega, A P; Villagra, N A; Urrutia, I M; Valenzuela, L M; Talamilla-Espinoza, A; Hidalgo, A A; Rodas, P I; Gil, F; Calderón, I L; Paredes-Sabja, D; Mora, G C; Fuentes, J A

    2016-11-01

    The difference in host range between Salmonella enterica serovar Typhimurium (S. Typhimurium) and Salmonella enterica serovar Typhi (S. Typhi) can be partially attributed to the gain of functions, to the loss of functions (i.e. pseudogenization), or to a combination of both processes. As previously reported, the loss of functions by pseudogenization may play a role in bacterial evolution, especially in host-restricted pathogens such as S. Typhi. The marT-fidL operon, located at the SPI-3, encodes the MarT transcriptional regulator and a hypothetical protein (i.e. FidL) with no significant similarities to known proteins, respectively. Even though predicted S. Typhimurium FidL exhibit 99.4% identity with S. Typhi FidL, marT has been annotated as a pseudogene in S. Typhi. In this work, we found that S. Typhi expressing S. Typhimurium marT-fidL exhibited an increased accumulation of reactive oxygen species (ROS), leading to a decreased survival in presence of H2O2. Moreover, we found that that the presence of a functional copy of S. Typhimurium marT-fidL in S. Typhi resulted in a repression of surV (STY4039), an ORF found in the S. Typhi SPI-3 but absent from S. Typhimurium SPI-3, that contribute to the resistance to H2O2 by decreasing the accumulation of ROS. Finally, we observed that the presence of S. Typhimurium marT-fidL in S. Typhi negatively affected the survival inside macrophage-like cells, but not in epithelial cells, after 24h post infection. Therefore, this work provides evidence arguing that marT pseudogenization in Salmonella Typhi contributed to the surV-dependent survival against H2O2, and inside human macrophage-like cells. This is a good example of how the loss of functions (marT pseudogenization) and the gain of functions (presence of surV) might contribute to phenotypic changes improving virulence.

  18. Multifaceted role of prohibitin in cell survival and apoptosis.

    Science.gov (United States)

    Peng, Ya-Ting; Chen, Ping; Ouyang, Ruo-Yun; Song, Lei

    2015-09-01

    Human eukaryotic prohibitin (prohibitin-1 and prohibitin-2) is a membrane protein with different cellular localizations. It is involved in multiple cellular functions, including energy metabolism, proliferation, apoptosis, and senescence. The subcellular localization of prohibitin may determine its functions. Membrane prohibitin regulate the cellular signaling of membrane transport, nuclear prohibitin control transcription activation and the cell cycle, and mitochondrial prohibitin complex stabilize the mitochondrial genome and modulate mitochondrial dynamics, mitochondrial morphology, mitochondrial biogenesis, and the mitochondrial intrinsic apoptotic pathway. Moreover, prohibitin can translocates into the nucleus or the mitochondria under apoptotic signals and the subcellular shuttling of prohibitin is necessary for apoptosis process. Apoptosis is the process of programmed cell death that is important for the maintenance of normal physiological functions. Consequently, any alteration in the content, post-transcriptional modification (i.e. phosphorylation) or the nuclear or mitochondrial translocation of prohibitin may influence cell fate. Understanding the mechanisms of the expression and regulation of prohibitin may be useful for future research. This review provides an overview of the multifaceted and essential roles played by prohibitin in the regulation of cell survival and apoptosis.

  19. Biphasic Dependence of Glioma Survival and Cell Migration on CD44 Expression Level.

    Science.gov (United States)

    Klank, Rebecca L; Decker Grunke, Stacy A; Bangasser, Benjamin L; Forster, Colleen L; Price, Matthew A; Odde, Thomas J; SantaCruz, Karen S; Rosenfeld, Steven S; Canoll, Peter; Turley, Eva A; McCarthy, James B; Ohlfest, John R; Odde, David J

    2017-01-03

    While several studies link the cell-surface marker CD44 to cancer progression, conflicting results show both positive and negative correlations with increased CD44 levels. Here, we demonstrate that the survival outcomes of genetically induced glioma-bearing mice and of high-grade human glioma patients are biphasically correlated with CD44 level, with the poorest outcomes occurring at intermediate levels. Furthermore, the high-CD44-expressing mesenchymal subtype exhibited a positive trend of survival with increased CD44 level. Mouse cell migration rates in ex vivo brain slice cultures were also biphasically associated with CD44 level, with maximal migration corresponding to minimal survival. Cell simulations suggest that cell-substrate adhesiveness is sufficient to explain this biphasic migration. More generally, these results highlight the potential importance of non-monotonic relationships between survival and biomarkers associated with cancer progression.

  20. Ras and Rheb Signaling in Survival and Cell Death

    Energy Technology Data Exchange (ETDEWEB)

    Ehrkamp, Anja [Molecular Neurobiochemistry, Ruhr University of Bochum, 44780 Bochum (Germany); Herrmann, Christian [Department of Physical Chemistry1, Protein Interaction, Ruhr University of Bochum, 44780 Bochum (Germany); Stoll, Raphael [Biomolecular NMR, Ruhr University of Bochum, 44780 Bochum (Germany); Heumann, Rolf, E-mail: rolf.heumann@rub.de [Molecular Neurobiochemistry, Ruhr University of Bochum, 44780 Bochum (Germany)

    2013-05-28

    One of the most obvious hallmarks of cancer is uncontrolled proliferation of cells partly due to independence of growth factor supply. A major component of mitogenic signaling is Ras, a small GTPase. It was the first identified human protooncogene and is known since more than three decades to promote cellular proliferation and growth. Ras was shown to support growth factor-independent survival during development and to protect from chemical or mechanical lesion-induced neuronal degeneration in postmitotic neurons. In contrast, for specific patho-physiological cases and cellular systems it has been shown that Ras may also promote cell death. Proteins from the Ras association family (Rassf, especially Rassf1 and Rassf5) are tumor suppressors that are activated by Ras-GTP, triggering apoptosis via e.g., activation of mammalian sterile 20-like (MST1) kinase. In contrast to Ras, their expression is suppressed in many types of tumours, which makes Rassf proteins an exciting model for understanding the divergent effects of Ras activity. It seems likely that the outcome of Ras signaling depends on the balance between the activation of its various downstream effectors, thus determining cellular fate towards either proliferation or apoptosis. Ras homologue enriched in brain (Rheb) is a protein from the Ras superfamily that is also known to promote proliferation, growth, and regeneration through the mammalian target of rapamycin (mTor) pathway. However, recent evidences indicate that the Rheb-mTor pathway may switch its function from a pro-growth into a cell death pathway, depending on the cellular situation. In contrast to Ras signaling, for Rheb, the cellular context is likely to modulate the whole Rheb-mTor pathway towards cellular death or survival, respectively.

  1. Autophagy-related proteins are functionally active in human spermatozoa and may be involved in the regulation of cell survival and motility

    Science.gov (United States)

    Aparicio, I. M.; Espino, J.; Bejarano, I.; Gallardo-Soler, A.; Campo, M. L.; Salido, G. M.; Pariente, J. A.; Peña, F. J.; Tapia, J. A.

    2016-01-01

    Macroautophagy (hereafter autophagy) is an evolutionarily highly conserved cellular process that participates in the maintenance of intracellular homeostasis through the degradation of most long-lived proteins and entire organelles. Autophagy participates in some reproductive events; however, there are not reports regarding the role of autophagy in the regulation of sperm physiology. Hence, the aim of this study was to investigate whether autophagy-related proteins are present and functionally active in human spermatozoa. Proteins related to autophagy/mitophagy process (LC3, Atg5, Atg16, Beclin 1, p62, m-TOR, AMPKα 1/2, and PINK1) were present in human spermatozoa. LC3 colocalized with p62 in the middle piece of the spermatozoa. Autophagy activation induced a significant increase in motility and a decrease in PINK1, TOM20 expression and caspase 3/7 activation. In contrast, autophagy inhibition resulted in decreased motility, viability, ATP and intracellular calcium concentration whereas PINK1, TOM20 expression, AMPK phosphorylation and caspase 3/7 activation were significantly increased. In conclusion our results show that autophagy related proteins and upstream regulators are present and functional in human spermatozoa. Modification of mitochondrial proteins expression after autophagy activation/inhibition may be indicating that a specialized form of autophagy named mitophagy may be regulating sperm function such as motility and viability and may be cooperating with apoptosis. PMID:27633131

  2. Properties of lewis lung carcinoma cells surviving curcumin toxicity.

    Science.gov (United States)

    Yan, Dejun; Geusz, Michael E; Jamasbi, Roudabeh J

    2012-01-01

    The anti-inflammatory agent curcumin can selectively eliminate malignant rather than normal cells. The present study examined the effects of curcumin on the Lewis lung carcinoma (LLC) cell line and characterized a subpopulation surviving curcumin treatments. Cell density was measured after curcumin was applied at concentrations between 10 and 60 μM for 30 hours. Because of the high cell loss at 60 μM, this dose was chosen to select for surviving cells that were then used to establish a new cell line. The resulting line had approximately 20% slower growth than the original LLC cell line and based on ELISA contained less of two markers, NF-κB and ALDH1A, used to identify more aggressive cancer cells. We also injected cells from the original and surviving lines subcutaneously into syngeneic C57BL/6 mice and monitored tumor development over three weeks and found that the curcumin surviving-line remained tumorigenic. Because curcumin has been reported to kill cancer cells more effectively when administered with light, we examined this as a possible way of enhancing the efficacy of curcumin against LLC cells. When LLC cells were exposed to curcumin and light from a fluorescent lamp source, cell loss caused by 20 μM curcumin was enhanced by about 50%, supporting a therapeutic use of curcumin in combination with white light. This study is the first to characterize a curcumin-surviving subpopulation among lung cancer cells. It shows that curcumin at a high concentration either selects for an intrinsically less aggressive cell subpopulation or generates these cells. The findings further support a role for curcumin as an adjunct to traditional chemical or radiation therapy of lung and other cancers.

  3. Stem cell death and survival in heart regeneration and repair.

    Science.gov (United States)

    Abdelwahid, Eltyeb; Kalvelyte, Audrone; Stulpinas, Aurimas; de Carvalho, Katherine Athayde Teixeira; Guarita-Souza, Luiz Cesar; Foldes, Gabor

    2016-03-01

    Cardiovascular diseases are major causes of mortality and morbidity. Cardiomyocyte apoptosis disrupts cardiac function and leads to cardiac decompensation and terminal heart failure. Delineating the regulatory signaling pathways that orchestrate cell survival in the heart has significant therapeutic implications. Cardiac tissue has limited capacity to regenerate and repair. Stem cell therapy is a successful approach for repairing and regenerating ischemic cardiac tissue; however, transplanted cells display very high death percentage, a problem that affects success of tissue regeneration. Stem cells display multipotency or pluripotency and undergo self-renewal, however these events are negatively influenced by upregulation of cell death machinery that induces the significant decrease in survival and differentiation signals upon cardiovascular injury. While efforts to identify cell types and molecular pathways that promote cardiac tissue regeneration have been productive, studies that focus on blocking the extensive cell death after transplantation are limited. The control of cell death includes multiple networks rather than one crucial pathway, which underlies the challenge of identifying the interaction between various cellular and biochemical components. This review is aimed at exploiting the molecular mechanisms by which stem cells resist death signals to develop into mature and healthy cardiac cells. Specifically, we focus on a number of factors that control death and survival of stem cells upon transplantation and ultimately affect cardiac regeneration. We also discuss potential survival enhancing strategies and how they could be meaningful in the design of targeted therapies that improve cardiac function.

  4. [Survival of the fattest: the key to human brain evolution].

    Science.gov (United States)

    Cunnane, Stephen C

    2006-01-01

    The circumstances of human brain evolution are of central importance to accounting for human origins, yet are still poorly understood. Human evolution is usually portrayed as having occurred in a hot, dry climate in East Africa where the earliest human ancestors became bipedal and evolved tool-making skills and language while struggling to survive in a wooded or savannah environment. At least three points need to be recognised when constructing concepts of human brain evolution : (1) The human brain cannot develop normally without a reliable supply of several nutrients, notably docosahexaenoic acid, iodine and iron. (2) At term, the human fetus has about 13 % of body weight as fat, a key form of energy insurance supporting brain development that is not found in other primates. (3) The genome of humans and chimpanzees is human brain become so much larger, and how was its present-day nutritional vulnerability circumvented during 5-6 million years of hominid evolution ? The abundant presence of fish bones and shellfish remains in many African hominid fossil sites dating to 2 million years ago implies human ancestors commonly inhabited the shores, but this point is usually overlooked in conceptualizing how the human brain evolved. Shellfish, fish and shore-based animals and plants are the richest dietary sources of the key nutrients needed by the brain. Whether on the shores of lakes, marshes, rivers or the sea, the consumption of most shore-based foods requires no specialized skills or tools. The presence of key brain nutrients and a rich energy supply in shore-based foods would have provided the essential metabolic and nutritional support needed to gradually expand the hominid brain. Abundant availability of these foods also provided the time needed to develop and refine proto-human attributes that subsequently formed the basis of language, culture, tool making and hunting. The presence of body fat in human babies appears to be the product of a long period of

  5. Long-term survival in small-cell lung cancer

    DEFF Research Database (Denmark)

    Lassen, U; Osterlind, K; Hansen, M

    1995-01-01

    PURPOSE: To describe in patients with small-cell lung cancer (SCLC) the characteristics of those who survive for > or = 5 years, to identify long-term prognostic factors, to analyze survival data of 5-year survivors, and to study 10-year survival in patients entered before 1981. PATIENTS AND METH......PURPOSE: To describe in patients with small-cell lung cancer (SCLC) the characteristics of those who survive for > or = 5 years, to identify long-term prognostic factors, to analyze survival data of 5-year survivors, and to study 10-year survival in patients entered before 1981. PATIENTS......, liver and bone marrow metastases, and elevated lactate dehydrogenase (LDH) and alkaline phosphatase levels were all negative prognostic factors. The 5-year survival rate was 3.5% (limited-stage disease, 4.8%; extensive-stage disease, 2.3%), and the 10-year survival rate was 1.8% (limited-stage disease......, especially tobacco-related cancers and other tobacco-related diseases....

  6. Stable shRNA Silencing of Lactate Dehydrogenase A (LDHA) in Human MDA-MB-231 Breast Cancer Cells Fails to Alter Lactic Acid Production, Glycolytic Activity, ATP or Survival.

    Science.gov (United States)

    Mack, Nzinga; Mazzio, Elizabeth A; Bauer, David; Flores-Rozas, Hernan; Soliman, Karam F A

    2017-03-01

    In the US, African Americans have a high death rate from triple-negative breast cancer (TNBC), characterized by lack of hormone receptors (ER, PR, HER2/ERRB2) which are otherwise valuable targets of chemotherapy. There is a need to identify novel targets that negatively impact TNBC tumorigenesis. TNBCs release an abundance of lactic acid, under normoxic, hypoxic and hyperoxic conditions; this referred to as the Warburg effect. Accumulated lactic acid sustains peri-cellular acidity which propels metastatic invasion and malignant aggressive transformation. The source of lactic acid is believed to be via conversion of pyruvate by lactate dehydrogenase (LDH) in the last step of glycolysis, with most studies focusing on the LDHA isoform. In this study, LDHA was silenced using long-term MISSION® shRNA lentivirus in human breast cancer MDA-MB-231 cells. Down-regulation of LDHA transcription and protein expression was confirmed by western blot, immunocytochemistry and qPCR. A number of parameters were measured in fully viable vector controls versus knock-down (KD) clones, including levels of lactic acid produced, glucose consumed, ATP and basic metabolic rates. The data show that lentivirus V-165 generated a knock-down clone most effective in reducing both gene and protein levels to less than 1% of vector controls. Stable KD showed absolutely no changes in cell viability, lactic acid production, ATP, glucose consumption or basic metabolic rate. Given the complete absence of impact on any observed parameter by LDH-A KD and this being somewhat contrary to findings in the literature, further analysis was required to determine why. Whole-transcriptome analytic profile on MDA-MB-231 for LDH subtypes using Agilent Human Genome 4×44k microarrays, where the data show the following component breakdown. Transcripts: 30.47 % LDHA, 69.36% LDHB, 0.12% LDHC and 0.05% LDHD. These findings underscore the importance of alternative isoforms of LDH in cancer cells to produce lactic acid

  7. Mitochondria in the Center of Human Eosinophil Apoptosis and Survival

    Directory of Open Access Journals (Sweden)

    Pinja Ilmarinen

    2014-03-01

    Full Text Available Eosinophils are abundantly present in most phenotypes of asthma and they contribute to the maintenance and exacerbations of the disease. Regulators of eosinophil longevity play critical roles in determining whether eosinophils accumulate into the airways of asthmatics. Several cytokines enhance eosinophil survival promoting eosinophilic airway inflammation while for example glucocorticoids, the most important anti-inflammatory drugs used to treat asthma, promote the intrinsic pathway of eosinophil apoptosis and by this mechanism contribute to the resolution of eosinophilic airway inflammation. Mitochondria seem to play central roles in both intrinsic mitochondrion-centered and extrinsic receptor-mediated pathways of apoptosis in eosinophils. Mitochondria may also be important for survival signalling. In addition to glucocorticoids, another important agent that regulates human eosinophil longevity via mitochondrial route is nitric oxide, which is present in increased amounts in the airways of asthmatics. Nitric oxide seems to be able to trigger both survival and apoptosis in eosinophils. This review discusses the current evidence of the mechanisms of induced eosinophil apoptosis and survival focusing on the role of mitochondria and clinically relevant stimulants, such as glucocorticoids and nitric oxide.

  8. Recombinant human erythropoietin increases survival and reduces neuronal apoptosis in a murine model of cerebral malaria

    DEFF Research Database (Denmark)

    Wiese, Lothar; Hempel, Casper; Penkowa, Milena;

    2008-01-01

    with recombinant human Epo (rhEpo; 50-5000 U/kg/OD, i.p.) at different time points. The effect on survival was measured. Brain pathology was investigated by TUNEL (Terminal deoxynucleotidyl transferase (TdT)-mediated deoxyuridine triphosphate (dUTP)-digoxigenin nick end labelling), as a marker of apoptosis. Gene...... expression in brain tissue was measured by real time PCR. RESULTS: Treatment with rhEpo increased survival in mice with CM in a dose- and time-dependent manner and reduced apoptotic cell death of neurons as well as the expression of pro-inflammatory cytokines in the brain. This neuroprotective effect...

  9. Cell Biology: ERADicating Survival with BOK.

    Science.gov (United States)

    Chipuk, Jerry Edward; Luna-Vargas, Mark P

    2016-06-01

    Mechanistic insights into the function of the pro-apoptotic BCL-2 family member BOK have remained elusive. A recent study shows that healthy cells constitutively degrade BOK via the ER-associated degradation and ubiquitin-proteasome pathways; following proteasome inhibition, BOK is stabilized to initiate a unique pro-apoptotic death program.

  10. DDX3X Biomarker Correlates with Poor Survival in Human Gliomas

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    Dueng-Yuan Hueng

    2015-07-01

    Full Text Available Primary high-grade gliomas possess invasive growth and lead to unfavorable survival outcome. The investigation of biomarkers for prediction of survival outcome in patients with gliomas is important for clinical assessment. The DEAD (Asp-Glu-Ala-Asp box helicase 3, X-linked (DDX3X controls tumor migration, proliferation, and progression. However, the role of DDX3X in defining the pathological grading and survival outcome in patients with human gliomas is not yet clarified. We analyzed the DDX3X gene expression, WHO pathological grading, and overall survival from de-linked data. Further validation was done using quantitative RT-PCR of cDNA from normal brain and glioma, and immunohistochemical (IHC staining of tissue microarray. Statistical analysis of GEO datasets showed that DDX3X mRNA expression demonstrated statistically higher in WHO grade IV (n = 81 than in non-tumor controls (n = 23, p = 1.13 × 10−10. Moreover, DDX3X level was also higher in WHO grade III (n = 19 than in non-tumor controls (p = 2.43 × 10−5. Kaplan–Meier survival analysis showed poor survival in patients with high DDX3X mRNA levels (n = 24 than in those with low DDX3X expression (n = 53 (median survival, 115 vs. 58 weeks, p = 0.0009, by log-rank test, hazard ratio: 0.3507, 95% CI: 0.1893–0.6496. Furthermore, DDX3X mRNA expression and protein production significantly increased in glioma cells compared with normal brain tissue examined by quantitative RT-PCR, and Western blot. IHC staining showed highly staining of high-grade glioma in comparison with normal brain tissue. Taken together, DDX3X expression level positively correlates with WHO pathologic grading and poor survival outcome, indicating that DDX3X is a valuable biomarker in human gliomas.

  11. Illicit survival of cancer cells during polyploidization and depolyploidization.

    Science.gov (United States)

    Vitale, I; Galluzzi, L; Senovilla, L; Criollo, A; Jemaà, M; Castedo, M; Kroemer, G

    2011-09-01

    Tetraploidy and the depolyploidization of tetraploid cells may contribute to oncogenesis. Several mechanisms have evolved to avoid the generation, survival, proliferation and depolyploidization of tetraploids. Cells that illicitly survive these checkpoints are prone to chromosomal instability and aneuploidization. Along with their replication, tetraploids constantly undergo chromosomal rearrangements that eventually lead to pseudodiploidy by two non-exclusive mechanisms: (i) multipolar divisions and (ii) illicit bipolar divisions in the presence of improper microtubule-kinetochore attachments. Here, we describe the regulation and the molecular mechanisms that underlie such a 'polyploidization-depolyploidization' cascade, while focusing on the role of oncogenes and tumor suppressor genes in tetraploidy-driven tumorigenesis. We speculate that the identification of signaling/metabolic cascades that are required for the survival of tetraploid or aneuploid (but not diploid) cancer cells may pave the way for the development of novel broad-spectrum anticancer agents.

  12. Homeostatic responses by surviving cortical pyramidal cells in neurodegenerative tauopathy.

    Science.gov (United States)

    Crimins, Johanna L; Rocher, Anne B; Peters, Alan; Shultz, Penny; Lewis, Jada; Luebke, Jennifer I

    2011-11-01

    Cortical neuron death is prevalent by 9 months in rTg(tau(P301L))4510 tau mutant mice (TG) and surviving pyramidal cells exhibit dendritic regression and spine loss. We used whole-cell patch-clamp recordings to investigate the impact of these marked structural changes on spontaneous excitatory and inhibitory postsynaptic currents (sEPSCs and sIPSCs) of layer 3 pyramidal cells in frontal cortical slices from behaviorally characterized TG and non-transgenic (NT) mice at this age. Frontal lobe function of TG mice was intact following a short delay interval but impaired following a long delay interval in an object recognition test, and cortical atrophy and cell loss were pronounced. Surviving TG cells had significantly reduced dendritic diameters, total spine density, and mushroom spines, yet sEPSCs were increased and sIPSCs were unchanged in frequency. Thus, despite significant regressive structural changes, synaptic responses were not reduced in TG cells, indicating that homeostatic compensatory mechanisms occur during progressive tauopathy. Consistent with this idea, surviving TG cells were more intrinsically excitable than NT cells, and exhibited sprouting of filopodia and axonal boutons. Moreover, the neuropil in TG mice showed an increased density of asymmetric synapses, although their mean size was reduced. Taken together, these data indicate that during progressive tauopathy, cortical pyramidal cells compensate for loss of afferent input by increased excitability and establishment of new synapses. These compensatory homeostatic mechanisms may play an important role in slowing the progression of neuronal network dysfunction during neurodegenerative tauopathies.

  13. Is Human Papillomavirus Associated with Prostate Cancer Survival?

    Directory of Open Access Journals (Sweden)

    Mariarosa Pascale

    2013-01-01

    Full Text Available The role of human papillomavirus (HPV in prostate carcinogenesis is highly controversial: some studies suggest a positive association between HPV infection and an increased risk of prostate cancer (PCa, whereas others do not reveal any correlation. In this study, we investigated the prognostic impact of HPV infection on survival in 150 primary PCa patients. One hundred twelve (74.67% patients had positive expression of HPV E7 protein, which was evaluated in tumour tissue by immunohistochemistry. DNA analysis on a subset of cases confirmed HPV infection and revealed the presence of genotype 16. In Kaplan-Meier analysis, HPV-positive cancer patients showed worse overall survival (OS (median 4.59 years compared to HPV-negative (median 8.24 years, P=0.0381. In multivariate analysis age (P<0.001, Gleason score (P<0.001, nuclear grading (P=0.002, and HPV status (P=0.034 were independent prognostic factors for OS. In our cohort, we observed high prevalence of HPV nuclear E7 oncoprotein and an association between HPV infection and PCa survival. In the debate about the oncogenic activity of HPV in PCa, our results further confirm the need for additional studies to clarify the possible role of HPV in prostate carcinogenesis.

  14. Human mesenchymal stem cells

    DEFF Research Database (Denmark)

    Abdallah, Basem; Kassem, Moustapha

    2008-01-01

    Mesenchymal stem cells (MSC) are a group of clonogenic cells present among the bone marrow stroma and capable of multilineage differentiation into mesoderm-type cells such as osteoblasts, adipocytes and chondrocytes. Due to their ease of isolation and their differentiation potential, MSC are being...... introduced into clinical medicine in variety of applications and through different ways of administration. Here, we discuss approaches for isolation, characterization and directing differentiation of human mesenchymal stem cells (hMSC). An update of the current clinical use of the cells is also provided....

  15. Veratridine increases the survival of retinal ganglion cells in vitro

    Directory of Open Access Journals (Sweden)

    S.P.F. Pereira

    1997-12-01

    Full Text Available Neuronal cell death is an important phenomenon involving many biochemical pathways. This degenerative event has been studied to understand how the cells activate the mechanisms that lead to self-destruction. Target cells and afferent cells play a relevant role in the regulation of natural cell death. We studied the effect of veratridine (1.5, 3.0, 4.5 and 6.0 µM on the survival of neonatal rat retinal ganglion cells in vitro. Veratridine (3.0 µM, a well-known depolarizing agent that opens the Na+ channel, promoted a two-fold increase in the survival of retinal ganglion cells kept in culture for 48 h. This effect was dose-dependent and was blocked by 1.0 µM tetrodotoxin (a classical voltage-dependent Na+ channel blocker and 30.0 µM flunarizine (a Na+ and Ca2+ channel blocker. These results indicate that electrical activity is also important for the maintenance of retinal ganglion cell survival in vitro

  16. Autophagy as a Survival Mechanism for Squamous Cell Carcinoma Cells in Endonuclease G-Mediated Apoptosis

    Science.gov (United States)

    Masui, Atsushi; Hamada, Masakazu; Kameyama, Hiroyasu; Wakabayashi, Ken; Takasu, Ayako; Imai, Tomoaki; Iwai, Soichi; Yura, Yoshiaki

    2016-01-01

    Safingol, L- threo-dihydrosphingosine, induces cell death in human oral squamous cell carcinoma (SCC) cells through an endonuclease G (endoG) -mediated pathway. We herein determined whether safingol induced apoptosis and autophagy in oral SCC cells. Safingol induced apoptotic cell death in oral SCC cells in a dose-dependent manner. In safingol-treated cells, microtubule-associated protein 1 light chain 3 (LC3)-I was changed to LC3-II and the cytoplasmic expression of LC3, amount of acidic vesicular organelles (AVOs) stained by acridine orange and autophagic vacuoles were increased, indicating the occurrence of autophagy. An inhibitor of autophagy, 3-methyladenine (3-MA), enhanced the suppressive effects of safingol on cell viability, and this was accompanied by an increase in the number of apoptotic cells and extent of nuclear fragmentation. The nuclear translocation of endoG was minimal at a low concentration of safingol, but markedly increased when combined with 3-MA. The suppressive effects of safingol and 3-MA on cell viability were reduced in endoG siRNA- transfected cells. The scavenging of reactive oxygen species (ROS) prevented cell death induced by the combinational treatment, whereas a pretreatment with a pan-caspase inhibitor z-VAD-fmk did not. These results indicated that safingol induced apoptosis and autophagy in SCC cells and that the suppression of autophagy by 3-MA enhanced apoptosis. Autophagy supports cell survival, but not cell death in the SCC cell system in which apoptosis occurs in an endoG-mediated manner. PMID:27658240

  17. Nanosecond pulsed electric field induced cytoskeleton, nuclear membrane and telomere damage adversely impact cell survival.

    Science.gov (United States)

    Stacey, M; Fox, P; Buescher, S; Kolb, J

    2011-10-01

    We investigated the effects of nanosecond pulsed electric fields (nsPEF) on three human cell lines and demonstrated cell shrinkage, breakdown of the cytoskeleton, nuclear membrane and chromosomal telomere damage. There was a differential response between cell types coinciding with cell survival. Jurkat cells showed cytoskeleton, nuclear membrane and telomere damage that severely impacted cell survival compared to two adherent cell lines. Interestingly, disruption of the actin cytoskeleton in adherent cells prior to nsPEF exposure significantly reduced cell survival. We conclude that nsPEF applications are able to induce damage to the cytoskeleton and nuclear membrane. Telomere sequences, regions that tether and stabilize DNA to the nuclear membrane, are severely compromised as measured by a pan-telomere probe. Internal pore formation following nsPEF applications has been described as a factor in induced cell death. Here we suggest that nsPEF induced physical changes to the cell in addition to pore formation need to be considered as an alternative method of cell death. We suggest nsPEF electrochemical induced depolymerization of actin filaments may account for cytoskeleton and nuclear membrane anomalies leading to sensitization.

  18. Survival

    Data.gov (United States)

    U.S. Geological Survey, Department of the Interior — These data provide information on the survival of California red-legged frogs in a unique ecosystem to better conserve this threatened species while restoring...

  19. Effects of Triclosan on Neural Stem Cell Viability and Survival.

    Science.gov (United States)

    Park, Bo Kyung; Gonzales, Edson Luck T; Yang, Sung Min; Bang, Minji; Choi, Chang Soon; Shin, Chan Young

    2016-01-01

    Triclosan is an antimicrobial or sanitizing agent used in personal care and household products such as toothpaste, soaps, mouthwashes and kitchen utensils. There are increasing evidence of the potentially harmful effects of triclosan in many systemic and cellular processes of the body. In this study, we investigated the effects of triclosan in the survivability of cultured rat neural stem cells (NSCs). Cortical cells from embryonic day 14 rat embryos were isolated and cultured in vitro. After stabilizing the culture, triclosan was introduced to the cells with concentrations ranging from 1 μM to 50 μM and in varied time periods. Thereafter, cell viability parameters were measured using MTT assay and PI staining. TCS decreased the cell viability of treated NSC in a concentration-dependent manner along with increased expressions of apoptotic markers, cleaved caspase-3 and Bax, while reduced expression of Bcl2. To explore the mechanisms underlying the effects of TCS in NSC, we measured the activation of MAPKs and intracellular ROS. TCS at 50 μM induced the activations of both p38 and JNK, which may adversely affect cell survival. In contrast, the activities of ERK, Akt and PI3K, which are positively correlated with cell survival, were inhibited. Moreover, TCS at this concentration augmented the ROS generation in treated NSC and depleted the glutathione activity. Taken together, these results suggest that TCS can induce neurodegenerative effects in developing rat brains through mechanisms involving ROS activation and apoptosis initiation.

  20. Astrocytes Upregulate Survival Genes in Tumor Cells and Induce Protection from Chemotherapy

    Directory of Open Access Journals (Sweden)

    Sun-Jin Kim

    2011-03-01

    Full Text Available In the United States, more than 40% of cancer patients develop brain metastasis. The median survival for untreated patients is 1 to 2 months, which may be extended to 6 months with conventional radiotherapy and chemotherapy. The growth and survival of metastasis depend on the interaction of tumor cells with host factors in the organ microenvironment. Brain metastases are surrounded and infiltrated by activated astrocytes and are highly resistant to chemotherapy. We report here that coculture of human breast cancer cells or lung cancer cells with murine astrocytes (but not murine fibroblasts led to the up-regulation of survival genes, including GSTA5, BCL2L1, and TWIST1, in the tumor cells. The degree of up-regulation directly correlated with increased resistance to all tested chemotherapeutic agents. We further show that the up-regulation of the survival genes and consequent resistance are dependent on the direct contact between the astrocytes and tumor cells through gap junctions and are therefore transient. Knocking down these genes with specific small interfering RNA rendered the tumor cells sensitive to chemotherapeutic agents. These data clearly demonstrate that host cells in the microenvironment influence the biologic behavior of tumor cells and reinforce the contention that the organ microenvironment must be taken into consideration during the design of therapy.

  1. The β-hemolysin and intracellular survival of Streptococcus agalactiae in human macrophages.

    Directory of Open Access Journals (Sweden)

    Anubha Sagar

    Full Text Available S. agalactiae (group B streptococci, GBS is a major microbial pathogen in human neonates and causes invasive infections in pregnant women and immunocompromised individuals. The S. agalactiae β-hemolysin is regarded as an important virulence factor for the development of invasive disease. To examine the role of β-hemolysin in the interaction with professional phagocytes, the THP-1 monocytic cell line and human granulocytes were infected with a serotype Ia S. agalactiae wild type strain and its isogenic nonhemolytic mutant. We could show that the nonhemolytic mutants were able to survive in significantly higher numbers than the hemolytic wild type strain, in THP-1 macrophage-like cells and in assays with human granulocytes. Intracellular bacterial multiplication, however, could not be observed. The hemolytic wild type strain stimulated a significantly higher release of Tumor Necrosis Factor-α than the nonhemolytic mutant in THP-1 cells, while similar levels of the chemokine Interleukin-8 were induced. In order to investigate bacterial mediators of IL-8 release in this setting, purified cell wall preparations from both strains were tested and found to exert a potent proinflammatory stimulus on THP-1 cells. In conclusion, our results indicate that the β-hemolysin has a strong influence on the intracellular survival of S. agalactiae and that a tightly controlled regulation of β-hemolysin expression is required for the successful establishment of S. agalactiae in different host niches.

  2. Bmi1 overexpression in the cerebellar granule cell lineage of mice affects cell proliferation and survival without initiating medulloblastoma formation

    Directory of Open Access Journals (Sweden)

    Hourinaz Behesti

    2013-01-01

    BMI1 is a potent inducer of neural stem cell self-renewal and neural progenitor cell proliferation during development and in adult tissue homeostasis. It is overexpressed in numerous human cancers – including medulloblastomas, in which its functional role is unclear. We generated transgenic mouse lines with targeted overexpression of Bmi1 in the cerebellar granule cell lineage, a cell type that has been shown to act as a cell of origin for medulloblastomas. Overexpression of Bmi1 in granule cell progenitors (GCPs led to a decrease in cerebellar size due to decreased GCP proliferation and repression of the expression of cyclin genes, whereas Bmi1 overexpression in postmitotic granule cells improved cell survival in response to stress by altering the expression of genes in the mitochondrial cell death pathway and of Myc and Lef-1. Although no medulloblastomas developed in ageing cohorts of transgenic mice, crosses with Trp53−/− mice resulted in a low incidence of medulloblastoma formation. Furthermore, analysis of a large collection of primary human medulloblastomas revealed that tumours with a BMI1high TP53low molecular profile are significantly enriched in Group 4 human medulloblastomas. Our data suggest that different levels and timing of Bmi1 overexpression yield distinct cellular outcomes within the same cellular lineage. Importantly, Bmi1 overexpression at the GCP stage does not induce tumour formation, suggesting that BMI1 overexpression in GCP-derived human medulloblastomas probably occurs during later stages of oncogenesis and might serve to enhance tumour cell survival.

  3. Genome-wide identification of genes essential for the survival of Streptococcus pneumoniae in human saliva.

    Directory of Open Access Journals (Sweden)

    Lilly M Verhagen

    Full Text Available Since Streptococcus pneumoniae transmits through droplet spread, this respiratory tract pathogen may be able to survive in saliva. Here, we show that saliva supports survival of clinically relevant S. pneumoniae strains for more than 24 h in a capsule-independent manner. Moreover, saliva induced growth of S. pneumoniae in growth-permissive conditions, suggesting that S. pneumoniae is well adapted for uptake of nutrients from this bodily fluid. By using Tn-seq, a method for genome-wide negative selection screening, we identified 147 genes potentially required for growth and survival of S. pneumoniae in saliva, among which genes predicted to be involved in cell envelope biosynthesis, cell transport, amino acid metabolism, and stress response predominated. The Tn-seq findings were validated by testing a panel of directed gene deletion mutants for their ability to survive in saliva under two testing conditions: at room temperature without CO2, representing transmission, and at 37 °C with CO2, representing in-host carriage. These validation experiments confirmed that the plsX gene and the amiACDEF and aroDEBC operons, involved in respectively fatty acid metabolism, oligopeptide transport, and biosynthesis of aromatic amino acids play an important role in the growth and survival of S. pneumoniae in saliva at 37 °C. In conclusion, this study shows that S. pneumoniae is well-adapted for growth and survival in human saliva and provides a genome-wide list of genes potentially involved in adaptation. This notion supports earlier evidence that S. pneumoniae can use human saliva as a vector for transmission.

  4. Genome-Wide Identification of Genes Essential for the Survival of Streptococcus pneumoniae in Human Saliva

    Science.gov (United States)

    Verhagen, Lilly M.; de Jonge, Marien I.; Burghout, Peter; Schraa, Kiki; Spagnuolo, Lorenza; Mennens, Svenja; Eleveld, Marc J.; van der Gaast-de Jongh, Christa E.; Zomer, Aldert; Hermans, Peter W. M.; Bootsma, Hester J.

    2014-01-01

    Since Streptococcus pneumoniae transmits through droplet spread, this respiratory tract pathogen may be able to survive in saliva. Here, we show that saliva supports survival of clinically relevant S. pneumoniae strains for more than 24 h in a capsule-independent manner. Moreover, saliva induced growth of S. pneumoniae in growth-permissive conditions, suggesting that S. pneumoniae is well adapted for uptake of nutrients from this bodily fluid. By using Tn-seq, a method for genome-wide negative selection screening, we identified 147 genes potentially required for growth and survival of S. pneumoniae in saliva, among which genes predicted to be involved in cell envelope biosynthesis, cell transport, amino acid metabolism, and stress response predominated. The Tn-seq findings were validated by testing a panel of directed gene deletion mutants for their ability to survive in saliva under two testing conditions: at room temperature without CO2, representing transmission, and at 37°C with CO2, representing in-host carriage. These validation experiments confirmed that the plsX gene and the amiACDEF and aroDEBC operons, involved in respectively fatty acid metabolism, oligopeptide transport, and biosynthesis of aromatic amino acids play an important role in the growth and survival of S. pneumoniae in saliva at 37°C. In conclusion, this study shows that S. pneumoniae is well-adapted for growth and survival in human saliva and provides a genome-wide list of genes potentially involved in adaptation. This notion supports earlier evidence that S. pneumoniae can use human saliva as a vector for transmission. PMID:24586856

  5. Metabolic pathways promoting cancer cell survival and growth.

    Science.gov (United States)

    Boroughs, Lindsey K; DeBerardinis, Ralph J

    2015-04-01

    Activation of oncogenes and loss of tumour suppressors promote metabolic reprogramming in cancer, resulting in enhanced nutrient uptake to supply energetic and biosynthetic pathways. However, nutrient limitations within solid tumours may require that malignant cells exhibit metabolic flexibility to sustain growth and survival. Here, we highlight these adaptive mechanisms and also discuss emerging approaches to probe tumour metabolism in vivo and their potential to expand the metabolic repertoire of malignant cells even further.

