Science.gov

Sample records for acid promotes cell

  1. Histone Deacetylase Inhibitor Valproic Acid Promotes the Differentiation of Human Induced Pluripotent Stem Cells into Hepatocyte-Like Cells

    PubMed Central

    Kondo, Yuki; Iwao, Takahiro; Yoshihashi, Sachimi; Mimori, Kayo; Ogihara, Ruri; Nagata, Kiyoshi; Kurose, Kouichi; Saito, Masayoshi; Niwa, Takuro; Suzuki, Takayoshi; Miyata, Naoki; Ohmori, Shigeru; Nakamura, Katsunori; Matsunaga, Tamihide

    2014-01-01

    In this study, we aimed to elucidate the effects and mechanism of action of valproic acid on hepatic differentiation from human induced pluripotent stem cell-derived hepatic progenitor cells. Human induced pluripotent stem cells were differentiated into endodermal cells in the presence of activin A and then into hepatic progenitor cells using dimethyl sulfoxide. Hepatic progenitor cells were matured in the presence of hepatocyte growth factor, oncostatin M, and dexamethasone with valproic acid that was added during the maturation process. After 25 days of differentiation, cells expressed hepatic marker genes and drug-metabolizing enzymes and exhibited drug-metabolizing enzyme activities. These expression levels and activities were increased by treatment with valproic acid, the timing and duration of which were important parameters to promote differentiation from human induced pluripotent stem cell-derived hepatic progenitor cells into hepatocytes. Valproic acid inhibited histone deacetylase activity during differentiation of human induced pluripotent stem cells, and other histone deacetylase inhibitors also enhanced differentiation into hepatocytes. In conclusion, histone deacetylase inhibitors such as valproic acid can be used to promote hepatic differentiation from human induced pluripotent stem cell-derived hepatic progenitor cells. PMID:25084468

  2. Gelatin-Hyaluronic Acid Hydrogels with Tuned Stiffness to Counterbalance Cellular Forces and Promote Cell Differentiation.

    PubMed

    Poveda-Reyes, Sara; Moulisova, Vladimira; Sanmartín-Masiá, Esther; Quintanilla-Sierra, Luis; Salmerón-Sánchez, Manuel; Ferrer, Gloria Gallego

    2016-09-01

    Cells interact mechanically with their environment, exerting mechanical forces that probe the extracellular matrix (ECM). The mechanical properties of the ECM determine cell behavior and control cell differentiation both in 2D and 3D environments. Gelatin (Gel) is a soft hydrogel into which cells can be embedded. This study shows significant 3D Gel shrinking due to the high traction cellular forces exerted by the cells on the matrix, which prevents cell differentiation. To modulate this process, Gel with hyaluronic acid (HA) has been combined in an injectable crosslinked hydrogel with controlled Gel-HA ratio. HA increases matrix stiffness. The addition of small amounts of HA leads to a significant reduction in hydrogel shrinking after cell encapsulation (C2C12 myoblasts). We show that hydrogel stiffness counterbalanced traction forces of cells and this was decisive in promoting cell differentiation and myotube formation of C2C12 encapsulated in the hybrid hydrogels.

  3. Promise of Retinoic Acid-Triazolyl Derivatives in Promoting Differentiation of Neuroblastoma Cells.

    PubMed

    Lone, Ali Mohd; Dar, Nawab John; Hamid, Abid; Shah, Wajaht Amin; Ahmad, Muzamil; Bhat, Bilal A

    2016-01-20

    Retinoic acid induces differentiation in various types of cells including skeletal myoblasts and neuroblasts and maintains differentiation of epithelial cells. The present study demonstrates synthesis and screening of a library of retinoic acid-triazolyl derivatives for their differentiation potential on neuroblastoma cells. Click chemistry approach using copper(I)-catalyzed azide-alkyne cycloaddition was adopted for the preparation of these derivatives. The neurite outgrowth promoting potential of retinoic acid-triazolyl derivatives was studied on neuroblastoma cells. Morphological examination revealed that compounds 8a, 8e, 8f, and 8k, among the various derivatives screened, exhibited promising neurite-outgrowth inducing activity at a concentration of 10 μM compared to undifferentiated and retinoic acid treated cells. Further on, to confirm this differentiation potential of these compounds, neuroblastoma cells were probed for expression of neuronal markers such as NF-H and NeuN. The results revealed a marked increase in the NF-H and NeuN protein expression when treated with 8a, 8e, 8f, and 8k compared to undifferentiated and retinoic acid treated cells. Thus, these compounds could act as potential leads in inducing neuronal differentiation for future studies.

  4. Perfluorooctanoic acid (PFOA) acts as a tumor promoter on Syrian hamster embryo (SHE) cells.

    PubMed

    Jacquet, N; Maire, M A; Rast, C; Bonnard, M; Vasseur, P

    2011-08-01

    Perfluorooctane sulfonate (PFOS) (C(8)F(17)SO(3)) and perfluorooctanoic acid (PFOA) (C(8)HF(15)O(2)) are synthetic chemicals widely used in industrial applications for their hydrophobic and oleophobic properties. They are persistent, bioaccumulative, and toxic to mammalian species. Their widespread distribution on earth and contamination of human serum raised concerns about long-term side effects. They are suspected to be carcinogenic through a nongenotoxic mode of action, a mechanism supported by recent findings that PFOS induced cell transformation but no genotoxicity in Syrian hamster embryo (SHE) cells. In the present study, we evaluated carcinogenic potential of PFOA using the cell transformation assay on SHE cells. The chemical was applied alone or in combination with a nontransformant concentration of benzo[a]pyrene (BaP, 0.4 μM) in order to detect PFOA ability to act as tumor initiator or tumor promoter. The results showed that PFOA tested alone in the range 3.7 × 10(-5) to 300 μM did not induce SHE cell transformation frequency in a 7-day treatment. On the other side, the combination BaP/PFOA induced cell transformation at all PFOA concentrations tested, which revealed synergistic effects. No genotoxicity of PFOA on SHE cells was detected using the comet assay after 5 and 24 h of exposure. No significant increase in DNA breakage was found in BaP-initiated cells exposed to PFOA in a 7-day treatment. The whole results showed that PFOA acts as a tumor promoter and a nongenotoxic carcinogen. Cell transformation in initiated cells was observed at concentrations equivalent to the ones found in human serum of nonoccupationally and occupationally exposed populations. An involvement of PFOA in increased incidence of cancer recorded in occupationally exposed population cannot be ruled out. PMID:22828883

  5. Perfluorooctanoic acid (PFOA) acts as a tumor promoter on Syrian hamster embryo (SHE) cells.

    PubMed

    Jacquet, N; Maire, M A; Rast, C; Bonnard, M; Vasseur, P

    2011-08-01

    Perfluorooctane sulfonate (PFOS) (C(8)F(17)SO(3)) and perfluorooctanoic acid (PFOA) (C(8)HF(15)O(2)) are synthetic chemicals widely used in industrial applications for their hydrophobic and oleophobic properties. They are persistent, bioaccumulative, and toxic to mammalian species. Their widespread distribution on earth and contamination of human serum raised concerns about long-term side effects. They are suspected to be carcinogenic through a nongenotoxic mode of action, a mechanism supported by recent findings that PFOS induced cell transformation but no genotoxicity in Syrian hamster embryo (SHE) cells. In the present study, we evaluated carcinogenic potential of PFOA using the cell transformation assay on SHE cells. The chemical was applied alone or in combination with a nontransformant concentration of benzo[a]pyrene (BaP, 0.4 μM) in order to detect PFOA ability to act as tumor initiator or tumor promoter. The results showed that PFOA tested alone in the range 3.7 × 10(-5) to 300 μM did not induce SHE cell transformation frequency in a 7-day treatment. On the other side, the combination BaP/PFOA induced cell transformation at all PFOA concentrations tested, which revealed synergistic effects. No genotoxicity of PFOA on SHE cells was detected using the comet assay after 5 and 24 h of exposure. No significant increase in DNA breakage was found in BaP-initiated cells exposed to PFOA in a 7-day treatment. The whole results showed that PFOA acts as a tumor promoter and a nongenotoxic carcinogen. Cell transformation in initiated cells was observed at concentrations equivalent to the ones found in human serum of nonoccupationally and occupationally exposed populations. An involvement of PFOA in increased incidence of cancer recorded in occupationally exposed population cannot be ruled out.

  6. Dimethylsulfoniopropionate Promotes Process Outgrowth in Neural Cells and Exerts Protective Effects against Tropodithietic Acid.

    PubMed

    Wichmann, Heidi; Brinkhoff, Thorsten; Simon, Meinhard; Richter-Landsberg, Christiane

    2016-01-01

    The marine environment harbors a plethora of bioactive substances, including drug candidates of potential value in the field of neuroscience. The present study was undertaken to investigate the effects of dimethylsulfoniopropionate (DMSP), produced by several algae, corals and higher plants, on cells of the mammalian nervous system, i.e., neuronal N2a and OLN-93 cells as model system for nerve cells and glia, respectively. Additionally, the protective capabilities of DMSP were assessed in cells treated with tropodithietic acid (TDA), a marine metabolite produced by several Roseobacter clade bacteria. Both cell lines, N2a and OLN-93, have previously been shown to be a sensitive target for the action of TDA, and cytotoxic effects of TDA have been connected to the induction of oxidative stress. Our data shows that DMSP promotes process outgrowth and microtubule reorganization and bundling, accompanied by an increase in alpha-tubulin acetylation. Furthermore, DMSP was able to prevent the cytotoxic effects exerted by TDA, including the breakdown of the mitochondrial membrane potential, upregulation of heat shock protein Hsp32 and activation of the extracellular signal-regulated kinases 1/2 (ERK1/2). Our study points to the conclusion that DMSP provides an antioxidant defense, not only in algae but also in mammalian neural cells. PMID:27164116

  7. Dimethylsulfoniopropionate Promotes Process Outgrowth in Neural Cells and Exerts Protective Effects against Tropodithietic Acid

    PubMed Central

    Wichmann, Heidi; Brinkhoff, Thorsten; Simon, Meinhard; Richter-Landsberg, Christiane

    2016-01-01

    The marine environment harbors a plethora of bioactive substances, including drug candidates of potential value in the field of neuroscience. The present study was undertaken to investigate the effects of dimethylsulfoniopropionate (DMSP), produced by several algae, corals and higher plants, on cells of the mammalian nervous system, i.e., neuronal N2a and OLN-93 cells as model system for nerve cells and glia, respectively. Additionally, the protective capabilities of DMSP were assessed in cells treated with tropodithietic acid (TDA), a marine metabolite produced by several Roseobacter clade bacteria. Both cell lines, N2a and OLN-93, have previously been shown to be a sensitive target for the action of TDA, and cytotoxic effects of TDA have been connected to the induction of oxidative stress. Our data shows that DMSP promotes process outgrowth and microtubule reorganization and bundling, accompanied by an increase in alpha-tubulin acetylation. Furthermore, DMSP was able to prevent the cytotoxic effects exerted by TDA, including the breakdown of the mitochondrial membrane potential, upregulation of heat shock protein Hsp32 and activation of the extracellular signal-regulated kinases 1/2 (ERK1/2). Our study points to the conclusion that DMSP provides an antioxidant defense, not only in algae but also in mammalian neural cells. PMID:27164116

  8. Promotion of Intestinal Epithelial Cell Turnover by Commensal Bacteria: Role of Short-Chain Fatty Acids

    PubMed Central

    Park, Jung-ha; Kotani, Takenori; Konno, Tasuku; Setiawan, Jajar; Kitamura, Yasuaki; Imada, Shinya; Usui, Yutaro; Hatano, Naoya; Shinohara, Masakazu; Saito, Yasuyuki; Murata, Yoji; Matozaki, Takashi

    2016-01-01

    The life span of intestinal epithelial cells (IECs) is short (3–5 days), and its regulation is thought to be important for homeostasis of the intestinal epithelium. We have now investigated the role of commensal bacteria in regulation of IEC turnover in the small intestine. The proliferative activity of IECs in intestinal crypts as well as the migration of these cells along the crypt-villus axis were markedly attenuated both in germ-free mice and in specific pathogen–free (SPF) mice treated with a mixture of antibiotics, with antibiotics selective for Gram-positive bacteria being most effective in this regard. Oral administration of chloroform-treated feces of SPF mice to germ-free mice resulted in a marked increase in IEC turnover, suggesting that spore-forming Gram-positive bacteria contribute to this effect. Oral administration of short-chain fatty acids (SCFAs) as bacterial fermentation products also restored the turnover of IECs in antibiotic-treated SPF mice as well as promoted the development of intestinal organoids in vitro. Antibiotic treatment reduced the phosphorylation levels of ERK, ribosomal protein S6, and STAT3 in IECs of SPF mice. Our results thus suggest that Gram-positive commensal bacteria are a major determinant of IEC turnover, and that their stimulatory effect is mediated by SCFAs. PMID:27232601

  9. Asparagine promotes cancer cell proliferation through use as an amino acid exchange factor.

    PubMed

    Krall, Abigail S; Xu, Shili; Graeber, Thomas G; Braas, Daniel; Christofk, Heather R

    2016-01-01

    Cellular amino acid uptake is critical for mTOR complex 1 (mTORC1) activation and cell proliferation. However, the regulation of amino acid uptake is not well-understood. Here we describe a role for asparagine as an amino acid exchange factor: intracellular asparagine exchanges with extracellular amino acids. Through asparagine synthetase knockdown and altering of media asparagine concentrations, we show that intracellular asparagine levels regulate uptake of amino acids, especially serine, arginine and histidine. Through its exchange factor role, asparagine regulates mTORC1 activity and protein synthesis. In addition, we show that asparagine regulation of serine uptake influences serine metabolism and nucleotide synthesis, suggesting that asparagine is involved in coordinating protein and nucleotide synthesis. Finally, we show that maintenance of intracellular asparagine levels is critical for cancer cell growth. Collectively, our results indicate that asparagine is an important regulator of cancer cell amino acid homeostasis, anabolic metabolism and proliferation. PMID:27126896

  10. Taurolithocholic acid promotes intrahepatic cholangiocarcinoma cell growth via muscarinic acetylcholine receptor and EGFR/ERK1/2 signaling pathway.

    PubMed

    Amonyingcharoen, Sumet; Suriyo, Tawit; Thiantanawat, Apinya; Watcharasit, Piyajit; Satayavivad, Jutamaad

    2015-01-01

    Cholangiocarcinoma (CCA) is a malignant cancer of the biliary tract and its occurrence is associated with chronic cholestasis which causes an elevation of bile acids in the liver and bile duct. The present study aimed to investigate the role and mechanistic effect of bile acids on the CCA cell growth. Intrahepatic CCA cell lines, RMCCA-1 and HuCCA-1, were treated with bile acids and their metabolites to determine the growth promoting effect. Cell viability, cell cycle analysis, EdU incorporation assays were conducted. Intracellular signaling proteins were detected by western immunoblotting. Among eleven forms of bile acids and their metabolites, only taurolithocholic acid (TLCA) concentration dependently (1-40 µM) increased the cell viability of RMCCA-1, but not HuCCA-1 cells. The cell cycle analysis showed induction of cells in the S phase and the EdU incorporation assay revealed induction of DNA synthesis in the TLCA-treated RMCCA-1 cells. Moreover, TLCA increased the phosphorylation of EGFR, ERK 1/2 and also increased the expression of cyclin D1 in RMCCA-1 cells. Furthermore, TLCA-induced RMCCA-1 cell growth could be inhibited by atropine, a non-selective muscarinic acetylcholine receptor (mAChR) antagonist, AG 1478, a specific EGFR inhibitor, or U 0126, a specific MEK 1/2 inhibitor. These results suggest that TLCA induces CCA cell growth via mAChR and EGFR/EKR1/2 signaling pathway. Moreover, the functional presence of cholinergic system plays a certain role in TLCA-induced CCA cell growth.

  11. Essential role for retinoic acid in the promotion of CD4+ T cell effector responses via retinoic acid receptor alpha

    PubMed Central

    Hall, J.A.; Cannons, J.L.; Grainger, J.R.; Santos, L.M. Dos; Hand, T.W.; Naik, S.; Wohlfert, E.A.; Chou, D.B.; Oldenhove, G.; Robinson, M.; Grigg, M.E.; Kastenmayer, R.; Schwartzberg, P.L.; Belkaid, Y.

    2012-01-01

    SUMMARY Vitamin A and its metabolite, retinoic acid (RA), have recently been implicated in the regulation of immune homeostasis via the peripheral induction of regulatory T cells. Here we show that RA is also required to elicit proinflammatory CD4+ helper T cell responses to infection and mucosal vaccination. Retinoic acid receptor alpha (RARα) is the critical mediator of these effects. Strikingly, antagonism of RAR signaling and deficiency in RARα(Rara−/−) results in a cell autonomous CD4+ T cell activation defect. Altogether, these findings reveal a fundamental role for the RA/RARα axis in the development of both regulatory and inflammatory arms of adaptive immunity and establish nutritional status as a broad regulator of adaptive T cell responses. PMID:21419664

  12. Fatty acid activated PPARγ promotes tumorigenicity of prostate cancer cells by up regulating VEGF via PPAR responsive elements of the promoter

    PubMed Central

    Forootan, Farzad S.; Forootan, Shiva S.; Gou, Xiaojun; Yang, Jin; Liu, Bichong; Chen, Danqing; Fayi, Majed Saad Al; Al-Jameel, Waseem; Rudland, Philip S.; Hussain, Syed A.; Ke, Youqiang

    2016-01-01

    In previous work, it is suggested that the excessive amount of fatty acids transported by FABP5 may facilitate the malignant progression of prostate cancer cells through a FABP5-PPARγ-VEGF signal transduction axis to increase angiogenesis. To further functionally characterise the FABP5-PPARγ-VEGF signal transduction pathway, we have, in this work, investigated the molecular mechanisms involved in its tumorigenicity promoting role in prostate cancer. Suppression of PPARγ in highly malignant prostate cancer cells produced a significant reduction (up to 53%) in their proliferation rate, invasiveness (up to 89%) and anchorage-independent growth (up to 94%) in vitro. Knockdown of PPARγ gene in PC3-M cells by siRNA significantly reduced the average size of tumours formed in nude mice by 99% and tumour incidence by 90%, and significantly prolonged the latent period by 3.5 fold. Results in this study combined with some previous results suggested that FABP5 promoted VEGF expression and angiogenesis through PPARγ which was activated by fatty acids transported by FABP5. Further investigations showed that PPARγ up-regulated VEGF expression through acting with the PPAR-responsive elements in the promoter region of VEGF gene in prostate cancer cells. Although androgen can modulate VEGF expression through Sp1/Sp3 binding site on VEGF promoter in androgen-dependent prostate cancer cells, this route, disappeared as the cells gradually lost their androgen dependency; was replaced by the FABP5-PPARγ-VEGF signalling pathway. These results suggested that the FABP5-PPARγ-VEGF signal transduction axis, rather than androgen modulated route, may be a more important novel therapeutic target for angiogenesis-suppression treatment of castration resistant prostate cancer. PMID:26814431

  13. Lysophosphatidic Acid Up-Regulates Hexokinase II and Glycolysis to Promote Proliferation of Ovarian Cancer Cells1

    PubMed Central

    Mukherjee, Abir; Ma, Yibao; Yuan, Fang; Gong, Yongling; Fang, Zhenyu; Mohamed, Esraa M.; Berrios, Erika; Shao, Huanjie; Fang, Xianjun

    2015-01-01

    Lysophosphatidic acid (LPA), a blood-borne lipid mediator, is present in elevated concentrations in ascites of ovarian cancer patients and other malignant effusions. LPA is a potent mitogen in cancer cells. The mechanism linking LPA signal to cancer cell proliferation is not well understood. Little is known about whether LPA affects glucose metabolism to accommodate rapid proliferation of cancer cells. Here we describe that in ovarian cancer cells, LPA enhances glycolytic rate and lactate efflux. A real time PCR-based miniarray showed that hexokinase II (HK2) was the most dramatically induced glycolytic gene to promote glycolysis in LPA-treated cells. Analysis of the human HK2 gene promoter identified the sterol regulatory element-binding protein as the primary mediator of LPA-induced HK2 transcription. The effects of LPA on HK2 and glycolysis rely on LPA2, an LPA receptor subtype overexpressed in ovarian cancer and many other malignancies. We further examined the general role of growth factor-induced glycolysis in cell proliferation. Like LPA, epidermal growth factor (EGF) elicited robust glycolytic and proliferative responses in ovarian cancer cells. Insulin-like growth factor 1 (IGF-1) and insulin, however, potently stimulated cell proliferation but only modestly induced glycolysis. Consistent with their differential effects on glycolysis, LPA and EGF-dependent cell proliferation was highly sensitive to glycolytic inhibition while the growth-promoting effect of IGF-1 or insulin was more resistant. These results indicate that LPA- and EGF-induced cell proliferation selectively involves up-regulation of HK2 and glycolytic metabolism. The work is the first to implicate LPA signaling in promotion of glucose metabolism in cancer cells. PMID:26476080

  14. Retinoic acid promotes the proliferation of primordial germ cell-like cells differentiated from mouse skin-derived stem cells in vitro.