  6. Cancer Cell Cannibalism: A Primeval Option to Survive.

    Science.gov (United States)

    Lozupone, F; Fais, S

    2015-01-01

    Cancer cell cannibalism is currently defined as a phenomenon in which an ensemble of a larger cell containing a smaller one, often in a big cytoplasmic vacuole, is detected in either cultured tumor cells or a tumor sample. After almost one century of considering this phenomenon as a sort of neglected curiosity, some recent studies have first proposed tumor cell cannibalism as a sort of "aberrant phagocytosis", making malignant cells very similar to professional phagocytes. Later, further research has shown that, differently to macrophages, exclusively ingesting exogenous material, apoptotic bodies, or cell debris, tumor cells are able to engulf other cells, including lymphocytes and erythrocytes, either dead or alive, with the main purpose to feed on them. This phenomenon has been associated to the malignancy of tumors, mostly exclusive of metastatic cells, and often associated to poor prognosis. The cannibalistic behavior increased depending on the microenvironmental condition of tumor cells, such as low nutrient supply or low pH, suggesting its key survival option for malignant cancers. However, the evidence that malignant cells may cannibalize tumor-infiltrating lymphocytes that act as their killers, suggests that tumor cell cannibalism could be a very direct and efficient way to neutralize immune response, as well. Tumor cell cannibalism may represent a sign of regression to a simpler, ancestral or primeval life style, similar to that of unicellular microorganisms, such as amoebas, where the goal is to survive and propagate in an overcrowded and very hostile microenvironment. In fact, we discovered that metastatic melanoma cells share with amoebas a transmembrane protein TM9SF4, indeed related to the cannibal behavior of these cells. This review attempts to provide a comprehensive description of the current knowledge about the role of TM9SF4 in cancer, highlighting its role as a key player in the cannibal behavior of malignant cancer cells. Moreover, we discuss

  7. Non-small cell lung cancer cell survival crucially depends on functional insulin receptors.

    Science.gov (United States)

    Frisch, Carolin Maria; Zimmermann, Katrin; Zilleßen, Pia; Pfeifer, Alexander; Racké, Kurt; Mayer, Peter

    2015-08-01

    Insulin plays an important role as a growth factor and its contribution to tumor proliferation is intensely discussed. It acts via the cognate insulin receptor (IR) but can also activate the IGF1 receptor (IGF1R). Apart from increasing proliferation, insulin might have additional effects in lung cancer. Therefore, we investigated insulin action and effects of IR knockdown (KD) in three (NCI-H292, NCI-H226 and NCI-H460) independent non-small cell lung cancer (NSCLC) cell lines. All lung cancer lines studied were found to express IR, albeit with marked differences in the ratio of the two variants IR-A and IR-B. Insulin activated the classical signaling pathway with IR autophosphorylation and Akt phosphorylation. Moreover, activation of MAPK was observed in H292 cells, accompanied by enhanced proliferation. Lentiviral shRNA IR KD caused strong decrease in survival of all three lines, indicating that the effects of insulin in lung cancer go beyond enhancing proliferation. Unspecific effects were ruled out by employing further shRNAs and different insulin-responsive cells (human pre-adipocytes) for comparison. Caspase assays demonstrated that IR KD strongly induced apoptosis in these lung cancer cells, providing the physiological basis of the rapid cell loss. In search for the underlying mechanism, we analyzed alterations in the gene expression profile in response to IR KD. A strong induction of certain cytokines (e.g. IL20 and tumour necrosis factor) became obvious and it turned out that these cytokines trigger apoptosis in the NSCLC cells tested. This indicates a novel role of IR in tumor cell survival via suppression of pro-apoptotic cytokines.

  8. Autophagy in response to photodynamic therapy: cell survival vs. cell death

    Science.gov (United States)

    Oleinick, Nancy L.; Xue, Liang-yan; Chiu, Song-mao; Joseph, Sheeba

    2009-02-01

    Autophagy (or more properly, macroautophagy) is a pathway whereby damaged organelles or other cell components are encased in a double membrane, the autophagosome, which fuses with lysosomes for digestion by lysosomal hydrolases. This process can promote cell survival by removing damaged organelles, but when damage is extensive, it can also be a mechanism of cell death. Similar to the Kessel and Agostinis laboratories, we have reported the vigorous induction of autophagy by PDT; this was found in human breast cancer MCF-7 cells whether or not they were able to efficiently induce apoptosis. One way to evaluate the role of autophagy in PDT-treated cells is to silence one of the essential genes in the pathway. Kessel and Reiners silenced the Atg7 gene of murine leukemia L1210 cells using inhibitory RNA and found sensitization to PDT-induced cell death at a low dose of PDT, implying that autophagy is protective when PDT damage is modest. We have examined the role of autophagy in an epithelium-derived cancer cell by comparing parental and Atg7-silenced MCF-7 cells to varying doses of PDT with the phthalocyanine photosensitizer Pc 4. In contrast to L1210 cells, autophagy-deficient MCF-7 cells were more resistant to the lethal effects of PDT, as judged by clonogenic assays. A possible explanation for the difference in outcome for L1210 vs. MCF-7 cells is the greatly reduced ability of the latter to undergo apoptosis, a deficiency that may convert autophagy into a cell-death process even at low PDT doses. Experiments to investigate the mechanism(s) responsible are in process.

  9. Coniferyl Aldehyde Ameliorates Radiation Intestine Injury via Endothelial Cell Survival

    Energy Technology Data Exchange (ETDEWEB)

    Jeong, Ye Ji; Jung, Myung Gu; Lee, Yoonjin; Lee, Haejune [Korea Institute of Radiological and Medical Sciences, Seoul (Korea, Republic of); Lee, Yunsil [Ewha Woman' s Univ., Seoul (Korea, Republic of); Ko, Younggyu [Korea Univ., Seoul (Korea, Republic of)

    2014-05-15

    Cancer treatments related gastrointestinal toxicity has also been recognized as a significant economic burden. Especially, extensive apoptosis of microvascular endothelial cell of the lamina propria is the primary lesion initiating intestinal radiation damage after abdominal radiation therapy. Coniferyl aldehyde (CA) is phenolic compounds isolated from cork stoppers, and one of the major pyrolysis products of lignin. Shi H. was support for the empirical use of CA as a medicinal food for cardiovascular diseases. CA has positive effect in broad way but there is no consequence in radiation induced intestine damage. Here, we investigate effect of CA on small intestine after abdominal IR to mice in this study. In this study, CA increased the survival rate in C3H mice against 13.5 Gy abdominal IR. We found CA protects small intestine via preventing endothelial cell apoptosis and enhancing their angiogenic activity. CA also showed protective effect on crypt cell survival. Endothelial cell survival may affect crypt cell protection against IR. From this data, we concluded that CA is effective for protection against abdominal radiation injury. CA could ameliorate side-effect of radiation therapy.

  10. Anti-Inflammatory Heat Shock Protein 70 Genes are Positively Associated with Human Survival

    DEFF Research Database (Denmark)

    Singh, Ripudaman; Kølvraa, Steen; Bross, Peter Gerd;

    2010-01-01

    with longevity. The involvement of heat shock protein 70 (Hsp70) in cellular maintenance and repair mechanisms, including its role as an anti-inflammatory protein, makes it a suitable candidate for studying such associations. We have studied the association of three single nucleotide polymorphisms, HSPA1A (-110A...... the opportunity to perform survival analysis on these subjects. Haplotype relative risk, and genotype relative risk were calculated to measure the effects of haplotypes and genotypes on human survival in a sex-specific manner. A significant association of HSPA1A-AA (RR=3.864; p=0.016) and HSPA1B-AA (RR=2.764; p=0...... observations from heat shock response (HSR) study where we had shown that after heat stimulation, mononuclear cells from the carriers of genotype HSPA1L-TT had better HSR than cells with the HSPA1L-CC genotype....

  11. Mast cell leukemia with prolonged survival on PKC412/midostaurin.

    Science.gov (United States)

    Xu, Xiangdong; Kreisel, Friederike H; Frater, John L; Hassan, Anjum

    2014-01-01

    Mast cell leukemia (MCL) is a rare and aggressive form of systemic mastocytosis. There are approximately 50 reported cases since 1950s. MCL is refractory to cytoreduction chemotherapy and the average survival is only six months. We report a MCL case in a 71 year-old woman with high tumor load at the initial presentation in 2005, who did not respond to either interleukin-2 or dasatinib therapy. After enrolled in a clinical trial of PKC412 (or Midostaurin) with a daily dose of 100 mg, the patient responded well to PKC412 and became transfusion independent in three months. Since then, her disease had been stably controlled. This is the first report of a high-tumor-load MCL case which achieved prolonged survival (101 months) by PKC 412. The 101-month overall survival is the longest among reported MCL cases in the English literature.

  12. Calcium-independent phospholipase A₂, group VIA, is critical for RPE cell survival

    DEFF Research Database (Denmark)

    Kolko, Miriam; Vohra, Rupali; Westlund, Barbro S.;

    2014-01-01

    PURPOSE: To investigate the significance of calcium-independent phospholipase A₂, group VIA (iPLA2-VIA), in RPE cell survival following responses to sodium iodate (SI) in cell cultures. METHODS: The human retinal pigment epithelium (RPE) cell line (ARPE-19) cells and primary mouse-RPE cultures were...... of iPLA₂-VIA after SI exposure. Inhibitors of iPLA₂-VIA were used to explore a potential protective role in cells exposed to SI. Primary RPE cell cultures were grown from iPLA₂-VIA knockout mice and wild-type mice. The cultures were exposed to SI to investigate a possible increased protection against......-VIA-specific inhibitors in ARPE-19 cell cultures. RPE cultures from iPLA₂-VIA knockout mice were less vulnerable to SI-induced cell death compared to RPE cultures from wild-type mice. CONCLUSIONS: SI -induced RPE cell death involves iPLA₂-VIA upregulation and activation, and amelioration of SI...

  13. Hypoxia enhances metastatic efficiency in HT1080 fibrosarcoma cells by increasing cell survival in lungs, not cell adhesion and invasion.

    Science.gov (United States)

    Zhang, Li; Hill, Richard P

    2007-08-15

    This study examined possible mechanisms for hypoxia-increased metastasis in a green fluorescent protein-labeled human fibrosarcoma cell line (HT1080). The efficiency of the lung arrest of tumor cells, which can be dependent on the adhesive potential of the tumor cells, was assessed by measuring the level of integrin alpha3beta1 protein and by adhesion assays, whereas the extravasation potential was examined by an invasion assay. These properties were not changed by exposure to hypoxia, indicating that lung arrest and extravasation are unlikely to play a major role in the effect of hypoxia on metastasis in this model. The main effect of hypoxic exposure was found to be increased survival after lung arrest as determined by clonogenic assay of tumor cells recovered from mouse lungs after i.v. injection. Concomitantly, apoptosis was identified as responsible for the death of lung-arrested cells, suggesting the involvement of an altered apoptotic response following hypoxic exposure of these cells. Consistent with this finding, we found that the effect of hypoxia on both increased metastasis and survival of arrested cells was inhibited by treatment with farnesylthiosalicylic acid. However, this effect was not due to down-regulation of hypoxia-inducible factor-1alpha, a mechanism of action of this drug reported by previous studies. Further detailed studies of the mechanisms of action of the drug are needed.

  14. Improved survival of children and adolescents with sickle cell disease.

    Science.gov (United States)

    Quinn, Charles T; Rogers, Zora R; McCavit, Timothy L; Buchanan, George R

    2010-04-29

    The survival of young children with sickle cell disease (SCD) has improved, but less is known about older children and adolescents. We studied the Dallas Newborn Cohort (DNC) to estimate contemporary 18-year survival for newborns with SCD and document changes in the causes and ages of death over time. We also explored whether improvements in the quality of medical care were temporally associated with survival. The DNC now includes 940 subjects with 8857 patient-years of follow-up. Most children with sickle cell anemia (93.9%) and nearly all children with milder forms of SCD (98.4%) now live to become adults. The incidence of death and the pattern of mortality changed over the duration of the cohort. Sepsis is no longer the leading cause of death. All the recent deaths in the cohort occurred in patients 18 years or older, most shortly after the transition to adult care. Quality of care in the DNC has improved over time, with significantly more timely initial visits and preventive interventions for young children. In summary, most children with SCD now survive the childhood years, but young adults who transition to adult medical care are at high risk for early death.

  15. Survival of human-associated bacteria in SLS

    Science.gov (United States)

    Fu, Yuming; Tikhomirov, Alexander A.; Nickolay Manukovsky, D..; Khizhnyak, Sergey; Kovalev, Vladimir

    2016-07-01

    Management of microbial communities to minimize the potential for risk to the crew and to the plants to be used for supporting the crew is an essential component of successful bioregenerative life support systems (BLSS). Previously it was shown that soil-like substrate (SLS), obtained as a result of bioconversion of non-edible plant biomass in the higher plants based BLSS, demonstrates strong anti-fungal activity against soil-borne plant pathogens (Nesterenko et al., 2009). The present study is devoted to the estimation of anti-bacterial activity of SLS against gram-negative (presented with Escherichia coli) and gram-positive (presented with Staphylococcus aureus) human-associated bacteria, both of which belong to the group of opportunistic pathogen. In vitro effects of different types of SLS on E. coli and S. aureus and in situ survival curves of the bacteria with corresponding math models are presented. Additionally we have examined the influence of community richness (the indigenous community of SLS) on the ability of introduced human-associated bacteria to persist within SLS. The work was carried out within the frames of the state task on the subject No 56.1.4 of the Basic Research Program (Section VI) of Russian State Academies for 2013-2020.

  16. Discrete dynamic modeling of T cell survival signaling networks

    Science.gov (United States)

    Zhang, Ranran

    2009-03-01

    Biochemistry-based frameworks are often not applicable for the modeling of heterogeneous regulatory systems that are sparsely documented in terms of quantitative information. As an alternative, qualitative models assuming a small set of discrete states are gaining acceptance. This talk will present a discrete dynamic model of the signaling network responsible for the survival and long-term competence of cytotoxic T cells in the blood cancer T-LGL leukemia. We integrated the signaling pathways involved in normal T cell activation and the known deregulations of survival signaling in leukemic T-LGL, and formulated the regulation of each network element as a Boolean (logic) rule. Our model suggests that the persistence of two signals is sufficient to reproduce all known deregulations in leukemic T-LGL. It also indicates the nodes whose inactivity is necessary and sufficient for the reversal of the T-LGL state. We have experimentally validated several model predictions, including: (i) Inhibiting PDGF signaling induces apoptosis in leukemic T-LGL. (ii) Sphingosine kinase 1 and NFκB are essential for the long-term survival of T cells in T-LGL leukemia. (iii) T box expressed in T cells (T-bet) is constitutively activated in the T-LGL state. The model has identified potential therapeutic targets for T-LGL leukemia and can be used for generating long-term competent CTL necessary for tumor and cancer vaccine development. The success of this model, and of other discrete dynamic models, suggests that the organization of signaling networks has an determining role in their dynamics. Reference: R. Zhang, M. V. Shah, J. Yang, S. B. Nyland, X. Liu, J. K. Yun, R. Albert, T. P. Loughran, Jr., Network Model of Survival Signaling in LGL Leukemia, PNAS 105, 16308-16313 (2008).

  17. Laser printing of skin cells and human stem cells.

    Science.gov (United States)

    Koch, Lothar; Kuhn, Stefanie; Sorg, Heiko; Gruene, Martin; Schlie, Sabrina; Gaebel, Ralf; Polchow, Bianca; Reimers, Kerstin; Stoelting, Stephanie; Ma, Nan; Vogt, Peter M; Steinhoff, Gustav; Chichkov, Boris

    2010-10-01

    Laser printing based on laser-induced forward transfer (LIFT) is a new biofabrication technique for the arrangement of biological materials or living cells in well-defined patterns. In the current study, skin cell lines (fibroblasts/keratinocytes) and human mesenchymal stem cells (hMSC) were chosen for laser printing experiments due to their high potential in regeneration of human skin and new application possibilities of stem cell therapy. To evaluate the influence of LIFT on the cells, their survival rate, their proliferation and apoptotic activity, and the DNA damages and modifications of their cell surface markers were assessed and statistically evaluated over several days. The cells survived the transfer procedure with a rate of 98%  +/- 1% standard error of the mean (skin cells) and 90%  +/- 10% (hMSC), respectively. All used cell types maintain their ability to proliferate after LIFT. Further, skin cells and hMSC did not show an increase of apoptosis or DNA fragmentation. In addition, the hMSC keep their phenotype as proven by fluorescence activated cell sorting (FACS) analysis. This study demonstrates LIFT as a suitable technique for unharmed computer-controlled positioning of different cell types and a promising tool for future applications in the ex vivo generation of tissue replacements.

  18. Molecular Imaging of Stem Cells: Tracking Survival, Biodistribution, Tumorigenicity, and Immunogenicity

    Directory of Open Access Journals (Sweden)

    Eugene Gu, Wen-Yi Chen, Jay Gu, Paul Burridge, Joseph C. Wu

    2012-01-01

    Full Text Available Being able to self-renew and differentiate into virtually all cell types, both human embryonic stem cells (hESCs and induced pluripotent stem cells (iPSCs have exciting therapeutic implications for myocardial infarction, neurodegenerative disease, diabetes, and other disorders involving irreversible cell loss. However, stem cell biology remains incompletely understood despite significant advances in the field. Inefficient stem cell differentiation, difficulty in verifying successful delivery to the target organ, and problems with engraftment all hamper the transition from laboratory animal studies to human clinical trials. Although traditional histopathological techniques have been the primary approach for ex vivo analysis of stem cell behavior, these postmortem examinations are unable to further elucidate the underlying mechanisms in real time and in vivo. Fortunately, the advent of molecular imaging has led to unprecedented progress in understanding the fundamental behavior of stem cells, including their survival, biodistribution, immunogenicity, and tumorigenicity in the targeted tissues of interest. This review summarizes various molecular imaging technologies and how they have advanced the current understanding of stem cell survival, biodistribution, immunogenicity, and tumorigenicity.

  19. Povidone-iodine Solutions Inhibit Cell Migration and Survival of Osteoblasts, Fibroblasts, and Myoblasts.

    Science.gov (United States)

    Liu, James X; Werner, Jordan A; Buza, John A; Kirsch, Thorsten; Zuckerman, Joseph D; Virk, Mandeep S

    2017-04-12

    In vitro laboratory study. The purpose of this study was to identify the effect of dilute povidone-iodine (PVI) solutions on human osteoblast, fibroblast and myoblast cells in vitro. Dilute PVI wound lavage has been used successfully in spine and joint arthroplasty procedures to prevent post-operative surgical site infection, but their biologic effect on host cells is largely unknown. Human primary osteoblasts, fibroblasts, and myoblasts were expanded in cell culture and subjected to various concentrations of PVI (0%, 0.001%, 0.01%, 0.1%, 0.35%, 1%) for 3 minutes. To assess the effect of PVI on cell migration, a scratch assay was performed, in which a "scratch" was made by a standard pipette tip in a cell monolayer following PVI exposure, and time to closure of the scratch was evaluated. Cell survival and proliferation was measured 48 hours post-PVI exposure using a cell viability and cytotoxicity assay. Closure of the scratch defect in all cell monolayers was achieved in PVI concentrations PVI concentrations of ≥ 0.1%. PVI concentrations PVI ≥ 0.1% had cell survival rates of less than 6% (p PVI (0.35%) exerts a pronounced cytotoxic effect on osteoblasts, fibroblast, and myoblasts in vitro. Further investigation is required to systematically study the effect of PVI on tissue healing in vivo and also determine a safe and clinically potent concentration for PVI lavage. N/A.

  20. The effect of selected factors on the survival of Bacillus cereus in the human gastrointestinal tract.

    Science.gov (United States)

    Berthold-Pluta, Anna; Pluta, Antoni; Garbowska, Monika

    2015-05-01

    Bacillus cereus is a Gram-positive bacterium widely distributed in soil and vegetation. This bacterial species can also contaminate raw or processed foods. Pathogenic B. cereus strains can cause a range of infections in humans, as well as food poisoning of an emetic (intoxication) or diarrheal type (toxico-infection). Toxico-infections are due to the action of the Hbl toxin, Nhe toxin, and cytotoxin K produced by the microorganism in the gastrointestinal tract. This occurs once the spores or vegetative B. cereus cells survive the pH barrier of the stomach and reach the small intestine where they produce toxins in sufficient amounts. This article discusses the effect of various factors on the survival of B. cereus in the gastrointestinal tract, including low pH and the presence of digestive enzymes in the stomach, bile salts in the small intestine, and indigenous microflora in the lower parts of the gastrointestinal tract. Additional aspects also reported to affect B. cereus survival and virulence in the gastrointestinal tract include the interaction of the spores and vegetative cells with enterocytes. In vitro studies revealed that both vegetative B. cereus and spores can survive in the gastrointestinal tract suggesting that the biological form of the microorganism may have less influence on the occurrence of the symptoms of infection than was once believed. It is most likely the interaction between the pathogen and enterocytes that is necessary for the diarrheal form of B. cereus food poisoning to develop. The adhesion of B. cereus to the intestinal epithelium allows the bacterium to grow and produce enterotoxins in the proximity of the epithelium. Recent studies suggest that the human intestinal microbiota inhibits the growth of vegetative B. cereus cells considerably.

  1. Stochastic modeling and experimental analysis of phenotypic switching and survival of cancer cells under stress

    Science.gov (United States)

    Zamani Dahaj, Seyed Alireza; Kumar, Niraj; Sundaram, Bala; Celli, Jonathan; Kulkarni, Rahul

    The phenotypic heterogeneity of cancer cells is critical to their survival under stress. A significant contribution to heterogeneity of cancer calls derives from the epithelial-mesenchymal transition (EMT), a conserved cellular program that is crucial for embryonic development. Several studies have investigated the role of EMT in growth of early stage tumors into invasive malignancies. Also, EMT has been closely associated with the acquisition of chemoresistance properties in cancer cells. Motivated by these studies, we analyze multi-phenotype stochastic models of the evolution of cancers cell populations under stress. We derive analytical results for time-dependent probability distributions that provide insights into the competing rates underlying phenotypic switching (e.g. during EMT) and the corresponding survival of cancer cells. Experimentally, we evaluate these model-based predictions by imaging human pancreatic cancer cell lines grown with and without cytotoxic agents and measure growth kinetics, survival, morphological changes and (terminal evaluation of) biomarkers with associated epithelial and mesenchymal phenotypes. The results derived suggest approaches for distinguishing between adaptation and selection scenarios for survival in the presence of external stresses.

  2. Auditor human capital and audit firm survival - The Dutch audit industry in 1930-1992

    NARCIS (Netherlands)

    Brocheler, [No Value; Maijoor, S; van Witteloostuijn, A; Bröcheler, V.

    2004-01-01

    This paper studies the relationship between auditor human capital and audit firm survival. Specifically, the effects are investigated of the human capital of auditors on the survival chances of newly established audit firms. Human capital is analyzed both at the time of entry of a new audit firm and

  3. Caspase inhibitors promote vestibular hair cell survival and function after aminoglycoside treatment in vivo

    Science.gov (United States)

    Matsui, Jonathan I.; Haque, Asim; Huss, David; Messana, Elizabeth P.; Alosi, Julie A.; Roberson, David W.; Cotanche, Douglas A.; Dickman, J. David; Warchol, Mark E.

    2003-01-01

    The sensory hair cells of the inner ear undergo apoptosis after acoustic trauma or aminoglycoside antibiotic treatment, causing permanent auditory and vestibular deficits in humans. Previous studies have demonstrated a role for caspase activation in hair cell death and ototoxic injury that can be reduced by concurrent treatment with caspase inhibitors in vitro. In this study, we examined the protective effects of caspase inhibition on hair cell death in vivo after systemic injections of aminoglycosides. In one series of experiments, chickens were implanted with osmotic pumps that administrated the pan-caspase inhibitor z-Val-Ala-Asp(Ome)-fluoromethylketone (zVAD) into inner ear fluids. One day after the surgery, the animals received a 5 d course of treatment with streptomycin, a vestibulotoxic aminoglycoside. Direct infusion of zVAD into the vestibule significantly increased hair cell survival after streptomycin treatment. A second series of experiments determined whether rescued hair cells could function as sensory receptors. Animals treated with streptomycin displayed vestibular system impairment as measured by a greatly reduced vestibulo-ocular response (VOR). In contrast, animals that received concurrent systemic administration of zVAD with streptomycin had both significantly greater hair cell survival and significantly increased VOR responses, as compared with animals treated with streptomycin alone. These findings suggest that inhibiting the activation of caspases promotes the survival of hair cells and protects against vestibular function deficits after aminoglycoside treatment.

  4. L-Arginine Modulates T Cell Metabolism and Enhances Survival and Anti-tumor Activity.

    Science.gov (United States)

    Geiger, Roger; Rieckmann, Jan C; Wolf, Tobias; Basso, Camilla; Feng, Yuehan; Fuhrer, Tobias; Kogadeeva, Maria; Picotti, Paola; Meissner, Felix; Mann, Matthias; Zamboni, Nicola; Sallusto, Federica; Lanzavecchia, Antonio

    2016-10-20

    Metabolic activity is intimately linked to T cell fate and function. Using high-resolution mass spectrometry, we generated dynamic metabolome and proteome profiles of human primary naive T cells following activation. We discovered critical changes in the arginine metabolism that led to a drop in intracellular L-arginine concentration. Elevating L-arginine levels induced global metabolic changes including a shift from glycolysis to oxidative phosphorylation in activated T cells and promoted the generation of central memory-like cells endowed with higher survival capacity and, in a mouse model, anti-tumor activity. Proteome-wide probing of structural alterations, validated by the analysis of knockout T cell clones, identified three transcriptional regulators (BAZ1B, PSIP1, and TSN) that sensed L-arginine levels and promoted T cell survival. Thus, intracellular L-arginine concentrations directly impact the metabolic fitness and survival capacity of T cells that are crucial for anti-tumor responses.

  5. Nifuroxazide inhibits survival of multiple myeloma cells by directly inhibiting STAT3.

    Science.gov (United States)

    Nelson, Erik A; Walker, Sarah R; Kepich, Alicia; Gashin, Laurie B; Hideshima, Teru; Ikeda, Hiroshi; Chauhan, Dharminder; Anderson, Kenneth C; Frank, David A

    2008-12-15

    Constitutive activation of the transcription factor STAT3 contributes to the pathogenesis of many cancers, including multiple myeloma (MM). Since STAT3 is dispensable in most normal tissue, targeted inhibition of STAT3 is an attractive therapy for patients with these cancers. To identify STAT3 inhibitors, we developed a transcriptionally based assay and screened a library of compounds known to be safe in humans. We found the drug nifuroxazide to be an effective inhibitor of STAT3 function. Nifuroxazide inhibits the constitutive phosphorylation of STAT3 in MM cells by reducing Jak kinase autophosphorylation, and leads to down-regulation of the STAT3 target gene Mcl-1. Nifuroxazide causes a decrease in viability of primary myeloma cells and myeloma cell lines containing STAT3 activation, but not normal peripheral blood mononuclear cells. Although bone marrow stromal cells provide survival signals to myeloma cells, nifuroxazide can overcome this survival advantage. Reflecting the interaction of STAT3 with other cellular pathways, nifuroxazide shows enhanced cytotoxicity when combined with either the histone deacetylase inhibitor depsipeptide or the MEK inhibitor UO126. Therefore, using a mechanistic-based screen, we identified the clinically relevant drug nifuroxazide as a potent inhibitor of STAT signaling that shows cytotoxicity against myeloma cells that depend on STAT3 for survival.

  6. Microbiota promotes systemic T-cell survival through suppression of an apoptotic factor.

    Science.gov (United States)

    Soto, Raymond; Petersen, Charisse; Novis, Camille L; Kubinak, Jason L; Bell, Rickesha; Stephens, W Zac; Lane, Thomas E; Fujinami, Robert S; Bosque, Alberto; O'Connell, Ryan M; Round, June L

    2017-05-23

    Symbiotic microbes impact the severity of a variety of diseases through regulation of T-cell development. However, little is known regarding the molecular mechanisms by which this is accomplished. Here we report that a secreted factor, Erdr1, is regulated by the microbiota to control T-cell apoptosis. Erdr1 expression was identified by transcriptome analysis to be elevated in splenic T cells from germfree and antibiotic-treated mice. Suppression of Erdr1 depends on detection of circulating microbial products by Toll-like receptors on T cells, and this regulation is conserved in human T cells. Erdr1 was found to function as an autocrine factor to induce apoptosis through caspase 3. Consistent with elevated levels of Erdr1, germfree mice have increased splenic T-cell apoptosis. RNA sequencing of Erdr1-overexpressing cells identified the up-regulation of genes involved in Fas-mediated cell death, and Erdr1 fails to induce apoptosis in Fas-deficient cells. Importantly, forced changes in Erdr1 expression levels dictate the survival of auto-reactive T cells and the clinical outcome of neuro-inflammatory autoimmune disease. Cellular survival is a fundamental feature regulating appropriate immune responses. We have identified a mechanism whereby the host integrates signals from the microbiota to control T-cell apoptosis, making regulation of Erdr1 a potential therapeutic target for autoimmune disease.

  7. Survival Motor Neuron Protein Regulates Stem Cell Division, Proliferation, and Differentiation in Drosophila

    OpenAIRE

    Stuart J Grice; Ji-Long Liu

    2011-01-01

    Spinal muscular atrophy is a severe neurogenic disease that is caused by mutations in the human survival motor neuron 1 (SMN1) gene. SMN protein is required for the assembly of small nuclear ribonucleoproteins and a dramatic reduction of the protein leads to cell death. It is currently unknown how the reduction of this ubiquitously essential protein can lead to tissue-specific abnormalities. In addition, it is still not known whether the disease is caused by developmental or degenerative defe...

  8. Cell encoding recombinant human erythropoietin

    Energy Technology Data Exchange (ETDEWEB)

    Beck, A.K.; Withy, R.M.; Zabrecky, J.R.; Masiello, N.C.

    1990-09-04

    This patent describes a C127 cell transformed with a recombinant DNA vector. It comprises: a DNA sequence encoding human erythropoietin, the transformed cell being capable of producing N-linked and O-linked glycosylated human erythropoietin.

  9. The Growing Complexity of Cancer Cell Response to DNA-Damaging Agents: Caspase 3 Mediates Cell Death or Survival?

    Science.gov (United States)

    Mirzayans, Razmik; Andrais, Bonnie; Kumar, Piyush; Murray, David

    2016-05-11

    It is widely stated that wild-type p53 either mediates the activation of cell cycle checkpoints to facilitate DNA repair and promote cell survival, or orchestrates apoptotic cell death following exposure to cancer therapeutic agents. This reigning paradigm has been challenged by numerous discoveries with different human cell types, including solid tumor-derived cell lines. Thus, activation of the p53 signaling pathway by ionizing radiation and other DNA-damaging agents hinders apoptosis and triggers growth arrest (e.g., through premature senescence) in some genetic backgrounds; such growth arrested cells remain viable, secrete growth-promoting factors, and give rise to progeny with stem cell-like properties. In addition, caspase 3, which is best known for its role in the execution phase of apoptosis, has been recently reported to facilitate (rather than suppress) DNA damage-induced genomic instability and carcinogenesis. This observation is consistent with an earlier report demonstrating that caspase 3 mediates secretion of the pro-survival factor prostaglandin E₂, which in turn promotes enrichment of tumor repopulating cells. In this article, we review these and related discoveries and point out novel cancer therapeutic strategies. One of our objectives is to demonstrate the growing complexity of the DNA damage response beyond the conventional "repair and survive, or die" hypothesis.

  10. Survival of free and encapsulated human and rat islet xenografts transplanted into the mouse bone marrow.

    Science.gov (United States)

    Meier, Raphael P H; Seebach, Jörg D; Morel, Philippe; Mahou, Redouan; Borot, Sophie; Giovannoni, Laurianne; Parnaud, Geraldine; Montanari, Elisa; Bosco, Domenico; Wandrey, Christine; Berney, Thierry; Bühler, Leo H; Muller, Yannick D

    2014-01-01

    Bone marrow was recently proposed as an alternative and potentially immune-privileged site for pancreatic islet transplantation. The aim of the present study was to assess the survival and rejection mechanisms of free and encapsulated xenogeneic islets transplanted into the medullary cavity of the femur, or under the kidney capsule of streptozotocin-induced diabetic C57BL/6 mice. The median survival of free rat islets transplanted into the bone marrow or under the kidney capsule was 9 and 14 days, respectively, whereas that of free human islets was shorter, 7 days (bone marrow) and 10 days (kidney capsule). Infiltrating CD8+ T cells and redistributed CD4+ T cells, and macrophages were detected around the transplanted islets in bone sections. Recipient mouse splenocytes proliferated in response to donor rat stimulator cells. One month after transplantation under both kidney capsule or into bone marrow, encapsulated rat islets had induced a similar degree of fibrotic reaction and still contained insulin positive cells. In conclusion, we successfully established a small animal model for xenogeneic islet transplantation into the bone marrow. The rejection of xenogeneic islets was associated with local and systemic T cell responses and macrophage recruitment. Although there was no evidence for immune-privilege, the bone marrow may represent a feasible site for encapsulated xenogeneic islet transplantation.

  11. Survival of free and encapsulated human and rat islet xenografts transplanted into the mouse bone marrow.

    Directory of Open Access Journals (Sweden)

    Raphael P H Meier

    Full Text Available Bone marrow was recently proposed as an alternative and potentially immune-privileged site for pancreatic islet transplantation. The aim of the present study was to assess the survival and rejection mechanisms of free and encapsulated xenogeneic islets transplanted into the medullary cavity of the femur, or under the kidney capsule of streptozotocin-induced diabetic C57BL/6 mice. The median survival of free rat islets transplanted into the bone marrow or under the kidney capsule was 9 and 14 days, respectively, whereas that of free human islets was shorter, 7 days (bone marrow and 10 days (kidney capsule. Infiltrating CD8+ T cells and redistributed CD4+ T cells, and macrophages were detected around the transplanted islets in bone sections. Recipient mouse splenocytes proliferated in response to donor rat stimulator cells. One month after transplantation under both kidney capsule or into bone marrow, encapsulated rat islets had induced a similar degree of fibrotic reaction and still contained insulin positive cells. In conclusion, we successfully established a small animal model for xenogeneic islet transplantation into the bone marrow. The rejection of xenogeneic islets was associated with local and systemic T cell responses and macrophage recruitment. Although there was no evidence for immune-privilege, the bone marrow may represent a feasible site for encapsulated xenogeneic islet transplantation.

  12. Bordetella pertussis entry into respiratory epithelial cells and intracellular survival.

    Science.gov (United States)

    Lamberti, Yanina; Gorgojo, Juan; Massillo, Cintia; Rodriguez, Maria E

    2013-12-01

    Bordetella pertussis is the causative agent of pertussis, aka whooping cough. Although generally considered an extracellular pathogen, this bacterium has been found inside respiratory epithelial cells, which might represent a survival strategy inside the host. Relatively little is known, however, about the mechanism of internalization and the fate of B. pertussis inside the epithelia. We show here that B. pertussis is able to enter those cells by a mechanism dependent on microtubule assembly, lipid raft integrity, and the activation of a tyrosine-kinase-mediated signaling. Once inside the cell, a significant proportion of the intracellular bacteria evade phagolysosomal fusion and remain viable in nonacidic lysosome-associated membrane-protein-1-negative compartments. In addition, intracellular B. pertussis was found able to repopulate the extracellular environment after complete elimination of the extracellular bacteria with polymyxin B. Taken together, these data suggest that B. pertussis is able to survive within respiratory epithelial cells and by this means potentially contribute to host immune system evasion.

  13. Oral squamous cell carcinoma: survival, recurrence and death

    Directory of Open Access Journals (Sweden)

    Antônio Camilo Souza Cruz

    2014-10-01

    Full Text Available This paper was based in data survey from macro and microscopic oral lesions characteristics, personal data and medical history of patients diagnosed with oral squamous cell carcinoma in the Lab of Pathological Anatomy from the Federal University of Alfenas from January 2000 to December 2010, establishing comparative parameters among clinical data, type of treatment, recurrence, survival and anatomic pathological characteristics of the lesions. Were analyzed the histopathological reports, dental and hospital records. The highest incidence was in white men, age between 50 and 60 years, married, with low education and socioeconomic levels. The beginning of treatment occurred in average 67 days after the histopathological diagnosis. The estimated survival of patients at five years was 42%. The consumption of alcohol and tobacco and the occurrence of metastasis were statistically significant for the increase of recurrence and lethality.

  14. Steroid hormone control of cell death and cell survival: molecular insights using RNAi.

    Directory of Open Access Journals (Sweden)

    Suganthi Chittaranjan

    2009-02-01

    Full Text Available The insect steroid hormone ecdysone triggers programmed cell death of obsolete larval tissues during metamorphosis and provides a model system for understanding steroid hormone control of cell death and cell survival. Previous genome-wide expression studies of Drosophila larval salivary glands resulted in the identification of many genes associated with ecdysone-induced cell death and cell survival, but functional verification was lacking. In this study, we test functionally 460 of these genes using RNA interference in ecdysone-treated Drosophila l(2mbn cells. Cell viability, cell morphology, cell proliferation, and apoptosis assays confirmed the effects of known genes and additionally resulted in the identification of six new pro-death related genes, including sorting nexin-like gene SH3PX1 and Sox box protein Sox14, and 18 new pro-survival genes. Identified genes were further characterized to determine their ecdysone dependency and potential function in cell death regulation. We found that the pro-survival function of five genes (Ras85D, Cp1, CG13784, CG32016, and CG33087, was dependent on ecdysone signaling. The TUNEL assay revealed an additional two genes (Kap-alpha3 and Smr with an ecdysone-dependent cell survival function that was associated with reduced cell death. In vitro, Sox14 RNAi reduced the percentage of TUNEL-positive l(2mbn cells (p<0.05 following ecdysone treatment, and Sox14 overexpression was sufficient to induce apoptosis. In vivo analyses of Sox14-RNAi animals revealed multiple phenotypes characteristic of aberrant or reduced ecdysone signaling, including defects in larval midgut and salivary gland destruction. These studies identify Sox14 as a positive regulator of ecdysone-mediated cell death and provide new insights into the molecular mechanisms underlying the ecdysone signaling network governing cell death and cell survival.

  15. Laser Phototherapy Enhances Mesenchymal Stem Cells Survival in Response to the Dental Adhesives

    Directory of Open Access Journals (Sweden)

    Ivana Márcia Alves Diniz

    2015-01-01

    Full Text Available Background. We investigated the influence of laser phototherapy (LPT on the survival of human mesenchymal stem cells (MSCs submitted to substances leached from dental adhesives. Method. MSCs were isolated and characterized. Oral mucosa fibroblasts and osteoblast-like cells were used as comparative controls. Cultured medium conditioned with two adhesive systems was applied to the cultures. Cell monolayers were exposed or not to LPT. Laser irradiations were performed using a red laser (GaAlAs, 780 nm, 0.04 cm2, 40 mW, 1 W/cm2, 0.4 J, 10 seconds, 1 point, 10 J/cm2. After 24 h, cell viability was assessed by the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide reduction assay. Data were statistically compared by ANOVA followed by Tukey’s test (P<0.05. Results. Different cell types showed different viabilities in response to the same materials. Substances leached from adhesives were less cytotoxic to MSCs than to other cell types. Substances leached from Clearfil SE Bond were highly cytotoxic to all cell types tested, except to the MSCs when applied polymerized and in association with LPT. LPT was unable to significantly increase the cell viability of fibroblasts and osteoblast-like cells submitted to the dental adhesives. Conclusion. LPT enhances mesenchymal stem cells survival in response to substances leached from dental adhesives.

  16. Control of neural stem cell survival by electroactive polymer substrates.

    Directory of Open Access Journals (Sweden)

    Vanessa Lundin

    Full Text Available Stem cell function is regulated by intrinsic as well as microenvironmental factors, including chemical and mechanical signals. Conducting polymer-based cell culture substrates provide a powerful tool to control both chemical and physical stimuli sensed by stem cells. Here we show that polypyrrole (PPy, a commonly used conducting polymer, can be tailored to modulate survival and maintenance of rat fetal neural stem cells (NSCs. NSCs cultured on PPy substrates containing different counter ions, dodecylbenzenesulfonate (DBS, tosylate (TsO, perchlorate (ClO(4 and chloride (Cl, showed a distinct correlation between PPy counter ion and cell viability. Specifically, NSC viability was high on PPy(DBS but low on PPy containing TsO, ClO(4 and Cl. On PPy(DBS, NSC proliferation and differentiation was comparable to standard NSC culture on tissue culture polystyrene. Electrical reduction of PPy(DBS created a switch for neural stem cell viability, with widespread cell death upon polymer reduction. Coating the PPy(DBS films with a gel layer composed of a basement membrane matrix efficiently prevented loss of cell viability upon polymer reduction. Here we have defined conditions for the biocompatibility of PPy substrates with NSC culture, critical for the development of devices based on conducting polymers interfacing with NSCs.

  17. Cyclophilin B Supports Myc and Mutant p53 Dependent Survival of Glioblastoma Multiforme Cells

    Science.gov (United States)

    Choi, Jae Won; Schroeder, Mark A.; Sarkaria, Jann N.; Bram, Richard J.

    2014-01-01

    Glioblastoma multiforme (GBM) is an aggressive, treatment-refractory type of brain tumor for which effective therapeutic targets remain important to identify. Here we report that cyclophilin B (CypB), a prolyl isomerase residing in the endoplasmic reticulum (ER), provides an essential survival signal in GBM cells. Analysis of gene expression databases revealed that CypB is upregulated in many cases of malignant glioma. We found that suppression of CypB reduced cell proliferation and survival in human GBM cells in vitro and in vivo. We also found that treatment with small molecule inhibitors of cyclophilins, including the approved drug cyclosporine, greatly reduced the viability of GBM cells. Mechanistically, depletion or pharmacologic inhibition of CypB caused hyperactivation of the oncogenic RAS-MAPK pathway, induction of cellular senescence signals, and death resulting from loss of MYC, mutant p53, Chk1 and JAK/STAT3 signaling. Elevated reactive oxygen species, ER expansion and abnormal unfolded protein responses in CypB-depleted GBM cells indicated that CypB alleviates oxidative and ER stresses and coordinates stress adaptation responses. Enhanced cell survival and sustained expression of multiple oncogenic proteins downstream of CypB may thus contribute to the poor outcome of GBM tumors. Our findings link chaperone-mediated protein folding in the ER to mechanisms underlying oncogenic transformation, and they make CypB an attractive and immediately targetable molecule for GBM therapy. PMID:24272483

  18. Heat shock genes – integrating cell survival and death

    Indian Academy of Sciences (India)

    Richa Arya; Moushami Mallik; Subhash C Lakhotia

    2007-04-01

    Heat shock induced gene expression and other cellular responses help limit the damage caused by stress and thus facilitate cellular recovery. Cellular damage also triggers apoptotic cell death through several pathways. This paper briefly reviews interactions of the major heat shock proteins with components of the apoptotic pathways. Hsp90, which acts as a chaperone for unstable signal transducers to keep them poised for activation, interacts with RIP and Akt and promotes NF-B mediated inhibition of apoptosis; in addition it also blocks some steps in the apoptotic pathways. Hsp70 is mostly anti-apoptotic and acts at several levels like inhibition of translocation of Bax into mitochondria, release of cytochrome c from mitochondria, formation of apoptosome and inhibition of activation of initiator caspases. Hsp70 also modulates JNK, NF-B and Akt signaling pathways in the apoptotic cascade. In contrast, Hsp60 has both anti- and pro-apoptotic roles. Cytosolic Hsp60 prevents translocation of the pro-apoptotic protein Bax into mitochondria and thus promotes cell survival but it also promotes maturation of procaspase-3, essential for caspase mediated cell death. Our recent in vivo studies show that RNAi for the Hsp60D in Drosophila melanogaster prevents induced apoptosis. Hsp27 exerts its anti-apoptotic influence by inhibiting cytochrome c and TNF-mediated cell death. crystallin suppresses caspase-8 and cytochrome c mediated activation of caspase-3. Studies in our laboratory also reveal that absence or reduced levels of the developmentally active as well as stress induced non-coding hsr transcripts, which are known to sequester diverse hnRNPs and related nuclear RNA-binding proteins, block induced apoptosis in Drosophila. Modulation of the apoptotic pathways by Hsps reflects their roles as ``weak links” between various ``hubs” in cellular networks. On the other hand, non-coding RNAs, by virtue of their potential to bind with multiple proteins, can act as ``hubs” in

  19. Neural stem cells improve neuronal survival in cultured postmortem brain tissue from aged and Alzheimer patients.

    Science.gov (United States)

    Wu, L; Sluiter, A A; Guo, Ho-Fu; Balesar, R A; Swaab, D F; Zhou, Jiang-Ning; Verwer, R W H

    2008-01-01

    Neurodegenerative diseases are progressive and incurable and are becoming ever more prevalent. To study whether neural stem cell can reactivate or rescue functions of impaired neurons in the human aging and neurodegenerating brain, we co-cultured postmortem slices from Alzheimer patients and control participants with rat embryonic day 14 (E14) neural stem cells. Viability staining based on the exclusion of ethidium bromide by intact plasma membranes showed that there were strikingly more viable cells and fewer dead cells in slices co-cultured with neural stem cells than in untreated slices. The presence of Alzheimer pathology in the brain slices did not influence this effect, although the slices from Alzheimer patients, in general, contained fewer viable cells. Co-culturing with rat E14 fibroblasts did not improve the viability of neurons in the human brain slices. Since the human slices and neural stem cells were separated by a membrane during co-culturing our data show for the first time that neural stem cells release diffusible factors that may improve the survival of aged and degenerating neurons in human brains.

  20. Differential bacterial survival, replication, and apoptosis-inducing ability of Salmonella serovars within human and murine macrophages.

    Science.gov (United States)

    Schwan, W R; Huang, X Z; Hu, L; Kopecko, D J

    2000-03-01

    Salmonella serovars are associated with human diseases that range from mild gastroenteritis to host-disseminated enteric fever. Human infections by Salmonella enterica serovar Typhi can lead to typhoid fever, but this serovar does not typically cause disease in mice or other animals. In contrast, S. enterica serovar Typhimurium and S. enterica serovar Enteritidis, which are usually linked to localized gastroenteritis in humans and some animal species, elicit a systemic infection in mice. To better understand these observations, multiple strains of each of several chosen serovars of Salmonella were tested for the ability in the nonopsonized state to enter, survive, and replicate within human macrophage cells (U937 and elutriated primary cells) compared with murine macrophage cells (J774A.1 and primary peritoneal cells); in addition, death of the infected macrophages was monitored. The serovar Typhimurium strains all demonstrated enhanced survival within J774A.1 cells and murine peritoneal macrophages, compared with the significant, almost 100-fold declines in viable counts noted for serovar Typhi strains. Viable counts for serovar Enteritidis either matched the level of serovar Typhi (J774A. 1 macrophages) or were comparable to counts for serovar Typhimurium (murine peritoneal macrophages). Apoptosis was significantly higher in J774A.1 cells infected with serovar Typhimurium strain LT2 compared to serovar Typhi strain Ty2. On the other hand, serovar Typhi survived at a level up to 100-fold higher in elutriated human macrophages and 2- to 3-fold higher in U937 cells compared to the serovar Typhimurium and Enteritidis strains tested. Despite the differential multiplication of serovar Typhi during infection of U937 cells, serovar Typhi caused significantly less apoptosis than infections with serovar Typhimurium. These observations indicate variability in intramacrophage survival and host cytotoxicity among the various serovars and are the first to show differences in

  1. Cell survival, cell death and cell cycle pathways are interconnected: Implications for cancer therapy

    DEFF Research Database (Denmark)

    Maddika, S; Ande, SR; Panigrahi, S

    2007-01-01

    The partial cross-utilization of molecules and pathways involved in opposing processes like cell survival, proliferation and cell death, assures that mutations within one signaling cascade will also affect the other opposite process at least to some extent, thus contributing to homeostatic...... both for their apoptosis-regulating capacity and also for their effect on the cell cycle progression. The PI3-K/Akt cell survival pathway is shown as regulator of cell metabolism and cell survival, but examples are also provided where aberrant activity of the pathway may contribute to the induction...

  2. Epidermal Growth Factor Receptor Cell Survival Signaling Requires Phosphatidylcholine Biosynthesis

    Directory of Open Access Journals (Sweden)

    Matt Crook

    2016-11-01

    Full Text Available Identification of pro-cell survival signaling pathways has implications for cancer, cardiovascular, and neurodegenerative disease. We show that the Caenorhabditis elegans epidermal growth factor receptor LET-23 (LET-23 EGFR has a prosurvival function in counteracting excitotoxicity, and we identify novel molecular players required for this prosurvival signaling. uv1 sensory cells in the C. elegans uterus undergo excitotoxic death in response to activation of the OSM-9/OCR-4 TRPV channel by the endogenous agonist nicotinamide. Activation of LET-23 EGFR can effectively prevent this excitotoxic death. We investigate the roles of signaling pathways known to act downstream of LET-23 EGFR in C. elegans and find that the LET-60 Ras/MAPK pathway, but not the IP3 receptor pathway, is required for efficient LET-23 EGFR activity in its prosurvival function. However, activation of LET-60 Ras/MAPK pathway does not appear to be sufficient to fully mimic LET-23 EGFR activity. We screen for genes that are required for EGFR prosurvival function and uncover a role for phosphatidylcholine biosynthetic enzymes in EGFR prosurvival function. Finally, we show that exogenous application of phosphatidylcholine is sufficient to prevent some deaths in this excitotoxicity model. Our work implicates regulation of lipid synthesis downstream of EGFR in cell survival and death decisions.

  3. Laser phototherapy enhances mesenchymal stem cells survival in response to the dental adhesives.

    Science.gov (United States)

    Diniz, Ivana Márcia Alves; Matos, Adriana Bona; Marques, Márcia Martins

    2015-01-01

    We investigated the influence of laser phototherapy (LPT) on the survival of human mesenchymal stem cells (MSCs) submitted to substances leached from dental adhesives. MSCs were isolated and characterized. Oral mucosa fibroblasts and osteoblast-like cells were used as comparative controls. Cultured medium conditioned with two adhesive systems was applied to the cultures. Cell monolayers were exposed or not to LPT. Laser irradiations were performed using a red laser (GaAlAs, 780 nm, 0.04 cm(2), 40 mW, 1 W/cm(2), 0.4 J, 10 seconds, 1 point, 10 J/cm(2)). After 24 h, cell viability was assessed by the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide reduction assay. Data were statistically compared by ANOVA followed by Tukey's test (P adhesives were less cytotoxic to MSCs than to other cell types. Substances leached from Clearfil SE Bond were highly cytotoxic to all cell types tested, except to the MSCs when applied polymerized and in association with LPT. LPT was unable to significantly increase the cell viability of fibroblasts and osteoblast-like cells submitted to the dental adhesives. LPT enhances mesenchymal stem cells survival in response to substances leached from dental adhesives.

  4. Oral cancer/endothelial cell fusion experiences nuclear fusion and acquisition of enhanced survival potential.

    Science.gov (United States)

    Song, Kai; Song, Yong; Zhao, Xiao-Ping; Shen, Hui; Wang, Meng; Yan, Ting-Lin; Liu, Ke; Shang, Zheng-Jun

    2014-10-15

    Most previous studies have linked cancer-macrophage fusion with tumor progression and metastasis. However, the characteristics of hybrid cells derived from oral cancer and endothelial cells and their involvement in cancer remained unknown. Double-immunofluorescent staining and fluorescent in situ hybridization (FISH) were performed to confirm spontaneous cell fusion between eGFP-labeled human umbilical vein endothelial cells (HUVECs) and RFP-labeled SCC9, and to detect the expression of vementin and cytokeratin 18 in the hybrids. The property of chemo-resistance of such hybrids was examined by TUNEL assay. The hybrid cells in xenografted tumor were identified by FISH and GFP/RFP dual-immunofluoresence staining. We showed that SCC9 cells spontaneously fused with cocultured endothelial cells, and the resultant hybrid cells maintained the division and proliferation activity after re-plating and thawing. Such hybrids expressed markers of both parental cells and became more resistant to chemotherapeutic drug cisplatin as compared to the parental SCC9 cells. Our in vivo data indicated that the hybrid cells contributed to tumor composition by using of immunostaining and FISH analysis, even though the hybrid cells and SCC9 cells were mixed with 1:10,000, according to the FACS data. Our study suggested that the fusion events between oral cancer and endothelial cells undergo nuclear fusion and acquire a new property of drug resistance and consequently enhanced survival potential. These experimental findings provide further supportive evidence for the theory that cell fusion is involved in cancer progression.

  5. Musashi1 regulates breast tumor cell proliferation and is a prognostic indicator of poor survival

    Directory of Open Access Journals (Sweden)

    Wang Xiao-Yang

    2010-08-01

    Full Text Available Abstract Background Musashi1 (Msi1 is a conserved RNA-binding protein that regulates the Notch and Wnt pathways, and serves as a stem cell marker in the breast and other tissues. It is unknown how Msi1 relates to other breast cancer markers, whether it denotes tumor initiating cells (TICs, and how it affects gene expression and tumor cell survival in breast cancer cells. Results Msi1 expression was analyzed in 20 breast cancer cell lines and in 140 primary breast tumors by western blotting and immunohistochemistry, respectively. Lentivirus RNA interference was used to reduce Msi1 expression in breast cancer cell lines MCF-7 and T47D grown as spheroid cultures and to assess stem cell gene expression and the growth of these cell lines as xenografts. In normal human breast tissue, Msi1 was expressed in 10.6% of myoepithelum and 1.2% of ductal epithelium in the terminal ductal lobular unit (TDLU, whereas, less than 0.05% of ductal epithelium and myoepithelium in large ducts outside the TDLU expressed Msi1. Msi1 was expressed in 55% of the breast cancer cell lines and correlated with ErbB2 expression in 50% of the cell lines. Msi1 was expressed in 68% of primary tumors and in 100% of lymph node metastases, and correlated with 5 year survival. Msi1 was enriched in CD133+ MCF-7 and T47D cells and in spheroid cultures of these cells, and Msi1 'knockdown' (KD with a lentivirus-expressed shRNA decreased the number and size of spheroid colonies. Msi1 KD reduced Notch1, c-Myc, ErbB2 and pERK1/2 expression, and increased p21CIP1 expression, which is consistent with known Msi1 target mRNAs. Msi1 KD also reduced the expression of the somatic and embryonic stem cell markers, CD133, Bmi1, Sox2, Nanog and Oct4. Xenografts of MCF-7 and T47D Msi1 KD cells resulted in a marked reduction of tumor growth, reduced Msi1 and Notch1 expression and increased p21CIP1 expression. Conclusion Msi1 is a negative prognostic indicator of breast cancer patient survival, and is

  6. Iron alters cell survival in a mitochondria-dependent pathway in ovarian cancer cells.

    Science.gov (United States)

    Bauckman, Kyle; Haller, Edward; Taran, Nicholas; Rockfield, Stephanie; Ruiz-Rivera, Abigail; Nanjundan, Meera

    2015-03-01

    The role of iron in the development of cancer remains unclear. We previously reported that iron reduces cell survival in a Ras/mitogen-activated protein kinase (MAPK)-dependent manner in ovarian cells; however, the underlying downstream pathway leading to reduced survival was unclear. Although levels of intracellular iron, ferritin/CD71 protein and reactive oxygen species did not correlate with iron-induced cell survival changes, we identified mitochondrial damage (via TEM) and reduced expression of outer mitochondrial membrane proteins (translocase of outer membrane: TOM20 and TOM70) in cell lines sensitive to iron. Interestingly, Ru360 (an inhibitor of the mitochondrial calcium uniporter) reversed mitochondrial changes and restored cell survival in HEY ovarian carcinoma cells treated with iron. Further, cells treated with Ru360 and iron also had reduced autophagic punctae with increased lysosomal numbers, implying cross-talk between these compartments. Mitochondrial changes were dependent on activation of the Ras/MAPK pathway since treatment with a MAPK inhibitor restored expression of TOM20/TOM70 proteins. Although glutathione antioxidant levels were reduced in HEY treated with iron, extracellular glutamate levels were unaltered. Strikingly, oxalomalate (inhibitor of aconitase, involved in glutamate production) reversed iron-induced responses in a similar manner to Ru360. Collectively, our results implicate iron in modulating cell survival in a mitochondria-dependent manner in ovarian cancer cells.

  7. β-Adrenergic Regulation of Cardiac Progenitor Cell Death Versus Survival and Proliferation

    Science.gov (United States)

    Khan, Mohsin; Mohsin, Sadia; Avitabile, Daniele; Siddiqi, Sailay; Nguyen, Jonathan; Wallach, Kathleen; Quijada, Pearl; McGregor, Michael; Gude, Natalie; Alvarez, Roberto; Tilley, Douglas G.; Koch, Walter J.; Sussman, Mark A.

    2013-01-01

    Rationale Short-term β-adrenergic stimulation promotes contractility in response to stress but is ultimately detrimental in the failing heart because of accrual of cardiomyocyte death. Endogenous cardiac progenitor cell (CPC) activation may partially offset cardiomyocyte losses, but consequences of long-term β-adrenergic drive on CPC survival and proliferation are unknown. Objective We sought to determine the relationship between β-adrenergic activity and regulation of CPC function. Methods and Results Mouse and human CPCs express only β2 adrenergic receptor (β2-AR) in conjunction with stem cell marker c-kit. Activation of β2-AR signaling promotes proliferation associated with increased AKT, extracellular signal-regulated kinase 1/2, and endothelial NO synthase phosphorylation, upregulation of cyclin D1, and decreased levels of G protein–coupled receptor kinase 2. Conversely, silencing of β2-AR expression or treatment with β2-antagonist ICI 118, 551 impairs CPC proliferation and survival. β1-AR expression in CPC is induced by differentiation stimuli, sensitizing CPC to isoproterenol-induced cell death that is abrogated by metoprolol. Efficacy of β1-AR blockade by metoprolol to increase CPC survival and proliferation was confirmed in vivo by adoptive transfer of CPC into failing mouse myocardium. Conclusions β-adrenergic stimulation promotes expansion and survival of CPCs through β2-AR, but acquisition of β1-AR on commitment to the myocyte lineage results in loss of CPCs and early myocyte precursors. PMID:23243208

  8. Neisseria gonorrhoeae survives within and modulates apoptosis and inflammatory cytokine production of human macrophages.

    Science.gov (United States)

    Château, Alice; Seifert, H Steven

    2016-04-01

    The human-adapted organism Neisseria gonorrhoeae is the causative agent of gonorrhoea, a sexually transmitted infection. It readily colonizes the genital, rectal and nasalpharyngeal mucosa during infection. While it is well established that N. gonorrhoeae recruits and modulates the functions of polymorphonuclear leukocytes during infection, how N. gonorrhoeae interacts with macrophages present in infected tissue is not fully defined. We studied the interactions of N. gonorrhoeae with two human monocytic cell lines, THP-1 and U937, and primary monocytes, all differentiated into macrophages. Most engulfed bacteria were killed in the phagolysosome, but a subset of bacteria was able to survive and replicate inside the macrophages suggesting that those cells may be an unexplored cellular reservoir for N. gonorrhoeae during infection. N. gonorrhoeae was able to modulate macrophage apoptosis: N. gonorrhoeae induced apoptosis in THP-1 cells whereas it inhibited induced apoptosis in U937 cells and primary human macrophages. Furthermore, N. gonorrhoeae induced expression of inflammatory cytokines in macrophages, suggesting a role for macrophages in recruiting polymorphonuclear leukocytes to the site of infection. These results indicate macrophages may serve as a significant replicative niche for N. gonorrhoeae and play an important role in gonorrheal pathogenesis.

  9. Regulation of cell survival and death during Flavivirus infections

    Institute of Scientific and Technical Information of China (English)

    Sounak; Ghosh; Roy; Beata; Sadigh; Emmanuel; Datan; Richard; A; Lockshin; Zahra; Zakeri

    2014-01-01

    Flaviviruses, ss(+) RNA viruses, include many of mankind’s most important pathogens. Their pathogenicity derives from their ability to infect many types of cells including neurons, to replicate, and eventually to kill the cells. Flaviviruses can activate tumor necrosis factor α and both intrinsic(Bax-mediated) and extrinsic pathways to apoptosis. Thus they can use many approaches for activating these pathways. Infection can lead to necrosis if viral load is extremely high or to other types of cell death if routes to apoptosis are blocked. Dengue and Japanese Encephalitis Virus can also activate autophagy. In this case the autophagy temporarily spares the infected cell, allowing a longer period of reproduction for the virus, and the autophagy further protects the cell against other stresses such as those caused by reactive oxygen species. Several of the viral proteins have been shown to induce apoptosis or autophagy on their own, independent of the presence of other viral proteins. Given the versatility of these viruses to adapt to and manipulate the metabolism, and thus to control the survival of, the infected cells, we need to understand much better how the specific viral proteins affect the pathways to apoptosis and autophagy. Only in this manner will we be able to minimize the pathology that they cause.

  10. Evidence for the involvement of NOD2 in regulating colonic epithelial cell growth and survival

    Institute of Scientific and Technical Information of China (English)

    Sheena M Cruickshank; Louise Wakenshaw; John Cardone; Peter D Howdle; Peter J Murray; Simon R Carding

    2008-01-01

    AIM: To investigate the function of NOD2 in colonic epithelial cells (CEC).METHODS: A combination of in vivo and in vitro analyses of epithelial cell turnover in the presence and absence of a functional NOD2 protein and, in response to enteric Salmonella typhimurium infection, were used. shRNA interference was also used to investigate the consequences of knocking down NOD2 gene expression on the growth and survival of colorectal carcinoma cell lines.RESULTS: In the colonic mucosa the highest levels of NOD2 expression were in proliferating crypt epithelial cells. Muramyl dipeptide (MDP), that is recognized by NOD2, promoted CEC growth in vitro. By contrast, the growth of NOD2-deficient CECs was impaired. In vivo CEC proliferation was also reduced and apoptosis increased in Nod2-/- mice, which were also evident following enteric Salmonella infection. Furthermore, neutralization of NOD2 mRNA expression in human colonic carcinoma cells by shRNA interference resulted in decreased survival due to increased levels of apoptosis.CONCLUSION: These findings are consistent with the involvement of NOD2 protein in promoting CEC growth and survival. Defects in proliferation by CECs in cases of CD may contribute to the underlying pathology of disrupted intestinal homeostasis and excessive inflammation.

  11. A comparative analysis of radiobiological models for cell surviving fractions at high doses.

    Science.gov (United States)

    Andisheh, B; Edgren, M; Belkić, Dž; Mavroidis, P; Brahme, A; Lind, B K

    2013-04-01

    For many years the linear-quadratic (LQ) model has been widely used to describe the effects of total dose and dose per fraction at low-to-intermediate doses in conventional fractionated radiotherapy. Recent advances in stereotactic radiosurgery (SRS) and stereotactic radiotherapy (SRT) have increased the interest in finding a reliable cell survival model, which will be accurate at high doses, as well. Different models have been proposed for improving descriptions of high dose survival responses, such as the Universal Survival Curve (USC), the Kavanagh-Newman (KN) and several generalizations of the LQ model, e.g. the Linear-Quadratic-Linear (LQL) model and the Pade Linear Quadratic (PLQ) model. The purpose of the present study is to compare a number of models in order to find the best option(s) which could successfully be used as a fractionation correction method in SRT. In this work, six independent experimental data sets were used: CHOAA8 (Chinese hamster fibroblast), H460 (non-small cell lung cancer, NSLC), NCI-H841 (small cell lung cancer, SCLC), CP3 and DU145 (human prostate carcinoma cell lines) and U1690 (SCLC). By detailed comparisons with these measurements, the performance of nine different radiobiological models was examined for the entire dose range, including high doses beyond the shoulder of the survival curves. Using the computed and measured cell surviving fractions, comparison of the goodness-of-fit for all the models was performed by means of the reduced χ (2)-test with a 95% confidence interval. The obtained results indicate that models with dose-independent final slopes and extrapolation numbers generally represent better choices for SRT. This is especially important at high doses where the final slope and extrapolation numbers are presently found to play a major role. The PLQ, USC and LQL models have the least number of shortcomings at all doses. The extrapolation numbers and final slopes of these models do not depend on dose. Their asymptotes

  12. Model of Protein Kinase B for Cell Survival/Death and its Equivalent Bio Circuit

    Science.gov (United States)

    Jain, Shruti; Naik, Pradeep K.; Bhooshan, Sunil V.

    2011-12-01

    Signaling pathways have traditionally focused on delineating immediate upstream and down stream interactions, and then organizing these interactions into linear cascades that relay and regulate information from cell surface receptors to cellular effectors such as metabolic enzymes, channels or transcription factors. However, recent analyses of signaling pathways have revealed that cellular signals do not necessarily propagate in a linear fashion. Because of their size and complexity, these networks are often too complicated for the human mind to organize and analyze. AkT (protein kinase B) is a central signaling molecule in the phosphatidylinositol 3-kinase pathway that is frequently activated in human cancer. Here we provide an overview of recent findings, how AkT promotes cell survival by inhibiting apoptosis by phosphorylating and inactivating several targets, including forkhead transcription factors, and caspase-9. The ability of AkT to promote survival was dependent on and proportional to its kinase activity. We have made computational model for AkT, on the basis of that model we have made the truth tables, Boolean equations and than implement the equations using logic circuits and Bio-circuits showing cell survival and death.

  13. Role of Ku80-dependent end-joining in delayed genomic instability in mammalian cells surviving ionizing radiation

    Energy Technology Data Exchange (ETDEWEB)

    Suzuki, Keiji, E-mail: kzsuzuki@nagasaki-u.ac.jp [Course of Life Sciences and Radiation Research, Graduate School of Biomedical Sciences, Nagasaki University, 1-12-4 Sakamoto, Nagasaki 852-8523 (Japan); Kodama, Seiji [Research Institute for Advanced Science and Technology, Osaka Prefecture University, 1-2 Gakuen-machi, Sakai 599-8570 (Japan); Watanabe, Masami [Kyoto University Research Reactor Institute, Kumatori-cho Sennan-gun, Osaka 590-0494 (Japan)

    2010-01-05

    Ionizing radiation induces delayed destabilization of the genome in the progenies of surviving cells. This phenomenon, which is called radiation-induced genomic instability, is manifested by delayed induction of radiation effects, such as cell death, chromosome aberration, and mutation in the progeny of cells surviving radiation exposure. Previously, there was a report showing that delayed cell death was absent in Ku80-deficient Chinese hamster ovary (CHO) cells, however, the mechanism of their defect has not been determined. We found that delayed induction of DNA double strand breaks and chromosomal breaks were intact in Ku80-deficient cells surviving X-irradiation, whereas there was no sign for the production of chromosome bridges between divided daughter cells. Moreover, delayed induction of dicentric chromosomes was significantly compromised in those cells compared to the wild-type CHO cells. Reintroduction of the human Ku86 gene complimented the defective DNA repair and recovered delayed induction of dicentric chromosomes and delayed cell death, indicating that defective Ku80-dependent dicentric induction was the cause of the absence of delayed cell death. Since DNA-PKcs-defective cells showed delayed phenotypes, Ku80-dependent illegitimate rejoining is involved in delayed impairment of the integrity of the genome in radiation-survived cells.

  14. Staying in Shape: the Impact of Cell Shape on Bacterial Survival in Diverse Environments.

    Science.gov (United States)

    Yang, Desirée C; Blair, Kris M; Salama, Nina R

    2016-03-01

    Bacteria display an abundance of cellular forms and can change shape during their life cycle. Many plausible models regarding the functional significance of cell morphology have emerged. A greater understanding of the genetic programs underpinning morphological variation in diverse bacterial groups, combined with assays of bacteria under conditions that mimic their varied natural environments, from flowing freshwater streams to diverse human body sites, provides new opportunities to probe the functional significance of cell shape. Here we explore shape diversity among bacteria, at the levels of cell geometry, size, and surface appendages (both placement and number), as it relates to survival in diverse environments. Cell shape in most bacteria is determined by the cell wall. A major challenge in this field has been deconvoluting the effects of differences in the chemical properties of the cell wall and the resulting cell shape perturbations on observed fitness changes. Still, such studies have begun to reveal the selective pressures that drive the diverse forms (or cell wall compositions) observed in mammalian pathogens and bacteria more generally, including efficient adherence to biotic and abiotic surfaces, survival under low-nutrient or stressful conditions, evasion of mammalian complement deposition, efficient dispersal through mucous barriers and tissues, and efficient nutrient acquisition.

  15. Inhibition of SIRT1 Catalytic Activity Increases p53 Acetylation but Does Not Alter Cell Survival following DNA Damage

    Science.gov (United States)

    Solomon, Jonathan M.; Pasupuleti, Rao; Xu, Lei; McDonagh, Thomas; Curtis, Rory; DiStefano, Peter S.; Huber, L. Julie

    2006-01-01

    Human SIRT1 is an enzyme that deacetylates the p53 tumor suppressor protein and has been suggested to modulate p53-dependent functions including DNA damage-induced cell death. In this report, we used EX-527, a novel, potent, and specific small-molecule inhibitor of SIRT1 catalytic activity to examine the role of SIRT1 in p53 acetylation and cell survival after DNA damage. Treatment with EX-527 dramatically increased acetylation at lysine 382 of p53 after different types of DNA damage in primary human mammary epithelial cells and several cell lines. Significantly, inhibition of SIRT1 catalytic activity by EX-527 had no effect on cell growth, viability, or p53-controlled gene expression in cells treated with etoposide. Acetyl-p53 was also increased by the histone deacetylase (HDAC) class I/II inhibitor trichostatin A (TSA). EX-527 and TSA acted synergistically to increase acetyl-p53 levels, confirming that p53 acetylation is regulated by both SIRT1 and HDACs. While TSA alone reduced cell survival after DNA damage, the combination of EX-527 and TSA had no further effect on cell viability and growth. These results show that, although SIRT1 deacetylates p53, this does not play a role in cell survival following DNA damage in certain cell lines and primary human mammary epithelial cells. PMID:16354677

  16. Expression of β-globin by cancer cells promotes cell survival during blood-borne dissemination

    Science.gov (United States)

    Zheng, Yu; Miyamoto, David T.; Wittner, Ben S.; Sullivan, James P.; Aceto, Nicola; Jordan, Nicole Vincent; Yu, Min; Karabacak, Nezihi Murat; Comaills, Valentine; Morris, Robert; Desai, Rushil; Desai, Niyati; Emmons, Erin; Milner, John D.; Lee, Richard J.; Wu, Chin-Lee; Sequist, Lecia V.; Haas, Wilhelm; Ting, David T.; Toner, Mehmet; Ramaswamy, Sridhar; Maheswaran, Shyamala; Haber, Daniel A.

    2017-01-01

    Metastasis-competent circulating tumour cells (CTCs) experience oxidative stress in the bloodstream, but their survival mechanisms are not well defined. Here, comparing single-cell RNA-Seq profiles of CTCs from breast, prostate and lung cancers, we observe consistent induction of β-globin (HBB), but not its partner α-globin (HBA). The tumour-specific origin of HBB is confirmed by sequence polymorphisms within human xenograft-derived CTCs in mouse models. Increased intracellular reactive oxygen species (ROS) in cultured breast CTCs triggers HBB induction, mediated through the transcriptional regulator KLF4. Depletion of HBB in CTC-derived cultures has minimal effects on primary tumour growth, but it greatly increases apoptosis following ROS exposure, and dramatically reduces CTC-derived lung metastases. These effects are reversed by the anti-oxidant N-Acetyl Cysteine. Conversely, overexpression of HBB is sufficient to suppress intracellular ROS within CTCs. Altogether, these observations suggest that β-globin is selectively deregulated in cancer cells, mediating a cytoprotective effect during blood-borne metastasis. PMID:28181495

  17. Measurement of posttransfusion red cell survival with the biotin label.

    Science.gov (United States)

    Mock, Donald M; Widness, John A; Veng-Pedersen, Peter; Strauss, Ronald G; Cancelas, Jose A; Cohen, Robert M; Lindsell, Christopher J; Franco, Robert S

    2014-07-01

    The goal of this review is to summarize and critically assess information concerning the biotin method to label red blood cells (RBC) for use in studies of RBC and transfusion biology-information that will prove useful to a broad audience of clinicians and scientists. A review of RBC biology, with emphasis on RBC senescence and in vivo survival, is included, followed by an analysis of the advantages and disadvantages of biotin-labeled RBC (BioRBC) for measuring circulating RBC volume, posttransfusion RBC recovery, RBC life span, and RBC age-dependent properties. The advantages of BioRBC over (51)Cr RBC labeling, the current reference method, are discussed. Because the biotin method is straightforward and robust, including the ability to follow the entire life spans of multiple RBC populations concurrently in the same subject, BioRBC offers distinct advantages for studying RBC biology and physiology, particularly RBC survival. The method for biotin labeling, validation of the method, and application of BioRBCs to studies of sickle cell disease, diabetes, and anemia of prematurity are reviewed. Studies documenting the safe use of BioRBC are reviewed; unanswered questions requiring future studies, remaining concerns, and regulatory barriers to broader application of BioRBC including adoption as a new reference method are also presented.

  18. Neuroendocrine differentiation as a survival prognostic factor in advanced non-small cell lung cancer

    Directory of Open Access Journals (Sweden)

    Petrović Marina

    2007-01-01

    Full Text Available Beckground/Aim. Neuroendrocine lung tumors are histologically heterogenous group of cancers with different clinical progression. In non-small cell lung cancer (NSCLC neuroendocrine differentiation exists in 10-30% of patients. The aim of this study was to determine the frequency and influence of neuroendocrine differentiation on survival of treated patients with advanced non-small cell lung cancer (NSCLC. Methods. A clinical trial included 158 patients (74% males and 26% females, with the diagnosis of NSCLC, determined by histological verification. The patients were treated by combined chemo - and X-ray therapy in stage III (without pleural effusion or chemotherapy only in stage III (with pleural effusion and stage IV. Chemotherapy was conducted until progression of the disease, but no more than six cycles. When the progression had been noted in stage III (without pleural effusion, the treatment was continued with X-ray therapy. Neuron specific enolase, chromogranin A, as well as synapthophysin expression in tissue examples were determined by immunohistochemical analysis with monoclonal mouse anti-human-bodies. Survival was assessed within a year and two years follow-up examination. Results. A total of 53 patients (34% had NSCLC with neuroendocrine differentiation, confirmed rather in large cell lung cancer and lung adenocarcinoma (66.7% and 40%, respectively. Neuron specific enolase, chromogranin A and synapthophysin expression was noted in 45 (28.5%, 34 (21.5% and 33 (20.1% patients, respectively. The one year and two years follow-up survival periods were confirmed in 39% and 17% of patients respectively. The median survival time in the patients with the neuroendocrine expression as compared to those without the expression was 15.6 vs 10.8 months; one year survival time with the expression compared to those without the expression achieved in 62% vs 27% of the patients, (p < 0.001; a two - year survival time noted in 30% of the patients (p = 0

  19. Regulation of cell survival by Na+/H+ exchanger-1.

    Science.gov (United States)

    Schelling, Jeffrey R; Abu Jawdeh, Bassam G

    2008-09-01

    Na(+)/H(+) exchanger-1 (NHE1) is a ubiquitous plasma membrane Na(+)/H(+) exchanger typically associated with maintenance of intracellular volume and pH. In addition to the NHE1 role in electroneutral Na(+)/H(+) transport, in renal tubular epithelial cells in vitro the polybasic, juxtamembrane NHE1 cytosolic tail domain acts as a scaffold, by binding with ezrin/radixin/moesin (ERM) proteins and phosphatidylinositol 4,5-bisphosphate, which initiates formation of a signaling complex that culminates in Akt activation and opposition to initial apoptotic stress. With robust apoptotic stimuli renal tubular epithelial cell NHE1 is a caspase substrate, and proteolytic cleavage may permit progression to apoptotic cell death. In vivo, genetic or pharmacological NHE1 loss of function causes renal tubule epithelial cell apoptosis and renal dysfunction following streptozotocin-induced diabetes, ureteral obstruction, and adriamycin-induced podocyte toxicity. Taken together, substantial in vivo and in vitro data demonstrate that NHE1 regulates tubular epithelial cell survival. In contrast to connotations of NHE1 as an unimportant "housekeeping" protein, this review highlights that NHE1 activity is critical for countering tubular atrophy and chronic renal disease progression.