    PubMed

    Tan, Hui; Wang, Jun-Jie; Cheng, Shun-Feng; Ge, Wei; Sun, Yuan-Chao; Sun, Xiao-Feng; Sun, Rui; Li, Lan; Li, Bo; Shen, Wei

    2016-02-01

    Skin-derived stem cells (SDSCs) have the potential to differentiate into gametes and are a potential resource for research and clinical applications. Sufficient amount of primordial germ cells (PGCs) is an important requirement for successful differentiation of SDSCs into gametes in vitro. Retinoic acid (RA), a vitamin A-derived small lipophilic molecule, promotes the growth of PGCs in vivo; however, the role of RA on the proliferation of PGC-like cells (PGCLCs) derived from SDSCs remains unknown. In this study, SDSCs were induced to differentiate into the embryoid body and cocultured with mouse fibroblasts to form PGCLCs. The proliferation of PGCLCs with the presence of various concentrations of RA was investigated in vitro. Immunofluorescence labeling showed that the 5-Bromo-2-deoxyUridine-positive ratio of PGCLCs was increased after the cells were treated with 5-μM RA, and flow cytometry results showed that the number of cells in the S phase was increased significantly. The messenger RNA expression levels of cell cycle-related genes, CCND1 and CDK2, were also increased. Furthermore, RA effectively promoted the external proliferation of endogenous PGCs when 11.5-days postcoitum fetal mouse genital ridges were cultured in vitro. In conclusion, 5-μM RA promoted the proliferation of SDSCs-derived PGCLCs and endogenous PGCs. Our study will provide a valuable model system for studying the differentiation of stem cells into gametes in vitro.

  15. Phosphatidic Acid (PA) can Displace PPARα/LXRα Binding to The EGFR Promoter Causing its Transrepression in Luminal Cancer Cells

    PubMed Central

    Mahankali, Madhu; Farkaly, Terry; Bedi, Shimpi; Hostetler, Heather A.; Gomez-Cambronero, Julian

    2015-01-01

    The expression of the epidermal growth factor receptor (EGFR) is highly regulated in normal cells, whereas some cancer cells have high constitutive levels. Understanding naturally-occurring ways of downregulating EGFR in cancer cells was investigated. Phosphatidic acid (PA) or Nuclear Receptors (NR) PPARα/RXRα/LXRα, enhance EGFR expression, mediated by the promoter region -856(A) to -226(T). Unexpectedly, the combination of NRs and PA caused repression. PA induces a conformational change in the nuclear receptor PPARα (increase of alpha-helices at the expense of decreasing beta-sheets), as evidenced by circular dichroism. This represses the naturally-enhancing capability of PPARα on EGFR transcription. PPARα-overexpressing cells in the presence of PA > 300 nM or the enzyme that produces it, phospholipase D (PLD), downregulate EGFR expression. The reasons are two-fold. First, PA displaces PPARα binding to the EGFR promoter at those concentrations. Second, NR heterodimer-dependent promoter activity is weakened in the presence of PA in vivo. Since other genes considered (β-catenin, cyclin D3, PLD2 and ACOX-1) are also downregulated with a PA + PPARα combination, the transrepression appears to be a global phenomenon. Lastly, the reported effect is greater in MCF-7 than in MDA-MB-231 breast cancer cells, which could provide a novel basis for regulating excessive expression of EGFR in luminal cancer cells. PMID:26493292

  16. Phosphatidic Acid (PA) can Displace PPARα/LXRα Binding to The EGFR Promoter Causing its Transrepression in Luminal Cancer Cells.

    PubMed

    Mahankali, Madhu; Farkaly, Terry; Bedi, Shimpi; Hostetler, Heather A; Gomez-Cambronero, Julian

    2015-10-23

    The expression of the epidermal growth factor receptor (EGFR) is highly regulated in normal cells, whereas some cancer cells have high constitutive levels. Understanding naturally-occurring ways of downregulating EGFR in cancer cells was investigated. Phosphatidic acid (PA) or Nuclear Receptors (NR) PPARα/RXRα/LXRα, enhance EGFR expression, mediated by the promoter region -856(A) to -226(T). Unexpectedly, the combination of NRs and PA caused repression. PA induces a conformational change in the nuclear receptor PPARα (increase of alpha-helices at the expense of decreasing beta-sheets), as evidenced by circular dichroism. This represses the naturally-enhancing capability of PPARα on EGFR transcription. PPARα-overexpressing cells in the presence of PA > 300 nM or the enzyme that produces it, phospholipase D (PLD), downregulate EGFR expression. The reasons are two-fold. First, PA displaces PPARα binding to the EGFR promoter at those concentrations. Second, NR heterodimer-dependent promoter activity is weakened in the presence of PA in vivo. Since other genes considered (β-catenin, cyclin D3, PLD2 and ACOX-1) are also downregulated with a PA + PPARα combination, the transrepression appears to be a global phenomenon. Lastly, the reported effect is greater in MCF-7 than in MDA-MB-231 breast cancer cells, which could provide a novel basis for regulating excessive expression of EGFR in luminal cancer cells.

  17. Adiponectin receptor 1 conserves docosahexaenoic acid and promotes photoreceptor cell survival

    PubMed Central

    Rice, Dennis S.; Calandria, Jorgelina M.; Gordon, William C.; Jun, Bokkyoo; Zhou, Yongdong; Gelfman, Claire M.; Li, Songhua; Jin, Minghao; Knott, Eric J.; Chang, Bo; Abuin, Alex; Issa, Tawfik; Potter, David; Platt, Kenneth A.; Bazan, Nicolas G.

    2015-01-01

    The identification of pathways necessary for photoreceptor and retinal pigment epithelium (RPE) function is critical to uncover therapies for blindness. Here we report the discovery of adiponectin receptor 1 (AdipoR1) as a regulator of these cells’ functions. Docosahexaenoic acid (DHA) is avidly retained in photoreceptors, while mechanisms controlling DHA uptake and retention are unknown. Thus, we demonstrate that AdipoR1 ablation results in DHA reduction. In situ hybridization reveals photoreceptor and RPE cell AdipoR1 expression, blunted in AdipoR1−/− mice. We also find decreased photoreceptor-specific phosphatidylcholine containing very long-chain polyunsaturated fatty acids and severely attenuated electroretinograms. These changes precede progressive photoreceptor degeneration in AdipoR1−/− mice. RPE-rich eyecup cultures from AdipoR1−/− reveal impaired DHA uptake. AdipoR1 overexpression in RPE cells enhances DHA uptake, whereas AdipoR1 silencing has the opposite effect. These results establish AdipoR1 as a regulatory switch of DHA uptake, retention, conservation and elongation in photoreceptors and RPE, thus preserving photoreceptor cell integrity. PMID:25736573

  18. Acidic extracellular pH promotes prostate cancer bone metastasis by enhancing PC-3 stem cell characteristics, cell invasiveness and VEGF-induced vasculogenesis of BM-EPCs.

    PubMed

    Huang, Sheng; Tang, Yubo; Peng, Xinsheng; Cai, Xingdong; Wa, Qingde; Ren, Dong; Li, Qiji; Luo, Jiaquan; Li, Liangping; Zou, Xuenong; Huang, Shuai

    2016-10-01

    Bone metastasis is a main cause of cancer-related mortality in patients with advanced prostate cancer. Emerging evidence suggests that the acidic extracellular microenvironment plays significant roles in the growth and metastasis of tumors. However, the effects of acidity on bone metastasis of PCa remain undefined. In the present study, PC-3 cells were cultured in acidic medium (AM; pH 6.5) or neutral medium (NM; pH 7.4), aiming to investigate the effects and possible mechanisms of acidic extracellular microenvironment in bone metastasis of PCa. Our results showed that AM can promote spheroid and colony formations, cell viability and expression of stem cell characteristic-related markers in PC-3 cells. Moreover, AM stimulates MMP-9 secretion and promotes invasiveness of PC-3 cells, and these effects can be inhibited by blocking of MMP-9. Furthermore, AM stimulates VEGF secretion of PC-3 and AM conditioned medium (CMAM) promotes vasculogenesis of BM-EPCs by increasing cell viability, migration, tube formation, which involved activating the phosphorylation of VEGFR-2, Akt and P38, when pH of NM conditioned medium (CMNM) was modulated the same as AM conditioned medium (CMAM). Further studies have shown that CMNM induced vasculogenesis of BM-EPCs can be inhibited by the inhibition of VEGFR2 with DMH4. These findings suggest that acidic extracellular microenvironment may have the potential to modulate prostate cancer bone metastasis by enhancing PC-3 stem cell characteristics, cell invasiveness and VEGF-induced vasculogenesis of BM-EPCs. Improved anticancer strategies should be designed to selectively target acidic tumor microenvironment.

  19. Unsaturated fatty acids promote bioaccessibility and basolateral secretion of carotenoids and α-tocopherol by Caco-2 cells.

    PubMed

    Failla, Mark L; Chitchumronchokchai, Chureeporn; Ferruzzi, Mario G; Goltz, Shellen R; Campbell, Wayne W

    2014-06-01

    Bioavailability of carotenoids and tocopherols from foods is determined by the efficiency of transfer from food/meal to mixed micelles during digestion, incorporation into chylomicrons for trans-epithelial transport to lymphatic/blood system, and distribution to target tissues. Fats and oils are important factors for facilitating the absorption of lipophilic compounds. However, dietary fats and oils are composed of various types of saturated and unsaturated fatty acids which may differentially impact the bioavailability of carotenoids and tocopherols from foods. We have investigated the effects of several common commercial lipids on bioavailability using an in vitro digestion model and Caco-2 human intestinal cells. Meals consisted of mixed salad vegetables containing a single test lipid. Micellarization and cellular uptake of β-carotene (βC) and lycopene (LYC) during small intestinal digestion was increased by lipids rich in unsaturated fatty acids: soybean oil > olive > canola > butter. In contrast, type of lipid minimally affected the bioaccessibility of lutein (LUT) and zeaxanthin (ZEA). To examine the influence of type of dietary triglyceride on uptake and basolateral secretion of carotenoids, Caco-2 cells grown on Transwell membranes were incubated with micellar mixtures of fatty acids (1.0 mM) mimicking the types and ratio of saturated to unsaturated (mono- + poly-unsaturated) fatty acids (FA) present in butter (70 : 30), olive oil (7 : 93) and soybean oil (11 : 89). Cells were exposed to micelles containing βC, LUT, α-tocopherol (α-TC) and a mixture of test fatty acids. Uptake and basolateral secretion of βC, LUT and α-TC were greater in cells pre-treated with mixtures enriched in unsaturated compared to saturated FA and these effects were mediated by increased assembly and secretion of chylomicrons. These results suggest that dietary fats/oils rich in unsaturated fatty acids promote carotenoid and α-TC bioavailability by enhancing their

  20. Select microtubule inhibitors increase lysosome acidity and promote lysosomal disruption in acute myeloid leukemia (AML) cells.

    PubMed

    Bernard, Dannie; Gebbia, Marinella; Prabha, Swayam; Gronda, Marcela; MacLean, Neil; Wang, Xiaoming; Hurren, Rose; Sukhai, Mahadeo A; Cho, Eunice E; Manolson, Morris F; Datti, Alessandro; Wrana, Jeffrey; Minden, Mark D; Al-Awar, Rima; Aman, Ahmed; Nislow, Corey; Giaever, Guri; Schimmer, Aaron D

    2015-07-01

    To identify new biological vulnerabilities in acute myeloid leukemia, we screened a library of natural products for compounds cytotoxic to TEX leukemia cells. This screen identified the novel small molecule Deoxysappanone B 7,4' dimethyl ether (Deox B 7,4), which possessed nanomolar anti-leukemic activity. To determine the anti-leukemic mechanism of action of Deox B 7,4, we conducted a genome-wide screen in Saccharomyces cerevisiae and identified enrichment of genes related to mitotic cell cycle as well as vacuolar acidification, therefore pointing to microtubules and vacuolar (V)-ATPase as potential drug targets. Further investigations into the mechanisms of action of Deox B 7,4 and a related analogue revealed that these compounds were reversible microtubule inhibitors that bound near the colchicine site. In addition, Deox B 7,4 and its analogue increased lysosomal V-ATPase activity and lysosome acidity. The effects on microtubules and lysosomes were functionally important for the anti-leukemic effects of these drugs. The lysosomal effects were characteristic of select microtubule inhibitors as only the Deox compounds and nocodazole, but not colchicine, vinca alkaloids or paclitaxel, altered lysosome acidity and induced lysosomal disruption. Thus, our data highlight a new mechanism of action of select microtubule inhibitors on lysosomal function. PMID:25832785

  1. Electrical stimulation promotes nerve cell differentiation on polypyrrole/poly (2-methoxy-5 aniline sulfonic acid) composites

    NASA Astrophysics Data System (ADS)

    Liu, Xiao; Gilmore, Kerry J.; Moulton, Simon E.; Wallace, Gordon G.

    2009-12-01

    The purpose of this work was to investigate for the first time the potential biomedical applications of novel polypyrrole (PPy) composites incorporating a large polyelectrolyte dopant, poly (2-methoxy-5 aniline sulfonic acid) (PMAS). The physical and electrochemical properties were characterized. The PPy/PMAS composites were found to be smooth and hydrophilic and have low electrical impedance. We demonstrate that PPy/PMAS supports nerve cell (PC12) differentiation, and that clinically relevant 250 Hz biphasic current pulses delivered via PPy/PMAS films significantly promote nerve cell differentiation in the presence of nerve growth factor (NGF). The capacity of PPy/PMAS composites to support and enhance nerve cell differentiation via electrical stimulation renders them valuable for medical implants for neurological applications.

  2. Unsaturated fatty acids promote proliferation via ERK1/2 and Akt pathway in bovine mammary epithelial cells

    SciTech Connect

    Yonezawa, Tomo Haga, Satoshi; Kobayashi, Yosuke; Katoh, Kazuo; Obara, Yoshiaki

    2008-03-21

    GPR40 has recently been identified as a G protein-coupled cell-surface receptor for long-chain fatty acids (LCFAs). The mRNA of the bovine ortholog of GPR40 (bGPR40) was detected by RT-PCR in cloned bovine mammary epithelial cells (bMEC) and in the bovine mammary gland at various stages of lactation. Oleate and linoleate caused an increase in intracellular Ca{sup 2+} concentrations in these cells, and significantly reduced forskolin-induced cAMP concentrations. Phosphorylation of extracellular signal-regulated kinase (ERK) 1/2 and Akt kinase, which regulates cell proliferation and survival, was rapidly increased by oleate. Incubation with oleate and linoleate for 24 h significantly promoted cell proliferation. Moreover, in serum-free medium, oleate significantly stimulated cell proliferation during a 7-day culture. These results suggest that bGPR40 mediates LCFA signaling in mammary epithelial cells and thereby plays an important role in cell proliferation and survival.

  3. Perfluorooctanoic acid induces gene promoter hypermethylation of glutathione-S-transferase Pi in human liver L02 cells.

    PubMed

    Tian, Meiping; Peng, Siyuan; Martin, Francis L; Zhang, Jie; Liu, Liangpo; Wang, Zhanlin; Dong, Sijun; Shen, Heqing

    2012-06-14

    Perfluorooctanoic acid (PFOA) is one of the most commonly used perfluorinated compounds. Being a persistent environmental pollutant, it can accumulate in human tissues via various exposure routes. PFOA may interfere in a toxic fashion on the immune system, liver, development, and endocrine systems. In utero human exposure had been associated with cord serum global DNA hypomethylation. In light of this, we investigated possible PFOA-induced DNA methylation alterations in L02 cells in order to shed light into its epigenetic-mediated mechanisms of toxicity in human liver. L02 cells were exposed to 5, 10, 25, 50 or 100 mg/L PFOA for 72h. Global DNA methylation levels were determined by LC/ESI-MS, glutathione-S-transferase Pi (GSTP) gene promoter DNA methylation was investigated by methylation-specific polymerase chain reaction (PCR) with bisulfite sequencing, and consequent mRNA expression levels were measured with quantitative real-time reverse transcriptase PCR. A dose-related increase of GSTP promoter methylation at the transcription factor specificity protein 1 (SP1) binding site was observed. However, PFOA did not significantly influence global DNA methylation; nor did it markedly alter the promoter gene methylation of p16 (cyclin-dependent kinase inhibitor 2A), ERα (estrogen receptor α) or PRB (progesterone receptor B). In addition, PFOA significantly elevated mRNA transcript levels of DNMT3A (which mediates de novo DNA methylation), Acox (lipid metabolism) and p16 (cell apoptosis). Considering the role of GSTP in detoxification, aberrant methylation may be pivotal in PFOA-mediated toxicity response via the inhibition of SP1 binding to GSTP promoter. PMID:22425687

  4. Tousled kinase activator, gallic acid, promotes homologous recombinational repair and suppresses radiation cytotoxicity in salivary gland cells.

    PubMed

    Timiri Shanmugam, Prakash Srinivasan; Nair, Renjith Parameshwaran; De Benedetti, Arrigo; Caldito, Gloria; Abreo, Fleurette; Sunavala-Dossabhoy, Gulshan

    2016-04-01

    Accidental or medical radiation exposure of the salivary glands can gravely impact oral health. Previous studies have shown the importance of Tousled-like kinase 1 (TLK1) and its alternate start variant TLK1B in cell survival against genotoxic stresses. Through a high-throughput library screening of natural compounds, the phenolic phytochemical, gallic acid (GA), was identified as a modulator of TLK1/1B. This small molecule possesses anti-oxidant and free radical scavenging properties, but in this study, we report that in vitro it promotes survival of human salivary acinar cells, NS-SV-AC, through repair of ionizing radiation damage. Irradiated cells treated with GA show improved clonogenic survival compared to untreated controls. And, analyses of DNA repair kinetics by alkaline single-cell gel electrophoresis and γ-H2AX foci immunofluorescence indicate rapid resolution of DNA breaks in drug-treated cells. Study of DR-GFP transgene repair indicates GA facilitates homologous recombinational repair to establish a functional GFP gene. In contrast, inactivation of TLK1 or its shRNA knockdown suppressed resolution of radiation-induced DNA tails in NS-SV-AC, and homology directed repair in DR-GFP cells. Consistent with our results in culture, animals treated with GA after exposure to fractionated radiation showed better preservation of salivary function compared to saline-treated animals. Our results suggest that GA-mediated transient modulation of TLK1 activity promotes DNA repair and suppresses radiation cytoxicity in salivary gland cells.

  5. CAP-D3 Promotes Bacterial Clearance in Human Intestinal Epithelial Cells by Repressing Expression of Amino Acid Transporters

    PubMed Central

    Kemp, Jacqueline R.; Nickerson, Kourtney P.; Deutschman, Emily; Kim, Yeojung; West, Gail; Sadler, Tammy; Stylianou, Eleni; Krokowski, Dawid; Hatzoglou, Maria; de la Motte, Carol; Rubin, Brian P.; Fiocchi, Claudio

    2015-01-01

    BACKGROUND & AIMS Defects in colonic epithelial barrier defenses are associated with ulcerative colitis (UC). The proteins that regulate bacterial clearance in the colonic epithelium have not been completely identified. The chromosome-associated protein D3 (dCAP-D3), regulates responses to bacterial infection. We examined whether CAP-D3 promotes bacterial clearance in human colonic epithelium. METHODS Clearance of Salmonella or adherent-invasive Escherichia coli LF82 was assessed by gentamycin protection assays in HT-29 and Caco-2 cells expressing small hairpin RNAs against CAP-D3. We used immunoblot assays to measure levels of CAP-D3 in colonic epithelial cells from patients with UC and healthy individuals (controls). RNA sequencing identified genes activated by CAP-D3. We analyzed the roles of CAP-D3 target genes in bacterial clearance using gentamycin protection and immunofluorescence assays and studies with pharmacologic inhibitors. RESULTS CAP-D3 expression was reduced in colonic epithelial cells from patients with active UC. Reduced CAP-D3 expression decreased autophagy and impaired intracellular bacterial clearance by HT-29 and Caco-2 colonic epithelial cells. Lower levels of CAP-D3 increased transcription of genes encoding SLC7A5 and SLC3A2, whose products heterodimerize to form an amino acid transporter in HT-29 cells following bacterial infection; levels of SLC7A5–SLC3A2 were increased in tissues from patients with UC, compared with controls. Reduced CAP-D3 in HT-29 cells resulted in earlier recruitment of SLC7A5 to Salmonella-containing vacuoles, increased activity of mTORC1, and increased survival of bacteria. Inhibition of SLC7A5–SLC3A2 or mTORC1 activity rescued the bacterial clearance defects of CAP-D3– deficient cells. CONCLUSIONS CAP-D3 downregulates transcription of genes that encode amino acid transporters (SLC7A5 and SLC3A2) to promote bacterial autophagy by colon epithelial cells. Levels of CAP-D3 protein are reduced in patients with

  6. BCAT1 promotes cell proliferation through amino acid catabolism in gliomas carrying wild-type IDH1

    PubMed Central

    Park, Yoon Jung; Wang, Wei; Schlotter, Magdalena; Lindroth, Anders M; Pleier, Sabrina V; Bai, Alfa H C; Karra, Daniela; Piro, Rosario M; Felsberg, Jörg; Addington, Adele; Lemke, Dieter; Weibrecht, Irene; Hovestadt, Volker; Rolli, Claudio G; Campos, Benito; Turcan, Sevin; Sturm, Dominik; Witt, Hendrik; Chan, Timothy A; Herold-Mende, Christel; Kemkemer, Ralf; König, Rainer; Schmidt, Kathrin; Hull, William-Edmund; Pfister, Stefan M; Jugold, Manfred; Hutson, Susan M; Plass, Christoph; Okun, Jürgen G; Reifenberger, Guido; Lichter, Peter; Radlwimmer, Bernhard

    2016-01-01

    Here we show that glioblastoma express high levels of branched-chain amino acid transaminase 1 (BCAT1), the enzyme that initiates the catabolism of branched-chain amino acids (BCAAs). Expression of BCAT1 was exclusive to tumors carrying wild-type isocitrate dehydrogenase 1 (IDH1) and IDH2 genes and was highly correlated with methylation patterns in the BCAT1 promoter region. BCAT1 expression was dependent on the concentration of α-ketoglutarate substrate in glioma cell lines and could be suppressed by ectopic overexpression of mutant IDH1 in immortalized human astrocytes, providing a link between IDH1 function and BCAT1 expression. Suppression of BCAT1 in glioma cell lines blocked the excretion of glutamate and led to reduced proliferation and invasiveness in vitro, as well as significant decreases in tumor growth in a glioblastoma xenograft model. These findings suggest a central role for BCAT1 in glioma pathogenesis, making BCAT1 and BCAA metabolism attractive targets for the development of targeted therapeutic approaches to treat patients with glioblastoma. PMID:23793099

  7. Colon cancer cell apoptosis is induced by combined exposure to the n-3 fatty acid docosahexaenoic acid and butyrate through promoter methylation.