  20. Triptolide increases transcript and protein levels of survival motor neurons in human SMA fibroblasts and improves survival in SMA-like mice.

    Science.gov (United States)

    Hsu, Ya-Yun; Jong, Yuh-Jyh; Tsai, Hsin-Hung; Tseng, Yu-Ting; An, Li-Mei; Lo, Yi-Ching

    2012-06-01

    Spinal muscular atrophy (SMA) is a progressive neuromuscular disease. Since disease severity is related to the amount of survival motor neuron (SMN) protein, up-regulated functional SMN protein levels from the SMN2 gene are considered a major SMA drug-discovery strategy. In this study, we investigated the possible effects of triptolide, a diterpene triepoxide purified from Tripterygium wilfordii Hook. F., as a new compound for increasing SMN protein. The effects and mechanisms of triptolide on the production of SMA protein were determined by cell-based assays using the motor neuronal cell line NSC34 and skin fibroblasts from SMA patients. Wild-type (Smn(+/+) SMN2(-/-) , C57BL/6) and SMA-like (Smn(-/-) SMN2) mice were injected with triptolide (0.01 or 0.1 mg·kg(-1) ·day(-1) , i.p.) and their survival rate and level of change in SMN protein in neurons and muscle tissue measured. In NSC34 cells and human SMA fibroblasts, pM concentrations of triptolide significantly increased SMN protein expression and the levels of SMN complex component (Gemin2 and Gemin3). In human SMA fibroblasts, triptolide increased SMN-containing nuclear gems and the ratio of full-length transcripts (FL-SMN2) to SMN2 transcripts lacking exon 7 (SMN2Δ7). Furthermore, in SMA-like mice, triptolide significantly increased SMN protein levels in the brain, spinal cord and gastrocnemius muscle. Furthermore, triptolide treatment increased survival and reduced weight loss in SMA-like mice. Triptolide enhanced SMN protein production by promoting SMN2 activation, exon 7 inclusion and increasing nuclear gems, and increased survival in SMA mice, which suggests triptolide might be a potential candidate for SMA therapy. © 2012 The Authors. British Journal of Pharmacology © 2012 The British Pharmacological Society.

  1. Survival-associated heterogeneity of marker-defined perivascular cells in colorectal cancer.

    Science.gov (United States)

    Mezheyeuski, Artur; Bradic Lindh, Maja; Guren, Tormod Kyrre; Dragomir, Anca; Pfeiffer, Per; Kure, Elin H; Ikdahl, Tone; Skovlund, Eva; Corvigno, Sara; Strell, Carina; Pietras, Kristian; Ponten, Fredrik; Mulder, Jan; Qvortrup, Camilla; Portyanko, Anna; Tveit, Kjell Magne; Glimelius, Bengt; Sorbye, Halfdan; Östman, Arne

    2016-07-05

    Perivascular cells (PC) were recently implied as regulators of metastasis and immune cell activity. Perivascular heterogeneity in clinical samples, and associations with other tumor features and outcome, remain largely unknown.Here we report a novel method for digital quantitative analyses of vessel characteristics and PC, which was applied to two collections of human metastatic colorectal cancer (mCRC).Initial analyses identified marker-defined subsets of PC, including cells expressing PDGFR-β or α-SMA or both markers. PC subsets were largely independently expressed in a manner unrelated to vessel density and size. Association studies implied specific oncogenic mutations in malignant cells as determinants of PC status. Semi-quantitative and digital-image-analyses-based scoring of the NORDIC-VII cohort identified significant associations between low expression of perivascular PDGFR-α and -β and shorter overall survival. Analyses of the SPCRC cohort confirmed these findings. Perivascular PDGFR-α and -β remained independent factors for survival in multivariate analyses.Overall, our study identified host vasculature and oncogenic status as determinants of tumor perivascular features. Perivascular PDGFR-α and -β were identified as novel independent markers predicting survival in mCRC. The novel methodology should be suitable for similar analyses in other tumor collections.

  2. Activation of Melatonin Signaling Promotes β-Cell Survival and Function.

    Science.gov (United States)

    Costes, Safia; Boss, Marti; Thomas, Anthony P; Matveyenko, Aleksey V

    2015-05-01

    Type 2 diabetes mellitus (T2DM) is characterized by pancreatic islet failure due to loss of β-cell secretory function and mass. Studies have identified a link between a variance in the gene encoding melatonin (MT) receptor 2, T2DM, and impaired insulin secretion. This genetic linkage raises the question whether MT signaling plays a role in regulation of β-cell function and survival in T2DM. To address this postulate, we used INS 832/13 cells to test whether activation of MT signaling attenuates proteotoxicity-induced β-cell apoptosis and through which molecular mechanism. We also used nondiabetic and T2DM human islets to test the potential of MT signaling to attenuate deleterious effects of glucotoxicity and T2DM on β-cell function. MT signaling in β-cells (with duration designed to mimic typical nightly exposure) significantly enhanced activation of the cAMP-dependent signal transduction pathway and attenuated proteotoxicity-induced β-cell apoptosis evidenced by reduced caspase-3 cleavage (∼40%), decreased activation of stress-activated protein kinase/Jun-amino-terminal kinase (∼50%) and diminished oxidative stress response. Activation of MT signaling in human islets was shown to restore glucose-stimulated insulin secretion in islets exposed to chronic hyperglycemia as well as in T2DM islets. Our data suggest that β-cell MT signaling is important for the regulation of β-cell survival and function and implies a preventative and therapeutic potential for preservation of β-cell mass and function in T2DM.

  3. Selective Mitochondrial Uptake of MKT-077 Can Suppress Medullary Thyroid Carcinoma Cell Survival and

    Directory of Open Access Journals (Sweden)

    Dmytro Starenki

    2015-12-01

    Full Text Available BackgroundMedullary thyroid carcinoma (MTC is a neuroendocrine tumor mainly caused by mutations in the rearranged during transfection (RET proto-oncogene. Not all patients with progressive MTC respond to current therapy inhibiting RET, demanding additional therapeutic strategies. We recently demonstrated that disrupting mitochondrial metabolism using a mitochondria-targeted agent or by depleting a mitochondrial chaperone effectively suppressed human MTC cells in culture and in mouse xenografts by inducing apoptosis and RET downregulation. These observations led us to hypothesize that mitochondria are potential therapeutic targets for MTC. This study further tests this hypothesis using1-ethyl-2-[[3-ethyl-5-(3-methylbenzothiazolin-2-yliden]-4-oxothiazolidin-2-ylidenemethyl] pyridinium chloride (MKT-077, a water-soluble rhodocyanine dye analogue, which can selectively accumulate in mitochondria.MethodsThe effects of MKT-077 on cell proliferation, survival, expression of RET and tumor protein 53 (TP53, and mitochondrial activity were determined in the human MTC lines in culture and in mouse xenografts.ResultsMKT-077 induced cell cycle arrest in TT and MZ-CRC-1. Intriguingly, MKT-077 also induced RET downregulation and strong cell death responses in TT cells, but not in MZ-CRC-1 cells. This discrepancy was mainly due to the difference between the capacities of these cell lines to retain MKT-077 in mitochondria. The cytotoxicity of MKT-077 in TT cells was mainly attributed to oxidative stress while being independent of TP53. MKT-077 also effectively suppressed tumor growth of TT xenografts.ConclusionMKT-077 can suppress cell survival of certain MTC subtypes by accumulating in mitochondria and interfering with mitochondrial activity although it can also suppress cell proliferation via other mechanisms. These results consistently support the hypothesis that mitochondrial targeting has therapeutic potential for MTC.

  4. Melanin dependent survival of Apergillus fumigatus conidia in lung epithelial cells.

    Science.gov (United States)

    Amin, Shayista; Thywissen, Andreas; Heinekamp, Thorsten; Saluz, Hans Peter; Brakhage, Axel A

    2014-07-01

    Aspergillus fumigatus is the most important air-borne pathogenic fungus of humans. Upon inhalation of conidia, the fungus makes close contact with lung epithelial cells, which only possess low phagocytic activity. These cells are in particular interesting to address the question whether there is some form of persistence of conidia of A. fumigatus in the human host. Therefore, by also using uracil-auxotrophic mutant strains, we were able to investigate the interaction of A549 lung epithelial cells and A. fumigatus conidia in detail for long periods. Interestingly, unlike professional phagocytes, our study showed that the presence of conidial dihydroxynaphthalene (DHN) melanin enhanced the uptake of A. fumigatus conidia by epithelial cells when compared with non-pigmented pksP mutant conidia. Furthermore, conidia of A. fumigatus were able to survive within epithelial cells. This was due to the presence of DHN melanin in the cell wall of conidia, because melanised wild-type conidia showed a higher survival rate inside epithelial cells and led to inhibition of acidification of phagolysosomes. Both effects were not observed for white (non-melanised) conidia of the pksP mutant strain. Moreover, in contrast to pksP mutant conidia, melanised wild-type conidia were able to inhibit the extrinsic apoptotic pathway in A549 lung epithelial cells even for longer periods. The anti-apoptotic effect was not restricted to conidia, because both conidia-derived melanin ghosts (cell-free DHN melanin) and a different type of melanin, dihydroxyphenylalanine (DOPA) melanin, acted anti-apoptotically. Taken together, these data indicate the possibility of melanin-dependent persistence of conidia in lung epithelial cells.

  5. Germ cells may survive clipping and division of the spermatic vessels in surgery for intra-abdominal testes

    DEFF Research Database (Denmark)

    Thorup, J M; Cortes, Dina; Visfeldt, J

    1999-01-01

    Laparoscopy is a well described modality that provides an accurate visual diagnosis upon which further management of intra-abdominal testes may be based. Laparoscopic ligation of spermatic vessels as stage 1 of the procedure is a natural extension of laparoscopy. A staged approach provides adequa...... viability of the intra-abdominal testis. However, it is uncertain whether the more sensitive germ cells survive this procedure in addition to the Sertoli and interstitial cells of the human testis. Survival of germ cells is a prerequisite of later fertility potential....

  6. Expression of HSF2 decreases in mitosis to enable stress-inducible transcription and cell survival.

    Science.gov (United States)

    Elsing, Alexandra N; Aspelin, Camilla; Björk, Johanna K; Bergman, Heidi A; Himanen, Samu V; Kallio, Marko J; Roos-Mattjus, Pia; Sistonen, Lea

    2014-09-15

    Unless mitigated, external and physiological stresses are detrimental for cells, especially in mitosis, resulting in chromosomal missegregation, aneuploidy, or apoptosis. Heat shock proteins (Hsps) maintain protein homeostasis and promote cell survival. Hsps are transcriptionally regulated by heat shock factors (HSFs). Of these, HSF1 is the master regulator and HSF2 modulates Hsp expression by interacting with HSF1. Due to global inhibition of transcription in mitosis, including HSF1-mediated expression of Hsps, mitotic cells are highly vulnerable to stress. Here, we show that cells can counteract transcriptional silencing and protect themselves against proteotoxicity in mitosis. We found that the condensed chromatin of HSF2-deficient cells is accessible for HSF1 and RNA polymerase II, allowing stress-inducible Hsp expression. Consequently, HSF2-deficient cells exposed to acute stress display diminished mitotic errors and have a survival advantage. We also show that HSF2 expression declines during mitosis in several but not all human cell lines, which corresponds to the Hsp70 induction and protection against stress-induced mitotic abnormalities and apoptosis.

  7. Expression of HSF2 decreases in mitosis to enable stress-inducible transcription and cell survival

    Science.gov (United States)

    Elsing, Alexandra N.; Aspelin, Camilla; Björk, Johanna K.; Bergman, Heidi A.; Himanen, Samu V.; Kallio, Marko J.; Roos-Mattjus, Pia

    2014-01-01

    Unless mitigated, external and physiological stresses are detrimental for cells, especially in mitosis, resulting in chromosomal missegregation, aneuploidy, or apoptosis. Heat shock proteins (Hsps) maintain protein homeostasis and promote cell survival. Hsps are transcriptionally regulated by heat shock factors (HSFs). Of these, HSF1 is the master regulator and HSF2 modulates Hsp expression by interacting with HSF1. Due to global inhibition of transcription in mitosis, including HSF1-mediated expression of Hsps, mitotic cells are highly vulnerable to stress. Here, we show that cells can counteract transcriptional silencing and protect themselves against proteotoxicity in mitosis. We found that the condensed chromatin of HSF2-deficient cells is accessible for HSF1 and RNA polymerase II, allowing stress-inducible Hsp expression. Consequently, HSF2-deficient cells exposed to acute stress display diminished mitotic errors and have a survival advantage. We also show that HSF2 expression declines during mitosis in several but not all human cell lines, which corresponds to the Hsp70 induction and protection against stress-induced mitotic abnormalities and apoptosis. PMID:25202032

  8. JAM-C maintains VEGR2 expression to promote retinal pigment epithelium cell survival under oxidative stress.

    Science.gov (United States)

    Jia, Xin; Zhao, Chen; Chen, Qishan; Du, Yuxiang; Huang, Lijuan; Ye, Zhimin; Ren, Xiangrong; Wang, Shasha; Lee, Chunsik; Tang, Zhongshu; Li, Xuri; Ju, Rong

    2017-04-03

    Junctional adhesion molecule-C (JAM-C) has been shown to play critical roles during development and in immune responses. However, its role in adult eyes under oxidative stress remains poorly understood. Here, we report that JAM-C is abundantly expressed in adult mouse retinae and choroids in vivo and in cultured retinal pigment epithelium (RPE) and photoreceptor cells in vitro. Importantly, both JAM-C expression and its membrane localisation are downregulated by H2O2-induced oxidative stress. Under H2O2-induced oxidative stress, JAM-C is critically required for the survival of human RPE cells. Indeed, loss of JAM-C by siRNA knockdown decreased RPE cell survival. Mechanistically, we show that JAM-C is required to maintain VEGFR2 expression in RPE cells, and VEGFR2 plays an important role in keeping the RPE cells viable since overexpression of VEGFR2 partially restored impaired RPE survival caused by JAM-C knockdown and increased RPE survival. We further show that JAM-C regulates VEGFR2 expression and, in turn, modulates p38 phosphorylation. Together, our data demonstrate that JAM-C plays an important role in maintaining VEGR2 expression to promote RPE cell survival under oxidative stress. Given the vital importance of RPE in the eye, approaches that can modulate JAM-C expression may have therapeutic values in treating diseases with impaired RPE survival.

  9. Oral cancer/endothelial cell fusion experiences nuclear fusion and acquisition of enhanced survival potential

    Energy Technology Data Exchange (ETDEWEB)

    Song, Kai [Department of Oral and Maxillofacial Surgery, The Affiliated Hospital of Qingdao University, Shandong Province (China); The State Key Laboratory Breeding Base of Basic Science of Stomatology (Hubei-MOST) and Key Laboratory for Oral Biomedicine Ministry of Education, Wuhan University, Wuhan (China); Song, Yong [The State Key Laboratory Breeding Base of Basic Science of Stomatology (Hubei-MOST) and Key Laboratory for Oral Biomedicine Ministry of Education, Wuhan University, Wuhan (China); Department of Stomatology, Liu Zhou People' s Hospital, Guangxi (China); Zhao, Xiao-Ping; Shen, Hui; Wang, Meng; Yan, Ting-lin [The State Key Laboratory Breeding Base of Basic Science of Stomatology (Hubei-MOST) and Key Laboratory for Oral Biomedicine Ministry of Education, Wuhan University, Wuhan (China); Liu, Ke, E-mail: liuke.1999@aliyun.com [Department of Oral and Maxillofacial-Head and Neck oncology, School and Hospital of Stomatology, Wuhan University, 237 Luoyu Road, Wuhan 430079 (China); The State Key Laboratory Breeding Base of Basic Science of Stomatology (Hubei-MOST) and Key Laboratory for Oral Biomedicine Ministry of Education, Wuhan University, Wuhan (China); Shang, Zheng-jun, E-mail: shangzhengjun@hotmail.com [Department of Oral and Maxillofacial-Head and Neck oncology, School and Hospital of Stomatology, Wuhan University, 237 Luoyu Road, Wuhan 430079 (China); The State Key Laboratory Breeding Base of Basic Science of Stomatology (Hubei-MOST) and Key Laboratory for Oral Biomedicine Ministry of Education, Wuhan University, Wuhan (China)

    2014-10-15

    Most previous studies have linked cancer–macrophage fusion with tumor progression and metastasis. However, the characteristics of hybrid cells derived from oral cancer and endothelial cells and their involvement in cancer remained unknown. Double-immunofluorescent staining and fluorescent in situ hybridization (FISH) were performed to confirm spontaneous cell fusion between eGFP-labeled human umbilical vein endothelial cells (HUVECs) and RFP-labeled SCC9, and to detect the expression of vementin and cytokeratin 18 in the hybrids. The property of chemo-resistance of such hybrids was examined by TUNEL assay. The hybrid cells in xenografted tumor were identified by FISH and GFP/RFP dual-immunofluoresence staining. We showed that SCC9 cells spontaneously fused with cocultured endothelial cells, and the resultant hybrid cells maintained the division and proliferation activity after re-plating and thawing. Such hybrids expressed markers of both parental cells and became more resistant to chemotherapeutic drug cisplatin as compared to the parental SCC9 cells. Our in vivo data indicated that the hybrid cells contributed to tumor composition by using of immunostaining and FISH analysis, even though the hybrid cells and SCC9 cells were mixed with 1:10,000, according to the FACS data. Our study suggested that the fusion events between oral cancer and endothelial cells undergo nuclear fusion and acquire a new property of drug resistance and consequently enhanced survival potential. These experimental findings provide further supportive evidence for the theory that cell fusion is involved in cancer progression. - Highlights: • The fusion events between oral cancer and endothelial cells undergo nuclear fusion. • The resulting hybrid cells acquire a new property of drug resistance. • The resulting hybrid cells express the markers of both parental cells (i.e. vimentin and cytokeratin 18). • The hybrid cells contribute to tumor repopulation in vivo.

  10. Nitric oxide mediates the survival action of IGF-1 and insulin in pancreatic beta cells.

    Science.gov (United States)

    Cahuana, Gladys M; Tejedo, Juan R; Hmadcha, Abdelkrim; Ramírez, Remedios; Cuesta, Antonio L; Soria, Bernat; Martin, Franz; Bedoya, Francisco J

    2008-02-01

    Generation of low levels of nitric oxide (NO) contributes to beta cell survival in vitro. The purpose of this study was to explore the link between NO and the survival pathway triggered by insulin-like growth factor-1 (IGF-1) and insulin in insulin producing RINm5F cells and in pancreatic islets. Results show that exposure of cells to IGF-1/insulin protects against serum deprivation-induced apoptosis. This action is prevented with inhibitors of NO generation, PI3K and Akt. Moreover, transfection with the negative dominant form of the tyrosine kinase c-Src abrogates the effect of IGF-1 and insulin on DNA fragmentation. An increase in the expression level of NOS3 protein and in the enzyme activity is observed following exposure of serum-deprived RINm5F cells to IGF-1 and insulin. Phosphorylation of IRS-1, IRS-2 and to less extent IRS-3 takes place when serum-deprived RINm5F cells and rat pancreatic islets are exposed to either IGF-1, insulin, or diethylenetriamine nitric oxide adduct (DETA/NO). In human islets, IRS-1 and IRS-2 proteins are present and tyrosine phosphorylated upon exposure to IGF-1, insulin and DETA/NO. Both rat and human pancreatic islets undergo DNA fragmentation when cultured in serum-free medium and IGF-1, insulin and DETA/NO protect efficiently from this damage. We then conclude that generation of NO participates in the activation of survival pathways by IGF-1 and insulin in beta cells.

  11. Leukemia cell microvesicles promote survival in umbilical cord blood hematopoietic stem cells.

    Science.gov (United States)

    Razmkhah, Farnaz; Soleimani, Masoud; Mehrabani, Davood; Karimi, Mohammad Hossein; Kafi-Abad, Sedigheh Amini

    2015-01-01

    Microvesicles can transfer their contents, proteins and RNA, to target cells and thereby transform them. This may induce apoptosis or survival depending on cell origin and the target cell. In this study, we investigate the effect of leukemic cell microvesicles on umbilical cord blood hematopoietic stem cells to seek evidence of apoptosis or cell survival. Microvesicles were isolated from both healthy donor bone marrow samples and Jurkat cells by ultra-centrifugation and were added to hematopoietic stem cells sorted from umbilical cord blood samples by magnetic associated cell sorting (MACS) technique. After 7 days, cell count, cell viability, flow cytometry analysis for hematopoietic stem cell markers and qPCR for P53 gene expression were performed. The results showed higher cell number, higher cell viability rate and lower P53 gene expression in leukemia group in comparison with normal and control groups. Also, CD34 expression as the most important hematopoietic stem cell marker, did not change during the treatment and lineage differentiation was not observed. In conclusion, this study showed anti-apoptotic effect of leukemia cell derived microvesicles on umbilical cord blood hematopoietic stem cells.

  12. Human T Cell Memory: A Dynamic View

    Directory of Open Access Journals (Sweden)

    Derek C. Macallan

    2017-02-01

    Full Text Available Long-term T cell-mediated protection depends upon the formation of a pool of memory cells to protect against future pathogen challenge. In this review we argue that looking at T cell memory from a dynamic viewpoint can help in understanding how memory populations are maintained following pathogen exposure or vaccination. For example, a dynamic view resolves the apparent paradox between the relatively short lifespans of individual memory cells and very long-lived immunological memory by focussing on the persistence of clonal populations, rather than individual cells. Clonal survival is achieved by balancing proliferation, death and differentiation rates within and between identifiable phenotypic pools; such pools correspond broadly to sequential stages in the linear differentiation pathway. Each pool has its own characteristic kinetics, but only when considered as a population; single cells exhibit considerable heterogeneity. In humans, we tend to concentrate on circulating cells, but memory T cells in non-lymphoid tissues and bone marrow are increasingly recognised as critical for immune defence; their kinetics, however, remain largely unexplored. Considering vaccination from this viewpoint shifts the focus from the size of the primary response to the survival of the clone and enables identification of critical system pinch-points and opportunities to improve vaccine efficacy.

  13. VILIP-1 downregulation in non-small cell lung carcinomas: mechanisms and prediction of survival.

    Directory of Open Access Journals (Sweden)

    Jian Fu

    Full Text Available VILIP-1, a member of the neuronal Ca++ sensor protein family, acts as a tumor suppressor gene in an experimental animal model by inhibiting cell proliferation, adhesion and invasiveness of squamous cell carcinoma cells. Western Blot analysis of human tumor cells showed that VILIP-1 expression was undetectable in several types of human tumor cells, including 11 out of 12 non-small cell lung carcinoma (NSCLC cell lines. The down-regulation of VILIP-1 was due to loss of VILIP-1 mRNA transcripts. Rearrangements, large gene deletions or mutations were not found. Hypermethylation of the VILIP-1 promoter played an important role in gene silencing. In most VILIP-1-silent cells the VILIP-1 promoter was methylated. In vitro methylation of the VILIP-1 promoter reduced its activity in a promoter-reporter assay. Transcriptional activity of endogenous VILIP-1 promoter was recovered by treatment with 5'-aza-2'-deoxycytidine (5'-Aza-dC. Trichostatin A (TSA, a histone deacetylase inhibitor, potently induced VILIP-1 expression, indicating that histone deacetylation is an additional mechanism of VILIP-1 silencing. TSA increased histone H3 and H4 acetylation in the region of the VILIP-1 promoter. Furthermore, statistical analysis of expression and promoter methylation (n = 150 primary NSCLC samples showed a significant relationship between promoter methylation and protein expression downregulation as well as between survival and decreased or absent VILIP-1 expression in lung cancer tissues (p<0.0001. VILIP-1 expression is silenced by promoter hypermethylation and histone deacetylation in aggressive NSCLC cell lines and primary tumors and its clinical evaluation could have a role as a predictor of short-term survival in lung cancer patients.

  14. Lipocalin-2-induced cytokine production enhances endometrial carcinoma cell survival and migration

    Directory of Open Access Journals (Sweden)

    Hsiu-Hsia Lin, Chi-Jr Liao, Ying-Chu Lee, Keng-Hsun Hu, Hsien-Wei Meng, Sin-Tak Chu

    2011-01-01

    Full Text Available Lipocalin-2 (Lcn-2 is an acute-phase protein that has been implicated in diverse physiological processes in mice, including: apoptosis, ion transport, inflammation, cell survival, and tumorigenesis. This study characterized the biological activity of Lcn-2 in human endometrial carcinoma cells (RL95-2. Exposure of RL95-2 cells to Lcn-2 for >24 h reduced Lcn-2-induced cell apoptosis, changed the cell proliferation and up-regulated cytokine secretions, including: interleukin-8 (IL-8, inteleukin-6 (IL-6, monocyte chemotatic protein-1 (MCP-1 and growth-related oncogene (GRO. However, IL-8 mRNA and protein levels were dramatically increased in Lcn-2-treated RL95-2 cells. To determine the IL-8 effect on Lcn-2-treated RL95-2 cells was our major focus. Adding recombinant IL-8 (rIL-8 resulted in decreased caspase-3 activity in Lcn-2-treated cells, whereas the addition of IL-8 antibodies resulted in significantly increased caspase-3 activity and decreased cell migration. Data indicate that IL-8 plays a crucial role in the induction of cell migration. Interestingly, Lcn-2-induced cytokines, secretion from RL95-2 cells, could not show the potent cell migration ability with the exception of IL-8. We conclude that Lcn-2 triggered cytokine secretions to prevent RL95-2 cells from undergoing apoptosis and subsequently increased cell migration. We hypothesize that Lcn-2 increased cytokine secretion by RL95-2 cells, which in turn activated a cellular defense system. This study suggests that Lcn-2 may play a role in the human female reproductive system or in endometrial cancer.

  15. Increased survival of normal cells during laser photodynamic therapy: implications for ex vivo autologous bone marrow purging

    Energy Technology Data Exchange (ETDEWEB)

    Gulliya, K.S.; Matthews, J.L.; Fay, J.W.; Dowben, R.M.

    1988-01-01

    Laser light-induced, dye-mediated photolysis of leukemic cells was tested in an in vitro model for its efficacy in eliminating occult tumor cells for ex vivo autologous bone marrow purging. Merocyanine 540 (MC540) was mixed with acute promyelocytic leukemia (HL-60) cells in the presence of human albumin. This cell-dye mixture was irradiated with 514 nm argon laser light. Results show that in the presence of 0.1%, 0.25% and 0.5% albumin, laser light doses of 62.4 J/cm/sup 2/, 93.6 J/cm/sup 2/ and 109.2 J/cm/sup 2/, respectively, were required for a 5 log reduction in the survival of leukemic cells. Under identical conditions, 80% to 84% of the normal bone marrow cells and 41% of the granulocyte-macrophage colony forming cells survived. The number of surviving stromal cells was reduced (1+) compared to the untreated control (4+). Mixing of irradiated bone marrow cells with equal number of HL-60 cells did not interfere with the killing of HL-60 cells treated with MC540 and laser light. The non-specific cytotoxicity of laser light alone was less than 6% for normal bone marrow cells. These results suggest that the concentration of human albumin plays an important role in laser light-induced phototoxicity. This laser light-induced selective photolysis of leukemic cells can be used in ex vivo purging of tumor cell-contaminated bone marrow grafts to achieve very high survival rates of normal bone marrow cells and granulocyte-macrophage colony forming cells.

  16. Protecting and rescuing the effectors: roles of differentiation and survival in the control of memory T cell development

    Directory of Open Access Journals (Sweden)

    Sema eKurtulus

    2013-01-01

    Full Text Available Vaccines, arguably the single most important intervention in improving human health, have exploited the phenomenon of immunological memory. The elicitation of memory T cells is often an essential part of successful long-lived protective immunity. Our understanding of T cell memory has been greatly aided by the development of TCR Tg mice and MHC tetrameric staining reagents that have allowed the precise tracking of antigen-specific T cell responses. Indeed, following acute infection or immunization, naïve T cells undergo a massive expansion culminating in the generation of a robust effector T cell population. This peak effector response is relatively short-lived and, while most effector T cells die by apoptosis, some remain and develop into memory cells. Although the molecular mechanisms underlying this cell fate decision remain incompletely defined, substantial progress has been made, particularly with regards to CD8+ T cells. For example, the effector CD8+ T cells generated during a response are heterogeneous, consisting of cells with more or less potential to develop into full-fledged memory cells. Development of CD8+ T cell memory is regulated by the transcriptional programs that control the differentiation and survival of effector T cells. While the type of antigenic stimulation and level of inflammation control effector CD8+ T cell differentiation, availability of cytokines and their ability to control expression and function of Bcl-2 family members governs their survival. These distinct differentiation and survival programs may allow for finer therapeutic intervention to control both the quality and quantity of CD8+ T cell memory. Effector to memory transition of CD4+ T cells is less well characterized than CD8+ T cells, emerging details will be discussed. This review will focus on the recent progress made in our understanding of the mechanisms underlying the development of T cell memory with an emphasis on factors controlling survival of

  17. Transcriptome profiling and genome-wide DNA binding define the differential role of fenretinide and all-trans RA in regulating the death and survival of human hepatocellular carcinoma Huh7 cells.

    Science.gov (United States)

    Hu, Ying; Liu, Hui-Xin; He, Yuqi; Fang, Yaping; Fang, Jianwen; Wan, Yu-Jui Yvonne

    2013-04-01

    Fenretinide is significantly more effective in inducing apoptosis in cancer cells than all-trans retinoic acid (ATRA). The current study uses a genome-wide approach to understand the differential role fenretinide and ATRA have in inducing apoptosis in Huh7 cells. Fenretinide and ATRA-induced gene expressions and DNA bindings were profiled using microarray and chromatin immunoprecipitation with anti-RXRα antibody. The data showed that fenretinide was not a strong transcription regulator. Fenretinide only changed the expressions of 1 093 genes, approximately three times less than the number of genes regulated by ATRA (2 811). Biological function annotation demonstrated that both fenretinide and ATRA participated in pathways that determine cell fate and metabolic processes. However, fenretinide specifically induced Fas/TNFα-mediated apoptosis by increasing the expression of pro-apoptotic genes i.e., DEDD2, CASP8, CASP4, and HSPA1A/B; whereas, ATRA induced the expression of BIRC3 and TNFAIP3, which inhibit apoptosis by interacting with TRAF2. In addition, fenretinide inhibited the expression of the genes involved in RAS/RAF/ERK-mediated survival pathway. In contrast, ATRA increased the expression of SOSC2, BRAF, MEK, and ERK genes. Most genes regulated by fenretinide and ATRA were bound by RXRα, suggesting a direct effect. This study revealed that by regulating fewer genes, the effects of fenretinide become more specific and thus has fewer side effects than ATRA. The data also suggested that fenretinide induces apoptosis via death receptor effector and by inhibiting the RAS/RAF/ERK pathway. It provides insight on how retinoid efficacy can be improved and how side effects in cancer therapy can be reduced.

  18. Recombinant human erythropoietin increases survival and reduces neuronal apoptosis in a murine model of cerebral malaria

    Directory of Open Access Journals (Sweden)

    Hempel Casper

    2008-01-01

    Full Text Available Abstract Background Cerebral malaria (CM is an acute encephalopathy with increased pro-inflammatory cytokines, sequestration of parasitized erythrocytes and localized ischaemia. In children CM induces cognitive impairment in about 10% of the survivors. Erythropoietin (Epo has – besides of its well known haematopoietic properties – significant anti-inflammatory, antioxidant and anti-apoptotic effects in various brain disorders. The neurobiological responses to exogenously injected Epo during murine CM were examined. Methods Female C57BL/6j mice (4–6 weeks, infected with Plasmodium berghei ANKA, were treated with recombinant human Epo (rhEpo; 50–5000 U/kg/OD, i.p. at different time points. The effect on survival was measured. Brain pathology was investigated by TUNEL (Terminal deoxynucleotidyl transferase (TdT-mediated deoxyuridine triphosphate (dUTP-digoxigenin nick end labelling, as a marker of apoptosis. Gene expression in brain tissue was measured by real time PCR. Results Treatment with rhEpo increased survival in mice with CM in a dose- and time-dependent manner and reduced apoptotic cell death of neurons as well as the expression of pro-inflammatory cytokines in the brain. This neuroprotective effect appeared to be independent of the haematopoietic effect. Conclusion These results and its excellent safety profile in humans makes rhEpo a potential candidate for adjunct treatment of CM.

  19. TPD52 represents a survival factor in ERBB2-amplified breast cancer cells.

    Science.gov (United States)

    Roslan, Nuruliza; Bièche, Ivan; Bright, Robert K; Lidereau, Rosette; Chen, Yuyan; Byrne, Jennifer A

    2014-10-01

    TPD52 and ERBB2 co-expression has been persistently reported in human breast cancer and animal models of this disease, but the significance of this is unknown. We identified significant positive associations between relative TPD52 and ERBB2 transcript levels in human diagnostic breast cancer samples, and maximal TPD52 expression in the hormone receptor (HR)- and ERBB2-positive sub-group. High-level TPD52 expression was associated with significantly reduced metastasis-free survival, within the overall cohort (log rank test, P = 8.6 × 10(-4), n = 375) where this was an independent predictor of metastasis-free survival (hazard ratio, 2.69, 95% confidence interval 1.59-4.54, P = 2.2 × 10(-4), n = 359), and the HR- and ERBB2-positive sub-group (log rank test, P = 0.035, n = 47). Transient TPD52 knock-down in the ERBB2-amplified breast cancer cell lines SK-BR-3 and BT-474 produced significant apoptosis, both singly and in combination with transient ERBB2 knock-down. Unlike ERBB2 knock-down, transient TPD52 knock-down produced no reduction in pAKT levels in SK-BR-3 or BT-474 cells. We then derived multiple SK-BR-3 cell lines in which TPD52 levels were stably reduced, and measured significant inverse correlations between pERBB2 and TPD52 levels in viable TPD52-depleted and control cell lines, all of which showed similar proliferative capacities. Our results therefore identify TPD52 as a survival factor in ERBB2-amplified breast cancer cells, and suggest complementary cellular functions for TPD52 and ERBB2.

  20. Fludarabine nucleoside modulates nuclear "survival and death" proteins in resistant chronic lymphocytic leukemia cells.

    Science.gov (United States)

    Henrich, Silke; Mactier, Swetlana; Best, Giles; Mulligan, Stephen P; Crossett, Ben; Christopherson, Richard Ian

    2011-12-01

    The nuclear mechanisms by which fludarabine nucleoside (F-ara-A) induces apoptosis have been investigated in human MEC1 cells derived from B-cell chronic lymphocytic leukemia. Upon treatment of cells with F-ara-A (100 μM, 72 hours), 15 nuclear proteins changed in abundance by more than 2-fold. Nuclear proteins up-regulated included calmodulin (4.3-fold), prohibitin (3.9-fold), β-actin variant (3.7-fold), and structure-specific recognition protein 1 (3.7-fold); those down-regulated included 60S ribosomal protein P2B (0.12-fold), fumarate hydratase (0.19-fold), splicing factor arginine/serine-rich 3 (0.35-fold), and replication protein A2 (0.42-fold). These changes in the levels of specific proteins promote survival or apoptosis; because the end result is apoptosis of MEC1 cells, apoptotic effects predominate.

  1. Survival of glucose phosphate isomerase null somatic cells and germ cells in adult mouse chimaeras.

    Science.gov (United States)

    Keighren, Margaret A; Flockhart, Jean H; West, John D

    2016-05-15

    The mouse Gpi1 gene encodes the glycolytic enzyme glucose phosphate isomerase. Homozygous Gpi1(-/-) null mouse embryos die but a previous study showed that some homozygous Gpi1(-/-) null cells survived when combined with wild-type cells in fetal chimaeras. One adult female Gpi1(-/-)↔Gpi1(c/c) chimaera with functional Gpi1(-/-) null oocytes was also identified in a preliminary study. The aims were to characterise the survival of Gpi1(-/-) null cells in adult Gpi1(-/-)↔Gpi1(c/c) chimaeras and determine if Gpi1(-/-) null germ cells are functional. Analysis of adult Gpi1(-/-)↔Gpi1(c/c) chimaeras with pigment and a reiterated transgenic lineage marker showed that low numbers of homozygous Gpi1(-/-) null cells could survive in many tissues of adult chimaeras, including oocytes. Breeding experiments confirmed that Gpi1(-/-) null oocytes in one female Gpi1(-/-)↔Gpi1(c/c) chimaera were functional and provided preliminary evidence that one male putative Gpi1(-/-)↔Gpi1(c/c) chimaera produced functional spermatozoa from homozygous Gpi1(-/-) null germ cells. Although the male chimaera was almost certainly Gpi1(-/-)↔Gpi1(c/c), this part of the study is considered preliminary because only blood was typed for GPI. Gpi1(-/-) null germ cells should survive in a chimaeric testis if they are supported by wild-type Sertoli cells. It is also feasible that spermatozoa could bypass a block at GPI, but not blocks at some later steps in glycolysis, by using fructose, rather than glucose, as the substrate for glycolysis. Although chimaera analysis proved inefficient for studying the fate of Gpi1(-/-) null germ cells, it successfully identified functional Gpi1(-/-) null oocytes and revealed that some Gpi1(-/-) null cells could survive in many adult tissues.

  2. Genome engineering in human cells.

    Science.gov (United States)

    Song, Minjung; Kim, Young-Hoon; Kim, Jin-Soo; Kim, Hyongbum

    2014-01-01

    Genome editing in human cells is of great value in research, medicine, and biotechnology. Programmable nucleases including zinc-finger nucleases, transcription activator-like effector nucleases, and RNA-guided engineered nucleases recognize a specific target sequence and make a double-strand break at that site, which can result in gene disruption, gene insertion, gene correction, or chromosomal rearrangements. The target sequence complexities of these programmable nucleases are higher than 3.2 mega base pairs, the size of the haploid human genome. Here, we briefly introduce the structure of the human genome and the characteristics of each programmable nuclease, and review their applications in human cells including pluripotent stem cells. In addition, we discuss various delivery methods for nucleases, programmable nickases, and enrichment of gene-edited human cells, all of which facilitate efficient and precise genome editing in human cells.