    PubMed

    Cho, Youngmi; Turner, Nancy D; Davidson, Laurie A; Chapkin, Robert S; Carroll, Raymond J; Lupton, Joanne R

    2014-03-01

    DNA methylation and histone acetylation contribute to the transcriptional regulation of genes involved in apoptosis. We have demonstrated that docosahexaenoic acid (DHA, 22:6 n-3) and butyrate enhance colonocyte apoptosis. To determine if DHA and/or butyrate elevate apoptosis through epigenetic mechanisms thereby restoring the transcription of apoptosis-related genes, we examined global methylation; gene-specific promoter methylation of 24 apoptosis-related genes; transcription levels of Cideb, Dapk1, and Tnfrsf25; and global histone acetylation in the HCT-116 colon cancer cell line. Cells were treated with combinations of (50 µM) DHA or linoleic acid (18:2 n-6), (5 mM) butyrate or an inhibitor of DNA methyltransferases, and 5-aza-2'-deoxycytidine (5-Aza-dC, 2 µM). Among highly methylated genes, the combination of DHA and butyrate significantly reduced methylation of the proapoptotic Bcl2l11, Cideb, Dapk1, Ltbr, and Tnfrsf25 genes compared to untreated control cells. DHA treatment reduced the methylation of Cideb, Dapk1, and Tnfrsf25. These data suggest that the induction of apoptosis by DHA and butyrate is mediated, in part, through changes in the methylation state of apoptosis-related genes.

  8. Ferulic acid inhibits proliferation and promotes apoptosis via blockage of PI3K/Akt pathway in osteosarcoma cell.

    PubMed

    Wang, Ting; Gong, Xia; Jiang, Rong; Li, Hongzhong; Du, Weimin; Kuang, Ge

    2016-01-01

    Ferulic acid, a ubiquitous phenolic acid abundant in corn, wheat and flax, has potent anti-tumor effect in various cancer cell lines. However, the anti-tumor effect of ferulic acid on osteosarcoma remains unclear. Therefore, we conduct current study to examine the effect of ferulic acid on osteosarcoma cells and explore the underlying mechanisms. In present study, ferulic acid inhibited proliferation and induced apoptosis in both 143B and MG63 osteosarcoma cells dose-dependently, indicated by MTT assay and Annexin V-FITC apoptosis detection. Additionally, ferulic acid induced G0/G1 phase arrest and down-regulated the expression of cell cycle-related protein, CDK 2, CDK 4, CDK 6, confirmed by flow cytometry assay and western blotting. Moreover, ferulic acid upregulated Bax, downregulated Bcl-2, and subsequently enhanced caspase-3 activity. More importantly, ferulic acid dose-dependently inhibited PI3K/Akt activation. Using adenoviruses expressing active Akt, the anti-proliferation and pro-apoptosis of ferulic acid were reverted. Our results demonstrated that ferulic acid might inhibit proliferation and induce apoptosis via inhibiting PI3K/Akt pathway in osteosarcoma cells. Ferulic acid is a novel therapeutic agent for osteosarcoma. PMID:27158383

  9. Photoelectrocatalytic oxidation of glucose at a ruthenium complex modified titanium dioxide electrode promoted by uric acid and ascorbic acid for photoelectrochemical fuel cells

    NASA Astrophysics Data System (ADS)

    Lu, Shuo-Jian; Ji, Shi-Bo; Liu, Jun-Chen; Li, Hong; Li, Wei-Shan

    2015-01-01

    The simultaneous presence of uric acid (UA) and ascorbic acid (AA) is first found to largely promote the photoelectrocatalytic oxidation of glucose (GLU) at an indium-tin oxide (ITO) or TiO2 nanoparticles/ITO electrode modified with [Ru(tatp)3]2+ (tatp = 1,4,8,9-tetra-aza-triphenylene) possessing good redox activity and nanoparticle size distribution. A well-defined electrocatalytic peak for GLU oxidation is shown at 0.265 V (vs. SCE) under approximate physiological conditions upon incorporation of UA and AA. The [Ru(tatp)3]2+/ITO electrode exhibits attractive amperometric oxidation responses towards GLU, UA and AA, while controlled potentiostatically at 0.3 V, 0.7 V and 1.0 V, respectively, indicating high sensitivity and excellent reproducibility. On basis of the photoelectrocatalysis of [Ru(tatp)3]2+/TiO2/ITO anode, a GLU concentration-dependent photoelectrochemical fuel cell vs. SCE is elaborately assembled. The proposed free-enzyme photoelectrochemical fuel cell employing 0.1 M GLU associated with 0.01 M UA and 0.01 M AA as fuel shows open-circuit photovoltage of 0.608 V, short-circuit photocurrent density of 124.5 μA cm-2 and maximum power density of 21.75 μW cm-2 at 0.455 V, fill factor of 0.32 and photoenergy conversion efficiency of 36.65%, respectively.

  10. Promotion of expression of interferon-stimulated genes in U937 monocytic cells by HIV RNAs, measured using stable isotope labeling with amino acids in cell culture (SILAC).

    PubMed

    Li, Yulan; Wen, Bin; Chen, Ran; Jiang, Feng; Zhao, Xiaofang; Deng, Xin

    2015-05-01

    Type I interferon (IFN) exerts strong antiviral activity, particularly against human immunodeficiency virus (HIV), and although several viral proteins have been shown to deregulate IFN induction, little is known about the induction of type I IFNs by HIV RNAs. In the present study, we used the stable isotope labeling with amino acids in cell culture (SILAC) method to determine the proteomic profile in U937 monocytic cells after transfection with viral RNA of HIV. We then used a western blot assay to validate the proteomic results. It was revealed by the SILAC method that there were 1624 non-redundant peptides with quantitative information and 281 proteins with quantitative information in the HIV-RNA-transfected U937 cells when compared to cells transfected with control RNA. In particular, 6, 8 or 12 hours post-transfection, HIV RNA transfection promoted the expression of such interferon stimulated genes (ISGs) as interferon-induced proteins with tetratricopeptide repeats (IFITs), interferon-induced transmembrane proteins (IFITMs), interferon-induced gene 15 protein (ISG15), myxovirus (influenza virus) resistance protein 1 (MX1), and interferon-induced guanylate-binding protein 1 (GBP1), and this was confirmed by western blot assay. In conclusion, HIV RNA is a strong stimulator of IFNs, promoting the expression of such ISGs as IFITs, IFITMs, ISG15, MX1 and GBP1.

  11. Melatonin promotes Bax sequestration to mitochondria reducing cell susceptibility to apoptosis via the lipoxygenase metabolite 5-hydroxyeicosatetraenoic acid.

    PubMed

    Radogna, F; Albertini, M C; De Nicola, M; Diederich, M; Bejarano, I; Ghibelli, L

    2015-03-01

    Extra-neurological functions of melatonin include control of the immune system and modulation of apoptosis. We previously showed that melatonin inhibits the intrinsic apoptotic pathway in leukocytes via stimulation of high affinity MT1/MT2 receptors, thereby promoting re-localization of the anti-apoptotic Bcl-2 protein to mitochondria. Here we show that Bcl-2 sequesters pro-apoptotic Bax into mitochondria in an inactive form after melatonin treatment, thus reducing cell propensity to apoptosis. Bax translocation and the anti-apoptotic effect of melatonin are strictly dependent on the presence of Bcl-2, and on the 5-lipoxygenase (5-LOX) metabolite 5-hydroxyeicosatetraenoic acid (5-HETE), which we have previously shown to be produced as a consequence of melatonin binding to its low affinity target calmodulin. Therefore, the anti-apoptotic effect of melatonin requires the simultaneous, independent interaction with high (MT1/MT2) and low (calmodulin) affinity targets, eliciting two independent signal transduction pathways converging into Bax sequestration and inactivation. MT1/MT2 vs. lipoxygenase pathways are activated by 10(-9) vs. 10(-5)M melatonin, respectively; the anti-apoptotic effect of melatonin is achieved at 10(-5)M, but drops to 10(-9)M upon addition of exogenous 5-HETE, revealing that lipoxygenase activation is the rate-limiting pathway. Therefore, in areas of inflammation with increased 5-HETE levels, physiological nanomolar concentrations of melatonin may suffice to maintain leukocyte viability.

  12. Transcriptional regulation of human mucin MUC4 by bile acids in oesophageal cancer cells is promoter-dependent and involves activation of the phosphatidylinositol 3-kinase signalling pathway.

    PubMed Central

    Mariette, Christophe; Perrais, Michaël; Leteurtre, Emmanuelle; Jonckheere, Nicolas; Hémon, Brigitte; Pigny, Pascal; Batra, Surinder; Aubert, Jean-Pierre; Triboulet, Jean-Pierre; Van Seuningen, Isabelle

    2004-01-01

    Abnormal gastro-oesophageal reflux and bile acids have been linked to the presence of Barrett's oesophageal premalignant lesion associated with an increase in mucin-producing goblet cells and MUC4 mucin gene overexpression. However, the molecular mechanisms underlying the regulation of MUC4 by bile acids are unknown. Since total bile is a complex mixture, we undertook to identify which bile acids are responsible for MUC4 up-regulation by using a wide panel of bile acids and their conjugates. MUC4 apomucin expression was studied by immunohistochemistry both in patient biopsies and OE33 oesophageal cancer cell line. MUC4 mRNA levels and promoter regulation were studied by reverse transcriptase-PCR and transient transfection assays respectively. We show that among the bile acids tested, taurocholic, taurodeoxycholic, taurochenodeoxycholic and glycocholic acids and sodium glycocholate are strong activators of MUC4 expression and that this regulation occurs at the transcriptional level. By using specific pharmacological inhibitors of mitogen-activated protein kinase, phosphatidylinositol 3-kinase, protein kinase A and protein kinase C, we demonstrate that bile acid-mediated up-regulation of MUC4 is promoter-specific and mainly involves activation of phosphatidylinositol 3-kinase. This new mechanism of regulation of MUC4 mucin gene points out an important role for bile acids as key molecules in targeting MUC4 overexpression in early stages of oesophageal carcinogenesis. PMID:14583090

  13. Delivery of iPS-NPCs to the Stroke Cavity within a Hyaluronic Acid Matrix Promotes the Differentiation of Transplanted Cells

    PubMed Central

    Lam, Jonathan; Lowry, William E.; Carmichael, S. Thomas; Segura, Tatiana

    2015-01-01

    Stroke is the leading cause of adult disability with ~80% being ischemic. Stem cell transplantation has been shown to improve functional recovery. However, the overall survival and differentiation of these cells is still low. The infarct cavity is an ideal location for transplantation as it is directly adjacent to the highly plastic peri-infarct region. Direct transplantation of cells near the infarct cavity has resulted in low cell viability. Here we deliver neural progenitor cells derived from induce pluripotent stem cells (iPS-NPC) to the infarct cavity of stroked mice encapsulated in a hyaluronic acid hydrogel matrix to protect the cells. To improve the overall viability of transplanted cells, each step of the transplantation process was optimized. Hydrogel mechanics and cell injection parameters were investigated to determine their effects on the inflammatory response of the brain and cell viability, respectively. Using parameters that balanced the desire to keep surgery invasiveness minimal and cell viability high, iPS-NPCs were transplanted to the stroke cavity of mice encapsulated in buffer or the hydrogel. While the hydrogel did not promote stem cell survival one week post-transplantation, it did promote differentiation of the neural progenitor cells to neuroblasts. PMID:26213530

  14. Carnosic acid, a component of rosemary (Rosmarinus officinalis L.), promotes synthesis of nerve growth factor in T98G human glioblastoma cells.

    PubMed

    Kosaka, Kunio; Yokoi, Toshio

    2003-11-01

    Nerve growth factor (NGF) is a factor vital for the growth and functional maintenance of nerve tissue. The authors found that a rosemary (Rosmarinus officinalis L.) extract enhanced the production of NGF in T98G human glioblastoma cells. Furthermore, the results indicated that carnosic acid and carnosol, which are major components of the rosemary extract, were able to promote markedly enhanced synthesis of NGF. PMID:14600414

  15. Inhibitory effect of α-lipoic acid on thioacetamide-induced tumor promotion through suppression of inflammatory cell responses in a two-stage hepatocarcinogenesis model in rats.

    PubMed

    Fujii, Yuta; Segawa, Risa; Kimura, Masayuki; Wang, Liyun; Ishii, Yuji; Yamamoto, Ryuichi; Morita, Reiko; Mitsumori, Kunitoshi; Shibutani, Makoto

    2013-09-25

    To investigate the protective effect of α-lipoic acid (a-LA) on the hepatocarcinogenic process promoted by thioacetamide (TAA), we used a two-stage liver carcinogenesis model in N-diethylnitrosamine (DEN)-initiated and TAA-promoted rats. We examined the modifying effect of co-administered a-LA on the liver tissue environment surrounding preneoplastic hepatocellular lesions, with particular focus on hepatic macrophages and the mechanism behind the decrease in apoptosis of cells surrounding preneoplastic hepatocellular lesions during the early stages of hepatocellular tumor promotion. TAA increased the number and area of glutathione S-transferase placental form (GST-P)(+) liver cell foci and the numbers of proliferating and apoptotic cells in the liver. Co-administration with a-LA suppressed these effects. TAA also increased the numbers of ED2(+), cyclooxygenase-2(+), and heme oxygenase-1(+) hepatic macrophages as well as the number of CD3(+) lymphocytes. These effects were also suppressed by a-LA. Transcript levels of some inflammation-related genes were upregulated by TAA and downregulated by a-LA in real-time RT-PCR analysis. Outside the GST-P(+) foci, a-LA reduced the numbers of apoptotic cells, active caspase-8(+) cells and death receptor (DR)-5(+) cells. These results suggest that hepatic macrophages producing proinflammatory factors may be activated in TAA-induced tumor promotion. a-LA may suppress tumor-promoting activity by suppressing the activation of these macrophages and the subsequent inflammatory responses. Furthermore, a-LA may suppress tumor-promoting activity by suppressing the DR5-mediated extrinsic pathway of apoptosis and the subsequent regeneration of liver cells outside GST-P(+) foci.

  16. TRIM32 promotes retinoic acid receptor {alpha}-mediated differentiation in human promyelogenous leukemic cell line HL60

    SciTech Connect

    Sato, Tomonobu; Okumura, Fumihiko; Iguchi, Akihiro; Ariga, Tadashi; Hatakeyama, Shigetsugu

    2012-01-06

    Highlights: Black-Right-Pointing-Pointer TRIM32 enhanced RAR{alpha}-mediated transcriptional activity even in the absence of RA. Black-Right-Pointing-Pointer TRIM32 stabilized RAR{alpha} in the human promyelogenous leukemic cell line HL60. Black-Right-Pointing-Pointer Overexpression of TRIM32 in HL60 cells induced granulocytic differentiation. Black-Right-Pointing-Pointer TRIM32 may function as a coactivator for RAR{alpha}-mediated transcription in APL cells. -- Abstract: Ubiquitination, one of the posttranslational modifications, appears to be involved in the transcriptional activity of nuclear receptors including retinoic acid receptor {alpha} (RAR{alpha}). We previously reported that an E3 ubiquitin ligase, TRIM32, interacts with several important proteins including RAR{alpha} and enhances transcriptional activity of RAR{alpha} in mouse neuroblastoma cells and embryonal carcinoma cells. Retinoic acid (RA), which acts as a ligand to nuclear receptors including RAR{alpha}, plays crucial roles in development, differentiation, cell cycles and apoptosis. In this study, we found that TRIM32 enhances RAR{alpha}-mediated transcriptional activity even in the absence of RA and stabilizes RAR{alpha} in the human promyelogenous leukemic cell line HL60. Moreover, we found that overexpression of TRIM32 in HL60 cells suppresses cellular proliferation and induces granulocytic differentiation even in the absence of RA. These findings suggest that TRIM32 functions as one of the coactivators for RAR{alpha}-mediated transcription in acute promyelogenous leukemia (APL) cells, and thus TRIM32 may become a potentially therapeutic target for APL.

  17. Activation of Transmembrane Bile Acid Receptor TGR5 Modulates Pancreatic Islet α Cells to Promote Glucose Homeostasis.

    PubMed

    Kumar, Divya P; Asgharpour, Amon; Mirshahi, Faridoddin; Park, So Hyun; Liu, Sichen; Imai, Yumi; Nadler, Jerry L; Grider, John R; Murthy, Karnam S; Sanyal, Arun J

    2016-03-25

    The physiological role of the TGR5 receptor in the pancreas is not fully understood. We previously showed that activation of TGR5 in pancreatic β cells by bile acids induces insulin secretion. Glucagon released from pancreatic α cells and glucagon-like peptide 1 (GLP-1) released from intestinal L cells regulate insulin secretion. Both glucagon and GLP-1 are derived from alternate splicing of a common precursor, proglucagon by PC2 and PC1, respectively. We investigated whether TGR5 activation in pancreatic α cells enhances hyperglycemia-induced PC1 expression thereby releasing GLP-1, which in turn increases β cell mass and function in a paracrine manner. TGR5 activation augmented a hyperglycemia-induced switch from glucagon to GLP-1 synthesis in human and mouse islet α cells by GS/cAMP/PKA/cAMP-response element-binding protein-dependent activation of PC1. Furthermore, TGR5-induced GLP-1 release from α cells was via an Epac-mediated PKA-independent mechanism. Administration of the TGR5 agonist, INT-777, to db/db mice attenuated the increase in body weight and improved glucose tolerance and insulin sensitivity. INT-777 augmented PC1 expression in α cells and stimulated GLP-1 release from islets of db/db mice compared with control. INT-777 also increased pancreatic β cell proliferation and insulin synthesis. The effect of TGR5-mediated GLP-1 from α cells on insulin release from islets could be blocked by GLP-1 receptor antagonist. These results suggest that TGR5 activation mediates cross-talk between α and β cells by switching from glucagon to GLP-1 to restore β cell mass and function under hyperglycemic conditions. Thus, INT-777-mediated TGR5 activation could be leveraged as a novel way to treat type 2 diabetes mellitus. PMID:26757816

  18. Residual Host Cell Protein Promotes Polysorbate 20 Degradation in a Sulfatase Drug Product Leading to Free Fatty Acid Particles.

    PubMed

    Dixit, Nitin; Salamat-Miller, Nazila; Salinas, Paul A; Taylor, Katherine D; Basu, Sujit K

    2016-05-01

    This study investigated the root cause behind an observed free fatty acid particle formation and resulting Polysorbate 20 (PS20) loss for a sulfatase drug product upon long-term storage at 5 ± 3°C. Reversed- phase chromatography with mass spectrometric analysis as well as charged aerosol detection was used to characterize the peaks associated with the intact and degraded PS20. Additionally, a proteomics study was undertaken to identify the residual host cell proteins in the sulfatase drug substance. PS20 stability studies were conducted in the presence of sulfatase, a sulfatase inhibitor, putative phospholipase B-like 2, and mock drug substance produced using a null cell line vector under experimental conditions optimized for PS20 degradation. This study provides the first published evidence where the residual host cell protein present in the drug substance was identified and experimentally shown to catalyze the breakdown of PS20 in a protein formulation over time, resulting in free fatty acid particles and PS20 loss. This study demonstrates the importance of early detection of potential impurities in the protein drug substance that may contribute to polysorbate degradation to make a judicious selection of the surfactant and its optimized concentration for the final drug product. PMID:27032893

  19. Cross-talk between lysophosphatidic acid receptor 1 and tropomyosin receptor kinase A promotes lung epithelial cell migration.

    PubMed

    Nan, Ling; Wei, Jianxin; Jacko, Anastasia M; Culley, Miranda K; Zhao, Jing; Natarajan, Viswanathan; Ma, Haichun; Zhao, Yutong

    2016-02-01

    Lysophosphatidic acid (LPA) is a bioactive lysophospholipid, which plays a crucial role in the regulation of cell proliferation, migration, and differentiation. LPA exerts its biological effects mainly through binding to cell-surface LPA receptors (LPA1-6), which belong to the G protein-coupled receptor (GPCR) family. Recent studies suggest that cross-talk between receptor tyrosine kinases (RTKs) and GPCRs modulates GPCRs-mediated signaling. Tropomyosin receptor kinase A (TrkA) is a RTK, which mediates nerve growth factor (NGF)-induced biological functions including cell migration in neuronal and non-neuronal cells. Here, we show LPA1 transactivation of TrkA in murine lung epithelial cells (MLE12). LPA induced tyrosine phosphorylation of TrkA in both time- and dose-dependent manners. Down-regulation of LPA1 by siRNA transfection attenuated LPA-induced phosphorylation of TrkA, suggesting a cross-talk between LPA1 and TrkA. To investigate the molecular regulation of the cross-talk, we focused on the interaction between LPA1 and TrkA. We found that LPA induced interaction between LPA1 and TrkA. The LPA1/TrkA complex was localized on the plasma membrane and in the cytoplasm. The C-terminus of LPA1 was identified as the binding site for TrkA. Inhibition of TrkA attenuated LPA-induced phosphorylation of TrkA and LPA1 internalization, as well as lung epithelial cell migration. These studies provide a molecular mechanism for the transactivation of TrkA by LPA, and suggest that the cross-talk between LPA1 and TrkA regulates LPA-induced receptor internalization and lung epithelial cell migration. PMID:26597701

  20. Thyroid hormone responsive (THRSP) promotes the synthesis of medium-chain fatty acids in goat mammary epithelial cells.

    PubMed

    Yao, D W; Luo, J; He, Q Y; Wu, M; Shi, H B; Wang, H; Wang, M; Xu, H F; Loor, J J

    2016-04-01

    In nonruminants, thyroid hormone responsive (THRSP) is a crucial protein for cellular de novo lipogenesis. However, the role of THRSP in regulating the synthesis of milk fatty acid composition in goat mammary gland remains unknown. In the present study, we compared gene expression of THRSP among different goat tissues. Results revealed that THRSP had the highest expression in subcutaneous fat, and expression was higher during lactation compared with the dry period. Overexpression of THRSP upregulated the expression of fatty acid synthase (FASN), stearoyl-coenzyme A desaturase 1 (SCD1), diacylglycerol acyltransferase 2 (DGAT2), and glycerol-3-phosphate acyltransferase (GPAM) in goat mammary epithelial cells. In contrast, overexpression of THRSP led to downregulation of thrombospondin receptor (CD36) and had no effect on the expression of acetyl-coenzyme A carboxylase α (ACACA) and sterol regulatory element binding transcription factor1 (SREBF1). In addition, overexpressing THRSP in vitro resulted in a significant increase in triacylglycerol (TAG) concentration and the concentrations of C12:0 and C14:0. Taken together, these results highlight an important role of THRSP in regulating lipogenesis in goat mammary epithelial cells. PMID:26851858

  1. Poly(γ-Glutamic Acid) as an Exogenous Promoter of Chondrogenic Differentiation of Human Mesenchymal Stem/Stromal Cells

    PubMed Central

    Antunes, Joana C.; Tsaryk, Roman; Gonçalves, Raquel M.; Pereira, Catarina Leite; Landes, Constantin; Brochhausen, Christoph; Ghanaati, Shahram

    2015-01-01

    Cartilage damage and/or aging effects can cause constant pain, which limits the patient's quality of life. Although different strategies have been proposed to enhance the limited regenerative capacity of cartilage tissue, the full production of native and functional cartilaginous extracellular matrix (ECM) has not yet been achieved. Poly(γ-glutamic acid) (γ-PGA), a naturally occurring polyamino acid, biodegradable into glutamate residues, has been explored for tissue regeneration. In this work, γ-PGA's ability to support the production of cartilaginous ECM by human bone marrow mesenchymal stem/stromal cells (MSCs) and nasal chondrocytes (NCs) was investigated. MSC and NC pellets were cultured in basal medium (BM), chondrogenic medium (CM), and CM-γ-PGA-supplemented medium (CM+γ-PGA) over a period of 21 days. Pellet size/shape was monitored with time. At 14 and 21 days of culture, the presence of sulfated glycosaminoglycans (sGAGs), type II collagen (Col II), Sox-9, aggrecan, type XI collagen (Col XI), type X collagen (Col X), calcium deposits, and type I collagen (Col I) was analyzed. After excluding γ-PGA's cytotoxicity, earlier cell condensation, higher sGAG content, Col II, Sox-9 (day 14), aggrecan, and Col X (day 14) production was observed in γ-PGA-supplemented MSC cultures, with no signs of mineralization or Col I. These effects were not evident with NCs. However, Sox-9 (at day 14) and Col X (at days 14 and 21) were increased, decreased, or absent, respectively. Overall, γ-PGA improved chondrogenic differentiation of MSCs, increasing ECM production earlier in culture. It is proposed that γ-PGA incorporation in novel biomaterials has a beneficial impact on future approaches for cartilage regeneration. PMID:25760236

  2. Poly(γ-Glutamic Acid) as an Exogenous Promoter of Chondrogenic Differentiation of Human Mesenchymal Stem/Stromal Cells.

    PubMed

    Antunes, Joana C; Tsaryk, Roman; Gonçalves, Raquel M; Pereira, Catarina Leite; Landes, Constantin; Brochhausen, Christoph; Ghanaati, Shahram; Barbosa, Mário A; Kirkpatrick, C James

    2015-06-01

    Cartilage damage and/or aging effects can cause constant pain, which limits the patient's quality of life. Although different strategies have been proposed to enhance the limited regenerative capacity of cartilage tissue, the full production of native and functional cartilaginous extracellular matrix (ECM) has not yet been achieved. Poly(γ-glutamic acid) (γ-PGA), a naturally occurring polyamino acid, biodegradable into glutamate residues, has been explored for tissue regeneration. In this work, γ-PGA's ability to support the production of cartilaginous ECM by human bone marrow mesenchymal stem/stromal cells (MSCs) and nasal chondrocytes (NCs) was investigated. MSC and NC pellets were cultured in basal medium (BM), chondrogenic medium (CM), and CM-γ-PGA-supplemented medium (CM+γ-PGA) over a period of 21 days. Pellet size/shape was monitored with time. At 14 and 21 days of culture, the presence of sulfated glycosaminoglycans (sGAGs), type II collagen (Col II), Sox-9, aggrecan, type XI collagen (Col XI), type X collagen (Col X), calcium deposits, and type I collagen (Col I) was analyzed. After excluding γ-PGA's cytotoxicity, earlier cell condensation, higher sGAG content, Col II, Sox-9 (day 14), aggrecan, and Col X (day 14) production was observed in γ-PGA-supplemented MSC cultures, with no signs of mineralization or Col I. These effects were not evident with NCs. However, Sox-9 (at day 14) and Col X (at days 14 and 21) were increased, decreased, or absent, respectively. Overall, γ-PGA improved chondrogenic differentiation of MSCs, increasing ECM production earlier in culture. It is proposed that γ-PGA incorporation in novel biomaterials has a beneficial impact on future approaches for cartilage regeneration.

  3. The retinoic acid-metabolizing enzyme CYP26A1 upregulates fascin and promotes the malignant behavior of breast carcinoma cells.

    PubMed

    Osanai, Makoto; Lee, Gang-Hong

    2015-08-01

    The retinoic acid (RA)-metabolizing enzyme CYP26A1 has been shown to efficiently enhance the oncogenic potential of breast cancer, suggesting a potential oncogenic function. We previously demonstrated that CYP26A1 confers unique cell survival properties by modulating the expression of a variety of genes and identified a number of genes that drive the cells into the oncogenic state. Accumulating evidence suggested that fascin is overexpressed in various types of cancer, primarily leading to increased cell motility. Therefore, in the present study, we examined fascin, an actin-bundling protein, using immunohistochemical and SA-β-gal staining as well as TUNEL and colony forming assays. The results of the present study showed that the expression levels of fascin increased significantly in response to CYP26A1 overexpression and, conversely, treatment with all-trans RA downregulated the expression of fascin. In addition, primary breast carcinoma samples, particularly hormone receptor-negative carcinomas and CYP26A1-overexpressing cancers, expressed elevated levels of fascin. Notably, fascin contributed to the ability of breast carcinoma cells to escape premature senescence and exhibit enhanced cell apoptotic resistance, promoting anchorage-independent growth properties. Fascin also promoted cell motility and the invasiveness of CYP26A1-expressing breast carcinoma cells. These data suggest that fascin expression is modulated by the intracellular RA status regulated by the expression of CYP26A1 and plays a significant role in the malignant behavior of CYP26A1-expressing breast carcinoma cells. CYP26A1 exerts oncogenic functions during breast carcinogenesis. Therefore, CYP26A1-mediated oncogenic characteristics may be partially responsible for the elevated expression of fascin. PMID:26058854

  4. Excessive feedback of Cyp26a1 promotes cell non-autonomous loss of retinoic acid signaling

    PubMed Central

    Rydeen, Ariel; Voisin, Norine; D’Aniello, Enrico; Ravisankar, Padmapriyadarshini; Devignes, Claire-Sophie; Waxman, Joshua S.

    2015-01-01

    Teratogenic levels of retinoic acid (RA) signaling can cause seemingly contradictory phenotypes indicative of both increases and decreases of RA signaling. However, the mechanisms underlying these contradictory phenotypes are not completely understood. Here, we report that using a hyperactive RA receptor to enhance RA signaling in zebrafish embryos leads to defects associated with gain and loss of RA signaling. While the gain-of-function phenotypes arise from an initial increase in RA signaling, using genetic epistasis analysis we found that the loss-of-function phenotypes result from a clearing of embryonic RA that requires a rapid and dramatic increase in cyp26a1 expression. Thus, the sensitivity of cyp26a1 expression to increased RA signaling causes an overcompensation of negative feedback and loss of embryonic RA signaling. Additionally, we used blastula transplantation experiments to test if Cyp26a1, despite its cellular localization, can limit RA exposure to neighboring cells. We find that enhanced Cyp26a1 expression limits RA signaling in the local environment, thus providing the first direct evidence that Cyp26 enzymes can have cell non-autonomous consequences on RA levels within tissues. Therefore, our results provide novel insights into the teratogenic mechanisms of RA signaling and the cellular mechanisms by which Cyp26a1 expression can shape a RA gradient. PMID:26116175

  5. Up-regulating the abscisic acid inactivation gene ZmABA8ox1b contributes to seed germination heterosis by promoting cell expansion

    PubMed Central

    Li, Yangyang; Wang, Cheng; Liu, Xinye; Song, Jian; Li, Hongjian; Sui, Zhipeng; Zhang, Ming; Fang, Shuang; Chu, Jinfang; Xin, Mingming; Xie, Chaojie; Zhang, Yirong; Sun, Qixin; Ni, Zhongfu

    2016-01-01

    Heterosis has been widely used in agriculture, but the underlying molecular principles are still largely unknown. During seed germination, we observed that maize (Zea mays) hybrid B73/Mo17 was less sensitive than its parental inbred lines to exogenous abscisic acid (ABA), and endogenous ABA content in hybrid embryos decreased more rapidly than in the parental inbred lines. ZmABA8ox1b, an ABA inactivation gene, was consistently more highly up-regulated in hybrid B73/Mo17 than in its parental inbred lines at early stages of seed germination. Moreover, ectopic expression of ZmABA8ox1b obviously promoted seed germination in Arabidopsis. Remarkably, microscopic observation revealed that cell expansion played a major role in the ABA-mediated maize seed germination heterosis, which could be attributed to the altered expression of cell wall-related genes. PMID:27034328

  6. Up-regulating the abscisic acid inactivation gene ZmABA8ox1b contributes to seed germination heterosis by promoting cell expansion.

    PubMed

    Li, Yangyang; Wang, Cheng; Liu, Xinye; Song, Jian; Li, Hongjian; Sui, Zhipeng; Zhang, Ming; Fang, Shuang; Chu, Jinfang; Xin, Mingming; Xie, Chaojie; Zhang, Yirong; Sun, Qixin; Ni, Zhongfu

    2016-04-01

    Heterosis has been widely used in agriculture, but the underlying molecular principles are still largely unknown. During seed germination, we observed that maize (Zea mays) hybrid B73/Mo17 was less sensitive than its parental inbred lines to exogenous abscisic acid (ABA), and endogenous ABA content in hybrid embryos decreased more rapidly than in the parental inbred lines. ZmABA8ox1b, an ABA inactivation gene, was consistently more highly up-regulated in hybrid B73/Mo17 than in its parental inbred lines at early stages of seed germination. Moreover, ectopic expression of ZmABA8ox1b obviously promoted seed germination in Arabidopsis Remarkably, microscopic observation revealed that cell expansion played a major role in the ABA-mediated maize seed germination heterosis, which could be attributed to the altered expression of cell wall-related genes.

  7. Sialic Acid-Binding Immunoglobulin-like Lectin G Promotes Atherosclerosis and Liver Inflammation by Suppressing the Protective Functions of B-1 Cells

    PubMed Central

    Gruber, Sabrina; Hendrikx, Tim; Tsiantoulas, Dimitrios; Ozsvar-Kozma, Maria; Göderle, Laura; Mallat, Ziad; Witztum, Joseph L.; Shiri-Sverdlov, Ronit; Nitschke, Lars; Binder, Christoph J.

    2016-01-01

    Summary Atherosclerosis is initiated and sustained by hypercholesterolemia, which results in the generation of oxidized LDL (OxLDL) and other metabolic byproducts that trigger inflammation. Specific immune responses have been shown to modulate the inflammatory response during atherogenesis. The sialic acid-binding immunoglobulin-like lectin G (Siglec-G) is a negative regulator of the functions of several immune cells, including myeloid cells and B-1 cells. Here, we show that deficiency of Siglec-G in atherosclerosis-prone mice inhibits plaque formation and diet-induced hepatic inflammation. We further demonstrate that selective deficiency of Siglec-G in B cells alone is sufficient to mediate these effects. Levels of B-1 cell-derived natural IgM with specificity for OxLDL were significantly increased in the plasma and peritoneal cavity of Siglec-G-deficient mice. Consistent with the neutralizing functions of OxLDL-specific IgM, Siglec-G-deficient mice were protected from OxLDL-induced sterile inflammation. Thus, Siglec-G promotes atherosclerosis and hepatic inflammation by suppressing protective anti-inflammatory effector functions of B cells. PMID:26947073

  8. Schwann cells promote endothelial cell migration

    PubMed Central

    Ramos, Tiago; Ahmed, Maqsood; Wieringa, Paul; Moroni, Lorenzo

    2015-01-01

    Directed cell migration is a crucial orchestrated process in embryonic development, wound healing, and immune response. The underlying substrate can provide physical and/or chemical cues that promote directed cell migration. Here, using electrospinning we developed substrates of aligned poly(lactic-co-glycolic acid) nanofibres to study the influence of glial cells on endothelial cells (ECs) in a 3-dimensional (3D) co-culture model. ECs build blood vessels and regulate their plasticity in coordination with neurons. Likewise, neurons construct nerves and regulate their circuits in coordination with ECs. In our model, the neuro-vascular cross-talk was assessed using a direct co-culture model of human umbilical vein endothelial cells (HUVECs) and rat Schwann cells (rSCs). The effect of rSCs on ECs behavior was demonstrated by earlier and higher velocity values and genetic expression profiles different of those of HUVECs when seeded alone. We observed 2 different gene expression trends in the co-culture models: (i) a later gene expression of angiogenic factors, such as interleukin-8 (IL-8) and vascular endothelial growth factor (VEGF), and (ii) an higher gene expression of genes involved in actin filaments rearrangement, such as focal adhesion kinase (FAK), Mitogen-activated protein kinase-activated protein kinase 13 (MAPKAPK13), Vinculin (VCL), and Profilin (PROF). These results suggested that the higher ECs migration is mainly due to proteins involved in the actin filaments rearrangement and in the directed cell migration rather than the effect of angiogenic factors. This co-culture model provides an approach to enlighten the neurovascular interactions, with particular focus on endothelial cell migration. PMID:26491999

  9. Long interspersed nucleotide acid element-1 ORF-1 protein promotes proliferation and invasion of human colorectal cancer LoVo cells through enhancing ETS-1 activity.

    PubMed

    Li, M Y; Zhu, M; Feng, F; Cai, F Y; Fan, K C; Jiang, H; Wang, Z Q; Linghu, E Q

    2014-04-14

    The human proto-oncogene long interspersed nucleotide acid element-1 (LINE-1) open reading frame-1 protein (ORF-1p) is involved in the progress of several cancers. The transcription factor ETS-1 can mediate the transcription of some downstream genes that play specific roles in the regulation of cancerous cell invasion and metastasis. In this study, the effects of LINE-1 ORF-1p on ETS-1 activity and on the proliferation and invasion of human colorectal cancer LoVo cells were investigated. Results showed that the overexpression of LINE-1 ORF-1p enhanced the transcription of ETS-1 downstream genes and increased their protein levels, and downregulation of the LINE-1 ORF-1p level by small interfering RNA (siRNA) reduced the transcriptional activation of ETS-1. In addition, overexpression of LINE-1 ORF-1p promoted LoVo cell proliferation and anchor-independent growth, and a knockdown of the LINE-1 protein level by siRNA reduced the proliferation and anchor-independent growth ability of LoVo cells. In vivo data revealed that LINE-1 ORF-1p overexpression increased LoVo tumor growth in nude mice, whereas the siRNA knockdown of endogenous LINE-1 ORF-1p expression decreased LoVo cell growth in nude mice. Therefore, LINE- 1 ORF-1p could promote LoVo cell proliferation and invasion both in vitro and in vivo, indicating that it might be a useful molecular target for the treatment of human colorectal cancer.

  10. Mapping the response of human fibroblast growth factor 21 (FGF21) promoter to serum availability and lipoic acid in HepG2 hepatoma cells.

    PubMed

    Xia, Mengna; Erickson, Anjeza; Yi, Xiaohua; Moreau, Régis

    2016-03-01

    The hormone-like polypeptide, fibroblast growth factor 21 (FGF21), is a major modulator of lipid and glucose metabolism and an exploratory treatment strategy for obesity related metabolic disorders. The costs of recombinant FGF21 and mode of delivery by injection are important constraints to its wide therapeutic use. The stimulation of endogenous FGF21 production through diet is being explored as an alternative approach. To that end, we examined the mechanism(s) by which serum manipulation and lipoic acid (a dietary activator of FGF21) induce FGF21 in human hepatocellular carcinoma HepG2 cells. Serum withdrawal markedly induced FGF21 mRNA levels (88 fold) and FGF21 secreted in the media (19 fold). Lipoic acid induced FGF21 mRNA 7 fold above DMSO-treated control cells and FGF21 secretion 3 fold. These effects were several-fold greater than those of PPARα agonist, Wy14643, which failed to induce FGF21 above and beyond the induction seen with serum withdrawal. The use of transcription inhibitor, actinomycin D, revealed that de novo mRNA synthesis drives FGF21 secretion in response to serum starvation. Four previously unrecognized loci in FGF21 promoter were nucleosome depleted and enriched in acetylated histone H3 revealing their role as transcriptional enhancers and putative transcription factor binding sites. FGF21 did not accumulate to a significant degree in induced HepG2 cells, which secreted FGF21 time dependently in media. We conclude that lipoic acid cell signaling connects with the transcriptional upregulation of FGF21 and it may prove to be a safe and affordable means to stimulate FGF21 production. PMID:26691139

  11. Blueberry diet derived 3-(3-hydroxyphenyl) propionic acid (PPA) suppresses osteoblastic cell senescence to promote bone accretion in mice

    Technology Transfer Automated Retrieval System (TEKTRAN)

    A blueberry (BB) supplemented diet has been previously shown to significantly stimulate bone formation in rapidly growing male and female rodents. Phenolic acids (PAs) are metabolites derived from polyphenols found in fruits and vegetables as a result of the actions of gut bacteria, and the levels o...

  12. A novel coating of type IV collagen and hyaluronic acid on stent material-titanium for promoting smooth muscle cell contractile phenotype.