  3. Induced Pluripotent Stem Cell-Derived Neural Cells Survive and Mature in the Nonhuman Primate Brain

    Directory of Open Access Journals (Sweden)

    Marina E. Emborg

    2013-03-01

    Full Text Available The generation of induced pluripotent stem cells (iPSCs opens up the possibility for personalized cell therapy. Here, we show that transplanted autologous rhesus monkey iPSC-derived neural progenitors survive for up to 6 months and differentiate into neurons, astrocytes, and myelinating oligodendrocytes in the brains of MPTP-induced hemiparkinsonian rhesus monkeys with a minimal presence of inflammatory cells and reactive glia. This finding represents a significant step toward personalized regenerative therapies.

  4. In vitro radiosensitivity of human leukemia cell lines

    Energy Technology Data Exchange (ETDEWEB)

    Weichselbaum, R.R.; Greenberger, J.S.; Schmidt, A.; Karpas, A.; Moloney, W.C.; Little, J.B.

    1981-05-01

    The in vitro radiobiologic survival values (anti n, D/sub 0/) of four tumor lines derived from human hematopoietic tumors were studied. These cell lines were HL60 promyelocytic leukemia; K562 erythroleukemia; 45 acute lymphocytic leukemia; and 176 acute monomyelogenous leukemia. More cell lines must be examined before the exact relationship between in vitro radiosensitivity and clinical radiocurability is firmly established.

  5. Intrinsic Constraint on Plasmablast Growth and Extrinsic Limits of Plasma Cell Survival

    OpenAIRE

    2000-01-01

    B cells recruited into splenic antibody responses grow exponentially, either in extrafollicular foci as plasmablasts, or in follicles where they form germinal centers. Both responses yield plasma cells. Although many splenic plasma cells survive

  6. Intramacrophage survival of uropathogenic Escherichia coli: Differences between diverse clinical isolates and between mouse and human macrophages

    DEFF Research Database (Denmark)

    Bokil, Nilesh J.; Totsika, Makrina; Carey, Alison J.;

    2011-01-01

    Uropathogenic E. coli (UPEC) are the primary cause of urinary tract infections. Recent studies have demonstrated that UPEC can invade and replicate within epithelial cells, suggesting that this bacterial pathogen may occupy an intracellular niche within the host. Given that many intracellular...... or initial uptake of bacteria. E. coli UTI89 localized to a Lamp1+ vesicular compartment within BMM. In contrast to survival within mouse BMM, intracellular bacterial loads of VR50 were low in both human monocyte-derived macrophages (HMDM) and in human T24 bladder epithelial cells. Collectively, these data......, originally isolated from patients with cystitis and asymptomatic bacteriuria respectively, showed elevated bacterial loads in BMM at 24h post-infection as compared to CFT073 and the asymptomatic bacteriuria strain 83972. These differences did not correlate with differential effects on macrophage survival...

  7. A Conceptual Framework for Representing Human Behavior Characteristics in a System of Systems Agent-Based Survivability Simulation

    Science.gov (United States)

    2010-11-22

    distribution is unlimited. A CONCEPTUAL FRAMEWORK FOR REPRESENTING HUMAN BEHAVIOR CHARACTERISTICS IN A SYSTEM OF SYSTEMS AGENT-BASED SURVIVABILITY...27411 -0001 ABSTRACT A CONCEPTUAL FRAMEWORK FOR REPRESENTING HUMAN BEHAVIOR CHARACTERISTICS IN A SYSTEM OF SYSTEMS AGENT-BASED SURVIVABILITY SIMULATION...TITLE AND SUBTITLE A CONCEPTUAL FRAMEWORK FOR REPRESENTING HUMAN BEHAVIOR CHARACTERISTICS IN A SYSTEM OF SYSTEMS AGENT-BASED SURVIVABILITY

  8. Stereotypical chronic lymphocytic leukemia B-cell receptors recognize survival promoting antigens on stromal cells.

    Directory of Open Access Journals (Sweden)

    Mascha Binder

    Full Text Available Chronic lymphocytic leukemia (CLL is the most common leukemia in the Western world. Survival of CLL cells depends on their close contact with stromal cells in lymphatic tissues, bone marrow and blood. This microenvironmental regulation of CLL cell survival involves the stromal secretion of chemo- and cytokines as well as the expression of adhesion molecules. Since CLL survival may also be driven by antigenic stimulation through the B-cell antigen receptor (BCR, we explored the hypothesis that these processes may be linked to each other. We tested if stromal cells could serve as an antigen reservoir for CLL cells, thus promoting CLL cell survival by stimulation through the BCR. As a proof of principle, we found that two CLL BCRs with a common stereotyped heavy chain complementarity-determining region 3 (previously characterized as "subset 1" recognize antigens highly expressed in stromal cells--vimentin and calreticulin. Both antigens are well-documented targets of autoantibodies in autoimmune disorders. We demonstrated that vimentin is displayed on the surface of viable stromal cells and that it is present and bound by the stereotyped CLL BCR in CLL-stroma co-culture supernatant. Blocking the vimentin antigen by recombinant soluble CLL BCR under CLL-stromal cell co-culture conditions reduces stroma-mediated anti-apoptotic effects by 20-45%. We therefore conclude that CLL BCR stimulation by stroma-derived antigens can contribute to the protective effect that the stroma exerts on CLL cells. This finding sheds a new light on the understanding of the pathobiology of this so far mostly incurable disease.

  9. Cell death versus cell survival instructed by supramolecular cohesion of nanostructures

    Science.gov (United States)

    Newcomb, Christina J.; Sur, Shantanu; Ortony, Julia H.; Lee, One-Sun; Matson, John B.; Boekhoven, Job; Yu, Jeong Min; Schatz, George C.; Stupp, Samuel I.

    2014-02-01

    Many naturally occurring peptides containing cationic and hydrophobic domains have evolved to interact with mammalian cell membranes and have been incorporated into materials for non-viral gene delivery, cancer therapy or treatment of microbial infections. Their electrostatic attraction to the negatively charged cell surface and hydrophobic interactions with the membrane lipids enable intracellular delivery or cell lysis. Although the effects of hydrophobicity and cationic charge of soluble molecules on the cell membrane are well known, the interactions between materials with these molecular features and cells remain poorly understood. Here we report that varying the cohesive forces within nanofibres of supramolecular materials with nearly identical cationic and hydrophobic structure instruct cell death or cell survival. Weak intermolecular bonds promote cell death through disruption of lipid membranes, while materials reinforced by hydrogen bonds support cell viability. These findings provide new strategies to design biomaterials that interact with the cell membrane.

  10. Modulating cancer cell survival by targeting intracellular cholesterol transport.

    Science.gov (United States)

    Kuzu, Omer F; Gowda, Raghavendra; Noory, Mohammad A; Robertson, Gavin P

    2017-08-08

    Demand for cholesterol is high in certain cancers making them potentially sensitive to therapeutic strategies targeting cellular cholesterol homoeostasis. A potential approach involves disruption of intracellular cholesterol transport, which occurs in Niemann-Pick disease as a result of acid sphingomyelinase (ASM) deficiency. Hence, a class of lysosomotropic compounds that were identified as functional ASM inhibitors (FIASMAs) might exhibit chemotherapeutic activity by disrupting cancer cell cholesterol homoeostasis. Here, the chemotherapeutic utility of ASM inhibition was investigated. The effect of FIASMAs on intracellular cholesterol levels, cholesterol homoeostasis, cellular endocytosis and signalling cascades were investigated. The in vivo efficacy of ASM inhibition was demonstrated using melanoma xenografts and a nanoparticle formulation was developed to overcome dose-limiting CNS-associated side effects of certain FIASMAs. Functional ASM inhibitors inhibited intracellular cholesterol transport leading to disruption of autophagic flux, cellular endocytosis and receptor tyrosine kinase signalling. Consequently, major oncogenic signalling cascades on which cancer cells were reliant for survival were inhibited. Two tested ASM inhibitors, perphenazine and fluphenazine that are also clinically used as antipsychotics, were effective in inhibiting xenografted tumour growth. Nanoliposomal encapsulation of the perphenazine enhanced its chemotherapeutic efficacy while decreasing CNS-associated side effects. This study suggests that disruption of intracellular cholesterol transport by targeting ASM could be utilised as a potential chemotherapeutic approach for treating cancer.

  11. Distance in cancer gene expression from stem cells predicts patient survival.

    Science.gov (United States)

    Riester, Markus; Wu, Hua-Jun; Zehir, Ahmet; Gönen, Mithat; Moreira, Andre L; Downey, Robert J; Michor, Franziska

    2017-01-01

    The degree of histologic cellular differentiation of a cancer has been associated with prognosis but is subjectively assessed. We hypothesized that information about tumor differentiation of individual cancers could be derived objectively from cancer gene expression data, and would allow creation of a cancer phylogenetic framework that would correlate with clinical, histologic and molecular characteristics of the cancers, as well as predict prognosis. Here we utilized mRNA expression data from 4,413 patient samples with 7 diverse cancer histologies to explore the utility of ordering samples by their distance in gene expression from that of stem cells. A differentiation baseline was obtained by including expression data of human embryonic stem cells (hESC) and human mesenchymal stem cells (hMSC) for solid tumors, and of hESC and CD34+ cells for liquid tumors. We found that the correlation distance (the degree of similarity) between the gene expression profile of a tumor sample and that of stem cells orients cancers in a clinically coherent fashion. For all histologies analyzed (including carcinomas, sarcomas, and hematologic malignancies), patients with cancers with gene expression patterns most similar to that of stem cells had poorer overall survival. We also found that the genes in all undifferentiated cancers of diverse histologies that were most differentially expressed were associated with up-regulation of specific oncogenes and down-regulation of specific tumor suppressor genes. Thus, a stem cell-oriented phylogeny of cancers allows for the derivation of a novel cancer gene expression signature found in all undifferentiated forms of diverse cancer histologies, that is competitive in predicting overall survival in cancer patients compared to previously published prediction models, and is coherent in that gene expression was associated with up-regulation of specific oncogenes and down-regulation of specific tumor suppressor genes associated with regulation of

  12. The Humanities in English Primary Schools: Struggling to Survive

    Science.gov (United States)

    Barnes, Jonathan; Scoffham, Stephen

    2017-01-01

    This article surveys the state of the humanities in English primary schools drawing on evidence from serving head teachers, current literature and policy documents. The findings suggest that whilst the humanities are highly valued in schools, there are serious challenges which threaten the "broad and balanced" curriculum. It is suggested…

  13. Combined cisplatin and aurora inhibitor treatment increase neuroblastoma cell death but surviving cells overproduce BDNF.

    Science.gov (United States)

    Polacchini, Alessio; Albani, Clara; Baj, Gabriele; Colliva, Andrea; Carpinelli, Patrizia; Tongiorgi, Enrico

    2016-07-15

    Drug-resistance to chemotherapics in aggressive neuroblastoma (NB) is characterized by enhanced cell survival mediated by TrkB and its ligand, brain-derived neurotrophic factor (BDNF); thus reduction in BDNF levels represent a promising strategy to overcome drug-resistance, but how chemotherapics regulate BDNF is unknown. Here, cisplatin treatment in SK-N-BE neuroblastoma upregulated multiple BDNF transcripts, except exons 5 and 8 variants. Cisplatin increased BDNF mRNA and protein, and enhanced translation of a firefly reporter gene flanked by BDNF 5'UTR exons 1, 2c, 4 or 6 and 3'UTR-long. To block BDNF translation we focused on aurora kinases inhibitors which are proposed as new chemotherapeutics. NB cell survival after 24 h treatment was 43% with cisplatin, and 22% by cisplatin+aurora kinase inhibitor PHA-680632, while the aurora kinases inhibitor alone was less effective; however the combined treatment induced a paradoxical increase of BDNF in surviving cells with strong translational activation of exon6-3'UTR-long transcript, while translation of BDNF transcripts 1, 2C and 4 was suppressed. In conclusion, combined cisplatin and aurora kinase inhibitor treatment increases cell death, but induces BDNF overproduction in surviving cells through an aurora kinase-independent mechanism.

  14. Injectable hydrogel promotes early survival of induced pluripotent stem cell-derived oligodendrocytes and attenuates longterm teratoma formation in a spinal cord injury model.

    Science.gov (United States)

    Führmann, T; Tam, R Y; Ballarin, B; Coles, B; Elliott Donaghue, I; van der Kooy, D; Nagy, A; Tator, C H; Morshead, C M; Shoichet, M S

    2016-03-01

    Transplantation of pluripotent stem cells and their differentiated progeny has the potential to preserve or regenerate functional pathways and improve function after central nervous system injury. However, their utility has been hampered by poor survival and the potential to form tumors. Peptide-modified biomaterials influence cell adhesion, survival and differentiation in vitro, but their effectiveness in vivo remains uncertain. We synthesized a peptide-modified, minimally invasive, injectable hydrogel comprised of hyaluronan and methylcellulose to enhance the survival and differentiation of human induced pluripotent stem cell-derived oligodendrocyte progenitor cells. Cells were transplanted subacutely after a moderate clip compression rat spinal cord injury. The hydrogel, modified with the RGD peptide and platelet-derived growth factor (PDGF-A), promoted early survival and integration of grafted cells. However, prolific teratoma formation was evident when cells were transplanted in media at longer survival times, indicating that either this cell line or the way in which it was cultured is unsuitable for human use. Interestingly, teratoma formation was attenuated when cells were transplanted in the hydrogel, where most cells differentiated to a glial phenotype. Thus, this hydrogel promoted cell survival and integration, and attenuated teratoma formation by promoting cell differentiation.

  15. Tumor cell "dead or alive": caspase and survivin regulate cell death, cell cycle and cell survival.

    Science.gov (United States)

    Suzuki, A; Shiraki, K

    2001-04-01

    Cell death and cell cycle progression are two sides of the same coin, and these two different phenomenons are regulated moderately to maintain the cellular homeostasis. Tumor is one of the disease states produced as a result of the disintegrated regulation and is characterized as cells showing an irreversible progression of cell cycle and a resistance to cell death signaling. Several investigations have been performed for the understanding of cell death or cell cycle, and cell death research has remarkably progressed in these 10 years. Caspase is a nomenclature referring to ICE/CED-3 cysteine proteinase family and plays a central role during cell death. Recently, several investigations raised some possible hypotheses that caspase is also involved in cell cycle regulation. In this issue, therefore, we review the molecular basis of cell death and cell cycle regulated by caspase in tumor, especially hepatocellular carcinoma cells.

  16. Hedgehog pathway regulators influence cervical cancer cell proliferation, survival and migration

    Energy Technology Data Exchange (ETDEWEB)

    Samarzija, Ivana [Ecole Polytechnique Federale Lausanne (EPFL), Department of Life Sciences, Swiss Institute for Experimental Cancer Research (ISREC), 1015 Lausanne (Switzerland); Beard, Peter, E-mail: peter.beard@epfl.ch [Ecole Polytechnique Federale Lausanne (EPFL), Department of Life Sciences, Swiss Institute for Experimental Cancer Research (ISREC), 1015 Lausanne (Switzerland)

    2012-08-17

    Highlights: Black-Right-Pointing-Pointer Unknown cellular mutations complement papillomavirus-induced carcinogenesis. Black-Right-Pointing-Pointer Hedgehog pathway components are expressed by cervical cancer cells. Black-Right-Pointing-Pointer Hedgehog pathway activators and inhibitors regulate cervical cancer cell biology. Black-Right-Pointing-Pointer Cell immortalization by papillomavirus and activation of Hedgehog are independent. -- Abstract: Human papillomavirus (HPV) infection is considered to be a primary hit that causes cervical cancer. However, infection with this agent, although needed, is not sufficient for a cancer to develop. Additional cellular changes are required to complement the action of HPV, but the precise nature of these changes is not clear. Here, we studied the function of the Hedgehog (Hh) signaling pathway in cervical cancer. The Hh pathway can have a role in a number of cancers, including those of liver, lung and digestive tract. We found that components of the Hh pathway are expressed in several cervical cancer cell lines, indicating that there could exists an autocrine Hh signaling loop in these cells. Inhibition of Hh signaling reduces proliferation and survival of the cervical cancer cells and induces their apoptosis as seen by the up-regulation of the pro-apoptotic protein cleaved caspase 3. Our results indicate that Hh signaling is not induced directly by HPV-encoded proteins but rather that Hh-activating mutations are selected in cells initially immortalized by HPV. Sonic Hedgehog (Shh) ligand induces proliferation and promotes migration of the cervical cancer cells studied. Together, these results indicate pro-survival and protective roles of an activated Hh signaling pathway in cervical cancer-derived cells, and suggest that inhibition of this pathway may be a therapeutic option in fighting cervical cancer.

  17. Progranulin is expressed within motor neurons and promotes neuronal cell survival

    Directory of Open Access Journals (Sweden)

    Kay Denis G

    2009-10-01

    Full Text Available Abstract Background Progranulin is a secreted high molecular weight growth factor bearing seven and one half copies of the cysteine-rich granulin-epithelin motif. While inappropriate over-expression of the progranulin gene has been associated with many cancers, haploinsufficiency leads to atrophy of the frontotemporal lobes and development of a form of dementia (frontotemporal lobar degeneration with ubiquitin positive inclusions, FTLD-U associated with the formation of ubiquitinated inclusions. Recent reports indicate that progranulin has neurotrophic effects, which, if confirmed would make progranulin the only neuroprotective growth factor that has been associated genetically with a neurological disease in humans. Preliminary studies indicated high progranulin gene expression in spinal cord motor neurons. However, it is uncertain what the role of Progranulin is in normal or diseased motor neuron function. We have investigated progranulin gene expression and subcellular localization in cultured mouse embryonic motor neurons and examined the effect of progranulin over-expression and knockdown in the NSC-34 immortalized motor neuron cell line upon proliferation and survival. Results In situ hybridisation and immunohistochemical techniques revealed that the progranulin gene is highly expressed by motor neurons within the mouse spinal cord and in primary cultures of dissociated mouse embryonic spinal cord-dorsal root ganglia. Confocal microscopy coupled to immunocytochemistry together with the use of a progranulin-green fluorescent protein fusion construct revealed progranulin to be located within compartments of the secretory pathway including the Golgi apparatus. Stable transfection of the human progranulin gene into the NSC-34 motor neuron cell line stimulates the appearance of dendritic structures and provides sufficient trophic stimulus to survive serum deprivation for long periods (up to two months. This is mediated at least in part through

  18. Adipose-derived mesenchymal stem cell administration does not improve corneal graft survival outcome.

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    Sherezade Fuentes-Julián

    Full Text Available The effect of local and systemic injections of mesenchymal stem cells derived from adipose tissue (AD-MSC into rabbit models of corneal allograft rejection with either normal-risk or high-risk vascularized corneal beds was investigated. The models we present in this study are more similar to human corneal transplants than previously reported murine models. Our aim was to prevent transplant rejection and increase the length of graft survival. In the normal-risk transplant model, in contrast to our expectations, the injection of AD-MSC into the graft junction during surgery resulted in the induction of increased signs of inflammation such as corneal edema with increased thickness, and a higher level of infiltration of leukocytes. This process led to a lower survival of the graft compared with the sham-treated corneal transplants. In the high-risk transplant model, in which immune ocular privilege was undermined by the induction of neovascularization prior to graft surgery, we found the use of systemic rabbit AD-MSCs prior to surgery, during surgery, and at various time points after surgery resulted in a shorter survival of the graft compared with the non-treated corneal grafts. Based on our results, local or systemic treatment with AD-MSCs to prevent corneal rejection in rabbit corneal models at normal or high risk of rejection does not increase survival but rather can increase inflammation and neovascularization and break the innate ocular immune privilege. This result can be partially explained by the immunomarkers, lack of immunosuppressive ability and immunophenotypical secretion molecules characterization of AD-MSC used in this study. Parameters including the risk of rejection, the inflammatory/vascularization environment, the cell source, the time of injection, the immunosuppression, the number of cells, and the mode of delivery must be established before translating the possible benefits of the use of MSCs in corneal transplants to clinical

  19. [Human pluripotent stem cell and neural differentiation].

    Science.gov (United States)

    Wataya, Takafumi; Muguruma, Keiko; Sasai, Yoshiki

    2008-10-01

    Recovery of lost brain function is an important issue in medical studies because neurons of the central nervous system (CNS) have poor potential for regeneration. Since few CNS diseases can be treated completely by medicines, regenerative therapy by using stem cells should be studied as a new type of therapeutic intervention. The efficacy of cell replacement therapy in Parkinson's disease has been well investigated. Several studies on fetal tissue transplantation have revealed that quantity and purity of transplanted cells are necessary for recovery of symptoms. SFEB (Serum-free floating culture of embryoid body-like aggregates) method is capable of inducing multi-potential CNS progenitors that can be steered to differentiate into region-specific tissues. On the basis of the existing knowledge of embryology, we have succeeded in the generating of various types of neurons such as telencephalic, cerebeller (Purkinje and granule cells), retinal (photoreceptor cells) and hypothalamic neurons. Application of this culture method to human ES (hES) cells is necessary for clinical purpose: however, poor survival of hES cells in SFEB culture might limit the possibility of using these cells for future medical applications. We found that a selective Rho-associated kinase (ROCK) inhibitor, Y-27632, markedly diminished the dissociation-induced apoptosis of hES cells and enabled the cells to form aggregates in SFEB culture. For both mouse and human ES cells, SFEB culture is a favorable method that can generate large amounts of region-specific neurons. However, stem cell-based therapy continues to face several obstacles. It is important that researchers in the basic sciences and clinical medicine should discuss these problems together to overcome both scientific and ethical issues related to stem cells.

  20. The Effects of GLUT1 on the Survival of Head and Neck Squamous Cell Carcinoma

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    Shengjiao Li

    2013-09-01

    Full Text Available Background/Aims: Cancer cells require increased nutrient uptake to support a high rate of proliferation, and the overexpression of glucose transporters, in particular GLUT1, is a common characteristic of human malignancies. Here, we investigated the relationship between the expression of GLUT1 and cell viability, colony forming ability and apoptosis of head and neck squamous cell carcinoma (HNSCC in vitro and in a xenograft mouse model in vivo. Methods: Lentiviral mediated overexpression and knock-down of GLUT1 was performed in two oral cancer cell lines (CAL27 and SCC25. QRT-PCR and Western blot analysis were used to detect the mRNA and protein expression of GLUT1 and nuclear factor-kappa B (NFκB p65 subunit. Cell viability and apoptosis were assessed by MTT and flow cytometry analyses, respectively. Colony formation assays were performed by staining with 0.5% crystal violet. The role of GLUT1 in HNSCC was examined in vivo through the generation of a CAL27 (or CAL27 with different transfections nude mice xenograft model of HNSCC. Results: GLUT1 overexpression promoted cell viability and colony formation whereas GLUT1 silencing had the opposite effect. GLUT1 knock-down significantly increased the number of Annexin V positive cells in both cell lines and GLUT1 overexpression had the opposite effect, indicating that GLUT1 modulates apoptosis. Xenograft mouse models of GLUT1 knockdown and overexpression showed that GLUT1 expression was associated with poor survival and increased tumor growth. GLUT1 overexpression significantly upregulated the expression of NFκB-p65, and this effect was reversed by inhibition of GLUT1 expression. Conclusions: GLUT1 expression plays an important role in the survival of HNSCC, and its effects may be associated with the activation of the NFκB pathway.

  1. Germ Cell Cancer and Multiple Relapses: Toxicity and Survival

    DEFF Research Database (Denmark)

    Lauritsen, Jakob; Kier, Maria G.G.; Mortensen, Mette S.

    2015-01-01

    Purpose: A small number of patients with germ cell cancer (GCC) receive more than one line of treatment for disseminated disease. The purpose of this study was to evaluate late toxicity and survival in an unselected cohort of patients who experienced relapse after receiving first-line treatment......, compared with patients treated with only orchiectomy, had an increased risk for a second cancer (hazard ratio [HR], 3.2; 95% CI, 1.9 to 5.5), major cardiovascular disease (HR, 1.9; 95% CI, 1.0 to 3.3), pulmonary disease (HR, 2.0; 95% CI, 1.0 to 3.8), GI disease (HR, 7.3; 95% CI, 3.6 to 14.8), renal...... for disseminated disease. Methods: From the Danish Testicular Cancer database, we identified all patients who received more than one line of treatment for disseminated disease. Information about late toxicity and mortality was obtained by means of linkage to national registers. Prognostic factors for relapse...

  2. An essential role for DNA methyltransferase DNMT3B in cancer cell survival.

    Science.gov (United States)

    Beaulieu, Normand; Morin, Steves; Chute, Ian C; Robert, Marie-France; Nguyen, Hannah; MacLeod, A Robert

    2002-08-02

    Abnormal methylation and associated silencing of tumor suppressor genes is a common feature of many types of cancers. The observation of persistent methylation in human cancer cells lacking the maintenance methyltransferase DNMT1 suggests the involvement of other DNA methyltransferases in gene silencing in cancer. To test this hypothesis, we have evaluated methylation and gene expression in cancer cells specifically depleted of DNMT3A or DNMT3B, de novo methyltransferases that are expressed in adult tissues. Here we have shown that depletion of DNMT3B, but not DNMT3A, induced apoptosis of human cancer cells but not normal cells. DNMT3B depletion reactivated methylation-silenced gene expression but did not induce global or juxtacentromeric satellite demethylation as did specific depletion of DNMT1. Furthermore, the effect of DNMT3B depletion was rescued by exogenous expression of either of the splice variants DNMT3B2 or DNMT3B3 but not DNMT1. These results indicate that DNMT3B has significant site selectivity that is distinct from DNMT1, regulates aberrant gene silencing, and is essential for cancer cell survival.

  3. Evidence that high-migration drug-surviving MOLT4 leukemia cells exhibit cancer stem cell-like properties.

    Science.gov (United States)

    Huang, Xiaoxing; Xiong, Meng; Jin, Yujie; Deng, Chaohua; Xu, Hui; An, Changqing; Hao, Ling; Yang, Xiangyong; Deng, Xinzhou; Tu, Zhenbo; Li, Xinran; Xiao, Ruijing; Zhang, Qiuping

    2016-07-01

    Leukemia represents a spectrum of hematological malignancies threatening human health. Resistance to treatments and metastasis of leukemia are the main causes of death in patients. Leukemia stem cells (LSCs) are the initiating cells of leukemia as well as the main source of drug resistance, invasion and metastasis. Consequently, eliminating LSCs is a prerequisite to eradicate leukemia. Preliminary studies in our laboratory have shown that chemokines and their related receptors play an important role in the drug resistance and metastasis of leukemic cells. In this study, we obtained high migration drug-surviving (short term) MOLT4 cells (hMDSCs-MOLT4) with treatment of doxorubicin (DOX) after Transwell assay. Then we detected stem cell-associated molecular markers on hMDSCs-MOLT4 cells and the parental MOLT4 cells by FCM, QPCR, western blotting, H&E staining and immunohisto-chemistry experimental techniques in vitro and in vivo. Moreover, we explored its impact on drug resistance and tumor formation. Then we found that compared with the parental MOLT4 cells, the mRNA expression levels of stem cell-related factors Sox2, Oct4, C-myc, Klf4, Nanog, Bmi-1, CXCR4 are increased in hMDSCs-MOLT4 cells, together with the protein expression levels of Sox2, Oct4, Klf4, Nanog, CXCR4 and CD34. Our results indicated that hMDSCs-MOLT4 cells exhibited strong drug resistance and certain cancer stem cell-like characteristics. It is the first indication that the targeting stemness factors such as Sox2, Oct4, Klf4, Nanog and CXCR4 may represent plausible options for eliminating T-ALL stem-like cells. The present findings shed light on the relationship between drug-tolerant leukemic cells and cancer stem cells.

  4. The modulatory effects of connexin 43 on cell death/survival beyond cell coupling.

    Science.gov (United States)

    Rodríguez-Sinovas, Antonio; Cabestrero, Alberto; López, Diego; Torre, Iratxe; Morente, Miriam; Abellán, Arancha; Miró, Elisabet; Ruiz-Meana, Marisol; García-Dorado, David

    2007-01-01

    Connexins form a diverse and ubiquitous family of integral membrane proteins. Characteristically, connexins are assembled into intercellular channels that aggregate into discrete cell-cell contact areas termed gap junctions (GJ), allowing intercellular chemical communication, and are essential for propagation of electrical impulses in excitable tissues, including, prominently, myocardium, where connexin 43 (Cx43) is the most important isoform. Previous studies have shown that GJ-mediated communication has an important role in the cellular response to stress or ischemia. However, recent evidence suggests that connexins, and in particular Cx43, may have additional effects that may be important in cell death and survival by mechanisms independent of cell to cell communication. Connexin hemichannels, located at the plasma membrane, may be important in paracrine signaling that could influence intracellular calcium and cell survival by releasing intracellular mediators as ATP, NAD(+), or glutamate. In addition, recent studies have shown the presence of connexins in cell structures other than the plasma membrane, including the cell nucleus, where it has been suggested that Cx43 influences cell growth and differentiation. In addition, translocation of Cx43 to mitochondria appears to be important for certain forms of cardioprotection. These findings open a new field of research of previously unsuspected roles of Cx43 intracellular signaling.

  5. The Non-Survival Effects of Glial Cell Line-Derived Neurotrophic Factor on Neural Cells

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    Daniel Cortés

    2017-08-01

    Full Text Available Glial cell line-derived neurotrophic factor (GDNF was first characterized as a survival-promoting molecule for dopaminergic neurons (DANs. Afterwards, other cells were also discovered to respond to GDNF not only as a survival factor but also as a protein supporting other cellular functions, such as proliferation, differentiation, maturation, neurite outgrowth and other phenomena that have been less studied than survival and are now more extendedly described here in this review article. During development, GDNF favors the commitment of neural precursors towards dopaminergic, motor, enteric and adrenal neurons; in addition, it enhances the axonal growth of some of these neurons. GDNF also induces the acquisition of a dopaminergic phenotype by increasing the expression of Tyrosine Hydroxylase (TH, Nurr1 and other proteins that confer this identity and promote further dendritic and electrical maturation. In motor neurons (MNs, GDNF not only promotes proliferation and maturation but also participates in regenerating damaged axons and modulates the neuromuscular junction (NMJ at both presynaptic and postsynaptic levels. Moreover, GDNF modulates the rate of neuroblastoma (NB and glioblastoma cancer cell proliferation. Additionally, the presence or absence of GDNF has been correlated with conditions such as depression, pain, muscular soreness, etc. Although, the precise role of GDNF is unknown, it extends beyond a survival effect. The understanding of the complete range of properties of this trophic molecule will allow us to investigate its broad mechanisms of action to accelerate and/or improve therapies for the aforementioned pathological conditions.

  6. Differential effects in cells exposed to ultra-short, high intensity electric fields: cell survival, DNA damage, and cell cycle analysis.

    Science.gov (United States)

    Stacey, M; Stickley, J; Fox, P; Statler, V; Schoenbach, K; Beebe, S J; Buescher, S

    2003-12-09

    High power, nanosecond pulsed electric field (nsPEF) effects have been focused on bacterial decontamination, but the impact on mammalian cells is now being revealed. During nsPEF applications, electrical pulses of 10, 60 or 300 ns durations were applied to cells using electric field amplitudes as high as 300 kV/cm. Because of the ultra-short pulse durations, the energy transferred to cells is negligible, and only non-thermal effects are observed. We investigated the genotoxicity of nsPEF on adherent and non-adherent cell lines including 10 human lines and one mouse cell line with different origin and growth characteristics. We present data examining the effects of nsPEF exposure on cell survival assessed by clonogenic formation or live cell count; DNA damage determined by the comet assay and chromosome aberrations; and cell cycle parameters by measuring the mitotic indices of exposed cells. Using each of these indicators, we observed differential effects among cell types with non-adherent cells being more sensitive to the genotoxic effects of nsPEF exposures than adherent cells. Non-adherent cultures showed a rapid decrease in cell viability (90%), induction of DNA damage, and a decrease in the number of cells reaching mitosis after one 60 ns pulse with an electric field intensity of 60 kV/cm. These effects were not observed in cells grown as adherent cultures, with the exception of the mouse 3T3 cell line, which showed survival characteristics similar to non-adherent cultures. These data suggest that nsPEF genotoxicity may be cell type specific, and therefore have potential applications in the selective removal of one cell type from another, for example, in diseased states.

  7. Diffusion inside living human cells

    DEFF Research Database (Denmark)

    Leijnse, N.; Jeon, J. -H.; Loft, Steffen

    2012-01-01

    Naturally occurring lipid granules diffuse in the cytoplasm and can be used as tracers to map out the viscoelastic landscape inside living cells. Using optical trapping and single particle tracking we found that lipid granules exhibit anomalous diffusion inside human umbilical vein endothelial...... cells. For these cells the exact diffusional pattern of a particular granule depends on the physiological state of the cell and on the localization of the granule within the cytoplasm. Granules located close to the actin rich periphery of the cell move less than those located towards to the center...... of the cell or within the nucleus. Also, granules in cells which are stressed by intense laser illumination or which have attached to a surface for a long period of time move in a more restricted fashion than those within healthy cells. For granules diffusing in healthy cells, in regions away from the cell...

  8. Suppression of Glioma-Cell Survival by Antisense and Dominant-Negative AKT2 RNA

    Institute of Scientific and Technical Information of China (English)

    Peiyu Pu; Chunsheng Kang; Jie Li; Guangxiu Wang

    2005-01-01

    OBJECTIVE Overexpression of growth factors and their receptors such as PDGF, FGF, VEGF, IGF, EGF, TGFα etc. Play a critical role in the development and progression of malignant gliomas. AKT, one of the most potent downstream signaling effectors of these growth factor receptors is usually overactivated in malignant gliomas. The present study was undertaken to investigate the effects of antisense and dominant negative AKT2 RNA on the survival of glioma cells with overexpression of AKT2.METHODS Antisense and dominant negative AKT2 constructs (AS-AKT2,DN-AKT2) were transfected into human glioblastoma cell line TJ905 with overexpression of AKT2. Using Western blotting, MTT assay, Ki67 labeling index (Ki67 LI), flow cytometry and the TUNEL method, the expression of AKT2 and GFAP, the proliferation rate and apoptosis of glioma cells transfected with AS-AKT2 or DN-AKT2 were compared to those characteristics of parental and glioma cells transfected with an empty vector.RESULTS Cell proliferation was inhibited in glioma cells transfected with ASAKT2 and DN-AKT2 RNA, while GFAP expression and apoptosis were markedly increased in those cells.CONCLUSION AKT is an important mediator in the growth signaling pathway of malignant gliomas and is a potential promising therapeutic target for malignant gliomas.

  9. Telomere dysfunction and cell survival: Roles for distinct TIN2-containing complexes

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Sahn-ho; Davalos, Albert R.; Heo, Seok-Jin; Rodier, Francis; Zou, Ying; Beausejour, Christian; Kaminker, Patrick; Yannone, Steven M.; Campisi, Judith

    2007-10-02

    Telomeres are maintained by three DNA binding proteins (TRF1, TRF2 and POT1), and several associated factors. One factor, TIN2, binds TRF1 and TRF2 directly and POT1 indirectly. Along with two other proteins, TPP1 and hRap1, these form a soluble complex that may be the core telomere maintenance complex. It is not clear whether sub-complexes also exist in vivo. We provide evidence for two TIN2 sub-complexes with distinct functions in human cells. We isolated these two TIN2 sub-complexes from nuclear lysates of unperturbed cells and cells expressing TIN2 mutants TIN2-13, TIN2-15C, which cannot bind TRF2 or TRF1, respectively. In cells with wild-type p53 function, TIN2-15C was more potent than TIN2-13 in causing telomere uncapping and eventual growth arrest. In cells lacking p53 function, TIN2-15C was more potent than TIN2-13 in causing telomere dysfunction and cell death. Our findings suggest that distinct TIN2 complexes exist, and that TIN2-15C-sensitive subcomplexes are particularly important for cell survival in the absence of functional p53.

  10. NF-κB2 mutation targets survival, proliferation and differentiation pathways in the pathogenesis of plasma cell tumors

    Directory of Open Access Journals (Sweden)

    McCarthy Brian A

    2012-05-01

    Full Text Available Abstract Background Abnormal NF-κB2 activation has been implicated in the pathogenesis of multiple myeloma, a cancer of plasma cells. However, a causal role for aberrant NF-κB2 signaling in the development of plasma cell tumors has not been established. Also unclear is the molecular mechanism that drives the tumorigenic process. We investigated these questions by using a transgenic mouse model with lymphocyte-targeted expression of p80HT, a lymphoma-associated NF-κB2 mutant, and human multiple myeloma cell lines. Methods We conducted a detailed histopathological characterization of lymphomas developed in p80HT transgenic mice and microarray gene expression profiling of p80HT B cells with the goal of identifying genes that drive plasma cell tumor development. We further verified the significance of our findings in human multiple myeloma cell lines. Results Approximately 40% of p80HT mice showed elevated levels of monoclonal immunoglobulin (M-protein in the serum and developed plasma cell tumors. Some of these mice displayed key features of human multiple myeloma with accumulation of plasma cells in the bone marrow, osteolytic bone lesions and/or diffuse osteoporosis. Gene expression profiling of B cells from M-protein-positive p80HT mice revealed aberrant expression of genes known to be important in the pathogenesis of multiple myeloma, including cyclin D1, cyclin D2, Blimp1, survivin, IL-10 and IL-15. In vitro assays demonstrated a critical role of Stat3, a key downstream component of IL-10 signaling, in the survival of human multiple myeloma cells. Conclusions These findings provide a mouse model for human multiple myeloma with aberrant NF-κB2 activation and suggest a molecular mechanism for NF-κB2 signaling in the pathogenesis of plasma cell tumors by coordinated regulation of plasma cell generation, proliferation and survival.

  11. Effects of common germ-line genetic variation in cell cycle genes on ovarian cancer survival

    DEFF Research Database (Denmark)

    Song, H.; Hogdall, E.; Ramus, S.J.

    2008-01-01

    PURPOSE: Somatic alterations have been shown to correlate with ovarian cancer prognosis and survival, but less is known about the effects on survival of common inherited genetic variation. Of particular interest are genes involved in cell cycle pathways, which regulate cell division and could pla...