    PubMed

    Li, Jingan; Zhang, Kun; Chen, Huiqing; Liu, Tao; Yang, Ping; Zhao, Yuancong; Huang, Nan

    2014-05-01

    The method of stent implantation is currently considered an effective means of treating atherosclerosis. However, implanting of cardiovascular stent often leads to intimal breakage and hyperplasia. The phenomenon that vascular smooth muscle cells (SMCs) transform from contractile to synthetic phenotype becomes a serious obstacle to intimal recovery. To improve how SMCs transform from a synthetic to contractile phenotype, a technique of coimmobilization was used to form type IV collagen (CoIV) and hyaluronic acid (HA) coating on the widely used stent material, titanium (Ti). In this work, several bio-functional coatings made of CoIV/HA mixtures in different ratios were fabricated on the Ti surface. The quantitative characterization of CoIV showed that introducing HA could enhance the amount of the immobilized CoIV on the alkali activated Ti (TiOH) surface. The immunofluorescence staining results of myosin heavy chain (MHC) and DAPI showed that the coating of CoIV/HA in ratios of 200 μg/ml (M200) and 500 μg/ml (M500) also could promote SMCs expressing more contractile phenotype compared with TiOH/CoIV control samples, while the AO/PI staining results indicated that SMCs on the M200 and M500 samples showed less apoptosis ratio. Thus, we hope that this study can provide more helpful exploration and application for promoting the SMC contractile phenotype on the cardiovascular stents. PMID:24656374

  13. All-trans retinoic acid impairs the vasculogenic mimicry formation ability of U87 stem-like cells through promoting differentiation

    PubMed Central

    LING, GENG-QIANG; LIU, YI-JING; KE, YI-QUAN; CHEN, LEI; JIANG, XIAO-DAN; JIANG, CHUAN-LU; YE, WEI

    2015-01-01

    The poor therapeutic effect of traditional antiangiogenic therapy on glioblastoma multiforme (GBM) may be attributed to vasculogenic mimicry (VM), which was previously reported to be promoted by cancer stem-like cells (SLCs). All-trans retinoic acid (ATRA), a potent reagent which drives differentiation, was reported to be able to eradicate cancer SLCs in certain malignancies. The aim of the present study was to investigate the effects of ATRA on the VM formation ability of U87 glioblastoma SLCs. The expression of cancer SLC markers CD133 and nestin was detected using immunocytochemistry in order to identify U87 SLCs. In addition, the differentiation of these SLCs was observed through detecting the expression of glial fibrillary acidic protein (GFAP), β-tubulin III and galactosylceramidase (Galc) using immunofluorescent staining. The results showed that the expression levels of GFAP, β-tubulin III and Galc were upregulated following treatment with ATRA in a dose-dependent manner. Furthermore, ATRA significantly reduced the proliferation, invasiveness, tube formation and vascular endothelial growth factor (VEGF) secretion of U87 SLCs. In conclusion, the VM formation ability of SLCs was found to be negatively correlated with differentiation. These results therefore suggested that ATRA may serve as a promising novel agent for the treatment of GBM due to its role in reducing VM formation. PMID:25760394

  14. A Lewis acid-promoted Pinner reaction

    PubMed Central

    Pfaff, Dominik; Nemecek, Gregor

    2013-01-01

    Summary Carbonitriles and alcohols react in a Lewis acid-promoted Pinner reaction to carboxylic esters. Best results are obtained with two equivalents of trimethylsilyl triflate as Lewis acid. Good yields are achieved with primary alcohols and aliphatic or benzylic carbonitriles, but the straightforward synthesis of acrylates and benzoates starting with acrylonitrile and benzonitrile, respectively, is similarly possible. Phenols are not acylated under these reaction conditions. The method has been used for the first total synthesis of the natural product monaspilosin. In the reaction of benzyl alcohols variable amounts of amides are formed in a Ritter-type side reaction. PMID:23946857

  15. Altered Tumor-Cell Glycosylation Promotes Metastasis

    PubMed Central

    Häuselmann, Irina; Borsig, Lubor

    2014-01-01

    Malignant transformation of cells is associated with aberrant glycosylation presented on the cell-surface. Commonly observed changes in glycan structures during malignancy encompass aberrant expression and glycosylation of mucins; abnormal branching of N-glycans; and increased presence of sialic acid on proteins and glycolipids. Accumulating evidence supports the notion that the presence of certain glycan structures correlates with cancer progression by affecting tumor-cell invasiveness, ability to disseminate through the blood circulation and to metastasize in distant organs. During metastasis tumor-cell-derived glycans enable binding to cells in their microenvironment including endothelium and blood constituents through glycan-binding receptors – lectins. In this review, we will discuss current concepts how tumor-cell-derived glycans contribute to metastasis with the focus on three types of lectins: siglecs, galectins, and selectins. Siglecs are present on virtually all hematopoietic cells and usually negatively regulate immune responses. Galectins are mostly expressed by tumor cells and support tumor-cell survival. Selectins are vascular adhesion receptors that promote tumor-cell dissemination. All lectins facilitate interactions within the tumor microenvironment and thereby promote cancer progression. The identification of mechanisms how tumor glycans contribute to metastasis may help to improve diagnosis, prognosis, and aid to develop clinical strategies to prevent metastasis. PMID:24592356

  16. CuII ions and the stilbene-chroman hybrid with a catechol moiety synergistically induced DNA damage, and cell cycle arrest and apoptosis of HepG2 cells: an interesting acid/base-promoted prooxidant reaction.

    PubMed

    Liu, Guo-Yun; Yang, Jie; Dai, Fang; Yan, Wen-Jing; Wang, Qi; Li, Xiu-Zhuang; Ding, De-Jun; Cao, Xiao-Yan; Zhou, Bo

    2012-08-27

    Development of potential cancer treatment strategies by using an exogenous reactive oxygen species (ROS)-generating agent (prooxidant) or redox intervention, has attracted much interest. One effective ROS generation method is to construct a prooxidant system by polyphenolic compounds and Cu(II) ions. This work demonstrates that Cu(II) and the stilbene-chroman hybrid with a catechol moiety could synergistically induce pBR322 plasmid DNA damage, as well as cell cycle arrest and apoptosis of HepG2 cells. Additionally, an interesting acid/base-promoted prooxidant reaction was found. The detailed chemical mechanisms for the reaction of the hybrid with Cu(II) in acid, neutral and base solutions are proposed based on UV/Vis spectral changes and identification of the related oxidative intermediates and products.

  17. Dose Dependent Activation of Retinoic Acid-Inducible Gene-I Promotes Both Proliferation and Apoptosis Signals in Human Head and Neck Squamous Cell Carcinoma

    PubMed Central

    Yan, Ming; Zhu, Chao; Ye, Weimin; Zhu, Hanguang; Chen, Wantao; Zhang, Chenping; Zhang, Zhiyuan

    2013-01-01

    The retinoic-acid-inducible gene (RIG)-like receptor (RLR) family proteins are major pathogen reorganization receptors (PRR) responsible for detection of viral RNA, which initiates antiviral response. Here, we evaluated the functional role of one RLR family member, RIG-I, in human head and neck squamous cell carcinoma (HNSCC). RIG-I is abundantly expressed both in poorly-differentiated primary cancer and lymph node metastasis, but not in normal adjacent tissues. Activation of RIG-I by transfection with low dose of 5′-triphosphate RNA (3p-RNA) induces low levels of interferon and proinflammatory cytokines and promotes NF-κB- and Akt-dependent cell proliferation, migration and invasion. In contrast, activation of RIG-I by a high dose of 3p-RNA induces robust mitochondria-derived apoptosis accompanied by decreased activation of Akt, which is independent of the interferon and TNFα receptor, but can be rescued by over-expression of constitutively active Akt. Furthermore, co-immunoprecipitation experiments indicate that the CARD domain of RIG-I is essential for inducing apoptosis by interacting with caspase-9. Together, our results reveal a dual role of RIG-I in HNSCC through regulating activation of Akt, in which RIG-I activation by low-dose viral dsRNA increases host cell surviral, whereas higher level of RIG-I activation leads to apopotosis. These findings highlight the therapeutic potential of dsRNA mediated RIG-I activation in the treatment of HNSCC. PMID:23484008

  18. Acid distribution in phosphoric acid fuel cells

    SciTech Connect

    Okae, I.; Seya, A.; Umemoto, M.

    1996-12-31

    Electrolyte acid distribution among each component of a cell is determined by capillary force when the cell is not in operation, but the distribution under the current load conditions had not been clear so far. Since the loss of electrolyte acid during operation is inevitable, it is necessary to store enough amount of acid in every cell. But it must be under the level of which the acid disturbs the diffusion of reactive gases. Accordingly to know the actual acid distribution during operation in a cell is very important. In this report, we carried out experiments to clarify the distribution using small single cells.

  19. Nrf2 promotes neuronal cell differentiation

    PubMed Central

    Zhao, Fei; Wu, Tongde; Lau, Alexandria; Jiang, Tao; Huang, Zheping; Wang, Xiao-Jun; Chen, Weimin; Wong, Pak Kin; Zhang, Donna D.

    2009-01-01

    The transcription factor Nrf2 has emerged as a master regulator for the endogenous antioxidant response, which is critical in defending cells against environmental insults and in maintaining intracellular redox balance. However, whether Nrf2 has any role in neuronal cell differentiation is largely unknown. In this report, we have examined the effects of Nrf2 on cell differentiation using a neuroblastoma cell line, SH-SY5Y. Retinoic acid (RA) and 12-O-tetradecanoylphorbol-13-acetate (TPA), two well-studied inducers for neuronal differentiation, are able to induce Nrf2 and its target gene NAD(P)H quinone oxidoreductase 1 (NQO1) in a dose- and time- dependent manner. RA-induced Nrf2 up-regulation is accompanied by neurite outgrowth and an induction of two neuronal differentiation markers, neurofilament-M (NF-M) and microtubule-associated protein 2 (MAP-2). Overexpression of Nrf2 in SH-SY5Y cells promotes neuronal differentiation whereas inhibition of endogenous Nrf2 expression inhibited neuronal differentiation. More remarkably, the positive role of Nrf2 in neuronal differentiation was verified ex vivo in primary neuron culture. Primary neurons isolated from Nrf2-null mice showed a retarded progress in differentiation, compared to that from wild-type mice. Collectively, our data demonstrate a novel role for Nrf2 in promoting neuronal cell differentiation, which will open new perspectives for therapeutic uses of Nrf2 activators in patients with neurodegenerative diseases. PMID:19573594

  20. Membrane lipid modification by polyunsaturated fatty acids sensitizes oligodendroglial OLN-93 cells against oxidative stress and promotes up-regulation of heme oxygenase-1 (HSP32).

    PubMed

    Brand, Annette; Bauer, Nina G; Hallott, Amanda; Goldbaum, Olaf; Ghebremeskel, Kebreab; Reifen, Ram; Richter-Landsberg, Christiane

    2010-04-01

    Polyunsaturated fatty acids (PUFA) are highly abundant in brain tissue, and docosahexaenoic acid (DHA) might protect cells from oxidative stress (OS) during inflammation and demyelinating disorders, but also might exert pro-oxidant effects. Here we investigated if PUFA supplements lead to heat shock protein induction, altered cell survival properties and stress responses to OS exerted by hydrogen peroxide in oligodendroglial OLN-93 cells. The data show that supplements of various fatty acids (FA) with 18-22 carbons chain length and 2-6 double bonds led to PUFA enrichment in cellular membranes. Depending on the degree of desaturation, FA-supplements caused the up-regulation of heme oxygenase-1 (HSP32), a stress protein inducible by OS, and an increase in sensitivity to hydrogen peroxide-treatment. DHA, with the highest number of double bonds, was most effective. Co-treatment with DHA and the lipophilic vitamin E analogue alpha-tocopherol, suppressed heme oxygenase-1 up-regulation and cell survival was restored. Analysis of the lipid profile demonstrates that alpha-tocopherol not only has antioxidant capacities, but also directly modified the PUFA profile in cell membranes. Enrichment with higher omega-3, -6 and -9 PUFA and an increase in the biosynthesis rate of very long chain fatty acids, mainly changed the FA profile of ethanolamine and serine phosphoglycerides.

  1. All trans-retinoic acid analogs promote cancer cell apoptosis through non-genomic Crabp1 mediating ERK1/2 phosphorylation

    PubMed Central

    Persaud, Shawna D.; Park, Sung Wook; Ishigami-Yuasa, Mari; Koyano-Nakagawa, Naoko; Kagechika, Hiroyuki; Wei, Li-Na

    2016-01-01

    All trans retinoic acid (atRA) is one of the most potent therapeutic agents, but extensive toxicity caused by nuclear RA receptors (RARs) limits its clinical application in treating cancer. AtRA also exerts non-genomic activities for which the mechanism remains poorly understood. We determine that cellular retinoic acid binding protein 1 (Crabp1) mediates the non-genomic activity of atRA, and identify two compounds as the ligands of Crabp1 to rapidly and RAR-independently activate extracellular signal regulated kinase 1/2 (ERK1/2). Non-canonically activated ERK activates protein phosphatase 2A (PP2A) and lengthens cell cycle duration in embryonic stem cells (ESC). This is abolished in Crabp1-null ESCs. Re-expressing Crabp1 in Crabp1-negative cancer cells also sensitizes their apoptotic induction by atRA. This study reveals a physiological relevance of the non-genomic action of atRA, mediated by Crabp1, in modulating cell cycle progression and apoptosis induction, and provides a new cancer therapeutic strategy whereby compounds specifically targeting Crabp1 can modulate cell cycle and cancer cell apoptosis in a RAR-independent fashion, thereby avoiding atRA’s toxicity caused by its genomic effects. PMID:26935534

  2. Role of Arachidonic Acid in Promoting Hair Growth

    PubMed Central

    Munkhbayar, Semchin; Jang, Sunhyae; Cho, A-Ri; Choi, Soon-Jin; Shin, Chang Yup; Eun, Hee Chul; Kim, Kyu Han

    2016-01-01

    Background Arachidonic acid (AA) is an omega-6 polyunsaturated fatty acid present in all mammalian cell membranes, and involved in the regulation of many cellular processes, including cell survival, angiogenesis, and mitogenesis. The dermal papilla, composed of specialized fibroblasts located in the bulb of the hair follicle, contributes to the control of hair growth and the hair cycle. Objective This study investigated the effect of AA on hair growth by using in vivo and in vitro models. Methods The effect of AA on human dermal papilla cells (hDPCs) and hair shaft elongation was evaluated by MTT assay and hair follicle organ culture, respectively. The expression of various growth and survival factors in hDPCs were investigated by western blot or immunohistochemistry. The ability of AA to induce and prolong anagen phase in C57BL/6 mice was analyzed. Results AA was found to enhance the viability of hDPCs and promote the expression of several factors responsible for hair growth, including fibroblast growth factor-7 (FGF-7) and FGF-10. Western blotting identified the role of AA in the phosphorylation of various transcription factors (ERK, CREB, and AKT) and increased expression of Bcl-2 in hDPCs. In addition, AA significantly promoted hair shaft elongation, with increased proliferation of matrix keratinocytes, during ex vivo hair follicle culture. It was also found to promote hair growth by induction and prolongation of anagen phase in telogen-stage C57BL/6 mice. Conclusion This study concludes that AA plays a role in promoting hair growth by increasing the expression of growth factors in hDPCs and enhancing follicle proliferation and survival. PMID:26848219

  3. Novel FTS-diamine/cinnamic acid hybrids inhibit tumor cell proliferation and migration and promote apoptosis via blocking Ras-related signaling in vitro.

    PubMed

    Ling, Yong; Zhao, Xinmei; Li, Xianghua; Wang, Xuemin; Yang, Yang; Wang, Zhiqiang; Wang, Xinyang; Zhang, Jie; Zhang, Yihua

    2015-02-01

    Novel FTS-diamine/cinnamic acid hybrids 7a-f were prepared, and their in vitro biological activities were evaluated. It was found that 7c showed the strongest anti-proliferation activities against cancer cells in vitro and significant growth inhibition of tumor in vivo, and more potential for inhibitory selectivity to tumor cells than intermediate 6 and FTS. Furthermore, the anti-proliferative effect of 7c in Lovo cell lines followed a similar pattern, which included a dose-dependent induction of cell apoptosis via the up-regulation of Bax as well as activated caspase-3 and down-regulation of Bcl-2, and the inhibition of cancer cells migration and invasion in a concentration-dependent way. More importantly, 7c could significantly block Ras-related signaling pathways, which may contribute to its pro-apoptotic induction of the cancer cell lines and its inhibition of carcinoma cell proliferation, migration, and invasion. Therefore, our novel findings may provide a new framework for the discovery of new FTS hybrids for the intervention of human carcinoma cells.

  4. Schwann cell proliferation and differentiation that is induced by ferulic acid through MEK1/ERK1/2 signalling promotes peripheral nerve remyelination following crush injury in rats

    PubMed Central

    Zhu, Xiaoyan; Li, Kun; Guo, Xin; Wang, Jian; Xiang, Yang

    2016-01-01

    Schwann cell proliferation and differentiation is critical for the remyelination of injured peripheral nerves. Ferulic acid (FA) is a widely used antioxidant agent with neuroprotective properties. However, the potentially beneficial effects of FA on Schwann cells are unknown. Therefore, the present study was designed to examine the effects of FA on Schwann cell proliferation and differentiation. By using the cultured primary Schwann cells and proliferation assay, the results identified that FA was capable of increasing Schwann cell proliferation and expression of myelin-associated glycoprotein (MAG) and myelin basic protein (MBP) in vitro. It was also observed that the beneficial effect of FA treatment on Schwann cells was mainly dependent on the activation of MEK1/ERK1/2 signalling. Furthermore, FA was intraperitoneally administered to rats with sciatic nerve crush injury, and the results revealed an increase in Schwann cell proliferation and differentiation, while the MAG and MBP expression levels in sciatic nerves were markedly upregulated following FA administration. In conclusion, the current results demonstrate that Schwann cell proliferation and differentiation is induced by FA through MEK1/ERK1/2 signalling and that FA may accelerate injured peripheral nerve remyelination. PMID:27588110

  5. Transgenesis of humanized fat1 promotes n-3 polyunsaturated fatty acid synthesis and expression of genes involved in lipid metabolism in goat cells.

    PubMed

    Fan, Yixuan; Ren, Caifang; Wang, Zhibo; Jia, Ruoxin; Wang, Dan; Zhang, Yanli; Zhang, Guomin; Wan, Yongjie; Huang, Mingrui; Wang, Feng

    2016-01-15

    The n-3 fatty acid desaturase gene fat1 codes for the n-3 desaturase enzyme, which can convert n-6 polyunsaturated fatty acids (PUFAs) to n-3 PUFAs. The n-3 PUFAs are essential components required for normal cellular function and have preventive and therapeutic effects on many diseases. Goat is an important domestic animal for human consumption of meat and milk. To elevate the concentrations of n-3 PUFAs and examine the regulatory mechanism of fat1 in PUFA metabolism in goat cells, we successfully constructed a humanized fat1 expression vector and confirmed the efficient expression of fat1 in goat ear skin-derived fibroblast cells (GEFCs) by qRT-PCR and Western blot analysis. Fatty acid analysis showed that fat1 overexpression significantly increased the levels of total n-3 PUFAs and decreased the levels of total n-6 PUFAs in GEFCs. In addition, qRT-PCR results indicate that the FADS1 and FADS2 desaturase genes, ELOV2 and ELOV5 elongase genes, ACO and CPT1 oxidation genes, and PPARa and PPARγ transcription factors are up-regulated, and transcription factors of SREBP-1c gene are down-regulated in the fat1 transgenic goat cells. Overall, fat1-overexpression resulted in an increase in the n-3 fatty acids and altered expression of PUFA synthesis related genes in GEFCs. This work lays a foundation for both the production of fat1 transgenic goats and further study of the mechanism of fat1 function in the PUFAs metabolism. PMID:26474750

  6. Transgenesis of humanized fat1 promotes n-3 polyunsaturated fatty acid synthesis and expression of genes involved in lipid metabolism in goat cells.

    PubMed

    Fan, Yixuan; Ren, Caifang; Wang, Zhibo; Jia, Ruoxin; Wang, Dan; Zhang, Yanli; Zhang, Guomin; Wan, Yongjie; Huang, Mingrui; Wang, Feng

    2016-01-15

    The n-3 fatty acid desaturase gene fat1 codes for the n-3 desaturase enzyme, which can convert n-6 polyunsaturated fatty acids (PUFAs) to n-3 PUFAs. The n-3 PUFAs are essential components required for normal cellular function and have preventive and therapeutic effects on many diseases. Goat is an important domestic animal for human consumption of meat and milk. To elevate the concentrations of n-3 PUFAs and examine the regulatory mechanism of fat1 in PUFA metabolism in goat cells, we successfully constructed a humanized fat1 expression vector and confirmed the efficient expression of fat1 in goat ear skin-derived fibroblast cells (GEFCs) by qRT-PCR and Western blot analysis. Fatty acid analysis showed that fat1 overexpression significantly increased the levels of total n-3 PUFAs and decreased the levels of total n-6 PUFAs in GEFCs. In addition, qRT-PCR results indicate that the FADS1 and FADS2 desaturase genes, ELOV2 and ELOV5 elongase genes, ACO and CPT1 oxidation genes, and PPARa and PPARγ transcription factors are up-regulated, and transcription factors of SREBP-1c gene are down-regulated in the fat1 transgenic goat cells. Overall, fat1-overexpression resulted in an increase in the n-3 fatty acids and altered expression of PUFA synthesis related genes in GEFCs. This work lays a foundation for both the production of fat1 transgenic goats and further study of the mechanism of fat1 function in the PUFAs metabolism.

  7. Promotion of granule cell survival by high K+ or excitatory amino acid treatment and Ca2+/calmodulin-dependent protein kinase activity.

    PubMed

    Hack, N; Hidaka, H; Wakefield, M J; Balázs, R

    1993-11-01

    Cerebellar granule cells in culture develop survival requirements which can be met either by chronic membrane depolarization (25 mM K+) or by stimulation of ionotropic excitatory amino acid receptors. We observed previously that this trophic effect is mediated via Ca2+ influx, either through dihydropyridine-sensitive, voltage-dependent calcium channels (activated directly by high K+ or indirectly by kainate) or through N-methyl-D-aspartate receptor-linked ion channels. Steps after Ca2+ entry in the transduction cascade mediating the survival-supporting effect of high K+ and excitatory amino acids have now been examined. Using protein kinase inhibitors (H-7, polymixin B and gangliosides), and modulating protein kinase C activity by treatment with the phorbol ester 12-O-tetradecanoylphorbol-13-acetate, we obtained evidence against the involvement of protein kinase C and cyclic nucleotide-dependent protein kinases in the transduction cascade. On the other hand, calmidazolium (employed as a calmodulin inhibitor) counteracted the trophic effect of elevated K+ with high potency (IC50 0.3 microM), which exceeded by approximately 10-fold the potency for the blockade by the drug of voltage-sensitive calcium channels. The potency of calmidazolium in interfering with the N-methyl-D-aspartate rescue of cells was also much higher in comparison with the inhibition of 45Ca2+ influx through N-methyl-D-aspartate receptor-linked channels. Our results indicated that after calmodulin the next step in the trophic effects involves Ca2+/calmodulin-dependent protein kinase II activity. KN-62, a fairly specific antagonist of this enzyme, compromised elevated K+ or excitatory amino acid-supported cell survival with high potency (IC50 2.5 microM). In the relevant concentration range, KN-62 had little or no effect on Ca2+ entry through either voltage- or N-methyl-D-aspartate receptor-gated channels. Combining information on the toxic action of glutamate in "mature" granule cells with the

  8. Delivery of ursolic acid (UA) in polymeric nanoparticles effectively promotes the apoptosis of gastric cancer cells through enhanced inhibition of cyclooxygenase 2 (COX-2).

    PubMed

    Zhang, Hao; Li, Xiaolin; Ding, Jing; Xu, Huae; Dai, Xinzheng; Hou, Zhibo; Zhang, Kai; Sun, Kun; Sun, Weihao

    2013-01-30

    It has been demonstrated that ursolic acid (UA) could effectively induces apoptosis of cancer cells by inhibiting the expression of cyclooxygenase 2 (COX-2), which constitutively expresses in gastric cancer. However, the hydrophobicity of UA increases the difficulty in its potential clinical application, which raises the possibility for its application as a novel model drug in nanoparticle-based delivery system. UA-loaded nanoparticles (UA-NPs) were prepared by a nano-precipitation method using amphilic methoxy poly(ethylene glycol)-polycaprolactone (mPEG-PCL) block copolymers as drug carriers. UA was effectively transported into SGC7901 cells by nanoparticles and localized around the nuclei in the cytoplasms. The in vitro cytotoxicity and apoptosis test indicated that UA-NPs significantly elicited more cell death at almost equivalent dose and corresponding incubation time. Moreover, UA-NPs led to more cell apoptosis through stronger inhibition of COX-2 and activation of caspase 3. The most powerful evidence from this report is that the significant differences between the cytotoxicity of free UA and UA-NPs are closely related to the expression levels of COX-2 and caspase-3, which demonstrates the superiority of UA-NPs over free UA through penetrating cell membrane. Therefore, the study offer an effective way to improve the anticancer efficiency of UA through nano-drug delivery system.

  9. Docosahexaenoic acid promotes dopaminergic differentiation in induced pluripotent stem cells and inhibits teratoma formation in rats with Parkinson-like pathology.

    PubMed

    Chang, Yuh-Lih; Chen, Shih-Jen; Kao, Chung-Lan; Hung, Shih-Chieh; Ding, Dah-Ching; Yu, Cheng-Chia; Chen, Yi-Jen; Ku, Hung-Hai; Lin, Chin-Po; Lee, Kun-Hsiung; Chen, Yu-Chih; Wang, Jhi-Joung; Hsu, Chuan-Chih; Chen, Liang-Kung; Li, Hsin-Yang; Chiou, Shih-Hwa

    2012-01-01

    Parkinson's disease (PD) is a neurodegenerative disorder characterized by the degeneration of dopaminergic (DA) neurons in the midbrain. Induced pluripotent stem (iPS) cells have shown potential for differentiation and may become a resource of functional neurons for the treatment of PD. However, teratoma formation is a major concern for transplantation-based therapies. This study examined whether functional neurons could be efficiently generated from iPS cells using a five-step induction procedure combined with docosahexaenoic acid (DHA) treatment. We demonstrated that DHA, a ligand for the RXR/Nurr1 heterodimer, significantly activated expression of the Nurr1 gene and the Nurr1-related pathway in iPS cells. DHA treatment facilitated iPS differentiation into tyrosine hydroxylase (TH)-positive neurons in vitro and in vivo and functionally increased dopamine release in transplanted grafts in PD-like animals. Furthermore, DHA dramatically upregulated the endogenous expression levels of neuroprotective genes (Bcl-2, Bcl-xl, brain-derived neurotrophic factor, and glial cell-derived neurotrophic factor) and protected against 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced apoptosis in iPS-derived neuronal precursor cells. DHA-treated iPS cells significantly improved the behavior of 6-hydroxydopamine (6-OHDA)-treated PD-like rats compared to control or eicosapentaenoic acid-treated group. Importantly, the in vivo experiment suggests that DHA induces the differentiation of functional dopaminergic precursors and improves the abnormal behavior of 6-OHDA-treated PD-like rats by 4 months after transplantation. Furthermore, we found that DHA treatment in iPS cell-grafted rats significantly downregulated the mRNA expression of embryonic stem cell-specific genes (Oct-4 and c-Myc) in the graft and effectively blocked teratoma formation. Importantly, 3 Tesla-magnetic resonance imaging and ex vivo green fluorescence protein imaging revealed that no teratomas were present

  10. Overexpression of SREBP1 (sterol regulatory element binding protein 1) promotes de novo fatty acid synthesis and triacylglycerol accumulation in goat mammary epithelial cells.

    PubMed

    Xu, H F; Luo, J; Zhao, W S; Yang, Y C; Tian, H B; Shi, H B; Bionaz, M

    2016-01-01

    Sterol regulatory element binding protein 1 (SREBP1; gene name SREBF1) is known to be the master regulator of lipid homeostasis in mammals, including milk fat synthesis. The major role of SREBP1 in controlling milk fat synthesis has been demonstrated in bovine mammary epithelial cells. Except for a demonstrated role in controlling the expression of FASN, a regulatory role of SREBP1 on milk fat synthesis is very likely, but has not yet been demonstrated in goat mammary epithelial cells (GMEC). To explore the regulatory function of SREBP1 on de novo fatty acids and triacylglycerol synthesis in GMEC, we overexpressed the mature form of SREBP1 (active NH2-terminal fragment) in GMEC using a recombinant adenovirus vector (Ad-nSREBP1), with Ad-GFP (recombinant adenovirus of green fluorescent protein) as control, and infected the GMEC for 48 h. In infected cells, we assessed the expression of 20 genes related to milk fat synthesis using real time-quantitative PCR, the protein abundance of SREBP1 and FASN by Western blot, the production of triacylglycerol, and the fatty acid profile. Expression of SREBF1 was modest in mammary compared with the other tissues in dairy goats but its expression increased approximately 30-fold from pregnancy to lactation. The overexpression of the mature form of SREBP1 was confirmed by >200-fold higher expression of SREBF1 in Ad-nSREBP1 compared with Ad-GFP. We observed no changes in amount of the precursor form of SREBP1 protein but a >10-fold increase of the mature form of SREBP1 protein with Ad-nSREBP1. Compared with Ad-GFP cells (control), Ad-nSREBP1 cells had a significant increase in expression of genes related to long-chain fatty acid activation (ACSL1), transport (FABP3), desaturation (SCD1), de novo synthesis of fatty acids (ACSS2, ACLY, IDH1, ACACA, FASN, and ELOVL6), and transcriptional factors (NR1H3 and PPARG). We observed a >10-fold increase in expression of INSIG1 but SCAP was downregulated by Ad-nSREBP1. Among genes related to

  11. Overexpression of SREBP1 (sterol regulatory element binding protein 1) promotes de novo fatty acid synthesis and triacylglycerol accumulation in goat mammary epithelial cells.

    PubMed

    Xu, H F; Luo, J; Zhao, W S; Yang, Y C; Tian, H B; Shi, H B; Bionaz, M

    2016-01-01

    Sterol regulatory element binding protein 1 (SREBP1; gene name SREBF1) is known to be the master regulator of lipid homeostasis in mammals, including milk fat synthesis. The major role of SREBP1 in controlling milk fat synthesis has been demonstrated in bovine mammary epithelial cells. Except for a demonstrated role in controlling the expression of FASN, a regulatory role of SREBP1 on milk fat synthesis is very likely, but has not yet been demonstrated in goat mammary epithelial cells (GMEC). To explore the regulatory function of SREBP1 on de novo fatty acids and triacylglycerol synthesis in GMEC, we overexpressed the mature form of SREBP1 (active NH2-terminal fragment) in GMEC using a recombinant adenovirus vector (Ad-nSREBP1), with Ad-GFP (recombinant adenovirus of green fluorescent protein) as control, and infected the GMEC for 48 h. In infected cells, we assessed the expression of 20 genes related to milk fat synthesis using real time-quantitative PCR, the protein abundance of SREBP1 and FASN by Western blot, the production of triacylglycerol, and the fatty acid profile. Expression of SREBF1 was modest in mammary compared with the other tissues in dairy goats but its expression increased approximately 30-fold from pregnancy to lactation. The overexpression of the mature form of SREBP1 was confirmed by >200-fold higher expression of SREBF1 in Ad-nSREBP1 compared with Ad-GFP. We observed no changes in amount of the precursor form of SREBP1 protein but a >10-fold increase of the mature form of SREBP1 protein with Ad-nSREBP1. Compared with Ad-GFP cells (control), Ad-nSREBP1 cells had a significant increase in expression of genes related to long-chain fatty acid activation (ACSL1), transport (FABP3), desaturation (SCD1), de novo synthesis of fatty acids (ACSS2, ACLY, IDH1, ACACA, FASN, and ELOVL6), and transcriptional factors (NR1H3 and PPARG). We observed a >10-fold increase in expression of INSIG1 but SCAP was downregulated by Ad-nSREBP1. Among genes related to

  12. Epithelial MUC1 promotes cell migration, reduces apoptosis and affects levels of mucosal modulators during acetylsalicylic acid (aspirin)-induced gastropathy.

    PubMed

    Banerjee, Debashish; Fernandez, Harvey Robert; Patil, Pradeep Bhatu; Premaratne, Pushpa; Quiding-Järbrink, Marianne; Lindén, Sara Katarina

    2015-02-01

    MUC1 is a transmembrane mucin highly expressed in the stomach. Although extensive research has uncovered many of its roles in cancer, knowledge about the functions of MUC1 in normal tissues is limited. In the present study, we showed that acetylsalicylic acid (ASA; aspirin) up-regulated MUC1/Muc1 expression in the gastric mucosa of humans and wild-type (WT) mice. ASA induced mucosal injury in all mice to a similar extent; however, WT animals and those chimaeras with Muc1 on the epithelia recovered faster than Muc1-knockout (KO) mice and chimaeras carrying Muc1 on haemopoietic but not epithelial cells. MUC1 enhanced proliferation and migration of the human gastric cell line MKN-7 and increased resistance to apoptosis. The repeated treatment regime used caused a reduction in cyclo-oxygenase-1 (Cox-1) expression, though WT animals returned faster towards pre-treatment levels and had increased Cox-2 and vascular endothelial growth factor levels during recovery. Thus we found that epithelial Muc1 is more important for the healing process than haemopoietic Muc1 and Muc1/MUC1 facilitates wound healing by enhancing cell migration and proliferation, protecting against apoptosis and mediating expression of mucosal modulators. Thus MUC1 plays essential roles during wound healing and development of treatment modalities targeting enhanced expression of MUC1 may be beneficial to treat mucosal wounds.

  13. Docosahexaenoic Acid Inhibits Tumor Promoter-Induced Urokinase-Type Plasminogen Activator Receptor by Suppressing PKCδ- and MAPKs-Mediated Pathways in ECV304 Human Endothelial Cells

    PubMed Central

    Lian, Sen; Xia, Yong; Nguyen, Thi Thinh; Ung, Trong Thuan; Yoon, Hyun Joong; Kim, Nam Ho; Kim, Kyung Keun; Jung, Young Do

    2016-01-01

    The overexpression of urokinase-type plasminogen activator receptor (uPAR) is associated with inflammation and virtually all human cancers. Despite the fact that docosahexaenoic acid (DHA) has been reported to possess anti-inflammatory and anti-tumor properties, the negative regulation of uPAR by DHA is still undefined. Here, we investigated the effect of DHA on 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced uPAR expression and the underlying molecular mechanisms in ECV304 human endothelial cells. DHA concentration-dependently inhibited TPA-induced uPAR. Specific inhibitors and mutagenesis studies showed that PKCδ, JNK1/2, Erk1/2, NF-κB, and AP-1 were critical for TPA-induced uPAR expression. Application of DHA suppressed TPA-induced translocation of PKCδ, activation of the JNK1/2 and Erk1/2 signaling pathways, and subsequent AP-1 and NF-κB transactivation. In conclusion, these observations suggest a novel role for DHA in reducing uPAR expression and cell invasion by inhibition of PKCδ, JNK1/2, and Erk1/2, and the reduction of AP-1 and NF-κB activation in ECV304 human endothelial cells. PMID:27654969

  14. New Role for Granulocyte Colony-Stimulating Factor-Induced Extracellular Signal-Regulated Kinase 1/2 in Histone Modification and Retinoic Acid Receptor α Recruitment to Gene Promoters: Relevance to Acute Promyelocytic Leukemia Cell Differentiation ▿

    PubMed Central

    Cassinat, B.; Zassadowski, F.; Ferry, C.; Llopis, L.; Bruck, N.; Lainey, E.; Duong, V.; Cras, A.; Despouy, G.; Chourbagi, O.; Beinse, G.; Fenaux, P.; Rochette Egly, C.; Chomienne, C.

    2011-01-01

    The induction of the granulocytic differentiation of leukemic cells by all-trans retinoic acid (RA) has been a major breakthrough in terms of survival for acute promyelocytic leukemia (APL) patients. Here we highlight the synergism and the underlying novel mechanism between RA and the granulocyte colony-stimulating factor (G-CSF) to restore differentiation of RA-refractory APL blasts. First, we show that in RA-refractory APL cells (UF-1 cell line), PML-RA receptor alpha (RARα) is not released from target promoters in response to RA, resulting in the maintenance of chromatin repression. Consequently, RARα cannot be recruited, and the RA target genes are not activated. We then deciphered how the combination of G-CSF and RA successfully restored the activation of RA target genes to levels achieved in RA-sensitive APL cells. We demonstrate that G-CSF restores RARα recruitment to target gene promoters through the activation of the extracellular signal-regulated kinase (ERK)/mitogen-activated protein kinase (MAPK) pathway and the subsequent derepression of chromatin. Thus, combinatorial activation of cytokines and RARs potentiates transcriptional activity through epigenetic modifications induced by specific signaling pathways. PMID:21262770

  15. Acid-functionalized polyolefin materials and their use in acid-promoted chemical reactions

    DOEpatents

    Oyola, Yatsandra; Tian, Chengcheng; Bauer, John Christopher; Dai, Sheng

    2016-06-07

    An acid-functionalized polyolefin material that can be used as an acid catalyst in a wide range of acid-promoted chemical reactions, wherein the acid-functionalized polyolefin material includes a polyolefin backbone on which acid groups are appended. Also described is a method for the preparation of the acid catalyst in which a precursor polyolefin is subjected to ionizing radiation (e.g., electron beam irradiation) of sufficient power and the irradiated precursor polyolefin reacted with at least one vinyl monomer having an acid group thereon. Further described is a method for conducting an acid-promoted chemical reaction, wherein an acid-reactive organic precursor is contacted in liquid form with a solid heterogeneous acid catalyst comprising a polyolefin backbone of at least 1 micron in one dimension and having carboxylic acid groups and either sulfonic acid or phosphoric acid groups appended thereto.

  16. Copper-promoted trifluoromethylation of primary and secondary alkylboronic acids.

    PubMed

    Xu, Jun; Xiao, Bin; Xie, Chuan-Qi; Luo, Dong-Fen; Liu, Lei; Fu, Yao

    2012-12-01

    New couple: The Cu-promoted trifluoromethylation of primary and secondary alkylboronic acids with TMSCF(3) extends the scope of transition-metal-catalyzed trifluoromethylation reactions to sp(3)-hybridized carbon centers. It also represents one of the first examples for Cu-catalyzed C-C cross-coupling reactions of alkylboronic acid derivatives. PMID:23143929

  17. Linoleic acid derivative DCP-LA ameliorates stress-induced depression-related behavior by promoting cell surface 5-HT1A receptor translocation, stimulating serotonin release, and inactivating GSK-3β.

    PubMed

    Kanno, Takeshi; Tanaka, Akito; Nishizaki, Tomoyuki

    2015-04-01

    Impairment of serotonergic neurotransmission is the major factor responsible for depression and glycogen synthase kinase 3β (GSK-3β) participates in serotonergic transmission-mediated signaling networks relevant to mental illnesses. In the forced-swim test to assess depression-like behavior, the immobility time for mice with restraint stress was significantly longer than that for nonstressed control mice. Postsynaptic cell surface localization of 5-HT1A receptor, but not 5-HT2A receptor, in the hypothalamus for mice with restraint stress was significantly reduced as compared with that for control mice, which highly correlated to prolonged immobility time, i.e., depression-like behavior. The linoleic acid derivative 8-[2-(2-pentyl-cyclopropylmethyl)-cyclopropyl]-octanoic acid (DCP-LA) restored restraint stress-induced reduction of cell surface 5-HT1A receptor and improved depression-like behavior in mice with restraint stress. Moreover, DCP-LA stimulated serotonin release from hypothalamic slices and cancelled restraint stress-induced reduction of GSK-3β phosphorylation at Ser9. Taken together, the results of the present study indicate that DCP-LA could ameliorate depression-like behavior by promoting translocation of 5-HT1A receptor to the plasma membrane on postsynaptic cells, stimulating serotonin release, and inactivating GSK-3β.

  18. Dysregulated hepatic bile acids collaboratively promote liver carcinogenesis.

    PubMed

    Xie, Guoxiang; Wang, Xiaoning; Huang, Fengjie; Zhao, Aihua; Chen, Wenlian; Yan, Jingyu; Zhang, Yunjing; Lei, Sha; Ge, Kun; Zheng, Xiaojiao; Liu, Jiajian; Su, Mingming; Liu, Ping; Jia, Wei

    2016-10-15

    Dysregulated bile acids (BAs) are closely associated with liver diseases and attributed to altered gut microbiota. Here, we show that the intrahepatic retention of hydrophobic BAs including deoxycholate (DCA), taurocholate (TCA), taurochenodeoxycholate (TCDCA), and taurolithocholate (TLCA) were substantially increased in a streptozotocin and high fat diet (HFD) induced nonalcoholic steatohepatitis-hepatocellular carcinoma (NASH-HCC) mouse model. Additionally chronic HFD-fed mice spontaneously developed liver tumors with significantly increased hepatic BA levels. Enhancing intestinal excretion of hydrophobic BAs in the NASH-HCC model mice by a 2% cholestyramine feeding significantly prevented HCC development. The gut microbiota alterations were closely correlated with altered BA levels in liver and feces. HFD-induced inflammation inhibited key BA transporters, resulting in sustained increases in intrahepatic BA concentrations. Our study also showed a significantly increased cell proliferation in BA treated normal human hepatic cell lines and a down-regulated expression of tumor suppressor gene CEBPα in TCDCA treated HepG2 cell line, suggesting that several hydrophobic BAs may collaboratively promote liver carcinogenesis.

  19. Targeting cancer cells with folic acid-iminoboronate fluorescent conjugates.

    PubMed

    Cal, Pedro M S D; Frade, Raquel F M; Chudasama, Vijay; Cordeiro, Carlos; Caddick, Stephen; Gois, Pedro M P

    2014-05-25

    Herein we present the synthesis of fluorescent 2-acetylbenzeneboronic acids that undergo B-N promoted conjugation with lysozyme and N-(2-aminoethyl) folic acid (EDA-FA), generating conjugates that are selectively recognized and internalized by cancer cells that over-express folic acid receptors.

  20. Meat and ascorbic acid can promote Fe availability from Fe-phytate but not from Fe-tannic acid complexes.

    PubMed

    Engle-Stone, Reina; Yeung, Andrew; Welch, Ross; Glahn, Raymond

    2005-12-28

    This study utilized an in vitro digestion/Caco-2 cell model to determine the levels of ascorbic acid (AA) and "meat factor" needed to promote Fe absorption from Fe complexed with phytic acid (PA) or tannic acid (TA). AA reversed the inhibition of Fe absorption by PA beginning at a molar ratio of 1:20:1 (Fe:PA:AA) but essentially had no effect on the Fe complexed with TA. Fish also reversed the inhibition of Fe uptake by PA but not by TA. TA and fish decreased total Fe solubility. Iron in the presence of PA was highly soluble. AA, but not fish, increased the percentage of soluble Fe as Fe2+ in the presence of both inhibitors. The results indicate that monoferric phytate is a form of Fe that can be available for absorption in the presence of uptake promoters. In contrast, a TA-Fe complex is much less soluble and unavailable in the presence of promoters.

  1. Promotion of adipogenesis by 15-(S)-hydroxyeicosatetraenoic acid.

    PubMed

    Song, Yong-Seok; Lee, Dong Hun; Yu, Jae-Hyuk; Oh, Deok-Kun; Hong, Jin Tae; Yoon, Do-Young

    2016-03-01

    Excess adipogenesis is a characteristic of obesity, which is associated with serious health problem, including type 2 diabetes. Here, to better understand the mechanisms for the development of adipocytes, we investigated the regulatory role of 15-(S)-hydroxyeicosatetraenoic acid (15(S)-HETE) in adipogenesis by treating 3T3-L1 murine preadipocytes and human bone marrow mesenchymal stem cells (hBMSCs) with 15(S)-HETE. In the 3T3-L1 study, 15(S)-HETE stimulated lipid accumulation and enhanced expression of adipogenic markers such as peroxisome proliferator-activated receptor gamma (PPARγ), yet reduced the activity of factor negatively controlling adipogenesis, such as the 5'-AMP-activated protein kinase. In the early stage of adipogenesis, 15(S)-HETE increased activation of protein kinase B and expression of CCAAT/enhancer-binding protein (C/EBP)β and C/EBPδ. Finally, 15(S)-HETE promoted adipogenesis in hBMSCs, and resulted in increased lipid accumulation and expression of adipogenic markers. In conclusion, 15(S)-HETE functions as a natural PPARγ agonist and enhances adipogenesis. Our findings may provide the basis for the development of novel therapeutic measures to treat obesity.