  12. Angiogenesis and mast cell density as predictors of patient survival in squamous cell carcinoma of lung

    Directory of Open Access Journals (Sweden)

    Ehsan Ullah

    2013-01-01

    Full Text Available Background: Measuring the microvascular and mast cell density in squamous cell carcinoma of lung and correlating them with the patient survival may be helpful to guide the use of cancer chemotherapeutic agents which target molecular mechanisms of tumour angiogenesis and mast cells. Materials and Methods: It was an observational study. It included 39 newly diagnosed, adult patients of pulmonary squamous cell carcinoma. Angiogenesis was determined by Chalkley′s method after immunohistochemical staining of micro-vessels with CD34. Mast cells per HPF were counted in Tolouidine blue stained sections. Results: Mean age of the patients was 58.33 ± 9.14 years. Male to female ratio was 9:1. Most (92.3% patients were current smokers. Majority of tumours (71.8% were localised to major bronchi and/or near to hilum and many of them (74.4% were poorly differentiated. Mean micro-vascular density was 11.80 ± 3.66 per HPF which showed strong negative correlation (r = -0.481, p =0.002 between microvascular density (MVD and tumour grade. Mean mast cell density was 1.60 ± 2.04 which showed strong negative correlation (r=-.683, p =0.0001 with grade. Angiogenesis and mast cell density were found to be positively correlated (r=0.439, p =0.005. High MVD, but not the MCD was associated with poor survival. Conclusion: Angiogenesis and mast cell density are positively correlated with each other however; only high MVD is associated with decreased survival. Thus, the anti-angiogenic agents may be useful in squamous cell carcinoma lung, especially the well differentiated tumours.

  13. BAP31 and its caspase cleavage product regulate cell surface expression of tetraspanins and integrin-mediated cell survival.

    Science.gov (United States)

    Stojanovic, Marina; Germain, Marc; Nguyen, Mai; Shore, Gordon C

    2005-08-26

    BAP31, a resident integral protein of the endoplasmic reticulum membrane, regulates the export of other integral membrane proteins to the downstream secretory pathway. Here we show that cell surface expression of the tetraspanins CD9 and CD81 is compromised in mouse cells from which the Bap31 gene has been deleted. CD9 and CD81 facilitate the function of multiprotein complexes at the plasma membrane, including integrins. Of note, BAP31 does not appear to influence the egress of alpha5beta1 or alpha(v)beta3 integrins to the cell surface, but in Bap31-null mouse cells, these integrins are not able to maintain cellular adhesion to the extracellular matrix in the presence of reduced serum. Consequently, Bap31-null cells are sensitive to serum starvation-induced apoptosis. Reconstitution of wild-type BAP31 into these Bap31-null cells restores integrin-mediated cell attachment and cell survival after serum stress, whereas interference with the functions of CD9, alpha5beta1, or alpha(v)beta3 by antagonizing antibodies makes BAP31 cells act similar to Bap31-null cells in these respects. Finally, in human KB epithelial cells protected from apoptosis by BCL-2, the caspase-8 cleavage product, p20 BAP31, inhibits egress of tetraspanin and integrin-mediated cell attachment. Thus, p20 BAP31 can operate upstream of BCL-2 in living cells to influence cell surface properties due to its effects on protein egress from the endoplasmic reticulum.

  14. [A test for sperm cell survival in peritoneal fluid].

    Science.gov (United States)

    Radwan, J; Niwald, W; Bielak, A; Pawlicki, J; Banaszczyk, R; Makuła, D

    1995-06-01

    The role of the peritoneal fluid in the physiology of reproduction, as well as in the transportation and survival of gametes, is little known. The authors have examined interactions between spermatozoa and the peritoneal fluid, collected during laparoscopy in the, so-called, survival test, from 42 infertile couples. The studied survival of spermatozoa in the peritoneal fluid was relatively high--19% after 48 hours--longer than in Menezo B2 fluid. Values of the test have been indicated, especially in cases of endometriosis-caused and idiopathic infertility.

  15. Improvement in autologous human fat transplant survival with SVF plus VEGF-PLA nano-sustained release microspheres.

    Science.gov (United States)

    Li, Liqun; Pan, Shengsheng; Ni, Binting; Lin, Yuanshao

    2014-08-01

    Early neovascularization is important for autologous fat transplant survival. SVF cells are ideal seed cells. Both vascular endothelial growth factor (VEGF) and SVF cells can promote neovascularization. However, the half-life (about 50 min) of VEGF is too short to sustain an adequate local concentration. We have investigated whether VEGF-polylactic acid (PLA) nano-sustained release microspheres plus SVF cells can improve neovascularization and survival of transplanted fat tissues. SVF cells were harvested and constructed VEGF-PLA nano-sustained release microspheres in vitro. Human fat tissues was mixed with SVF cells plus VEGF-PLA, SVF cells alone or Dulbecco's modified Eagle's medium as the control. These three mixtures were injected into random sites in 18 nude mice. Two months later, the transplants were weighed and examined histologically; and capillaries were counted to quantify neovascularization. Hematoxylin-eosin (HE) and anti-VEGF stains were applied to reveal cell infiltration. The mean wet weight of fat in the SVF plus VEGF-PLA, SVF alone, and control transplants were 0.18 ± 0.013 g, 0.16 ± 0.015 g, and 0.071 ± 0.12 g, respectively; the differences between groups were statistically significant. More vessels were present in the SVF plus VEGF-PLA transplants than in the other two types. Transplants mixed with SVF cells also had an acceptable density of capillaries. Histological analysis revealed that both the SVF plus VEGF-PLA and SVF alone transplants, but not the control transplants, were composed of adipose tissue, and had less fat necrosis and less fibrosis than control specimens. SVF plus VEGF-PLA transplants had significantly greater capillary density and VEGF expression than the other two transplant groups. Thus transplanted fat tissue survival and quality can be enhanced by the addition of VEGF-PLA nano-sustained release microspheres plus SVF cells.

  16. Signal transducer and activator of transcription 3 activation is associated with bladder cancer cell growth and survival

    Directory of Open Access Journals (Sweden)

    Hsieh Fu-Chuan

    2008-10-01

    Full Text Available Abstract Background Constitutive activation of signal transducer and activator of transcription 3 (Stat3 signaling pathway plays an important role in several human cancers. Activation of Stat3 is dependent on the phosphorylation at the tyrosine residue 705 by upstream kinases and subsequent nuclear translocation after dimerization. It remains unclear whether oncogenic Stat3 signaling pathway is involved in the oncogenesis of bladder cancer. Results We found that elevated Stat3 phosphorylation in 19 of 100 (19% bladder cancer tissues as well as bladder cancer cell lines, WH, UMUC-3 and 253J. To explore whether Stat3 activation is associated with cell growth and survival of bladder cancer, we targeted the Stat3 signaling pathway in bladder cancer cells using an adenovirus-mediated dominant-negative Stat3 (Y705F and a small molecule compound, STA-21. Both prohibited cell growth and induction of apoptosis in these bladder cancer cell lines but not in normal bladder smooth muscle cell (BdSMC. The survival inhibition might be mediated through apoptotic caspase 3, 8 and 9 pathways. Moreover, down-regulation of anti-apoptotic genes (Bcl-2, Bcl-xL and survivin and a cell cycle regulating gene (cyclin D1 was associated with the cell growth inhibition and apoptosis. Conclusion These results indicated that activation of Stat3 is crucial for bladder cancer cell growth and survival. Therefore, interference of Stat3 signaling pathway emerges as a potential therapeutic approach for bladder cancer.

  17. Survival of pathogenic and lactobacilli species of fermented olives during simulated human digestion

    OpenAIRE

    Francisco Noé eArroyo López; Stephanie eBlanquet-Diot; Sylvain eDenis; Jonathan eThevenot; Sandrine eChalancon; Monique eAlric; Francisco Rodriguez Gomez; Verónica eRomero-Gil; Rufino eJimenez Diaz; Antonio eGarrido Fernández

    2014-01-01

    The present survey uses a dynamic gastric and small intestinal model to assess the survival of one pathogenic (Escherichia coli O157:H7 EDL 933) and three lactobacilli bacteria with probiotic potential (Lactobacillus rhamnosus GG, Lactobacillus pentosus TOMC-LAB2 and Lactobacillus pentosus TOMC-LAB4) during their passage through the human gastrointestinal tract using fermented olives as the food matrix. The data showed that the survival of the E. coli strain in the stomach and duodenum was ve...

  18. Overexpression of human SOD1 improves survival of mice susceptible to endotoxic shock

    Directory of Open Access Journals (Sweden)

    Charchaflieh J

    2012-07-01

    Full Text Available Jean Charchaflieh,1,2 Georges I Labaze,1 Pulsar Li,1 Holly Van Remmen,3 Haekyung Lee,1 Helen Stutz,1 Arlan Richardson,3 Asher Emanuel,1 Ming Zhang1,41Department of Anesthesiology, State University of New York (SUNY Downstate Medical Center, New York, NY, USA; 2Department of Anesthesiology, Yale University School of Medicine, New Haven, CT, USA; 3Barshop Center for Longevity and Aging Studies, University of Texas Health Science Center at San Antonio, San Antonio, TX, USA; 4Department of Cell Biology, State University of New York (SUNY Downstate Medical Center, New York, NY, USABackground: Protective effects of the antioxidant enzyme Cu-Zn superoxide dismutase (SOD1 against endotoxic shock have not been demonstrated in animal models. We used a murine model to investigate whether overexpression of SOD1 protects against endotoxic shock, and whether the genetic background of SOD1 affects its effective protective effects and susceptibility to endotoxic shock.Methods: Transgenic (tg mice overexpressing human SOD1 and control mice were divided into four groups based on their genetic background: (1 tg mice with mixed genetic background (tg-JAX; (2 wild-type (WT littermates of tg-JAX strain (WT-JAX; (3 tg mice with C57BL/6J background (tg-TX; (4 WT littermates of tg-TX strain (WT-TX. Activity of SOD1 in the intestine, heart, and liver of tg and control mice was confirmed using a polyacrylamide activity gel. Endotoxic shock was induced by intraperitoneal injection of lipopolysaccharide. Survival rates over 120 hours (mean, 95% confidence interval were analyzed using Kaplan–Meier survival curves.Results: Human SOD1 enzymatic activities were significantly higher in the intestine, heart, and liver of both tg strains (tg-JAX and tg-TX compared with their WT littermates (WT-JAX and WT-TX, respectively. Interestingly, the endogenous SOD1 activities in tg-JAX mice were decreased compared with their WT littermates (WT-JAX, but such aberrant changes were not

  19. M2 Polarization of Human Macrophages Favors Survival of the Intracellular Pathogen Chlamydia pneumoniae.

    Directory of Open Access Journals (Sweden)

    Tanja Buchacher

    Full Text Available Intracellular pathogens have developed various strategies to escape immunity to enable their survival in host cells, and many bacterial pathogens preferentially reside inside macrophages, using diverse mechanisms to penetrate their defenses and to exploit their high degree of metabolic diversity and plasticity. Here, we characterized the interactions of the intracellular pathogen Chlamydia pneumoniae with polarized human macrophages. Primary human monocytes were pre-differentiated with granulocyte macrophage colony-stimulating factor or macrophage colony-stimulating factor for 7 days to yield M1-like and M2-like macrophages, which were further treated with interferon-γ and lipopolysaccharide or with interleukin-4 for 48 h to obtain fully polarized M1 and M2 macrophages. M1 and M2 cells exhibited distinct morphology with round or spindle-shaped appearance for M1 and M2, respectively, distinct surface marker profiles, as well as different cytokine and chemokine secretion. Macrophage polarization did not influence uptake of C. pneumoniae, since comparable copy numbers of chlamydial DNA were detected in M1 and M2 at 6 h post infection, but an increase in chlamydial DNA over time indicating proliferation was only observed in M2. Accordingly, 72±5% of M2 vs. 48±7% of M1 stained positive for chlamydial lipopolysaccharide, with large perinuclear inclusions in M2 and less clearly bordered inclusions for M1. Viable C. pneumoniae was present in lysates from M2, but not from M1 macrophages. The ability of M1 to restrict chlamydial replication was not observed in M1-like macrophages, since chlamydial load showed an equal increase over time for M1-like and M2-like macrophages. Our findings support the importance of macrophage polarization for the control of intracellular infection, and show that M2 are the preferred survival niche for C. pneumoniae. M1 did not allow for chlamydial proliferation, but failed to completely eliminate chlamydial infection

  20. Cell cycle arrest and cell survival induce reverse trends of cardiolipin remodeling.

    Directory of Open Access Journals (Sweden)

    Yu-Jen Chao

    Full Text Available Cell survival from the arrested state can be a cause of the cancer recurrence. Transition from the arrest state to the growth state is highly regulated by mitochondrial activity, which is related to the lipid compositions of the mitochondrial membrane. Cardiolipin is a critical phospholipid for the mitochondrial integrity and functions. We examined the changes of cardiolipin species by LC-MS in the transition between cell cycle arrest and cell reviving in HT1080 fibrosarcoma cells. We have identified 41 cardiolipin species by MS/MS and semi-quantitated them to analyze the detailed changes of cardiolipin species. The mass spectra of cardiolipin with the same carbon number form an envelope, and the C64, C66, C68, C70 C72 and C74 envelopes in HT1080 cells show a normal distribution in the full scan mass spectrum. The cardiolipin quantity in a cell decreases while entering the cell cycle arrest, but maintains at a similar level through cell survival. While cells awakening from the arrested state and preparing itself for replication, the groups with short acyl chains, such as C64, C66 and C68 show a decrease of cardiolipin percentage, but the groups with long acyl chains, such as C70 and C72 display an increase of cardiolipin percentage. Interestingly, the trends of the cardiolipin species changes during the arresting state are completely opposite to cell growing state. Our results indicate that the cardiolipin species shift from the short chain to long chain cardiolipin during the transition from cell cycle arrest to cell progression.

  1. HSP90 promotes Burkitt lymphoma cell survival by maintaining tonic B-cell receptor signaling.

    Science.gov (United States)

    Walter, Roland; Pan, Kuan-Ting; Doebele, Carmen; Comoglio, Federico; Tomska, Katarzyna; Bohnenberger, Hanibal; Young, Ryan M; Jacobs, Laura; Keller, Ulrich; Bönig, Halvard; Engelke, Michael; Rosenwald, Andreas; Urlaub, Henning; Staudt, Louis M; Serve, Hubert; Zenz, Thorsten; Oellerich, Thomas

    2017-02-02

    Burkitt lymphoma (BL) is an aggressive B-cell neoplasm that is currently treated by intensive chemotherapy in combination with anti-CD20 antibodies. Because of their toxicity, current treatment regimens are often not suitable for elderly patients or for patients in developing countries where BL is endemic. Targeted therapies for BL are therefore needed. In this study, we performed a compound screen in 17 BL cell lines to identify small molecule inhibitors affecting cell survival. We found that inhibitors of heat shock protein 90 (HSP90) induced apoptosis in BL cells in vitro at concentrations that did not affect normal B cells. By global proteomic and phosphoproteomic profiling, we show that, in BL, HSP90 inhibition compromises the activity of the pivotal B-cell antigen receptor (BCR)-proximal effector spleen tyrosine kinase (SYK), which we identified as an HSP90 client protein. Consistently, expression of constitutively active TEL-SYK counteracted the apoptotic effect of HSP90 inhibition. Together, our results demonstrate that HSP90 inhibition impairs BL cell survival by interfering with tonic BCR signaling, thus providing a molecular rationale for the use of HSP90 inhibitors in the treatment of BL.

  2. Human stromal (mesenchymal) stem cells

    DEFF Research Database (Denmark)

    Aldahmash, Abdullah; Zaher, Walid; Al-Nbaheen, May

    2012-01-01

    Human stromal (mesenchymal) stem cells (hMSC) represent a group of non-hematopoietic stem cells present in the bone marrow stroma and the stroma of other organs including subcutaneous adipose tissue, placenta, and muscles. They exhibit the characteristics of somatic stem cells of self-renewal and......Human stromal (mesenchymal) stem cells (hMSC) represent a group of non-hematopoietic stem cells present in the bone marrow stroma and the stroma of other organs including subcutaneous adipose tissue, placenta, and muscles. They exhibit the characteristics of somatic stem cells of self...... of clinical applications, e.g., non-healing bone fractures and defects and also non-skeletal degenerative diseases like heart failure. Currently, the numbers of clinical trials that employ MSC are increasing. However, several biological and biotechnological challenges need to be overcome to benefit from...

  3. Survival motor neuron protein regulates stem cell division, proliferation, and differentiation in Drosophila.

    Directory of Open Access Journals (Sweden)

    Stuart J Grice

    2011-04-01

    Full Text Available Spinal muscular atrophy is a severe neurogenic disease that is caused by mutations in the human survival motor neuron 1 (SMN1 gene. SMN protein is required for the assembly of small nuclear ribonucleoproteins and a dramatic reduction of the protein leads to cell death. It is currently unknown how the reduction of this ubiquitously essential protein can lead to tissue-specific abnormalities. In addition, it is still not known whether the disease is caused by developmental or degenerative defects. Using the Drosophila system, we show that SMN is enriched in postembryonic neuroblasts and forms a concentration gradient in the differentiating progeny. In addition to the developing Drosophila larval CNS, Drosophila larval and adult testes have a striking SMN gradient. When SMN is reduced in postembryonic neuroblasts using MARCM clonal analysis, cell proliferation and clone formation defects occur. These SMN mutant neuroblasts fail to correctly localise Miranda and have reduced levels of snRNAs. When SMN is removed, germline stem cells are lost more frequently. We also show that changes in SMN levels can disrupt the correct timing of cell differentiation. We conclude that highly regulated SMN levels are essential to drive timely cell proliferation and cell differentiation.

  4. PDK1 regulates VDJ recombination, cell-cycle exit and survival during B-cell development.

    Science.gov (United States)

    Venigalla, Ram K C; McGuire, Victoria A; Clarke, Rosemary; Patterson-Kane, Janet C; Najafov, Ayaz; Toth, Rachel; McCarthy, Pierre C; Simeons, Frederick; Stojanovski, Laste; Arthur, J Simon C

    2013-04-03

    Phosphoinositide-dependent kinase-1 (PDK1) controls the activation of a subset of AGC kinases. Using a conditional knockout of PDK1 in haematopoietic cells, we demonstrate that PDK1 is essential for B cell development. B-cell progenitors lacking PDK1 arrested at the transition of pro-B to pre-B cells, due to a cell autonomous defect. Loss of PDK1 decreased the expression of the IgH chain in pro-B cells due to impaired recombination of the IgH distal variable segments, a process coordinated by the transcription factor Pax5. The expression of Pax5 in pre-B cells was decreased in PDK1 knockouts, which correlated with reduced expression of the Pax5 target genes IRF4, IRF8 and Aiolos. As a result, Ccnd3 is upregulated in PDK1 knockout pre-B cells and they have an impaired ability to undergo cell-cycle arrest, a necessary event for Ig light chain rearrangement. Instead, these cells underwent apoptosis that correlated with diminished expression of the pro-survival gene Bcl2A1. Reintroduction of both Pax5 and Bcl2A1 together into PDK1 knockout pro-B cells restored their ability to differentiate in vitro into mature B cells.

  5. Chymase activities and survival in endotoxin-induced human chymase transgenic mice.

    Science.gov (United States)

    Rafiq, Kazi; Fan, Yu-Yan; Sherajee, Shamshad J; Takahashi, Yoshimasa; Matsuura, Junji; Hase, Naoki; Mori, Hirohito; Nakano, Daisuke; Kobara, Hideki; Hitomi, Hirofumi; Masaki, Tsutomu; Urata, Hidenori; Nishiyama, Akira

    2014-01-01

    We examined the effects of overexpressed human chymase on survival and activity in lipopolysaccharide (LPS)-treated mice. Human chymase transgenic (Tg) and wild-type C57BL/6 (WT) mice were treated with LPS (0.03, 0.1 and 0.3 mg/day; intraperitoneal) for 2 weeks. Treatment with 0.03 mg LPS did not affect survival in either WT or Tg mice. WT mice were not affected by 0.1 mg/day of LPS, whereas 25% of Tg mice died. Survival of mice treated with 0.3 mg/day of LPS was 87.5% and 0% in WT and Tg, respectively. LPS-induced increases in chymase activity in the heart and skin were significantly greater in Tg than WT mice. These data suggest a possible contribution of human chymase activation to LPS-induced mortality.

  6. Imbalance between pro-apoptotic and pro-survival factors in human retinal pericytes in diabetic-like conditions.

    Science.gov (United States)

    Beltramo, Elena; Arroba, Ana I; Mazzeo, Aurora; Valverde, Angela M; Porta, Massimo

    2017-01-27

    Loss of pericytes is one the key events in the pathogenesis of diabetic retinopathy. We have previously demonstrated that human retinal pericytes (HRP) are more vulnerable to intermittent than stable high glucose concentrations, with an increase in apoptosis. Our aim was to explore the expression of molecules involved in pro-apoptotic and survival pathways in pericytes cultured in stable/intermittent high glucose and/or hypoxia, to clarify the mechanisms of action of these diabetic-like stressing stimuli. Human retinal pericytes (HRP) were exposed intermittently at 48-hr intervals to high/physiological glucose for 8 days (intHG) and/or hypoxia over the last 48 hr. Control cells were kept in stable physiological and high glucose. Cell proliferation and apoptosis were assessed. The expression of pro-apoptotic and pro-survival molecules was evaluated by Western blotting. Caspase-8 translocation from the cytoplasm into the nucleus was checked by Western blotting of nuclear versus cytoplasmic fractions and immunofluorescence. Hypoxia, alone and combined with intHG, increased HRP apoptosis and decreased proliferation. Pro-apoptotic molecules increased in HRP cultured in these conditions, while some survival markers decreased. Conversely, in stable HG, pro-apoptotic molecules were stable or even decreased, and survival factors increased. Translocation of caspase-8 from cytoplasm into nucleus indicates a primary role for this molecule in inducing apoptosis. Diabetic-like conditions are able to stimulate pericyte apoptosis through activation of pro-apoptotic molecules, leading to an imbalance between pro-apoptotic and survival signalling pathways, with caspase-8 playing a pivotal role. Our identification of such intermediates could help finding new therapeutic approaches for the prevention of diabetic retinopathy. © 2017 Acta Ophthalmologica Scandinavica Foundation. Published by John Wiley & Sons Ltd.

  7. Intramacrophage survival of uropathogenic Escherichia coli: Differences between diverse clinical isolates and between mouse and human macrophages

    KAUST Repository

    Bokil, Nilesh J.

    2011-11-01

    Uropathogenic E. coli (UPEC) are the primary cause of urinary tract infections. Recent studies have demonstrated that UPEC can invade and replicate within epithelial cells, suggesting that this bacterial pathogen may occupy an intracellular niche within the host. Given that many intracellular pathogens target macrophages, we assessed the interactions between UPEC and macrophages. Colonization of the mouse bladder by UPEC strain CFT073 resulted in increased expression of myeloid-restricted genes, consistent with the recruitment of inflammatory macrophages to the site of infection. In in vitro assays, CFT073 was able to survive within primary mouse bone marrow-derived macrophages (BMM) up to 24h post-infection. Three additional well-characterized clinical UPEC isolates associated with distinct UTI symptomatologies displayed variable long-term survival within BMM. UPEC strains UTI89 and VR50, originally isolated from patients with cystitis and asymptomatic bacteriuria respectively, showed elevated bacterial loads in BMM at 24h post-infection as compared to CFT073 and the asymptomatic bacteriuria strain 83972. These differences did not correlate with differential effects on macrophage survival or initial uptake of bacteria. E. coli UTI89 localized to a Lamp1 + vesicular compartment within BMM. In contrast to survival within mouse BMM, intracellular bacterial loads of VR50 were low in both human monocyte-derived macrophages (HMDM) and in human T24 bladder epithelial cells. Collectively, these data suggest that some UPEC isolates may subvert macrophage anti-microbial pathways, and that host species differences may impact on intracellular UPEC survival. © 2011 Elsevier GmbH.

  8. Intramacrophage survival of uropathogenic Escherichia coli: differences between diverse clinical isolates and between mouse and human macrophages.

    Science.gov (United States)

    Bokil, Nilesh J; Totsika, Makrina; Carey, Alison J; Stacey, Katryn J; Hancock, Viktoria; Saunders, Bernadette M; Ravasi, Timothy; Ulett, Glen C; Schembri, Mark A; Sweet, Matthew J

    2011-11-01

    Uropathogenic E. coli (UPEC) are the primary cause of urinary tract infections. Recent studies have demonstrated that UPEC can invade and replicate within epithelial cells, suggesting that this bacterial pathogen may occupy an intracellular niche within the host. Given that many intracellular pathogens target macrophages, we assessed the interactions between UPEC and macrophages. Colonization of the mouse bladder by UPEC strain CFT073 resulted in increased expression of myeloid-restricted genes, consistent with the recruitment of inflammatory macrophages to the site of infection. In in vitro assays, CFT073 was able to survive within primary mouse bone marrow-derived macrophages (BMM) up to 24h post-infection. Three additional well-characterized clinical UPEC isolates associated with distinct UTI symptomatologies displayed variable long-term survival within BMM. UPEC strains UTI89 and VR50, originally isolated from patients with cystitis and asymptomatic bacteriuria respectively, showed elevated bacterial loads in BMM at 24h post-infection as compared to CFT073 and the asymptomatic bacteriuria strain 83972. These differences did not correlate with differential effects on macrophage survival or initial uptake of bacteria. E. coli UTI89 localized to a Lamp1(+) vesicular compartment within BMM. In contrast to survival within mouse BMM, intracellular bacterial loads of VR50 were low in both human monocyte-derived macrophages (HMDM) and in human T24 bladder epithelial cells. Collectively, these data suggest that some UPEC isolates may subvert macrophage anti-microbial pathways, and that host species differences may impact on intracellular UPEC survival.

  9. Opa+ Neisseria gonorrhoeae exhibits reduced survival in human neutrophils via Src family kinase-mediated bacterial trafficking into mature phagolysosomes.

    Science.gov (United States)

    Johnson, M Brittany; Ball, Louise M; Daily, Kylene P; Martin, Jennifer N; Columbus, Linda; Criss, Alison K

    2015-05-01

    During gonorrhoeal infection, there is a heterogeneous population of Neisseria gonorrhoeae (Gc) varied in their expression of opacity-associated (Opa) proteins. While Opa proteins are important for bacterial attachment and invasion of epithelial cells, Opa+ Gc has a survival defect after exposure to neutrophils. Here, we use constitutively Opa- and OpaD+ Gc in strain background FA1090 to show that Opa+ Gc is more sensitive to killing inside adherent, chemokine-treated primary human neutrophils due to increased bacterial residence in mature, degradative phagolysosomes that contain primary and secondary granule antimicrobial contents. Although Opa+ Gc stimulates a potent oxidative burst, neutrophil killing of Opa+ Gc was instead attributable to non-oxidative components, particularly neutrophil proteases and the bactericidal/permeability-increasing protein. Blocking interaction of Opa+ Gc with carcinoembryonic antigen-related cell adhesion molecules (CEACAMs) or inhibiting Src family kinase signalling, which is downstream of CEACAM activation, enhanced the survival of Opa+ Gc in neutrophils. Src family kinase signalling was required for fusion of Gc phagosomes with primary granules to generate mature phagolysosomes. Conversely, ectopic activation of Src family kinases or coinfection with Opa+ Gc resulted in decreased survival of Opa- Gc in neutrophils. From these results, we conclude that Opa protein expression is an important modulator of Gc survival characteristics in neutrophils by influencing phagosome dynamics and thus bacterial exposure to neutrophils' full antimicrobial arsenal.

  10. Effect of Bcl-2 and Bax on survival of side population cells from hepatocellular carcinoma cells

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    AIM: To understand the role and significance of side population (SP) cells from hepatocellular carcinoma (HCC) in hepatocarcinogenesis, development, relapse and metastasis, we simulated the denutrition conditions that cancer cells experience in clinical therapy, observed the different anti-apoptosis ability of SP cells and non-SP cells under such conditions, and established the possible effects of P53, Bcl-2 and Bax on survival of SP cells.METHODS: We used flow cytometry to analyze and sort the SP and non-SP cells in established HCC lines MHCC97and hHCC. We evaluated cell proliferation by methyl thiazolyl tetrazolium (MTT) assay and investigated the expression of p53, bd-2 and bax genes during denutrition,by RT-PCR and immunofluorescence staining.RESULTS: The percentage of SP cells in the two established HCC lines was 0.25% and 0.5%, respectively.SP cells had greater anti-apoptosis and proliferation ability than non-SP cells. Expression of Bcl-2 and Bax in SP and non-SP cells differed during denutrition. The former was up-regulated in SP cells, and the latter was up-regulated in non-SP cells.CONCLUSION: It may be that different upstream molecules acted and led to different expression levels of Bcl-2 and Bax in these two cell lines. There was a direct relationship between up-regulation of Bcl-2 and down-regulation of Bax and higher anti-apoptosis ability in SP cells. It may be that the existence and activity of SP cells are partly responsible for some of the clinical phenomena which are seen in HCC, such as relapse or metastasis. Further research on SP cells may have potential applications in the field of anticancer therapy.

  11. Thirst for Power: Energy, Water and Human Survival

    Science.gov (United States)

    Webber, M.

    2015-12-01

    Energy and water are precious resources, and they are interconnected. The energy sector uses a lot of water -- the thermoelectric power sector alone is the largest user of water in the U.S., withdrawing 200 billion gallons daily for powerplant cooling. Conversely, the water sector is responsible for over twelve percent of national energy consumption for moving, pumping, treating, and heating water. This interdependence means that droughts can cause energy shortages, and power outages can bring the water system to a halt. It also means that water efficiency is a pathway to energy efficiency and vice versa. This talk will give a big-picture overview of global energy and water trends to describe how they interact, what conflicts are looming, and how they can work together. This talk will include the vulnerabilities and cross-cutting solutions such as efficient markets and smart technologies that embed more information about resource management. It will include discussion of how population growth, economic growth, climate change, and short-sighted policies are likely to make things worse. Yet, more integrated planning with long-term sustainability in mind along with cultural shifts, advanced technologies, and better design can avert such a daunting future. Combining anecdotes and personal stories with insights into the latest science of energy and water, this talk will identify a hopeful path toward wise, long-range water-energy decisions and a more reliable and abundant future for humanity.

  12. Monte Carlo based protocol for cell survival and tumour control probability in BNCT

    Science.gov (United States)

    Ye, Sung-Joon

    1999-02-01

    A mathematical model to calculate the theoretical cell survival probability (nominally, the cell survival fraction) is developed to evaluate preclinical treatment conditions for boron neutron capture therapy (BNCT). A treatment condition is characterized by the neutron beam spectra, single or bilateral exposure, and the choice of boron carrier drug (boronophenylalanine (BPA) or boron sulfhydryl hydride (BSH)). The cell survival probability defined from Poisson statistics is expressed with the cell-killing yield, the (n, ) reaction density, and the tolerable neutron fluence. The radiation transport calculation from the neutron source to tumours is carried out using Monte Carlo methods: (i) reactor-based BNCT facility modelling to yield the neutron beam library at an irradiation port; (ii) dosimetry to limit the neutron fluence below a tolerance dose (10.5 Gy-Eq); (iii) calculation of the (n, ) reaction density in tumours. A shallow surface tumour could be effectively treated by single exposure producing an average cell survival probability of - for probable ranges of the cell-killing yield for the two drugs, while a deep tumour will require bilateral exposure to achieve comparable cell kills at depth. With very pure epithermal beams eliminating thermal, low epithermal and fast neutrons, the cell survival can be decreased by factors of 2-10 compared with the unmodified neutron spectrum. A dominant effect of cell-killing yield on tumour cell survival demonstrates the importance of choice of boron carrier drug. However, these calculations do not indicate an unambiguous preference for one drug, due to the large overlap of tumour cell survival in the probable ranges of the cell-killing yield for the two drugs. The cell survival value averaged over a bulky tumour volume is used to predict the overall BNCT therapeutic efficacy, using a simple model of tumour control probability (TCP).

  13. Always follow your nose: the functional significance of social chemosignals in human reproduction and survival.

    Science.gov (United States)

    Lübke, Katrin T; Pause, Bettina M

    2015-02-01

    This article is part of a Special Issue "Chemosignals and Reproduction" Across phyla, chemosensory communication is crucial for mediating a variety of social behaviors, which form the basis for ontogenetic and phylogenetic survival. In the present paper, evidence on chemosensory communication in humans, with special reference to reproduction and survival, will be presented. First, the impact of chemosignals on human reproduction will be reviewed. Work will be presented, showing how chemosensory signals are involved in mate choice and partnership formation by communicating attractiveness and facilitating a partner selection, which is of evolutionary advantage, and furthermore providing information about the level of sexual hormones. In addition to direct effects on phylogenetic survival, chemosignals indirectly aid reproductive success by fostering harm protection. Results will be presented, showing that chemosensory communication aids the emotional bond between mother and child, which in turn motivates parental caretaking and protection, leading to infant survival. Moreover, the likelihood of group survival can be increased through the use of stress-related chemosignals. Stress-related chemosignals induce a stress-related physiology in the perceiver, thereby priming a fight-flight-response, which is necessary for an optimum adaption to environmental harm. Finally, effects of sexual orientation on chemosensory communication will be discussed in terms of their putative role in stabilizing social groups, which might indirectly provide harm protection and foster survival. An integrative model of the presented data will be introduced. In conclusion, an outlook, focusing on the involvement of chemosensory communication in human social behavior and illustrating a novel approach to the significance of chemosensory signals in human survival, will be given.

  14. Thirst for Power: Energy, Water and Human Survival

    Science.gov (United States)

    Webber, M.

    2016-12-01

    Energy, food and water are precious resources, and they are interconnected. The energy sector uses a lot of water, the food sector uses a lot of energy and water, the water sector uses a lot of energy, and as a nation we are contemplating a biofuels policy that uses food for energy. The thermoelectric power sector alone is the largest user of water in the U.S., withdrawing 200 billion gallons daily for powerplant cooling. Conversely, the water sector is responsible for over twelve percent of national energy consumption for moving, pumping, treating, and heating water. The food system uses over ten percent of national energy consumption. This interdependence means that droughts can cause energy shortages, and power outages can bring the water system to a halt, while energy and water challenges pose constraints to our food system. It also means that water efficiency is a pathway to energy efficiency and vice versa. This talk will give a big-picture overview of global food, energy and water trends to describe how they interact, what conflicts are looming, and how they can work together. This talk will include the vulnerabilities and cross-cutting solutions such as efficient markets and smart technologies that embed more information about resource management. It will include discussion of how population growth, economic growth, climate change, and short-sighted policies are likely to make things worse. Yet, more integrated planning with long-term sustainability in mind along with cultural shifts, advanced technologies, and better design can avert such a daunting future. Combining anecdotes and personal stories with insights into the latest science of energy and water, this talk will identify a hopeful path toward wise, long-range water-energy decisions and a more reliable and abundant future for humanity.

  15. γ-secretase directly sheds the survival receptor BCMA from plasma cells

    Science.gov (United States)

    Laurent, Sarah A.; Hoffmann, Franziska S.; Kuhn, Peer-Hendrik; Cheng, Qingyu; Chu, Yuanyuan; Schmidt-Supprian, Marc; Hauck, Stefanie M.; Schuh, Elisabeth; Krumbholz, Markus; Rübsamen, Heike; Wanngren, Johanna; Khademi, Mohsen; Olsson, Tomas; Alexander, Tobias; Hiepe, Falk; Pfister, Hans-Walter; Weber, Frank; Jenne, Dieter; Wekerle, Hartmut; Hohlfeld, Reinhard; Lichtenthaler, Stefan F.; Meinl, Edgar

    2015-01-01

    Survival of plasma cells is regulated by B-cell maturation antigen (BCMA), a membrane-bound receptor activated by its agonist ligands BAFF and APRIL. Here we report that γ-secretase directly cleaves BCMA, without prior truncation by another protease. This direct shedding is facilitated by the short length of BCMA's extracellular domain. In vitro, γ-secretase reduces BCMA-mediated NF-κB activation. In addition, γ-secretase releases soluble BCMA (sBCMA) that acts as a decoy neutralizing APRIL. In vivo, inhibition of γ-secretase enhances BCMA surface expression in plasma cells and increases their number in the bone marrow. Furthermore, in multiple sclerosis, sBCMA levels in spinal fluid are elevated and associated with intracerebral IgG production; in systemic lupus erythematosus, sBCMA levels in serum are elevated and correlate with disease activity. Together, shedding of BCMA by γ-secretase controls plasma cells in the bone marrow and yields a potential biomarker for B-cell involvement in human autoimmune diseases. PMID:26065893

  16. Low concentrations of nitric oxide delay the differentiation of embryonic stem cells and promote their survival.

    Science.gov (United States)

    Tejedo, J R; Tapia-Limonchi, R; Mora-Castilla, S; Cahuana, G M; Hmadcha, A; Martin, F; Bedoya, F J; Soria, B

    2010-10-07

    Nitric oxide (NO) is an intracellular messenger in several cell systems, but its contribution to embryonic stem cell (ESC) biology has not been characterized. Exposure of ESCs to low concentrations (2-20 μM) of the NO donor diethylenetriamine NO adduct confers protection from apoptosis elicited by leukaemia inhibitory factor (LIF) withdrawal. NO blocked caspase 3 activation, PARP degradation, downregulation of the pro-apoptotic genes Casp7, Casp9, Bax and Bak1 and upregulation of the anti-apoptotic genes Bcl-2 111, Bcl-2 and Birc6. These effects were also observed in cells overexpressing eNOS. Exposure of LIF-deprived mESCs to low NO prevented the loss of expression of self-renewal genes (Oct4, Nanog and Sox2) and the SSEA marker. Moreover, NO blocked the differentiation process promoted by the absence of LIF and bFGF in mouse and human ESCs. NO treatment decreased the expression of differentiation markers, such as Brachyury, Gata6 and Gata4. Constitutive overexpression of eNOS in cells exposed to LIF deprivation maintained the expression of self-renewal markers, whereas the differentiation genes were repressed. These effects were reversed by addition of the NOS inhibitor L-NMMA. Altogether, the data suggest that low NO has a role in the regulation of ESC differentiation by delaying the entry into differentiation, arresting the loss of self-renewal markers and promoting cell survival by inhibiting apoptosis.