  2. Functional analysis of a Lemna gibba rbcS promoter regulated by abscisic acid and sugar.

    PubMed

    Wang, Youru

    2013-04-01

    Photosynthesis-associated nuclear genes (PhANGs) are able to respond to multiple environmental and developmental signals, including light, sugar and abscisic acid (ABA). PhANGs have been extensively studied at the level of transcriptional regulation, and several cis-acting elements important for light responsiveness have been identified in their promoter sequences. However, the regulatory elements involved in sugar and ABA regulation of PhANGs have not been completely characterized. A ribulose-1,5-bisphosphate carboxylase small subunit gene (rbcS) promoter (SSU5C promoter) was isolated from duckweed (Lemna gibba). A series of SSU5C promoter 5' deletion fragments were fused to an intron-gus gene, and transgenic tobacco suspension cell lines were generated. Assay of tobacco suspension cell line harbouring the complete promoter in the fusion construct indicated that SSU5C promoter was negatively regulated by sugar and ABA under the condition of regular photoperiod. 5' deletion analysis of SSU5C promoter in transgenic tobacco suspension cell lines confirmed that a region between positions -310 and -152 included the ABA-response region, and that sugar-response cis-acting elements might be located in the region between -152 and -117. Taken together, our results confirmed that the cis-regulatory region responsible for repression by ABA and sugar in the SSU5C promoter was located between -310 and -117. PMID:23640406

  3. Microbes promote amino acid harvest to rescue undernutrition in Drosophila

    PubMed Central

    Yamada, Ryuichi; Deshpande, Sonali A.; Bruce, Kimberley D.; Mak, Elizabeth M.; Ja, William W.

    2015-01-01

    Summary Microbes play an important role in the pathogenesis of nutritional disorders such as protein-specific malnutrition. However, the precise contribution of microbes to host energy balance during undernutrition is unclear. Here, we show that Issatchenkia orientalis, a fungal microbe isolated from field-caught Drosophila melanogaster, promotes amino acid harvest to rescue the lifespan of undernourished flies. Using radioisotope-labeled dietary components (amino acids, nucleotides, and sucrose) to quantify nutrient transfer from food to microbe to fly, we demonstrate that I. orientalis extracts amino acids directly from nutrient-poor diets and increases protein flux to the fly. This microbial association restores body mass, protein, glycerol, and ATP levels and phenocopies the metabolic profile of adequately fed flies. Our study uncovers amino acid harvest as a fundamental mechanism linking microbial and host metabolism, and highlights Drosophila as a platform for quantitative studies of host-microbe relationships. PMID:25683709

  4. Acid-Base Pairs in Lewis Acidic Zeolites Promote Direct Aldol Reactions by Soft Enolization.

    PubMed

    Lewis, Jennifer D; Van de Vyver, Stijn; Román-Leshkov, Yuriy

    2015-08-17

    Hf-, Sn-, and Zr-Beta zeolites catalyze the cross-aldol condensation of aromatic aldehydes with acetone under mild reaction conditions with near quantitative yields. NMR studies with isotopically labeled molecules confirm that acid-base pairs in the Si-O-M framework ensemble promote soft enolization through α-proton abstraction. The Lewis acidic zeolites maintain activity in the presence of water and, unlike traditional base catalysts, in acidic solutions.

  5. Neuroprotection of lipoic acid treatment promotes angiogenesis and reduces the glial scar formation after brain injury.

    PubMed

    Rocamonde, B; Paradells, S; Barcia, J M; Barcia, C; García Verdugo, J M; Miranda, M; Romero Gómez, F J; Soria, J M

    2012-11-01

    After trauma brain injury, a large number of cells die, releasing neurotoxic chemicals into the extracellular medium, decreasing cellular glutathione levels and increasing reactive oxygen species that affect cell survival and provoke an enlargement of the initial lesion. Alpha-lipoic acid is a potent antioxidant commonly used as a treatment of many degenerative diseases such as multiple sclerosis or diabetic neuropathy. Herein, the antioxidant effects of lipoic acid treatment after brain cryo-injury in rat have been studied, as well as cell survival, proliferation in the injured area, gliogenesis and angiogenesis. Thus, it is shown that newborn cells, mostly corresponded with blood vessels and glial cells, colonized the damaged area 15 days after the lesion. However, lipoic acid was able to stimulate the synthesis of glutathione, decrease cell death, promote angiogenesis and decrease the glial scar formation. All those facts allow the formation of new neural tissue. In view of the results herein, lipoic acid might be a plausible pharmacological treatment after brain injury, acting as a neuroprotective agent of the neural tissue, promoting angiogenesis and reducing the glial scar formation. These findings open new possibilities for restorative strategies after brain injury, stroke or related disorders.

  6. Differential Radiosensitizing Effect of Valproic Acid in Differentiation Versus Self-Renewal Promoting Culture Conditions

    SciTech Connect

    Debeb, Bisrat G.; Xu Wei; Mok, Henry; Li Li; Robertson, Fredika; Ueno, Naoto T.; Reuben, Jim; Lucci, Anthony; Cristofanilli, Massimo; Woodward, Wendy A.

    2010-03-01

    Purpose: It has been shown that valproic acid (VA) enhances the proliferation and self-renewal of normal hematopoietic stem cells and that breast cancer stem/progenitor cells can be resistant to radiation. From these data, we hypothesized that VA would fail to radiosensitize breast cancer stem/progenitor cells grown to three-dimensional (3D) mammospheres. Methods and Materials: We used the MCF7 breast cancer cell line grown under stem cell-promoting culture conditions (3D mammosphere) and standard nonstem cell monolayer culture conditions (two-dimensional) to examine the effect of pretreatment with VA on radiation sensitivity in clonogenic survival assays and on the expression of embryonic stem cell transcription factors. Results: 3D-cultured MCF-7 cells expressed higher levels of Oct4, Nanog, and Sox2. The 3D passage enriched self-renewal and increased radioresistance in the 3D mammosphere formation assays. VA radiosensitized adherent cells but radioprotected 3D cells in single-fraction clonogenic assays. Moreover, fractionated radiation sensitized VA-treated adherent MCF7 cells but did not have a significant effect on VA-treated single cells grown to mammospheres. Conclusion: We have concluded that VA might preferentially radiosensitize differentiated cells compared with those expressing stem cell surrogates and that stem cell-promoting culture is a useful tool for in vitro evaluation of novel cancer therapeutic agents and radiosensitizers.

  7. Promotora de Salud: Promoting Folic Acid Use Among Hispanic Women

    PubMed Central

    deRosset, Leslie; Mullenix, Amy; Flores, Alina; Mattia-Dewey, Daniel; Mai, Cara T.

    2015-01-01

    Background The U.S. Public Health Service recommends that all women in the United States capable of becoming pregnant consume 400 μg of folic acid daily to reduce their risk of having a pregnancy affected by a neural tube defect (NTD). However, disparities exist in the consumption of folic acid, with Hispanic women having lower rates of folic acid consumption than non-Hispanic white women. Methods A community-based feasibility study was designed to assess the utility of the promotora de salud model to promote consumption of multivitamins containing folic acid for the prevention of NTDs among Spanish-speaking Hispanic women in North Carolina. The study consisted of an educational intervention given by a promotora (a lay, community health worker), with data collection occurring at baseline and four months post-intervention to measure changes in knowledge and behavior. Overall, 52% (n = 303) of participants completed all components of the study. Results Self-reported daily multivitamin consumption increased from 24% at baseline to 71% four months post-intervention. During the same time frame, awareness of folic acid increased from 78% to 98% and knowledge of the role of folic acid in the prevention of birth defects increased from 82% to 92%. Conclusions The results of this study indicate that the promotora de salud model may be effective in reaching a subpopulation of women with the folic acid message. Additional studies with larger population sizes are warranted to validate these findings. PMID:24707879

  8. TiO2 and Al2O3 promoted Pt/C nanocomposites as low temperature fuel cell catalysts for electro oxidation of methanol in acidic media

    NASA Astrophysics Data System (ADS)

    Naeem, Rabia; Ahmed, Riaz; Shahid Ansari, Muhammad

    2014-06-01

    Carbon corrosion and platinum dissolution are the two major catalyst layer degradation problems in polymer electrolyte membrane fuel cells (PEMFC). Ceramic addition can reduce the corrosion of carbon and increase the stability of catalysts. Pt/TiO2, Pt/TiO2-C, Pt/Al2O3 and Pt/Al2O3-C catalysts were synthesized and characterized. Electrochemical surface area of Pt/TiO2-C and Pt/Al2O3-C nanocomposite catalysts was much higher than the Pt/TiO2 and Pt/Al2O3 catalysts. Peak current, specific activity and mass activity of the catalysts was also determined by cyclic voltammetry and were much higher for the carbon nanocomposites. Exchange current densities were determined from Tafel plots. Heterogeneous rates of reaction of electro oxidation of methanol were determined for all the catalysts and were substantially higher for titania catalysts as compared to alumina added catalysts. Mass activity of Pt/TiO2-C was much higher than mass activity of Pt/Al2O3-C. Stability studies showed that addition of ceramics have increased the catalytic activity and durability of the catalysts considerably.

  9. Promotion of Hepatocarcinogenesis by Perfluoroalkyl Acids in Rainbow Trout

    PubMed Central

    Benninghoff, Abby D.; Orner, Gayle A.; Buchner, Clarissa H.; Hendricks, Jerry D.; Duffy, Aaron M.; Williams, David E.

    2012-01-01

    Previously, we reported that perfluorooctanoic acid (PFOA) promotes liver cancer in a manner similar to that of 17β-estradiol (E2) in rainbow trout. Also, other perfluoroalkyl acids (PFAAs) are weakly estrogenic in trout and bind the trout liver estrogen receptor. The primary objective of this study was to determine whether multiple PFAAs enhance hepatic tumorigenesis in trout, an animal model that represents human insensitivity to peroxisome proliferation. A two-stage chemical carcinogenesis model was employed in trout to evaluate PFOA, perfluorononanoic acid (PFNA), perfluorodecanoic acid (PFDA), perfluorooctane sulfonate (PFOS), and 8:2 fluorotelomer alcohol (8:2FtOH) as complete carcinogens or promoters of aflatoxin B1 (AFB1)- and/or N-methyl-N′-nitro-N-nitrosoguanidine (MNNG)-induced liver cancer. A custom trout DNA microarray was used to assess hepatic transcriptional response to these dietary treatments in comparison with E2 and the classic peroxisome proliferator, clofibrate (CLOF). Incidence, multiplicity, and size of liver tumors in trout fed diets containing E2, PFOA, PFNA, and PFDA were significantly higher compared with AFB1-initiated animals fed control diet, whereas PFOS caused a minor increase in liver tumor incidence. E2 and PFOA also enhanced MNNG-initiated hepatocarcinogenesis. Pearson correlation analyses, unsupervised hierarchical clustering, and principal components analyses showed that the hepatic gene expression profiles for E2 and PFOA, PFNA, PFDA, and PFOS were overall highly similar, though distinct patterns of gene expression were evident for each treatment, particularly for PFNA. Overall, these data suggest that multiple PFAAs can promote liver cancer and that the mechanism of promotion may be similar to that of E2. PMID:21984479

  10. Phototherapy promotes cell migration in the presence of hydroxyurea.

    PubMed

    Zungu, I L; Mbene, A B; Hawkins Evans, D H; Houreld, N N; Abrahamse, H

    2009-03-01

    Phototherapy has been shown to cause an increase in cell proliferation and migration. This study focused on viability (trypan blue), proliferation [sodium 3'-(1-(phenylaminocarbonyl)-3,4-tetrazolium)-bis(4-methoxy-6-nitro)-benzene sulphonic acid hydrate (XTT) and adenosine triphosphate (ATP)] and migration of WS1 cells following irradiation in the presence of hydroxyurea (HU), which is an inhibitor of proliferation. Wounded cells were irradiated on days 1 and 4 with a fluence of 5 J/cm(2) with a helium-neon (He-Ne) laser at 632.8 nm. After a repair time of 24 h, cellular responses were assessed. Wounded irradiated cells without HU showed an increase in cell viability and proliferation, which was confirmed by complete wound closure by day 4. Although wounded irradiated cells treated with 5 mM HU showed incomplete wound closure, these cells showed increased migration compared with that of control cells. This study showed that laser irradiation using an He-Ne laser with a fluence of 5 J/cm(2) stimulates cell viability. The HU results confirmed that laser irradiation promotes cell migration and proliferation. PMID:18214574

  11. 5-Aminolevulinic Acid-Mediated Sonodynamic Therapy Promotes Phenotypic Switching from Dedifferentiated to Differentiated Phenotype via Reactive Oxygen Species and p38 Mitogen-Activated Protein Kinase in Vascular Smooth Muscle Cells.

    PubMed

    Dan, Juhua; Sun, Xin; Li, Wanlu; Zhang, Yun; Li, Xuesong; Xu, Haobo; Li, Zhitao; Tian, Zhen; Guo, Shuyuan; Yao, Jianting; Gao, Weidong; Tian, Ye

    2015-06-01

    Sonodynamic therapy (SDT) has been found to inhibit in-stent restenosis in animal models. However, the mechanism is not fully elucidated. Here, we investigated the effects of 5-aminolevulinic acid (ALA)-mediated SDT (ALA-SDT) on vascular smooth muscle cells (VSMCs), a cause of restenosis, with a focus on SDT-induced phenotypic switching. Serum-induced dedifferentiated VSMCs were cultured with ALA (1 mm, 24 h) and exposed to ultrasound (0.8 W/cm(2)) for 5 min. Results indicated that ALA-SDT inhibited the migration and proliferation of VSMCs and enhanced the expression of differentiated phenotypic markers in VSMCs. Additionally, ALA-SDT increased intracellular reactive oxygen species accumulation and phosphorylated p38 mitogen-activated protein kinase in VSMCs. Inhibition of reactive oxygen species elevation or p38 mitogen-activated protein kinase activity abolished the expression of smooth muscle 22α (SM22α) in VSMCs induced by ALA-SDT. Taken together, these results suggest that ALA-SDT promotes transformation of the VSMC phenotype from the dedifferentiated to differentiated status via reactive oxygen species and activated p38 mitogen-activated protein kinase.

  12. Synchronized Cell Cycle Arrest Promotes Osteoclast Differentiation

    PubMed Central

    Kwon, Minsuk; Kim, Jin-Man; Lee, Kyunghee; Park, So-Young; Lim, Hyun-Sook; Kim, Taesoo; Jeong, Daewon

    2016-01-01

    Osteoclast progenitors undergo cell cycle arrest before differentiation into osteoclasts, induced by exposure to macrophage colony-stimulating factor (M-CSF) and receptor activator of nuclear factor-κB ligand (RANKL). The role of such cell cycle arrest in osteoclast differentiation has remained unclear, however. We here examined the effect of synchronized cell cycle arrest on osteoclast formation. Osteoclast progenitors deprived of M-CSF in culture adopted a uniform morphology and exhibited cell cycle arrest at the G0–G1 phase in association with both down-regulation of cyclins A and D1 as well as up-regulation of the cyclin-dependent kinase inhibitor p27Kip1. Such M-CSF deprivation also promoted the differentiation of osteoclast progenitors into multinucleated osteoclasts expressing high levels of osteoclast marker proteins such as NFATc1, c-Fos, Atp6v0d2, cathepsin K, and integrin β3 on subsequent exposure to M-CSF and RANKL. Our results suggest that synchronized arrest and reprogramming of osteoclast progenitors renders them poised to respond to inducers of osteoclast formation. Further characterization of such effects may facilitate induction of the differentiation of heterogeneous and multipotent cells into desired cell lineages. PMID:27517906

  13. Synchronized Cell Cycle Arrest Promotes Osteoclast Differentiation.

    PubMed

    Kwon, Minsuk; Kim, Jin-Man; Lee, Kyunghee; Park, So-Young; Lim, Hyun-Sook; Kim, Taesoo; Jeong, Daewon

    2016-01-01

    Osteoclast progenitors undergo cell cycle arrest before differentiation into osteoclasts, induced by exposure to macrophage colony-stimulating factor (M-CSF) and receptor activator of nuclear factor-κB ligand (RANKL). The role of such cell cycle arrest in osteoclast differentiation has remained unclear, however. We here examined the effect of synchronized cell cycle arrest on osteoclast formation. Osteoclast progenitors deprived of M-CSF in culture adopted a uniform morphology and exhibited cell cycle arrest at the G₀-G₁ phase in association with both down-regulation of cyclins A and D1 as well as up-regulation of the cyclin-dependent kinase inhibitor p27(Kip1). Such M-CSF deprivation also promoted the differentiation of osteoclast progenitors into multinucleated osteoclasts expressing high levels of osteoclast marker proteins such as NFATc1, c-Fos, Atp6v0d2, cathepsin K, and integrin β3 on subsequent exposure to M-CSF and RANKL. Our results suggest that synchronized arrest and reprogramming of osteoclast progenitors renders them poised to respond to inducers of osteoclast formation. Further characterization of such effects may facilitate induction of the differentiation of heterogeneous and multipotent cells into desired cell lineages. PMID:27517906

  14. Promoting Cell Proliferation Using Water Dispersible Germanium Nanowires

    PubMed Central

    Bezuidenhout, Michael; Liu, Pai; Singh, Shalini; Kiely, Maeve

    2014-01-01

    Group IV Nanowires have strong potential for several biomedical applications. However, to date their use remains limited because many are synthesised using heavy metal seeds and functionalised using organic ligands to make the materials water dispersible. This can result in unpredicted toxic side effects for mammalian cells cultured on the wires. Here, we describe an approach to make seedless and ligand free Germanium nanowires water dispersible using glutamic acid, a natural occurring amino acid that alleviates the environmental and health hazards associated with traditional functionalisation materials. We analysed the treated material extensively using Transmission electron microscopy (TEM), High resolution-TEM, and scanning electron microscope (SEM). Using a series of state of the art biochemical and morphological assays, together with a series of complimentary and synergistic cellular and molecular approaches, we show that the water dispersible germanium nanowires are non-toxic and are biocompatible. We monitored the behaviour of the cells growing on the treated germanium nanowires using a real time impedance based platform (xCELLigence) which revealed that the treated germanium nanowires promote cell adhesion and cell proliferation which we believe is as a result of the presence of an etched surface giving rise to a collagen like structure and an oxide layer. Furthermore this study is the first to evaluate the associated effect of Germanium nanowires on mammalian cells. Our studies highlight the potential use of water dispersible Germanium Nanowires in biological platforms that encourage anchorage-dependent cell growth. PMID:25237816

  15. Promoting cell proliferation using water dispersible germanium nanowires.

    PubMed

    Bezuidenhout, Michael; Liu, Pai; Singh, Shalini; Kiely, Maeve; Ryan, Kevin M; Kiely, Patrick A

    2014-01-01

    Group IV Nanowires have strong potential for several biomedical applications. However, to date their use remains limited because many are synthesised using heavy metal seeds and functionalised using organic ligands to make the materials water dispersible. This can result in unpredicted toxic side effects for mammalian cells cultured on the wires. Here, we describe an approach to make seedless and ligand free Germanium nanowires water dispersible using glutamic acid, a natural occurring amino acid that alleviates the environmental and health hazards associated with traditional functionalisation materials. We analysed the treated material extensively using Transmission electron microscopy (TEM), High resolution-TEM, and scanning electron microscope (SEM). Using a series of state of the art biochemical and morphological assays, together with a series of complimentary and synergistic cellular and molecular approaches, we show that the water dispersible germanium nanowires are non-toxic and are biocompatible. We monitored the behaviour of the cells growing on the treated germanium nanowires using a real time impedance based platform (xCELLigence) which revealed that the treated germanium nanowires promote cell adhesion and cell proliferation which we believe is as a result of the presence of an etched surface giving rise to a collagen like structure and an oxide layer. Furthermore this study is the first to evaluate the associated effect of Germanium nanowires on mammalian cells. Our studies highlight the potential use of water dispersible Germanium Nanowires in biological platforms that encourage anchorage-dependent cell growth.

  16. The EGF Receptor Promotes the Malignant Potential of Glioma by Regulating Amino Acid Transport System xc(-).

    PubMed

    Tsuchihashi, Kenji; Okazaki, Shogo; Ohmura, Mitsuyo; Ishikawa, Miyuki; Sampetrean, Oltea; Onishi, Nobuyuki; Wakimoto, Hiroaki; Yoshikawa, Momoko; Seishima, Ryo; Iwasaki, Yoshimi; Morikawa, Takayuki; Abe, Shinya; Takao, Ayumi; Shimizu, Misato; Masuko, Takashi; Nagane, Motoo; Furnari, Frank B; Akiyama, Tetsu; Suematsu, Makoto; Baba, Eishi; Akashi, Koichi; Saya, Hideyuki; Nagano, Osamu

    2016-05-15

    Extracellular free amino acids contribute to the interaction between a tumor and its microenvironment through effects on cellular metabolism and malignant behavior. System xc(-) is composed of xCT and CD98hc subunits and functions as a plasma membrane antiporter for the uptake of extracellular cystine in exchange for intracellular glutamate. Here, we show that the EGFR interacts with xCT and thereby promotes its cell surface expression and function in human glioma cells. EGFR-expressing glioma cells manifested both enhanced antioxidant capacity as a result of increased cystine uptake, as well as increased glutamate, which promotes matrix invasion. Imaging mass spectrometry also revealed that brain tumors formed in mice by human glioma cells stably overexpressing EGFR contained higher levels of reduced glutathione compared with those formed by parental cells. Targeted inhibition of xCT suppressed the EGFR-dependent enhancement of antioxidant capacity in glioma cells, as well as tumor growth and invasiveness. Our findings establish a new functional role for EGFR in promoting the malignant potential of glioma cells through interaction with xCT at the cell surface. Cancer Res; 76(10); 2954-63. ©2016 AACR.