  17. Low concentrations of nitric oxide delay the differentiation of embryonic stem cells and promote their survival

    Science.gov (United States)

    Tejedo, J R; Tapia-Limonchi, R; Mora-Castilla, S; Cahuana, G M; Hmadcha, A; Martin, F; Bedoya, F J; Soria, B

    2010-01-01

    Nitric oxide (NO) is an intracellular messenger in several cell systems, but its contribution to embryonic stem cell (ESC) biology has not been characterized. Exposure of ESCs to low concentrations (2–20 μM) of the NO donor diethylenetriamine NO adduct confers protection from apoptosis elicited by leukaemia inhibitory factor (LIF) withdrawal. NO blocked caspase 3 activation, PARP degradation, downregulation of the pro-apoptotic genes Casp7, Casp9, Bax and Bak1 and upregulation of the anti-apoptotic genes Bcl-2 111, Bcl-2 and Birc6. These effects were also observed in cells overexpressing eNOS. Exposure of LIF-deprived mESCs to low NO prevented the loss of expression of self-renewal genes (Oct4, Nanog and Sox2) and the SSEA marker. Moreover, NO blocked the differentiation process promoted by the absence of LIF and bFGF in mouse and human ESCs. NO treatment decreased the expression of differentiation markers, such as Brachyury, Gata6 and Gata4. Constitutive overexpression of eNOS in cells exposed to LIF deprivation maintained the expression of self-renewal markers, whereas the differentiation genes were repressed. These effects were reversed by addition of the NOS inhibitor L-NMMA. Altogether, the data suggest that low NO has a role in the regulation of ESC differentiation by delaying the entry into differentiation, arresting the loss of self-renewal markers and promoting cell survival by inhibiting apoptosis. PMID:21368853

  18. Survival of cord blood haematopoietic stem cells in a hyaluronan hydrogel for ex vivo biomimicry.

    Science.gov (United States)

    Demange, Elise; Kassim, Yusra; Petit, Cyrille; Buquet, Catherine; Dulong, Virginie; Cerf, Didier Le; Buchonnet, Gérard; Vannier, Jean-Pierre

    2013-11-01

    Haematopoietic stem cells (HSCs) and haematopoietic progenitor cells (HPCs) grow in a specified niche in close association with the microenvironment, the so-called 'haematopoietic niche'. Scaffolds have been introduced to overcome the liquid culture limitations, mimicking the presence of the extracellular matrix (ECM). In the present study the hyaluronic acid scaffold, already developed in the laboratory, has been used for the first time to maintain long-term cultures of CD34⁺ haematopoietic cells obtained from human cord blood. One parameter investigated was the impact on ex vivo survival of CD34⁺ cord blood cells (CBCs) on the hyaluronic acid surface, immobilized with peptides containing the RGD motif. This peptide was conjugated by coating the hyaluronan hydrogel and cultured in serum-free liquid phase complemented with stem cell factor (SCF), a commonly indispensable cytokine for haematopoiesis. Our work demonstrated that these hyaluronan hydrogels were superior to traditional liquid cultures by maintaining and expanding the HPCs without the need for additional cytokines, and a colonization of 280-fold increment in the hydrogel compared with liquid culture after 28 days of ex vivo expansion. Copyright © 2012 John Wiley & Sons, Ltd.

  19. Inflammatory cytokines and survival factors from serum modulate tweak-induced apoptosis in PC-3 prostate cancer cells.

    Directory of Open Access Journals (Sweden)

    Ana Belen Sanz

    Full Text Available Tumor necrosis factor-like weak inducer of apoptosis (TWEAK, TNFSF12 is a member of the tumor necrosis factor superfamily. TWEAK activates the Fn14 receptor, and may regulate cell death, survival and proliferation in tumor cells. However, there is little information on the function and regulation of this system in prostate cancer. Fn14 expression and TWEAK actions were studied in two human prostate cancer cell lines, the androgen-independent PC-3 cell line and androgen-sensitive LNCaP cells. Additionally, the expression of Fn14 was analyzed in human biopsies of prostate cancer. Fn14 expression is increased in histological sections of human prostate adenocarcinoma. Both prostate cancer cell lines express constitutively Fn14, but, the androgen-independent cell line PC-3 showed higher levels of Fn14 that the LNCaP cells. Fn14 expression was up-regulated in PC-3 human prostate cancer cells in presence of inflammatory cytokines (TNFα/IFNγ as well as in presence of bovine fetal serum. TWEAK induced apoptotic cell death in PC-3 cells, but not in LNCaP cells. Moreover, in PC-3 cells, co-stimulation with TNFα/IFNγ/TWEAK induced a higher rate of apoptosis. However, TWEAK or TWEAK/TNFα/IFNγ did not induce apoptosis in presence of bovine fetal serum. TWEAK induced cell death through activation of the Fn14 receptor. Apoptosis was associated with activation of caspase-3, release of mitochondrial cytochrome C and an increased Bax/BclxL ratio. TWEAK/Fn14 pathway activation promotes apoptosis in androgen-independent PC-3 cells under certain culture conditions. Further characterization of the therapeutic target potential of TWEAK/Fn14 for human prostate cancer is warranted.

  20. Cell survival signalling through PPARδ and arachidonic acid metabolites in neuroblastoma.

    Directory of Open Access Journals (Sweden)

    Emma Bell

    Full Text Available Retinoic acid (RA has paradoxical effects on cancer cells: promoting cell death, differentiation and cell cycle arrest, or cell survival and proliferation. Arachidonic acid (AA release occurs in response to RA treatment and, therefore, AA and its downstream metabolites may be involved in cell survival signalling. To test this, we inhibited phospholipase A2-mediated AA release, cyclooxygenases and lipoxygenases with small-molecule inhibitors to determine if this would sensitise cells to cell death after RA treatment. The data suggest that, in response to RA, phospholipase A2-mediated release of AA and subsequent metabolism by lipoxygenases is important for cell survival. Evidence from gene expression reporter assays and PPARδ knockdown suggests that lipoxygenase metabolites activate PPARδ. The involvement of PPARδ in cell survival is supported by results of experiments with the PPARδ inhibitor GSK0660 and siRNA-mediated knockdown. Quantitative reverse transcriptase PCR studies demonstrated that inhibition of 5-lipoxygenase after RA treatment resulted in a strong up-regulation of mRNA for PPARδ2, a putative inhibitory PPARδ isoform. Over-expression of PPARδ2 using a tetracycline-inducible system in neuroblastoma cells reduced proliferation and induced cell death. These data provide evidence linking lipoxygenases and PPARδ in a cell survival-signalling mechanism and suggest new drug-development targets for malignant and hyper-proliferative diseases.

  1. Acrolein activates cell survival and apoptotic death responses involving the endoplasmic reticulum in A549 lung cells.

    Science.gov (United States)

    Tanel, André; Pallepati, Pragathi; Bettaieb, Ahmed; Morin, Patrick; Averill-Bates, Diana A

    2014-05-01

    Acrolein, a highly reactive α,β-unsaturated aldehyde, is a product of endogenous lipid peroxidation. It is a ubiquitous environmental pollutant that is generated mainly by smoke, overheated cooking oil and vehicle exhaust. Acrolein damages cellular proteins, which could lead to accumulation of aberrantly-folded proteins in the endoplasmic reticulum (ER). This study determines the mechanisms involved in acrolein-induced apoptosis mediated by the ER and possible links with the ER stress response in human A549 lung cells. The exposure of cells to acrolein (15-50μM) for shorter times of 15 to 30min activated several ER stress markers. These included the ER chaperone protein BiP and the three ER sensors: (i) the survival/rescue molecules protein kinase RNA (PKR)-like ER kinase (PERK) and eukaryotic initiation factor 2 alpha (eIF2α) were phosphorylated; (ii) cleavage of activating transcription factor 6 (ATF6) occurred, and (iii) inositol-requiring protein-1 alpha (IRE1α) was phosphorylated. Acrolein (25-50μM) caused apoptotic cell death mediated by the ER after 2h, which was characterised by the induction of CHOP and activation of ER proteases calpain and caspase-4. Calpain and caspase-7 were the initiating factors for caspase-4 activation in acrolein-induced apoptosis. These results increase our knowledge about cellular responses to acrolein in lung cells, which have implications for human health.

  2. The P2X7 receptor regulates cell survival, migration and invasion of pancreatic ductal adenocarcinoma cells

    DEFF Research Database (Denmark)

    Giannuzzo, Andrea; Pedersen, Stine Helene Falsig; Novak, Ivana

    2015-01-01

    BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) is presently one of the cancers with the worst survival rates and least effective treatments. Moreover, total deaths due to PDAC are predicted to increase in the next 15 years. Therefore, novel insights into basic mechanism of PDAC development...... and therapies are needed. PDAC is characterized by a complex microenvironment, in which cancer and stromal cells release different molecules, such as ATP. ATP can be transported and/or exocytosed from active cancer cells and released from dying cells in the necrotic core of the cancer. We hypothesized that one...... of the ATP receptors, the P2X7 receptor (P2X7R) could be an important player in PDAC behaviour. METHODS: We determined the expression (real time PCR and Western blot) and localization (immunofluorescence) of P2X7R in human PDAC cell lines (AsPC-1, BxPC-3, Capan-1, MiaPaCa-2, Panc-1) and a "normal" human...

  3. Nonlinearity in MCF7 Cell Survival Following Exposure to Modulated 6 MV Radiation Fields

    Directory of Open Access Journals (Sweden)

    Laetitia Lacoste-Collin MD, PhD

    2015-10-01

    Full Text Available The study of cell survival following exposure to nonuniform radiation fields is taking on particular interest because of the increasing evidence of a nonlinear relationship at low doses. We conducted in vitro experiments using the MCF7 breast cancer cell line. A 2.4 × 2.4 cm2 square area of a T25 flask was irradiated by a Varian Novalis accelerator delivering 6 MV photons. Cell survival inside the irradiation field, in the dose gradient zone and in the peripheral zone, was determined using a clonogenic assay for different radiation doses at the isocenter. Increased cell survival was observed inside the irradiation area for doses of 2, 10, and 20 Gy when nonirradiated cells were present at the periphery, while the cells at the periphery showed decreased survival compared to controls. Increased survival was also observed at the edge of the dose gradient zone for cells receiving 0.02 to 0.01 Gy when compared with cells at the periphery of the same flask, whatever the isocenter dose. These data are the first to report cell survival in the dose gradient zone. Radiotherapists must be aware of this nonlinearity in dose response.

  4. Nonlinearity in MCF7 Cell Survival Following Exposure to Modulated 6 MV Radiation Fields

    Science.gov (United States)

    Castiella, Marion; Franceries, Xavier; Cassol, Emmanuelle; Vieillevigne, Laure; Pereda, Veronica; Bardies, Manuel; Courtade-Saïdi, Monique

    2015-01-01

    The study of cell survival following exposure to nonuniform radiation fields is taking on particular interest because of the increasing evidence of a nonlinear relationship at low doses. We conducted in vitro experiments using the MCF7 breast cancer cell line. A 2.4 × 2.4 cm2 square area of a T25 flask was irradiated by a Varian Novalis accelerator delivering 6 MV photons. Cell survival inside the irradiation field, in the dose gradient zone and in the peripheral zone, was determined using a clonogenic assay for different radiation doses at the isocenter. Increased cell survival was observed inside the irradiation area for doses of 2, 10, and 20 Gy when nonirradiated cells were present at the periphery, while the cells at the periphery showed decreased survival compared to controls. Increased survival was also observed at the edge of the dose gradient zone for cells receiving 0.02 to 0.01 Gy when compared with cells at the periphery of the same flask, whatever the isocenter dose. These data are the first to report cell survival in the dose gradient zone. Radiotherapists must be aware of this nonlinearity in dose response. PMID:26740805

  5. Conditioned medium from activated spleen cells supports the survival of rat retinal cells in vitro

    Directory of Open Access Journals (Sweden)

    A. Sholl-Franco

    1997-11-01

    Full Text Available Cytokines are a heterogeneous group of molecules that have been associated with several functions in the nervous system, such as survival and differentiation of neuronal and glial cells. In the present study, we demonstrated that conditioned medium from spleen cells activated with concanavalin A increased neuritogenesis and survival of retinal cells, as measured by biochemical and morphological criteria. Our data showed that conditioned medium induced a five-fold increase in the amount of protein after 120 h in vitro. This effect was not inhibited by the blockade of voltage-dependent L-type calcium channels with 5.0 µM nifedipine. However, the use of an intracellular calcium chelator (15.0 µM BAPTA-AM inhibited this effect. Our results support the idea that factors secreted by activated lymphocytes, such as cytokines, can modulate the maintenance and the differentiation of rat retinal cells in vitro, indicating a possible role of these molecules in the development of retinal cells, as well as in its protection against pathological conditions

  6. Track-event theory of cell survival with second-order repair.

    Science.gov (United States)

    Besserer, Jürgen; Schneider, Uwe

    2015-05-01

    When fractionation schemes for hypofractionation and stereotactic body radiotherapy are considered, a reliable cell survival model at high dose is needed for calculating doses of similar biological effectiveness. In this work, a simple model for cell survival which is valid also at high dose is developed from Poisson statistics. It is assumed that a cell is killed by an event that is defined by two double-strand breaks on the same or different chromosomes. Two different mechanisms can produce events. A one-track event is always represented by two simultaneous double-strand breaks. A two-track event results in one double-strand break. Therefore, at least two two-track events on the same or different chromosomes are necessary to produce an event. It is assumed that two double-strand breaks can be repaired with a certain repair probability. Both the one-track events and the two-track events are statistically independent. From the stochastic nature of cell killing which is described by the Poisson distribution, the cell survival probability was derived. The model was fitted to experimental data. It was shown that a solution based on Poisson statistics exists for cell survival. It exhibits exponential cell survival at high dose and a finite gradient of cell survival at vanishing dose, which is in agreement with experimental cell studies. The model fits the experimental data as well as the LQ model and is based on two free parameters. It was shown that cell survival can be described with a simple analytical formula on the basis of Poisson statistics. This solution represents in the limit of large dose the typical exponential behavior and predicts cell survival as well as the LQ model.

  7. Interleukin polymorphisms associated with overall survival, disease-free survival, and recurrence in non-small cell lung cancer patients.

    Science.gov (United States)

    Woods, Nicholas T; Monteiro, Alvaro N; Thompson, Zachary J; Amankwah, Ernest K; Naas, Nina; Haura, Eric B; Beg, Amer A; Schabath, Matthew B

    2015-06-01

    Biomarkers based on germline DNA variations could have translational implications by identifying prognostic factors and sub-classifying patients to tailored, patient-specific treatment. To investigate the association between germline variations in interleukin (IL) genes and lung cancer outcomes, we genotyped 251 single nucleotide polymorphisms (SNPs) from 33 different IL genes in 651 non-small cell lung cancer (NSCLC) patients. Analyses were performed to investigate overall survival, disease-free survival, and recurrence. Our analyses revealed 24 different IL SNPs significantly associated with one or more of the lung cancer outcomes of interest. The GG genotype of IL16:rs7170924 was significantly associated with disease-free survival (HR = 0.65; 95% CI 0.50-0.83) and was the only SNP that produced a false discovery rate (FDR) of modest confidence that the association is unlikely to represent a false-positive result (FDR = 0.142). Classification and regression tree (CART) analyses were used to identify potential higher-order interactions. We restricted the CART analyses to the five SNPs that were significantly associated with multiple endpoints (IL1A:rs1800587, IL1B:rs1143634, IL8:s12506479, IL12A:rs662959, and IL13:rs1881457) and IL16:rs7170924 which had the lowest FDR. CART analyses did not yield a tree structure for overall survival; separate CART tree structures were identified for recurrence, based on three SNPs (IL13:rs1881457, IL1B:rs1143634, and IL12A:rs662959), and for disease-free survival, based on two SNPs (IL12A:rs662959 and IL16:rs7170924), which may suggest that these candidate IL SNPs have a specific impact on lung cancer progression and recurrence. These data suggest that germline variations in IL genes are associated with clinical outcomes in NSCLC patients. © 2015 Wiley Periodicals, Inc.

  8. Ten-year survival of patients with oesophageal squamous cell ...

    African Journals Online (AJOL)

    [1] The outcome for ... survival after surgical resection of oesophageal SCC. ... the upper chest were performed for all tumours of the lower third ... clinicopathological characteristics of patients who were alive more than 10 years after oesophagectomy and .... series, 72 patients (6.9%) underwent neo-adjuvant therapy, and.

  9. Cell Survival and DNA Damage in Normal Prostate Cells Irradiated Out-of-Field.

    LENUS (Irish Health Repository)

    Shields, L

    2014-10-31

    Interest in out-of-field radiation dose has been increasing with the introduction of new techniques, such as volumetric modulated arc therapy (VMAT). These new techniques offer superior conformity of high-dose regions to the target compared to conventional techniques, however more normal tissue is exposed to low-dose radiation with VMAT. There is a potential increase in radiobiological effectiveness associated with lower energy photons delivered during VMAT as normal cells are exposed to a temporal change in incident photon energy spectrum. During VMAT deliveries, normal cells can be exposed to the primary radiation beam, as well as to transmission and scatter radiation. The impact of low-dose radiation, radiation-induced bystander effect and change in energy spectrum on normal cells are not well understood. The current study examined cell survival and DNA damage in normal prostate cells after exposure to out-of-field radiation both with and without the transfer of bystander factors. The effect of a change in energy spectrum out-of-field compared to in-field was also investigated. Prostate cancer (LNCaP) and normal prostate (PNT1A) cells were placed in-field and out-of-field, respectively, with the PNT1A cells being located 1 cm from the field edge when in-field cells were being irradiated with 2 Gy. Clonogenic and γ-H2AX assays were performed postirradiation to examine cell survival and DNA damage. The assays were repeated when bystander factors from the LNCaP cells were transferred to the PNT1A cells and also when the PNT1A cells were irradiated in-field to a different energy spectrum. An average out-of-field dose of 10.8 ± 4.2 cGy produced a significant reduction in colony volume and increase in the number of γ-H2AX foci\\/cell in the PNT1A cells compared to the sham-irradiated control cells. An adaptive response was observed in the PNT1A cells having first received a low out-of-field dose and then the bystander factors. The PNT1A cells showed a significant

  10. Midkine is a NF-κB-inducible gene that supports prostate cancer cell survival

    Directory of Open Access Journals (Sweden)

    You Zongbing

    2008-02-01

    Full Text Available Abstract Background Midkine is a heparin-binding growth factor that is over-expressed in various human cancers and plays important roles in cell transformation, growth, survival, migration, and angiogenesis. However, little is known about the upstream factors and signaling mechanisms that regulate midkine gene expression. Methods Two prostate cancer cell lines LNCaP and PC3 were studied for their expression of midkine. Induction of midkine expression in LNCaP cells by serum, growth factors and cytokines was determined by Western blot analysis and/or real-time quantitative reverse-transcription – polymerase chain reaction (RT-PCR. The cell viability was determined by the trypan blue exclusion assay when the LNCaP cells were treated with tumor necrosis factor alpha (TNFα and/or recombinant midkine. When the LNCaP cells were treated with recombinant midkine, activation of intracellular signalling pathways was determined by Western blot analysis. Prostate tissue microarray slides containing 129 cases (18 normal prostate tissues, 40 early stage cancers, and 71 late stage cancers were assessed for midkine expression by immunohistochemical staining. Results We identified that fetal bovine serum, some growth factors (epidermal growth factor, androgen, insulin-like growth factor-I, and hepatocyte growth factor and cytokines (TNFα and interleukin-1beta induced midkine expression in a human prostate cancer cell line LNCaP cells. TNFα also induced midkine expression in PC3 cells. TNFα was the strongest inducer of midkine expression via nuclear factor-kappa B pathway. Midkine partially inhibited TNFα-induced apoptosis in LNCaP cells. Knockdown of endogenous midkine expression by small interfering RNA enhanced TNFα-induced apoptosis in LNCaP cells. Midkine activated extracellular signal-regulated kinase 1/2 and p38 mitogen-activated protein kinase pathways in LNCaP cells. Furthermore, midkine expression was significantly increased in late stage

  11. Cytokine-Induced Killer Cells Kill Chemo-surviving Melanoma Cancer Stem Cells.

    Science.gov (United States)

    Gammaitoni, Loretta; Giraudo, Lidia; Macagno, Marco; Leuci, Valeria; Mesiano, Giulia; Rotolo, Ramona; Sassi, Francesco; Sanlorenzo, Martina; Zaccagna, Alessandro; Pisacane, Alberto; Senetta, Rebecca; Cangemi, Michela; Cattaneo, Giulia; Martin, Valentina; Coha, Valentina; Gallo, Susanna; Pignochino, Ymera; Sapino, Anna; Grignani, Giovanni; Carnevale-Schianca, Fabrizio; Aglietta, Massimo; Sangiolo, Dario

    2017-05-01

    Purpose: The MHC-unrestricted activity of cytokine-induced killer (CIK) cells against chemo-surviving melanoma cancer stem cells (mCSC) was explored, as CSCs are considered responsible for chemoresistance and relapses.Experimental Design: Putative mCSCs were visualized by engineering patient-derived melanoma cells (MC) with a lentiviral vector encoding eGFP under expression control by stemness gene promoter oct4 Their stemness potential was confirmed in vivo by limiting dilution assays. We explored the sensitivity of eGFP(+) mCSCs to chemotherapy (CHT), BRAF inhibitor (BRAFi) or CIK cells, as single agents or in sequence, in vitro First, we treated MCs in vitro with fotemustine or dabrafenib (BRAF-mutated cases); then, surviving MCs, enriched in mCSCs, were challenged with autologous CIK cells. CIK cell activity against chemoresistant mCSCs was confirmed in vivo in two distinct immunodeficient murine models.Results: We visualized eGFP(+) mCSCs (14% ± 2.1%) in 11 MCs. The tumorigenic precursor rate in vivo was higher within eGFP(+) MCs (1/42) compared with the eGFP(-) counterpart (1/4,870). In vitro mCSCs were relatively resistant to CHT and BRAFi, but killed by CIK cells (n = 11, 8/11 autologous), with specific lysis ranging from 95% [effector:tumor ratio (E:T), 40:1] to 20% (E:T 1:3). In vivo infusion of autologous CIK cells into mice bearing xenografts from three distinct melanomas demonstrated significant tumor responses involving CHT-spared eGFP(+) mCSCs (P = 0.001). Sequential CHT-immunotherapy treatment retained antitumor activity (n = 12, P = 0.001) reducing mCSC rates (P = 0.01).Conclusions: These findings are the first demonstration that immunotherapy with CIK cells is active against autologous mCSCs surviving CHT or BRAFi. An experimental platform for mCSC study and rationale for CIK cells in melanoma clinical study is provided. Clin Cancer Res; 23(9); 2277-88. ©2016 AACR. ©2016 American Association for Cancer Research.

  12. Promotion of Survival and Engraftment of Transplanted Adipose Tissue-Derived Stromal and Vascular Cells by Overexpression of Manganese Superoxide Dismutase

    Directory of Open Access Journals (Sweden)

    Silvia Baldari

    2016-07-01

    Full Text Available Short-term persistence of transplanted cells during early post-implant period limits clinical efficacy of cell therapy. Poor cell survival is mainly due to the harsh hypoxic microenvironment transplanted cells face at the site of implantation and to anoikis, driven by cell adhesion loss. We evaluated the hypothesis that viral-mediated expression of a gene conferring hypoxia resistance to cells before transplant could enhance survival of grafted cells in early stages after implant. We used adipose tissue as cell source because it consistently provides high yields of adipose-tissue-derived stromal and vascular cells (ASCs, suitable for regenerative purposes. Luciferase positive cells were transduced with lentiviral vectors expressing either green fluorescent protein as control or human manganese superoxide dismutase (SOD2. Cells were then exposed in vitro to hypoxic conditions, mimicking cell transplantation into an ischemic site. Cells overexpressing SOD2 displayed survival rates significantly greater compared to mock transduced cells. Similar results were also obtained in vivo after implantation into syngeneic mice and assessment of cell engraftment by in vivo bioluminescent imaging. Taken together, these findings suggest that ex vivo gene transfer of SOD2 into ASCs before implantation confers a cytoprotective effect leading to improved survival and engraftment rates, therefore enhancing cell therapy regenerative potential.

  13. NGF is an essential survival factor for bronchial epithelial cells during respiratory syncytial virus infection.

    Directory of Open Access Journals (Sweden)

    Sreekumar Othumpangat

    Full Text Available Overall expression of neurotrophins in the respiratory tract is upregulated in infants infected by the respiratory syncytial virus (RSV, but it is unclear where (structural vs. inflammatory cells, upper vs. lower airways and why, these changes occur. We analyzed systematically the expression of neurotrophic factors and receptors following RSV infection of human nasal, tracheal, and bronchial epithelial cells, and tested the hypothesis that neurotrophins work as innate survival factors for infected respiratory epithelia.Expression of neurotrophic factors (nerve growth factor, NGF; brain-derived neurotrophic factor, BDNF and receptors (trkA, trkB, p75 was analyzed at the protein level by immunofluorescence and flow cytometry and at the mRNA level by real-time PCR. Targeted siRNA was utilized to blunt NGF expression, and its effect on virus-induced apoptosis/necrosis was evaluated by flow cytometry following annexin V/7-AAD staining.RSV infection was more efficient in cells from more distal (bronchial vs. more proximal origin. In bronchial cells, RSV infection induced transcript and protein overexpression of NGF and its high-affinity receptor trkA, with concomitant downregulation of the low-affinity p75(NTR. In contrast, tracheal cells exhibited an increase in BDNF, trkA and trkB, and nasal cells increased only trkA. RSV-infected bronchial cells transfected with NGF-specific siRNA exhibited decreased trkA and increased p75(NTR expression. Furthermore, the survival of bronchial epithelial cells was dramatically decreased when their endogenous NGF supply was depleted prior to RSV infection.RSV infection of the distal airway epithelium, but not of the more proximal sections, results in overexpression of NGF and its trkA receptor, while the other p75(NTR receptor is markedly downregulated. This pattern of neurotrophin expression confers protection against virus-induced apoptosis, and its inhibition amplifies programmed cell death in the infected

  14. IL-12 is required for anti-OX40-mediated CD4 T cell survival.

    Science.gov (United States)

    Ruby, Carl E; Montler, Ryan; Zheng, Rongxui; Shu, Suyu; Weinberg, Andrew D

    2008-02-15

    Engagement of OX40 greatly improves CD4 T cell function and survival. Previously, we showed that both OX40 engagement and CTLA-4 blockade led to enhanced CD4 T cell expansion, but only OX40 signaling increased survival. To identify pathways associated with OX40-mediated survival, the gene expression of Ag-activated CD4 T cells isolated from mice treated with anti-OX40 and -CTLA-4 was compared. This comparison revealed a potential role for IL-12 through increased expression of the IL-12R-signaling subunit (IL-12Rbeta2) on T cells activated 3 days previously with Ag and anti-OX40. The temporal expression of IL-12Rbeta2 on OX40-stimulated CD4 T cells was tightly regulated and peaked approximately 4-6 days after initial activation/expansion, but before the beginning of T cell contraction. IL-12 signaling, during this window of IL-12Rbeta2 expression, was required for enhanced T cell survival and survival was associated with STAT4-specific signaling. The findings from these observations were exploited in several different mouse tumor models where we found that the combination of anti-OX40 and IL-12 showed synergistic therapeutic efficacy. These results may lead to the elucidation of the molecular pathways involved with CD4 T cell survival that contribute to improved memory, and understanding of these pathways could lead to greater efficacy of immune stimulatory Abs in tumor-bearing individuals.

  15. Regulatory T cell expressed MyD88 is critical for prolongation of allograft survival.

    Science.gov (United States)

    Borges, Christopher M; Reichenbach, Dawn K; Kim, Beom Seok; Misra, Aditya; Blazar, Bruce R; Turka, Laurence A

    2016-08-01

    MyD88 signaling directly promotes T-cell survival and is required for optimal T-cell responses to pathogens. To examine the role of T-cell-intrinsic MyD88 signals in transplantation, we studied mice with targeted T-cell-specific MyD88 deletion. Contrary to expectations, we found that these mice were relatively resistant to prolongation of graft survival with anti-CD154 plus rapamycin in a class II-mismatched system. To specifically examine the role of MyD88 in Tregs, we created a Treg-specific MyD88-deficient mouse. Transplant studies in these animals replicated the findings observed with a global T-cell MyD88 knockout. Surprisingly, given the role of MyD88 in conventional T-cell survival, we found no defect in the survival of MyD88-deficient Tregs in vitro or in the transplant recipients and also observed intact cell homing and expression of Treg effector molecules. MyD88-deficient Tregs also fail to protect allogeneic bone marrow transplant recipients from chronic graft-versus-host disease, confirming the observations of defective regulation seen in a solid organ transplant system. Together, our data define MyD88 as having a divergent requirement for cell survival in non-Tregs and Tregs, and a yet-to-be defined survival-independent requirement for Treg function during the response to alloantigen.

  16. The survival of mankind and human speciation in a complex astrobiological context

    Directory of Open Access Journals (Sweden)

    I. Valentin Petrescu-Mag

    2009-12-01

    Full Text Available The migration of Homo sapiens in the outer space brings one of the most significanenvironmental transformations in the history of a species. The astronomic isolation barrier is certainlystronger than the known geographic isolation. The future human colonies will split into new races,subspecies and finally new species. The survival of human civilization depends in a large extent onspread of these colonies outside our solar system and outside Milky Way.

  17. Human embryonic stem cells handbook

    Directory of Open Access Journals (Sweden)

    Carlo Alberto Redi

    2013-03-01

    Full Text Available After the Nobel prize in physiology or medicine was awarded jointly to Sir John Gurdon and Shinya Yamanaka for the discovery that mature cells can be reprogrammed to become pluripotent it became imperative to write down the review for a book entirely devoted to human embryonic stem cells (hES, those cells that are a urgent need for researchers, those cells that rekindle the ethical debates and finally, last but not least, those cells whose study paved the way to obtain induced pluripotent stem cells by the OSKC’s Yamanaka method (the OSKC acronim refers, for those not familiar with the topic, to the four stemness genes used to transfect somatic fibroblasts: Oct4, Sox2, Klf4 and c-Myc....

  18. PI3K/AKT Signaling Pathway Is Essential for Survival of Induced Pluripotent Stem Cells.

    Directory of Open Access Journals (Sweden)

    Amir M Hossini

    Full Text Available Apoptosis is a highly conserved biochemical mechanism which is tightly controlled in cells. It contributes to maintenance of tissue homeostasis and normally eliminates highly proliferative cells with malignant properties. Induced pluripotent stem cells (iPSCs have recently been described with significant functional and morphological similarities to embryonic stem cells. Human iPSCs are of great hope for regenerative medicine due to their broad potential to differentiate into specialized cell types in culture. They may be useful for exploring disease mechanisms and may provide the basis for future cell-based replacement therapies. However, there is only poor insight into iPSCs cell signaling as the regulation of apoptosis. In this study, we focused our attention on the apoptotic response of Alzheimer fibroblast-derived iPSCs and two other Alzheimer free iPSCs to five biologically relevant kinase inhibitors as well as to the death ligand TRAIL. To our knowledge, we are the first to report that the relatively high basal apoptotic rate of iPSCs is strongly suppressed by the pancaspase inhibitor QVD-Oph, thus underlining the dependency on proapoptotic caspase cascades. Furthermore, wortmannin, an inhibitor of phosphoinositid-3 kinase / Akt signaling (PI3K-AKT, dramatically and rapidly induced apoptosis in iPSCs. In contrast, parental fibroblasts as well as iPSC-derived neuronal cells were not responsive. The resulting condensation and fragmentation of DNA and decrease of the membrane potential are typical features of apoptosis. Comparable effects were observed with an AKT inhibitor (MK-2206. Wortmannin resulted in disappearance of phosphorylated AKT and activation of the main effector caspase-3 in iPSCs. These results clearly demonstrate for the first time that PI3K-AKT represents a highly essential survival signaling pathway in iPSCs. The findings provide improved understanding on the underlying mechanisms of apoptosis regulation in iPSCs.

  19. Human BLyS facilitates engraftment of human PBL derived B cells in immunodeficient mice.

    Directory of Open Access Journals (Sweden)

    Madelyn R Schmidt

    Full Text Available The production of fully immunologically competent humanized mice engrafted with peripheral lymphocyte populations provides a model for in vivo testing of new vaccines, the durability of immunological memory and cancer therapies. This approach is limited, however, by the failure to efficiently engraft human B lymphocytes in immunodeficient mice. We hypothesized that this deficiency was due to the failure of the murine microenvironment to support human B cell survival. We report that while the human B lymphocyte survival factor, B lymphocyte stimulator (BLyS/BAFF enhances the survival of human B cells ex vivo, murine BLyS has no such protective effect. Although human B cells bound both human and murine BLyS, nuclear accumulation of NF-kappaB p52, an indication of the induction of a protective anti-apoptotic response, following stimulation with human BLyS was more robust than that induced with murine BLyS suggesting a fundamental disparity in BLyS receptor signaling. Efficient engraftment of both human B and T lymphocytes in NOD rag1(-/- Prf1(-/- immunodeficient mice treated with recombinant human BLyS is observed after adoptive transfer of human PBL relative to PBS treated controls. Human BLyS treated recipients had on average 40-fold higher levels of serum Ig than controls and mounted a de novo antibody response to the thymus-independent antigens in pneumovax vaccine. The data indicate that production of fully immunologically competent humanized mice from PBL can be markedly facilitated by providing human BLyS.

  20. Progression-free survival, post-progression survival, and tumor response as surrogate markers for overall survival in patients with extensive small cell lung cancer

    Directory of Open Access Journals (Sweden)

    Hisao Imai

    2015-01-01

    Full Text Available Objectives: The effects of first-line chemotherapy on overall survival (OS might be confounded by subsequent therapies in patients with small cell lung cancer (SCLC. We examined whether progression-free survival (PFS, post-progression survival (PPS, and tumor response could be valid surrogate endpoints for OS after first-line chemotherapies for patients with extensive SCLC using individual-level data. Methods: Between September 2002 and November 2012, we analyzed 49 cases of patients with extensive SCLC who were treated with cisplatin and irinotecan as first-line chemotherapy. The relationships of PFS, PPS, and tumor response with OS were analyzed at the individual level. Results: Spearman rank correlation analysis and linear regression analysis showed that PPS was strongly correlated with OS (r = 0.97, p < 0.05, R 2 = 0.94, PFS was moderately correlated with OS (r = 0.58, p < 0.05, R 2 = 0.24, and tumor shrinkage was weakly correlated with OS (r = 0.37, p < 0.05, R 2 = 0.13. The best response to second-line treatment, and the number of regimens employed after progression beyond first-line chemotherapy were both significantly associated with PPS ( p ≤ 0.05. Conclusion: PPS is a potential surrogate for OS in patients with extensive SCLC. Our findings also suggest that subsequent treatment after disease progression following first-line chemotherapy may greatly influence OS.

  1. Dclk1, a tumor stem cell marker, regulates pro-survival signaling and self-renewal of intestinal tumor cells.

    Science.gov (United States)

    Chandrakesan, Parthasarathy; Yao, Jiannan; Qu, Dongfeng; May, Randal; Weygant, Nathaniel; Ge, Yang; Ali, Naushad; Sureban, Sripathi M; Gude, Modhi; Vega, Kenneth; Bannerman-Menson, Eddie; Xia, Lijun; Bronze, Michael; An, Guangyu; Houchen, Courtney W

    2017-02-01

    More than 80% of intestinal neoplasia is associated with the adenomatous polyposis coli (APC) mutation. Doublecortin-like kinase 1 (Dclk1), a kinase protein, is overexpressed in colorectal cancer and specifically marks tumor stem cells (TSCs) that self-renew and increased the tumor progeny in Apc (Min/+) mice. However, the role of Dclk1 expression and its contribution to regulating pro-survival signaling for tumor progression in Apc mutant cancer is poorly understood. We analyzed DCLK1 and pro-survival signaling gene expression datasets of 329 specimens from TCGA Colon Adenocarcinoma Cancer Data. The network of DCLK1 and pro-survival signaling was analyzed utilizing the GeneMANIA database. We examined the expression levels of Dclk1 and other stem cell-associated markers, pro-survival signaling pathways, cell self-renewal in the isolated intestinal epithelial cells of Apc (Min/+) mice with high-grade dysplasia and adenocarcinoma. To determine the functional role of Dclk1 for tumor progression, we knocked down Dclk1 and determined the pro-survival signaling pathways and stemness. We used siRNA technology to gene silence pro-survival signaling in colon cancer cells in vitro. We utilized FACS, IHC, western blot, RT-PCR, and clonogenic (self-renewal) assays. We found a correlation between DCLK1 and pro-survival signaling expression. The expression of Dclk1 and stem cell-associated markers Lgr5, Bmi1, and Musashi1 were significantly higher in the intestinal epithelial cells of Apc (Min/+) mice than in wild-type controls. Intestinal epithelial cells of Apc (Min/+) mice showed increased expression of pro-survival signaling, pluripotency and self-renewal ability. Furthermore, the enteroids formed from the intestinal Dclk1(+) cells of Apc (Min/+) mice display higher pluripotency and pro-survival signaling. Dclk1 knockdown in Apc (Min/+) mice attenuates intestinal adenomas and adenocarcinoma, and decreases pro-survival signaling and self-renewal. Knocking down RELA and NOTCH1

  2. Suofu Qin’s work on studies of cell survival signaling in cancer and epithelial cells

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    Reactive oxygen species (ROS) encompass a variety of diverse chemical species including superoxide anions, hydrogen peroxide, hydroxyl radicals and peroxynitrite, which are mainly produced via mitochondrial oxidative metabolism, enzymatic reactions, and light-initiated lipid peroxidation. Over-production of ROS and/or decrease in the antioxidant capacity cause cells to undergo oxi- dative stress that damages cellular macromolecules such as proteins, lipids, and DNA. Oxidative stress is associated with ageing and the development of agerelated diseases such as cancer and age-related macular degeneration. ROS activate signaling pathways that promote cell survival or lead to cell death, depending on the source and site of ROS production, the specific ROS generated, the concentration and kinetics of ROS generation, and the cell types being challenged. However, how the nature and compartmentalization of ROS contribute to the pathogenesis of individual diseases is poorly understood. Consequently, it is crucial to gain a comprehensive understanding of the molecular bases of cell oxidative stress signaling, which will then provide novel therapeutic opportunities to interfere with disease progression via targeting specific signaling pathways.Currently, Dr. Qin’s work is focused on inflammatory and oxidative stress responses using the retinal pigment epithelial (RPE) cells as a model. The study of RPE cell inflammatory and oxidative stress responses has successfully led to a better understanding of RPE cell biology and identification of potential therapeutic targets.

  3. Multi-OMIC profiling of survival and metabolic signaling networks in cells subjected to photodynamic therapy.

    Science.gov (United States)

    Weijer, Ruud; Clavier, Séverine; Zaal, Esther A; Pijls, Maud M E; van Kooten, Robert T; Vermaas, Klaas; Leen, René; Jongejan, Aldo; Moerland, Perry D; van Kampen, Antoine H C; van Kuilenburg, André B P; Berkers, Celia R; Lemeer, Simone; Heger, Michal

    2017-03-01

    Photodynamic therapy (PDT) is an established palliative treatment for perihilar cholangiocarcinoma that is clinically promising. However, tumors tend to regrow after PDT, which may result from the PDT-induced activation of survival pathways in sublethally afflicted tumor cells. In this study, tumor-comprising cells (i.e., vascular endothelial cells, macrophages, perihilar cholangiocarcinoma cells, and EGFR-overexpressing epidermoid cancer cells) were treated with the photosensitizer zinc phthalocyanine that was encapsulated in cationic liposomes (ZPCLs). The post-PDT survival pathways and metabolism were studied following sublethal (LC50) and supralethal (LC90) PDT. Sublethal PDT induced survival signaling in perihilar cholangiocarcinoma (SK-ChA-1) cells via mainly HIF-1-, NF-кB-, AP-1-, and heat shock factor (HSF)-mediated pathways. In contrast, supralethal PDT damage was associated with a dampened survival response. PDT-subjected SK-ChA-1 cells downregulated proteins associated with EGFR signaling, particularly at LC90. PDT also affected various components of glycolysis and the tricarboxylic acid cycle as well as metabolites involved in redox signaling. In conclusion, sublethal PDT activates multiple pathways in tumor-associated cell types that transcriptionally regulate cell survival, proliferation, energy metabolism, detoxification, inflammation/angiogenesis, and metastasis. Accordingly, tumor cells sublethally afflicted by PDT are a major therapeutic culprit. Our multi-omic analysis further unveiled multiple druggable targets for pharmacological co-intervention.

  4. VH1-44 gene usage defines a subset of canine B-cell lymphomas associated with better patient survival.

    Science.gov (United States)

    Chen, Hsiao-Wei; Small, George W; Motsinger-Reif, Alison; Suter, Steven E; Richards, Kristy L

    2014-02-15

    The use of specific immunoglobulin heavy chain variable region (VH) genes has been associated with increased patient survival in human B-cell lymphomas (hBCL). Given the similarity of human and canine BCL (cBCL) in morphology and clinical treatment, we examined the choice of VH in cBCL and determined whether VH gene selection was a distinct feature associated with survival time in dogs. VH gene selection and mutational status in 52 cBCL, including 29 diffuse large B-cell lymphomas (cDLBCL, the most common subtype of cBCL), were analyzed by comparison with the 80 published canine germline VH gene sequences. We further examined the prognostic impact of the subgroups defined by these features on canine survival. We found that VH1-44 was preferentially expressed in the majority of the 52 cBCLs (60%) as well as in the majority of the cDLBCL subset (59%). VH1-44 gene expression was associated with a statistically better overall survival (p=0.039) in cBCL patients, as well as in the cDLBCL subset of patients (p=0.038). These findings suggest that VH gene selection in cBCL is not random and may therefore have functional implications for cBCL lymphomagenesis, in addition to being a useful prognostic biomarker.

  5. IR-induced autophagy plays a role in survival of HeLa cells

    Energy Technology Data Exchange (ETDEWEB)

    Kang, Mi Young; Jang, Eun Yeong; Ryu, Tae Ho; Chung, Dong Min; Kim, Jin Hong; Kim, Jin Kyu [Advanced Radiation Technology Institute, Korea Atomic Energy Research Institute, Jeongeup (Korea, Republic of)

    2014-04-15

    Cells respond to stress with repair, or are diverted into irreversible cell cycle exit (senescence) or are eliminated through programmed cell death. There are two major morphologically distinctive forms of programmed cell death, apoptosis and autophagic cell death. Apoptosis contribute to cell death, whereas autophagy can play a dual role in mediating either cell survival or death in response to various stress stimuli. Here we analysed cellular responses induced by IR. The understanding of an appropriate cellular stress response is of crucial importance in foreseeing the cell fate. Apoptotic feagures were not detected in HeLa under our experimental irradiation condition. Autophagic cell death in HeLa may play an important role in cell protection and can result in cell survival.

  6. HTLV-1 tax stabilizes MCL-1 via TRAF6-dependent K63-linked polyubiquitination to promote cell survival and transformation.

    Science.gov (United States)

    Choi, Young Bong; Harhaj, Edward William

    2014-10-01

    The human T-cell leukemia virus type 1 (HTLV-1) Tax protein hijacks the host ubiquitin machinery to activate IκB kinases (IKKs) and NF-κB and promote cell survival; however, the key ubiquitinated factors downstream of Tax involved in cell transformation are unknown. Using mass spectrometry, we undertook an unbiased proteome-wide quantitative survey of cellular proteins modified by ubiquitin in the presence of Tax or a Tax mutant impaired in IKK activation. Tax induced the ubiquitination of 22 cellular proteins, including the anti-apoptotic BCL-2 family member MCL-1, in an IKK-dependent manner. Tax was found to promote the nondegradative lysine 63 (K63)-linked polyubiquitination of MCL-1 that was dependent on the E3 ubiquitin ligase TRAF6 and the IKK complex. Tax interacted with and activated TRAF6, and triggered its mitochondrial localization, where it conjugated four carboxyl-terminal lysine residues of MCL-1 with K63-linked polyubiquitin chains, which stabilized and protected MCL-1 from genotoxic stress-induced degradation. TRAF6 and MCL-1 played essential roles in the survival of HTLV-1 transformed cells and the immortalization of primary T cells by HTLV-1. Therefore, K63-linked polyubiquitination represents a novel regulatory mechanism controlling MCL-1 stability that has been usurped by a viral oncogene to precipitate cell survival and transformation.

  7. Stathmin/oncoprotein 18, a microtubule regulatory protein, is required for survival of both normal and cancer cell lines lacking the tumor suppressor, p53.

    Science.gov (United States)

    Carney, Bruce K; Cassimeris, Lynne

    2010-05-01

    Stathmin, a microtubule regulatory protein, is overexpressed in many cancers and required for survival of several cancer lines. In a study of breast cancer cell lines(1) proposed that stathmin is required for survival of cells lacking p53, but this hypothesis was not tested directly. Here we tested their hypothesis by examining cell survival in cells depleted of stathmin, p53 or both proteins. Comparing HCT116 colon cancer cell lines differing in TP53 genotype, stathmin depletion resulted in significant death only in cells lacking p53. As a second experimental system, we compared the effects of stathmin depletion from HeLa cells, which normally lack detectable levels of p53 due to expression of the HPV E6 protein. Stathmin depletion caused a large percentage of HeLa cells to die. Restoring p53, by depletion of HPV E6, rescued HeLa cells from stathmin-depletion induced death. Cleaved PARP was detected in HCT116(p53-/-) cells depleted of stathmin and cell death in stathmin-depleted HeLa cells was blocked by the caspase inhibitor Z-VAD-FMK, consistent with apoptotic death. The stathmin-dependent survival of cells lacking p53 was not confined to cancerous cells because both proteins were required for survival of normal human fibroblasts. In HCT116 and HeLa cells, depletion of both stathmin and p53 leads to a cell cycle delay through G(2). Our results demonstrate that stathmin is required for cell survival in cells lacking p53, suggesting that stathmin depletion could be used therapeutically to induce apoptosis in tumors without functional p53.

  8. Analyzing age-specific genetic effects on human extreme age survival in cohort-based longitudinal studies

    DEFF Research Database (Denmark)

    Tan, Qihua; Jacobsen, Rune; Sørensen, Mette

    2013-01-01

    The analysis of age-specific genetic effects on human survival over extreme ages is confronted with a deceleration pattern in mortality that deviates from traditional survival models and sparse genetic data available. As human late life is a distinct phase of life history, exploring the genetic...... effects on extreme age survival can be of special interest to evolutionary biology and health science. We introduce a non-parametric survival analysis approach that combines population survival information with individual genotype data in assessing the genetic effects in cohort-based longitudinal studies...

  9. Physical skill training increases the number of surviving new cells in the adult hippocampus.

    Directory of Open Access Journals (Sweden)

    Daniel M Curlik

    Full Text Available The dentate gyrus is a major site of plasticity in the adult brain, giving rise to thousands of new neurons every day, through the process of adult neurogenesis. Although the majority of these cells die within two weeks of their birth, they can be rescued from death by various forms of learning. Successful acquisition of select types of associative and spatial memories increases the number of these cells that survive. Here, we investigated the possibility that an entirely different form of learning, physical skill learning, could rescue new hippocampal cells from death. To test this possibility, rats were trained with a physically-demanding and technically-difficult version of a rotarod procedure. Acquisition of the physical skill greatly increased the number of new hippocampal cells that survived. The number of surviving cells positively correlated with performance on the task. Only animals that successfully mastered the task retained the cells that would have otherwise died. Animals that failed to learn, and those that did not learn well did not retain any more cells than those that were untrained. Importantly, acute voluntary exercise in activity wheels did not increase the number of surviving cells. These data suggest that acquisition of a physical skill can increase the number of surviving hippocampal cells. Moreover, learning an easier version of the task did not increase cell survival. These results are consistent with previous reports revealing that learning only rescues new neurons from death when acquisition is sufficiently difficult to achieve. Finally, complete hippocampal lesions did not disrupt acquisition of this physical skill. Therefore, physical skill training that does not depend on the hippocampus can effectively increase the number of surviving cells in the adult hippocampus, the vast majority of which become mature neurons.

  10. Physical skill training increases the number of surviving new cells in the adult hippocampus.

    Science.gov (United States)

    Curlik, Daniel M; Maeng, Lisa Y; Agarwal, Prateek R; Shors, Tracey J

    2013-01-01

    The dentate gyrus is a major site of plasticity in the adult brain, giving rise to thousands of new neurons every day, through the process of adult neurogenesis. Although the majority of these cells die within two weeks of their birth, they can be rescued from death by various forms of learning. Successful acquisition of select types of associative and spatial memories increases the number of these cells that survive. Here, we investigated the possibility that an entirely different form of learning, physical skill learning, could rescue new hippocampal cells from death. To test this possibility, rats were trained with a physically-demanding and technically-difficult version of a rotarod procedure. Acquisition of the physical skill greatly increased the number of new hippocampal cells that survived. The number of surviving cells positively correlated with performance on the task. Only animals that successfully mastered the task retained the cells that would have otherwise died. Animals that failed to learn, and those that did not learn well did not retain any more cells than those that were untrained. Importantly, acute voluntary exercise in activity wheels did not increase the number of surviving cells. These data suggest that acquisition of a physical skill can increase the number of surviving hippocampal cells. Moreover, learning an easier version of the task did not increase cell survival. These results are consistent with previous reports revealing that learning only rescues new neurons from death when acquisition is sufficiently difficult to achieve. Finally, complete hippocampal lesions did not disrupt acquisition of this physical skill. Therefore, physical skill training that does not depend on the hippocampus can effectively increase the number of surviving cells in the adult hippocampus, the vast majority of which become mature neurons.

  11. IL-15 Expression on RA Synovial Fibroblasts Promotes B Cell Survival

    Science.gov (United States)

    Benito-Miguel, Marta; García-Carmona, Yolanda; Balsa, Alejandro; Bautista-Caro, María-Belén; Arroyo-Villa, Irene; Cobo-Ibáñez, Tatiana; Bonilla-Hernán, María Gema; de Ayala, Carlos Pérez; Sánchez-Mateos, Paloma; Martín-Mola, Emilio; Miranda-Carús, María-Eugenia

    2012-01-01

    Introduction The purpose of this study was to examine the role of RA Synovial Fibroblast (RASFib) IL-15 expression on B cell survival. Methods Magnetically sorted peripheral blood memory B cells from 15 healthy subjects were cocultured with RASFib. Results RASFib constitutively expressed membrane IL-15. Survival of isolated B cells cultured for 6 days, below 5%, was extended in coculture with RASFib to 52+/−8% (p<0.001). IL-15 neutralizing agents but not isotype controls, reduced this rate to 31+/−6% (p<0.05). Interestingly, rhIL-15 had no effect on isolated B cells but significantly increased their survival in coculture with RASFib. In parallel, B cell IL-15R chains were upregulated in cocultures. BAFF and VCAM-1, that are expressed on RASFib, were tested as potential candidates involved in upregulating B cell IL-15R. Culture of B cells in the presence of rhBAFF or rhVCAM-1 resulted in significantly increased survival, together with upregulation of all three IL-15R chains; in parallel, rhIL-15 potentiated the anti-apoptotic effect of BAFF and VCAM-1. Both BAFF and VCAM-1 neutralizing agents downmodulated the effect of RASFib on B cell survival and IL-15R expression. In parallel, rhIL-15 had a lower effect on the survival of B cells cocultured with RASFib in the presence of BAFF or VCAM-1 neutralizing agents. Peripheral blood B cells from 15 early RA patients demonstrated an upregulated IL-15R and increased survival in cocultures. Conclusion IL-15 expression on RASFib significantly contributes to the anti-apoptotic effect of RASFib on B cells. IL-15 action is facilitated by BAFF and VCAM-1 expressed on RASFib, through an upregulation of IL-15R chains. PMID:22792388

  12. IL-15 expression on RA synovial fibroblasts promotes B cell survival.

    Directory of Open Access Journals (Sweden)

    Marta Benito-Miguel

    Full Text Available INTRODUCTION: The purpose of this study was to examine the role of RA Synovial Fibroblast (RASFib IL-15 expression on B cell survival. METHODS: Magnetically sorted peripheral blood memory B cells from 15 healthy subjects were cocultured with RASFib. RESULTS: RASFib constitutively expressed membrane IL-15. Survival of isolated B cells cultured for 6 days, below 5%, was extended in coculture with RASFib to 52+/-8% (p<0.001. IL-15 neutralizing agents but not isotype controls, reduced this rate to 31+/-6% (p<0.05. Interestingly, rhIL-15 had no effect on isolated B cells but significantly increased their survival in coculture with RASFib. In parallel, B cell IL-15R chains were upregulated in cocultures. BAFF and VCAM-1, that are expressed on RASFib, were tested as potential candidates involved in upregulating B cell IL-15R. Culture of B cells in the presence of rhBAFF or rhVCAM-1 resulted in significantly increased survival, together with upregulation of all three IL-15R chains; in parallel, rhIL-15 potentiated the anti-apoptotic effect of BAFF and VCAM-1. Both BAFF and VCAM-1 neutralizing agents downmodulated the effect of RASFib on B cell survival and IL-15R expression. In parallel, rhIL-15 had a lower effect on the survival of B cells cocultured with RASFib in the presence of BAFF or VCAM-1 neutralizing agents. Peripheral blood B cells from 15 early RA patients demonstrated an upregulated IL-15R and increased survival in cocultures. CONCLUSION: IL-15 expression on RASFib significantly contributes to the anti-apoptotic effect of RASFib on B cells. IL-15 action is facilitated by BAFF and VCAM-1 expressed on RASFib, through an upregulation of IL-15R chains.

  13. The P2X7 receptor regulates cell survival, migration and invasion of pancreatic ductal adenocarcinoma cells

    DEFF Research Database (Denmark)

    Giannuzzo, Andrea; Pedersen, Stine Helene Falsig; Novak, Ivana

    2015-01-01

    BACKGROUND: Pancreatic ductal adenocarcinoma (PDAC) is presently one of the cancers with the worst survival rates and least effective treatments. Moreover, total deaths due to PDAC are predicted to increase in the next 15 years. Therefore, novel insights into basic mechanism of PDAC development...... of the ATP receptors, the P2X7 receptor (P2X7R) could be an important player in PDAC behaviour. METHODS: We determined the expression (real time PCR and Western blot) and localization (immunofluorescence) of P2X7R in human PDAC cell lines (AsPC-1, BxPC-3, Capan-1, MiaPaCa-2, Panc-1) and a "normal" human...... pancreatic duct epithelial cell line (HPDE). The function of P2X7R in proliferation (BrdU assay), migration (wound assay) and invasion (Boyden chamber with matrigel) was characterized. Furthermore, we studied P2X7R-dependent pore formation (YoPro-1 assay) and cell death (caspase and annexin V / propidium...

  14. Cell survival and chromosomal aberrations in CHO-K1 cells irradiated by carbon ions.

    Science.gov (United States)

    Czub, J; Banaś, D; Błaszczyk, A; Braziewicz, J; Buraczewska, I; Choiński, J; Górak, U; Jaskóła, M; Korman, A; Lankoff, A; Lisowska, H; Łukaszek, A; Szefliński, Z; Wójcik, A

    2009-03-01

    Chinese hamster ovary CHO-K1 cells were exposed to high LET (12)C-beam (LET: 830 keV/microm) in the dose range of 0-6 Gy and to (60)Co irradiation and the RBE value was obtained. Effects of (12)C-beam exposure on cell survival and chromosomal aberrations were calculated. The chromosomal aberration data were fitted with linear equation. The distribution of aberration in cells was examined with a standard u-test and used to evaluate the data according to Poisson probabilities. The variance to the mean ratio sigma(2)/Y and the dispersion index (u) were determined. Overdispersion was significant (p<0.05) when the value of u exceeded 1.96.

  15. The regulation of erythrocyte survival and suicidal cell death

    OpenAIRE

    Föller, Michael

    2008-01-01

    The life span of erythrocytes is tightly regulated. Therefore, a mechanism is required to remove senescent or damaged erythrocytes without rupture of the cell membrane resulting in the release of hemoglobin which may impair kidney function. The mechanism of suicidal erythrocyte death is called eryptosis and shares similarities with apoptosis of nucleated cells such as exposure of phosphatidylserine at the cell surface, increase in cytosolic Ca2+ concentration, blebbing of the membrane, cell s...

  16. Early Natural Killer Cell Reconstitution Predicts Overall Survival in T Cell-Replete Allogeneic Hematopoietic Stem Cell Transplantation

    DEFF Research Database (Denmark)

    Minculescu, Lia; Marquart, Hanne Vibeke; Friis, Lone Smidstrup

    2016-01-01

    Early immune reconstitution plays a critical role in clinical outcome after allogeneic hematopoietic stem cell transplantation (HSCT). Natural killer (NK) cells are the first lymphocytes to recover after transplantation and are considered powerful effector cells in HSCT. We aimed to evaluate...... the clinical impact of early NK cell recovery in T cell-replete transplant recipients. Immune reconstitution was studied in 298 adult patients undergoing HSCT for acute myeloid leukemia, acute lymphoblastic leukemia, and myelodysplastic syndrome from 2005 to 2013. In multivariate analysis NK cell numbers...... on day 30 (NK30) > 150 cells/µL were independently associated with superior overall survival (hazard ratio, .79; 95% confidence interval, .66 to .95; P = .01). Cumulative incidence analyses showed that patients with NK30 > 150 cells/µL had significantly less transplant-related mortality (TRM), P = .01...

  17. Conditional survival of patients with diffuse large B-cell lymphoma

    DEFF Research Database (Denmark)

    Møller, Michael Boe; Pedersen, Niels Tinggaard; Christensen, Bjarne E

    2006-01-01

    a period of time after treatment. Conditional survival data have not been reported for lymphoma patients. METHODS: Conditional survival was estimated for 1209 patients with diffuse large B-cell lymphoma (DLBCL) from the population-based LYFO registry of the Danish Lymphoma Group. The Kaplan-Meier method...... was also significant at diagnosis, but 2 years after diagnosis only age had prognostic impact. Multivariate analysis of patients who survived > or = 3 years identified only age as a prognostic factor. CONCLUSION: For patients with DLBCL who have survived more than 1 year after diagnosis, the conditional......BACKGROUND: Prognosis of lymphoma patients is usually estimated at the time of diagnosis and the estimates are guided by the International Prognostic Index (IPI). However, conditional survival estimates are more informative clinically, as they consider those patients only who have already survived...

  18. Prolonged Survival of Transplanted Osteoblastic Cells Does Not Directly Accelerate the Healing of Calvarial Bone Defects.

    Science.gov (United States)

    Kitami, Megumi; Kaku, Masaru; Rocabado, Juan Marcelo Rosales; Ida, Takako; Akiba, Nami; Uoshima, Katsumi

    2016-09-01

    Considering the increased interest in cell-based bone regeneration, it is necessary to reveal the fate of transplanted cells and their substantive roles in bone regeneration. The aim of this study was to analyze the fate of transplanted cells and the effect of osteogenic cell transplantation on calvarial bone defect healing. An anti-apoptotic protein, heat shock protein (HSP) 27, was overexpressed in osteoblasts. Then, the treated osteoblasts were transplanted to calvarial bone defect and their fate was analyzed to evaluate the significance of transplanted cell survival. Transient overexpression of Hsp27 rescued MC3T3-E1 osteoblastic cells from H2 O2 -induced apoptosis without affecting osteoblastic differentiation in culture. Transplantation of Hsp27-overexpressing cells, encapsulated in collagen gel, showed higher proliferative activity, and fewer apoptotic cells in comparison with control cells. After 4-week of transplantation, both control cell- and Hsp27 overexpressed cell-transplanted groups showed significantly higher new bone formation in comparison with cell-free gel-transplantation group. Interestingly, the prolonged survival of transplanted osteoblastic cells by Hsp27 did not provide additional effect on bone healing. The transplanted cells in collagen gel survived for up to 4-week but did not differentiate into bone-forming osteoblasts. In conclusion, cell-containing collagen gel accelerated calvarial bone defect healing in comparison with cell-free collagen gel. However, prolonged survival of transplanted cells by Hsp27 overexpression did not provide additional effect. These results strongly indicate that cell transplantation-based bone regeneration cannot be explained only by the increment of osteogenic cells. Further studies are needed to elucidate the practical roles of transplanted cells that will potentiate successful bone regeneration. J. Cell. Physiol. 231: 1974-1982, 2016. © 2016 Wiley Periodicals, Inc.

  19. HSP70 mediates survival in apoptotic cells-Boolean network prediction and experimental validation.

    Science.gov (United States)

    Vasaikar, Suhas V; Ghosh, Sourish; Narain, Priyam; Basu, Anirban; Gomes, James

    2015-01-01

    Neuronal stress or injury results in the activation of proteins, which regulate the balance between survival and apoptosis. However, the complex mechanism of cell signaling involving cell death and survival, activated in response to cellular stress is not yet completely understood. To bring more clarity about these mechanisms, a Boolean network was constructed that represented the apoptotic pathway in neuronal cells. FasL and neurotrophic growth factor (NGF) were considered as inputs in the absence and presence of heat shock proteins known to shift the balance toward survival by rescuing pro-apoptotic cells. The probabilities of survival, DNA repair and apoptosis as cellular fates, in the presence of either the growth factor or FasL, revealed a survival bias encoded in the network. Boolean predictions tested by measuring the mRNA level of caspase-3, caspase-8, and BAX in neuronal Neuro2a (N2a) cell line with NGF and FasL as external input, showed positive correlation with the observed experimental results for survival and apoptotic states. It was observed that HSP70 contributed more toward rescuing cells from apoptosis in comparison to HSP27, HSP40, and HSP90. Overexpression of HSP70 in N2a transfected cells showed reversal of cellular fate from FasL-induced apoptosis to survival. Further, the pro-survival role of the proteins BCL2, IAP, cFLIP, and NFκB determined by vertex perturbation analysis was experimentally validated through protein inhibition experiments using EM20-25, Embelin and Wedelolactone, which resulted in 1.27-, 1.26-, and 1.46-fold increase in apoptosis of N2a cells. The existence of a one-to-one correspondence between cellular fates and attractor states shows that Boolean networks may be employed with confidence in qualitative analytical studies of biological networks.

  20. Regulatory T Cell-Enriching Microparticles for Promoting Vascularized Composite Allotransplant Survival

    Science.gov (United States)

    2016-10-01

    AWARD NUMBER: W81XWH-15-1-0244 TITLE: Regulatory T Cell-Enriching Microparticles for Promoting Vascularized Composite Allotransplant Survival...2016 4. TITLE AND SUBTITLE 5a. CONTRACT NUMBER Regulatory T Cell-Enriching Microparticles for Promoting Vascularized Composite Allotransplant Survival... trauma , sepsis/disease, cancer, and congenital defects. In most cases, current reconstructive strategies are sub-optimal or fail to provide optimal

  1. Cell surface lactate receptor GPR81 is crucial for cancer cell survival.

    Science.gov (United States)

    Roland, Christina L; Arumugam, Thiruvengadam; Deng, Defeng; Liu, Shi He; Philip, Bincy; Gomez, Sobeyda; Burns, William R; Ramachandran, Vijaya; Wang, Huamin; Cruz-Monserrate, Zobeida; Logsdon, Craig D

    2014-09-15

    The mechanisms that allow cancer cells to adapt to the typical tumor microenvironment of low oxygen and glucose and high lactate are not well understood. GPR81 is a lactate receptor recently identified in adipose and muscle cells that has not been investigated in cancer. In the current study, we examined GPR81 expression and function in cancer cells. We found that GPR81 was present in colon, breast, lung, hepatocellular, salivary gland, cervical, and pancreatic carcinoma cell lines. Examination of tumors resected from patients with pancreatic cancer indicated that 94% (148 of 158) expressed high levels of GPR81. Functionally, we observed that the reduction of GPR81 levels using shRNA-mediated silencing had little effect on pancreatic cancer cells cultured in high glucose, but led to the rapid death of cancer cells cultured in conditions of low glucose supplemented with lactate. We also observed that lactate addition to culture media induced the expression of genes involved in lactate metabolism, including monocarboxylase transporters in control, but not in GPR81-silenced cells. In vivo, GPR81 expression levels correlated with the rate of pancreatic cancer tumor growth and metastasis. Cells in which GPR81 was silenced showed a dramatic decrease in growth and metastasis. Implantation of cancer cells in vivo was also observed to lead to greatly elevated levels of GPR81. These data support that GPR81 is important for cancer cell regulation of lactate transport mechanisms. Furthermore, lactate transport is important for the survival of cancer cells in the tumor microenvironment. Cancer Res; 74(18); 5301-10. ©2014 AACR. ©2014 American Association for Cancer Research.

  2. Microsomal PGE2 synthase-1 regulates melanoma cell survival and associates with melanoma disease progression.

    Science.gov (United States)

    Kim, Sun-Hee; Hashimoto, Yuuri; Cho, Sung-Nam; Roszik, Jason; Milton, Denái R; Dal, Fulya; Kim, Sangwon F; Menter, David G; Yang, Peiying; Ekmekcioglu, Suhendan; Grimm, Elizabeth A

    2016-05-01

    COX-2 and its product PGE2 enhance carcinogenesis and tumor progression, which has been previously reported in melanoma. As most COX inhibitors cause much toxicity, the downstream microsomal PGE2 synthase-1 (mPGES1) is a consideration for targeting. Human melanoma TMAs were employed for testing mPGES1 protein staining intensity and percentage levels, and both increased with clinical stage; employing a different Stage III TMA, mPGES1 intensity (not percentage) associated with reduced patient survival. Our results further show that iNOS was also highly expressed in melanoma tissues with high mPGES1 levels, and iNOS-mediated NO promoted mPGES1 expression and PGE2 production. An mPGES1-specific inhibitor (CAY10526) as well as siRNA attenuated cell survival and increased apoptosis. CAY10526 significantly suppressed tumor growth and increased apoptosis in melanoma xenografts. Our findings support the value of a prognostic and predictive role for mPGES1, and suggest targeting this molecule in the PGE2 pathway as another avenue toward improving melanoma therapy.

  3. Stanniocalcin 2 enhances mesenchymal stem cell survival by suppressing oxidative stress.

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    Kim, Pyung-Hwan; Na, Sang-Su; Lee, Bomnaerin; Kim, Joo-Hyun; Cho, Je-Yoel

    2015-12-01

    To overcome the disadvantages of stem cell-based cell therapy like low cell survival at the disease site, we used stanniocalcin 2 (STC2), a family of secreted glycoprotein hormones that function to inhibit apoptosis and oxidative damage and to induce proliferation. STC2 gene was transfected into two kinds of stem cells to prolong cell survival and protect the cells from the damage by oxidative stress. The stem cells expressing STC2 exhibited increased cell viability and improved cell survival as well as elevated expression of the pluripotency and self-renewal markers (Oct4 and Nanog) under sub-lethal oxidative conditions. Up-regulation of CDK2 and CDK4 and down-regulation of cell cycle inhibitors p16 and p21 were observed after the delivery of STC2. Furthermore, STC2 transduction activated pAKT and pERK 1/2 signal pathways. Taken together, the STC2 can be used to enhance cell survival and maintain long-term stemness in therapeutic use of stem cells.

  4. Survival of Human Neurofibroma in Immunodeficient Mice and Initial Results of Therapy With Pirfenidone

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    Babovic-Vuksanovic Dusica

    2004-01-01

    Full Text Available Neurofibromatosis type I is a common tumor predisposing disease in humans. Surgical therapy can be applied only in selected patients with resectable masses. Hence, development of new therapies for this disease is urgent. We used human neurofibroma implants in mice with severe combined immunodeficiency (SCID as a model to test the toxicity and potential efficacy of pirfenidone, a new therapeutic agent. Two hundred twelve human neurofibromas were transplanted into various locations in 59 experimental animals, and 30 mice with implants received oral pirfenidone for up to six weeks. Survival of neurofibromas in animals treated with pirfenidone was lower than in the control group ( P=.02 . Tumors did not change histologic appearance or vascularization in response to pirfenidone. Treatment with pirfenidone, a new antifibrotic agent, inhibits survival of some tumors without causing toxicity in animals.

  5. BibA: a novel immunogenic bacterial adhesin contributing to group B Streptococcus survival in human blood.

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    Santi, Isabella; Scarselli, Maria; Mariani, Massimo; Pezzicoli, Alfredo; Masignani, Vega; Taddei, Annarita; Grandi, Guido; Telford, John L; Soriani, Marco

    2007-02-01

    By the analysis of the recently sequenced genomes of Group B Streptococcus (GBS) we have identified a novel immunogenic adhesin with anti-phagocytic activity, named BibA. The bibA gene is present in 100% of the 24 GBS strains analysed. BibA-specific IgG were found in human sera from normal healthy donors. The putative protein product is a polypeptide of 630 amino acids containing a helix-rich N-terminal domain, a proline-rich region and a canonical LPXTG cell wall-anchoring domain. BibA is expressed on the surface of several GBS strains, but is also recovered in GBS culture supernatants. BibA specifically binds to human C4-binding protein, a regulator of the classic complement pathway. Deletion of the bibA gene severely reduced the capacity of GBS to survive in human blood and to resist opsonophagocytic killing by human neutrophils. In addition, BibA expression increased the virulence of GBS in a mouse infection model. The role of BibA in GBS adhesion was demonstrated by the impaired ability of a bibA knockout mutant strain to adhere to both human cervical and lung epithelial cells. Furthermore, we calculated that recombinant BibA bound to human epithelial cells of distinct origin with an affinity constant of approximately 10(-8) M for cervical epithelial cells. Hence BibA is a novel multifunctional protein involved in both resistance to phagocytic killing and adhesion to host cells. The identification of this potential new virulence factor represents an important step in the development of strategies to combat GBS-associated infections.

  6. Gender Specific Mutation Incidence and Survival Associations in Clear Cell Renal Cell Carcinoma (CCRCC.

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    Christopher J Ricketts

    Full Text Available Renal cell carcinoma (RCC is diagnosed in >200,000 individuals worldwide each year, accounting for ~2% of all cancers, but the spread of this disease amongst genders is distinctly uneven. In the U.S. the male:female incidence ratio is approximately 2:1. A potential hypothesis is mutation spectra may differ between tumors dependent upon the gender of the patient, such as mutations of X chromosome encoded genes being more prevalent in male-derived tumors. Combined analysis of three recent large-scale clear cell renal cell carcinoma (CCRCC mutation sequencing projects identified a significantly increased mutation frequency of PBRM1 and the X chromosome encoded KDM5C in tumors from male patients and BAP1 in tumors from female patients. Mutation of BAP1 had previously been significantly associated with poorer overall survival; however, when stratified by gender, mutation of BAP1 only significantly affected overall survival in female patients. Mutation of chromatin remodeling genes alters gene regulation, but the overall effect of these alterations may also be modified by the presence of other gender specific factors. Thus, the combination of gender and mutation of a specific gene, such as BAP1, may have implications not only for prognosis but also for understanding the role of chromatin remodeling gene mutations in kidney cancer progression.

  7. TNFAIP3 promotes survival of CD4 T cells by restricting MTOR and promoting autophagy.

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    Matsuzawa, Yu; Oshima, Shigeru; Takahara, Masahiro; Maeyashiki, Chiaki; Nemoto, Yasuhiro; Kobayashi, Masanori; Nibe, Yoichi; Nozaki, Kengo; Nagaishi, Takashi; Okamoto, Ryuichi; Tsuchiya, Kiichiro; Nakamura, Tetsuya; Ma, Averil; Watanabe, Mamoru

    2015-01-01

    Autophagy plays important roles in metabolism, differentiation, and survival in T cells. TNFAIP3/A20 is a ubiquitin-editing enzyme that is thought to be a negative regulator of autophagy in cell lines. However, the role of TNFAIP3 in autophagy remains unclear. To determine whether TNFAIP3 regulates autophagy in CD4 T cells, we first analyzed Tnfaip3-deficient naïve CD4 T cells in vitro. We demonstrated that Tnfaip3-deficient CD4 T cells exhibited reduced MAP1LC3/LC3 (microtubule-associated protein 1 light chain 3) puncta formation, increased mitochondrial content, and exaggerated reactive oxygen species (ROS) production. These results indicate that TNFAIP3 promotes autophagy after T cell receptor (TCR) stimulation in CD4 T cells. We then investigated the mechanism by which TNFAIP3 promotes autophagy signaling. We found that TNFAIP3 bound to the MTOR (mechanistic target of rapamycin) complex and that Tnfaip3-deficient cells displayed enhanced ubiquitination of the MTOR complex and MTOR activity. To confirm the effects of enhanced MTOR activity in Tnfaip3-deficient cells, we analyzed cell survival following treatment with Torin1, an MTOR inhibitor. Tnfaip3-deficient CD4 T cells exhibited fewer cell numbers than the control cells in vitro and in vivo. In addition, the impaired survival of Tnfaip3-deficient cells was ameliorated with Torin1 treatment in vitro and in vivo. The effect of Torin1 was abolished by Atg5 deficiency. Thus, enhanced MTOR activity regulates the survival of Tnfaip3-deficient CD4 T cells. Taken together, our findings illustrate that TNFAIP3 restricts MTOR signaling and promotes autophagy, providing new insight into the manner in which MTOR and autophagy regulate survival in CD4 T cells.

  8. Inflammatory cytokines regulate endothelial cell survival and tissue repair functions via NF-κB signaling

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    Kanaji N

    2011-09-01

    Full Text Available Nobuhiro Kanaji1, Tadashi Sato2, Amy Nelson3, Xingqi Wang3, YingJi Li4, Miok Kim5, Masanori Nakanishi6, Hesham Basma3, Joel Michalski3, Maha Farid3, Michael Chandler3, William Pease3, Amol Patil3, Stephen I Rennard3, Xiangde Liu31Division of Hematology, Rheumatology and Respiratory Medicine, Kagawa University, Kagawa, Japan; 2Department of Respiratory Medicine, Juntendo University School of Medicine, Tokyo, Japan; 3Pulmonary and Critical Care Medicine, University of Nebraska Medical Center, Omaha, Nebraska; 4Department of Hygiene and Public Health, Nippon Medical School, Tokyo, Japan; 5Third Department of Internal Medicine, Wakayama Medical University School of Medicine, Wakayama, Japan; 6Department of Internal Medicine, Jeju Medical College, Jeju, Republic of KoreaAbstract: Inflammation contributes to the development of fibrotic and malignant diseases. We assessed the ability of inflammatory cytokines to modulate endothelial cell survival and functions related to tissue repair/remodeling. Treatment with interleukin (IL-1ß or tumor necrosis factor (TNF-α (2 ng/mL led to human pulmonary artery endothelial cells becoming spindle-shaped fibroblast-like cells. However, immunoblot and DNA microarray showed no change in most endothelial and mesenchymal markers. In the presence of IL-1ß or TNF-α, cells were resistant to apoptosis induced by deprivation of serum and growth factor, and were more migratory. In addition, cells treated with IL-1ß or TNF-α contracted collagen gels more robustly. In contrast, transforming growth factor-ß1 did not induce these responses. RNA interference targeting nuclear factor (NF- κB p65 blocked the effects of IL-1ß or TNF-α on cell morphologic change, survival, migration, and collagen gel contraction. These results suggest that endothelial cells may contribute to tissue repair/remodeling via the NF-κB signaling in a milieu of airway inflammation.Keywords: NF-κB, IL-1ß, TNF-α, apoptosis, tissue repair

  9. ATP promotes cell survival via regulation of cytosolic [Ca2+] and Bcl-2/Bax ratio in lung cancer cells.

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    Song, Shanshan; Jacobson, Krista N; McDermott, Kimberly M; Reddy, Sekhar P; Cress, Anne E; Tang, Haiyang; Dudek, Steven M; Black, Stephen M; Garcia, Joe G N; Makino, Ayako; Yuan, Jason X-J

    2016-01-15

    Adenosine triphosphate (ATP) is a ubiquitous extracellular messenger elevated in the tumor microenvironment. ATP regulates cell functions by acting on purinergic receptors (P2X and P2Y) and activating a series of intracellular signaling pathways. We examined ATP-induced Ca(2+) signaling and its effects on antiapoptotic (Bcl-2) and proapoptotic (Bax) proteins in normal human airway epithelial cells and lung cancer cells. Lung cancer cells exhibited two phases (transient and plateau phases) of increase in cytosolic [Ca(2+)] ([Ca(2+)]cyt) caused by ATP, while only the transient phase was observed in normal cells. Removal of extracellular Ca(2+) eliminated the plateau phase increase of [Ca(2+)]cyt in lung cancer cells, indicating that the plateau phase of [Ca(2+)]cyt increase is due to Ca(2+) influx. The distribution of P2X (P2X1-7) and P2Y (P2Y1, P2Y2, P2Y4, P2Y6, P2Y11) receptors was different between lung cancer cells and normal cells. Proapoptotic P2X7 was nearly undetectable in lung cancer cells, which may explain why lung cancer cells showed decreased cytotoxicity when treated with high concentration of ATP. The Bcl-2/Bax ratio was increased in lung cancer cells following treatment with ATP; however, the antiapoptotic protein Bcl-2 demonstrated more sensitivity to ATP than proapoptotic protein Bax. Decreasing extracellular Ca(2+) or chelating intracellular Ca(2+) with BAPTA-AM significantly inhibited ATP-induced increase in Bcl-2/Bax ratio, indicating that a rise in [Ca(2+)]cyt through Ca(2+) influx is the critical mediator for ATP-mediated increase in Bcl-2/Bax ratio. Therefore, despite high ATP levels in the tumor microenvironment, which would induce cell apoptosis in normal cells, the decreased P2X7 and elevated Bcl-2/Bax ratio in lung cancer cells may enable tumor cells to survive. Increasing the Bcl-2/Bax ratio by exposure to high extracellular ATP may, therefore, be an important selective pressure promoting transf