  17. The Campylobacter jejuni Ferric Uptake Regulator Promotes Acid Survival and Cross-Protection against Oxidative Stress

    PubMed Central

    Askoura, Momen; Sarvan, Sabina; Couture, Jean-François

    2016-01-01

    Campylobacter jejuni is a prevalent cause of bacterial gastroenteritis in humans worldwide. The mechanisms by which C. jejuni survives stomach acidity remain undefined. In the present study, we demonstrated that the C. jejuni ferric uptake regulator (Fur) plays an important role in C. jejuni acid survival and acid-induced cross-protection against oxidative stress. A C. jejuni Δfur mutant was more sensitive to acid than the wild-type strain. Profiling of the acid stimulon of the C. jejuni Δfur mutant allowed us to uncover Fur-regulated genes under acidic conditions. In particular, Fur was found to upregulate genes involved in flagellar and cell envelope biogenesis upon acid stress, and mutants with deletions of these genes were found to be defective in surviving acid stress. Interestingly, prior acid exposure of C. jejuni cross-protected against oxidative stress in a catalase (KatA)- and Fur-dependent manner. Western blotting and reverse transcription-quantitative PCR revealed increased expression of KatA upon acid stress. Electrophoretic mobility shift assays (EMSAs) demonstrated that the binding affinity between Fur and the katA promoter is reduced in vitro under conditions of low pH, rationalizing the higher levels of expression of katA under acidic conditions. Strikingly, the Δfur mutant exhibited reduced virulence in both human epithelial cells and the Galleria mellonella infection model. Altogether, this is the first study showing that, in addition to its role in iron metabolism, Fur is an important regulator of C. jejuni acid responses and this function cross-protects against oxidative stress. Moreover, our results clearly demonstrate Fur's important role in C. jejuni pathogenesis. PMID:26883589

  18. Leukocyte protease binding to nucleic acids promotes nuclear localization and cleavage of nucleic acid binding proteins.

    PubMed

    Thomas, Marshall P; Whangbo, Jennifer; McCrossan, Geoffrey; Deutsch, Aaron J; Martinod, Kimberly; Walch, Michael; Lieberman, Judy

    2014-06-01

    Killer lymphocyte granzyme (Gzm) serine proteases induce apoptosis of pathogen-infected cells and tumor cells. Many known Gzm substrates are nucleic acid binding proteins, and the Gzms accumulate in the target cell nucleus by an unknown mechanism. In this study, we show that human Gzms bind to DNA and RNA with nanomolar affinity. Gzms cleave their substrates most efficiently when both are bound to nucleic acids. RNase treatment of cell lysates reduces Gzm cleavage of RNA binding protein targets, whereas adding RNA to recombinant RNA binding protein substrates increases in vitro cleavage. Binding to nucleic acids also influences Gzm trafficking within target cells. Preincubation with competitor DNA and DNase treatment both reduce Gzm nuclear localization. The Gzms are closely related to neutrophil proteases, including neutrophil elastase (NE) and cathepsin G. During neutrophil activation, NE translocates to the nucleus to initiate DNA extrusion into neutrophil extracellular traps, which bind NE and cathepsin G. These myeloid cell proteases, but not digestive serine proteases, also bind DNA strongly and localize to nuclei and neutrophil extracellular traps in a DNA-dependent manner. Thus, high-affinity nucleic acid binding is a conserved and functionally important property specific to leukocyte serine proteases. Furthermore, nucleic acid binding provides an elegant and simple mechanism to confer specificity of these proteases for cleavage of nucleic acid binding protein substrates that play essential roles in cellular gene expression and cell proliferation.

  19. Knockdown of delta-5-desaturase promotes the anti-cancer activity of dihomo-γ-linolenic acid and enhances the efficacy of chemotherapy in colon cancer cells expressing COX-2

    PubMed Central

    Xu, Yi; Yang, Xiaoyu; Zhao, Pinjing; Yang, Zhongyu; Yan, Changhui; Guo, Bin; Qian, Steven Y.

    2016-01-01

    Cyclooxygenase (COX), commonly overexpressed in cancer cells, is a major lipid peroxidizing enzyme that metabolizes polyunsaturated fatty acids (ω-3s and ω-6s). The COX-catalyzed free radical peroxidation of arachidonic acid (ω-6) can produce deleterious metabolites (e.g. 2-series prostaglandins) that are implicated in cancer development. Thus, COX inhibition has been intensively investigated as a complementary therapeutic strategy for cancer. However, our previous study has demonstrated that a free radical-derived by product (8-hydroxyoctanoic acid) formed from COX-catalyzed peroxidation of dihomo-γ-linolenic acid (DGLA, the precursor of arachidonic acid) can inhibit colon cancer cell growth. We thus hypothesize that the commonly overexpressed COX in cancer (~90% of colon cancer patients) can be taken advantage to suppress cell growth by knocking down delta-5-desaturase (D5D, a key enzyme that converts DGLA to arachidonic acid). In addition, D5D knockdown along with DGLA supplement may enhance the efficacy of chemotherapeutic drugs. After knocking down D5D in HCA-7 colony 29 cells and HT-29 cells (human colon cancer cell lines with high and low COX levels, respectively), the antitumor activity of DGLA was significantly enhanced along with the formation of a threshold range (~0.5–1.0 µM) of 8-hydroxyoctanoic acid. In contrast, DGLA treatment did not inhibit cell growth when D5D was not knocked down and only limited amount of 8-hydroxyoctanoic acid was formed. D5D knockdown along with DGLA treatment also enhanced the cytotoxicities of various chemotherapeutic drugs, including 5-fluorouracil, regorafenib, and irinotecan, potentially through the activation of pro-apoptotic proteins, e.g. p53 and caspase 9. For the first time, we have demonstrated that the overexpressed COX in cancer cells can be utilized in suppressing cancer cell growth. This finding may provide a new option besides COX inhibition to optimize cancer therapy. The outcome of this translational

  20. Leptin promotes cell proliferation and survival of trophoblastic cells.

    PubMed

    Magariños, María Paula; Sánchez-Margalet, Víctor; Kotler, Mónica; Calvo, Juan Carlos; Varone, Cecilia L

    2007-02-01

    Leptin, the 16-kDa protein product of the obese gene, was originally considered as an adipocyte-derived signaling molecule for the central control of metabolism. However, leptin has been suggested to be involved in other functions during pregnancy, particularly in placenta. In the present work, we studied a possible effect of leptin on trophoblastic cell proliferation, survival, and apoptosis. Recombinant human leptin added to JEG-3 and BeWo choriocarcinoma cell lines showed a stimulatory effect on cell proliferation up to 3 and 2.4 times, respectively, measured by (3)H-thymidine incorporation and cell counting. These effects were time and dose dependent. Maximal effect was achieved at 250 ng leptin/ml for JEG-3 cells and 50 ng leptin/ml for BeWo cells. Moreover, by inhibiting endogenous leptin expression with 2 microM of an antisense oligonucleotide (AS), cell proliferation was diminished. We analyzed cell population distribution during the different stages of cell cycle by fluorescence-activated cell sorting, and we found that leptin treatment displaced the cells towards a G2/M phase. We also found that leptin upregulated cyclin D1 expression, one of the key cell cycle-signaling proteins. Since proliferation and death processes are intimately related, the effect of leptin on cell apoptosis was investigated. Treatment with 2 microM leptin AS increased the number of apoptotic cells 60 times, as assessed by annexin V-fluorescein isothiocyanate/propidium iodide staining, and the caspase-3 activity was increased more than 2 fold. This effect was prevented by the addition of 100 ng leptin/ml. In conclusion, we provide evidence that suggests that leptin is a trophic and mitogenic factor for trophoblastic cells by virtue of its inhibiting apoptosis and promoting proliferation. PMID:17021346

  1. Root-Shoot Signaling crosstalk involved in the shoot growth promoting action of rhizospheric humic acids.

    PubMed

    Olaetxea, Maite; Mora, Verónica; García, Andrés Calderin; Santos, Leandro Azevedo; Baigorri, Roberto; Fuentes, Marta; Garnica, María; Berbara, Ricardo Luis Louro; Zamarreño, Angel Maria; Garcia-Mina, Jose M

    2016-01-01

    Numerous studies have shown the ability of humic substances to improve plant development. This action is normally reflected in an enhancement of crop yields and quality. However, the mechanisms responsible for this action of humic substances remain rather unknown. Our studies have shown that the shoot promoting action of sedimentary humic acids is dependent of its ability to increase root hydraulic conductivity through signaling pathways related to ABA, which in turn is affected in roots by humic acids in an IAA-NO dependent way. Furthermore, these studies also indicate that the primary action of humic acids in roots might also be physical, resulting from a transient mild stress caused by humic acids associated with a fouling-cleaning cycle of wall cell pores. Finally the role of alternative signal molecules, such as ROS, and corresponding signaling pathways are also discussed and modeled in the context of the above-mentioned framework. PMID:26966789

  2. Root-Shoot Signaling crosstalk involved in the shoot growth promoting action of rhizospheric humic acids

    PubMed Central

    Olaetxea, Maite; Mora, Verónica; García, Andrés Calderin; Santos, Leandro Azevedo; Baigorri, Roberto; Fuentes, Marta; Garnica, María; Berbara, Ricardo Luis Louro; Zamarreño, Angel Maria; Garcia-Mina, Jose M.

    2016-01-01

    ABSTRACT Numerous studies have shown the ability of humic substances to improve plant development. This action is normally reflected in an enhancement of crop yields and quality. However, the mechanisms responsible for this action of humic substances remain rather unknown. Our studies have shown that the shoot promoting action of sedimentary humic acids is dependent of its ability to increase root hydraulic conductivity through signaling pathways related to ABA, which in turn is affected in roots by humic acids in an IAA-NO dependent way. Furthermore, these studies also indicate that the primary action of humic acids in roots might also be physical, resulting from a transient mild stress caused by humic acids associated with a fouling-cleaning cycle of wall cell pores. Finally the role of alternative signal molecules, such as ROS, and corresponding signaling pathways are also discussed and modeled in the context of the above-mentioned framework. PMID:26966789

  3. Structural and functional characterization of the mouse fatty acid translocase promoter: activation during adipose differentiation.

    PubMed Central

    Teboul, L; Febbraio, M; Gaillard, D; Amri, E Z; Silverstein, R; Grimaldi, P A

    2001-01-01

    Fatty acid translocase (FAT/CD36) is a cell-surface glycoprotein that functions as a receptor/transporter for long-chain fatty acids (LCFAs), and interacts with other protein and lipid ligands. FAT/CD36 is expressed by various cell types, including platelets, monocytes/macrophages and endothelial cells, and tissues with an active LCFA metabolism, such as adipose, small intestine and heart. FAT/CD36 expression is induced during adipose cell differentiation and is transcriptionally up-regulated by LCFAs and thiazolidinediones in pre-adipocytes via a peroxisome-proliferator-activated receptor (PPAR)-mediated process. We isolated and analysed the murine FAT/CD36 promoter employing C(2)C(12)N cells directed to differentiate to either adipose or muscle. Transient transfection studies revealed that the 309 bp upstream from the start of exon 1 confer adipose specific activity. Sequence analysis of this DNA fragment revealed the presence of two imperfect direct repeat-1 elements. Electrophoretic mobility-shift assay demonstrated that these elements were peroxisome-proliferator-responsive elements (PPREs). Mutagenesis and transfection experiments indicated that both PPREs co-operate to drive strong promoter activity in adipose cells. We conclude that murine FAT/CD36 expression in adipose tissue is dependent upon transcriptional activation via PPARs through binding to two PPREs located at -245 to -233 bp and -120 to -108 bp from the transcription start site. PMID:11716758

  4. Abeta40 promotes neuronal cell fate in neural progenitor cells.

    PubMed

    Chen, Y; Dong, C

    2009-03-01

    Sequential cleavage of the amyloid precursor protein (APP) by beta- and then gamma- secretase gives rise to Abeta(1-40) (Abeta40), a major species of Abeta (beta-amyloid) produced by neurons under physiological conditions. Abeta(1-42) (Abeta42), a minor species of Abeta, is also produced by a similar but less understood mechanism of the gamma-secretase. The physiological functions of these Abeta species remain to be defined. In this report, we demonstrate that freshly prepared soluble Abeta40 significantly promotes neurogenesis in primary neural progenitor cells (NPCs). First, Abeta40 increases neuronal markers as determined by NeuN expression and Tuj1 promoter activity, differing from Abeta42, which induces astrocyte markers in NPCs. Second, Abeta40 induces neuronal differentiation at the end of S-phase in the cell cycle. Third, Abeta40 promotes NPC entry into S-phase, playing a role in NPC self-renewal. Interestingly, Abeta40 does not significantly increase apoptotic indexes such as DNA condensation and DNA fragmentation. In addition, Abeta40 does not augment caspase-3 activation in NeuN(+) or nestin(+) cells. Collectively, this report provides strong evidence that Abeta40 is a neurogenic factor and suggests that the debilitated function of Abeta40 in neurogenesis may account for the shortage of neurons in Alzheimer's disease.

  5. Promotion of viral internal ribosomal entry site-mediated translation under amino acid starvation.

    PubMed

    Licursi, Maria; Komatsu, Yumiko; Pongnopparat, Theerawat; Hirasawa, Kensuke

    2012-05-01

    Cap-dependent and internal ribosomal entry site (IRES)-mediated translation are regulated differently within cells. Viral IRES-mediated translation often remains active when cellular cap-dependent translation is severely impaired under cellular stresses induced by virus infection. To investigate how cellular stresses influence the efficiency of viral IRES-mediated translation, we used a bicistronic luciferase reporter construct harbouring IRES elements from the following viruses: encephalomyocarditis virus (EMCV), foot-and-mouth disease virus (FMDV), hepatitis C virus (HCV) or human rhinovirus (HRV). NIH3T3 cells transfected with these bicistronic reporter constructs were subjected to different cellular stresses. Increased translation initiation was only observed under amino acid starvation when EMCV or FMDV IRES elements were present. To identify cellular mechanisms that promoted viral IRES-mediated translation, we tested the involvement of eukaryotic initiation factor 4E-binding protein (4E-BP), general control non-depressed 2 (GCN2) and eukaryotic initiation factor 2B (eIF2B), as these are known to be modulated under amino acid starvation. Knockdown of 4E-BP1 impaired the promotion of EMCV and FMDV IRES-mediated translation under amino acid starvation, whereas GCN2 and eIF2B were not involved. To further investigate how 4E-BP1 regulates translation initiated by EMCV and FMDV IRES elements, we used a phosphoinositide kinase-3 inhibitor (LY294002), an mTOR inhibitor (Torin1) or leucine starvation to mimic 4E-BP1 dephosphorylation induced by amino acid starvation. 4E-BP1 dephosphorylation induced by the treatments was not sufficient to promote viral IRES-mediated translation. These results suggest that 4E-BP1 regulates EMCV and FMDV IRES-mediated translation under amino acid starvation, but not via its dephosphorylation. PMID:22302880

  6. Homophilic Protocadherin Cell-Cell Interactions Promote Dendrite Complexity.

    PubMed

    Molumby, Michael J; Keeler, Austin B; Weiner, Joshua A

    2016-05-01

    Growth of a properly complex dendrite arbor is a key step in neuronal differentiation and a prerequisite for neural circuit formation. Diverse cell surface molecules, such as the clustered protocadherins (Pcdhs), have long been proposed to regulate circuit formation through specific cell-cell interactions. Here, using transgenic and conditional knockout mice to manipulate γ-Pcdh repertoire in the cerebral cortex, we show that the complexity of a neuron's dendritic arbor is determined by homophilic interactions with other cells. Neurons expressing only one of the 22 γ-Pcdhs can exhibit either exuberant or minimal dendrite complexity, depending only on whether surrounding cells express the same isoform. Furthermore, loss of astrocytic γ-Pcdhs, or disruption of astrocyte-neuron homophilic matching, reduces dendrite complexity cell non-autonomously. Our data indicate that γ-Pcdhs act locally to promote dendrite arborization via homophilic matching, and they confirm that connectivity in vivo depends on molecular interactions between neurons and between neurons and astrocytes.

  7. Fatty acids identified in the Burmese python promote beneficial cardiac growth.

    PubMed

    Riquelme, Cecilia A; Magida, Jason A; Harrison, Brooke C; Wall, Christopher E; Marr, Thomas G; Secor, Stephen M; Leinwand, Leslie A

    2011-10-28

    Burmese pythons display a marked increase in heart mass after a large meal. We investigated the molecular mechanisms of this physiological heart growth with the goal of applying this knowledge to the mammalian heart. We found that heart growth in pythons is characterized by myocyte hypertrophy in the absence of cell proliferation and by activation of physiological signal transduction pathways. Despite high levels of circulating lipids, the postprandial python heart does not accumulate triglycerides or fatty acids. Instead, there is robust activation of pathways of fatty acid transport and oxidation combined with increased expression and activity of superoxide dismutase, a cardioprotective enzyme. We also identified a combination of fatty acids in python plasma that promotes physiological heart growth when injected into either pythons or mice.

  8. Biotransformation of cinnamic acid, p-coumaric acid, caffeic acid, and ferulic acid by plant cell cultures of Eucalyptus perriniana.

    PubMed

    Katsuragi, Hisashi; Shimoda, Kei; Kubota, Naoji; Nakajima, Nobuyoshi; Hamada, Hatsuyuki; Hamada, Hiroki

    2010-01-01

    Biotransformations of phenylpropanoids such as cinnamic acid, p-coumaric acid, caffeic acid, and ferulic acid were investigated with plant-cultured cells of Eucalyptus perriniana. The plant-cultured cells of E. perriniana converted cinnamic acid into cinnamic acid β-D-glucopyranosyl ester, p-coumaric acid, and 4-O-β-D-glucopyranosylcoumaric acid. p-Coumaric acid was converted into 4-O-β-D-glucopyranosylcoumaric acid, p-coumaric acid β-D-glucopyranosyl ester, 4-O-β-D-glucopyranosylcoumaric acid β-D-glucopyranosyl ester, a new compound, caffeic acid, and 3-O-β-D-glucopyranosylcaffeic acid. On the other hand, incubation of caffeic acid with cultured E. perriniana cells gave 3-O-β-D-glucopyranosylcaffeic acid, 3-O-(6-O-β-D-glucopyranosyl)-β-D-glucopyranosylcaffeic acid, a new compound, 3-O-β-D-glucopyranosylcaffeic acid β-D-glucopyranosyl ester, 4-O-β-D-glucopyranosylcaffeic acid, 4-O-β-D-glucopyranosylcaffeic acid β-D-glucopyranosyl ester, ferulic acid, and 4-O-β-D-glucopyranosylferulic acid. 4-O-β-D-Glucopyranosylferulic acid, ferulic acid β-D-glucopyranosyl ester, and 4-O-β-D-glucopyranosylferulic acid β-D-glucopyranosyl ester were isolated from E. perriniana cells treated with ferulic acid.

  9. The Polyunsaturated Fatty Acids Arachidonic Acid and Docosahexaenoic Acid Induce Mouse Dendritic Cells Maturation but Reduce T-Cell Responses In Vitro

    PubMed Central

    Carlsson, Johan A.; Wold, Agnes E.; Sandberg, Ann-Sofie; Östman, Sofia M.

    2015-01-01

    Long-chain polyunsaturated fatty acids (PUFAs) might regulate T-cell activation and lineage commitment. Here, we measured the effects of omega-3 (n-3), n-6 and n-9 fatty acids on the interaction between dendritic cells (DCs) and naïve T cells. Spleen DCs from BALB/c mice were cultured in vitro with ovalbumin (OVA) with 50 μM fatty acids; α-linolenic acid, arachidonic acid (AA), eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA), linoleic acid or oleic acid and thereafter OVA-specific DO11.10 T cells were added to the cultures. Fatty acids were taken up by the DCs, as shown by gas chromatography analysis. After culture with arachidonic acid or DHA CD11c+ CD11b+ and CD11c+ CD11bneg DCs expressed more CD40, CD80, CD83, CD86 and PDL-1, while IAd remained unchanged. However, fewer T cells co-cultured with these DCs proliferated (CellTrace Violetlow) and expressed CD69 or CD25, while more were necrotic (7AAD+). We noted an increased proportion of T cells with a regulatory T cell (Treg) phenotype, i.e., when gating on CD4+ FoxP3+ CTLA-4+, CD4+ FoxP3+ Helios+ or CD4+ FoxP3+ PD-1+, in co-cultures with arachidonic acid- or DHA-primed DCs relative to control cultures. The proportion of putative Tregs was inversely correlated to T-cell proliferation, indicating a suppressive function of these cells. With arachidonic acid DCs produced higher levels of prostaglandin E2 while T cells produced lower amounts of IL-10 and IFNγ. In conclusion arachidonic acid and DHA induced up-regulation of activation markers on DCs. However arachidonic acid- and DHA-primed DCs reduced T-cell proliferation and increased the proportion of T cells expressing FoxP3, indicating that these fatty acids can promote induction of regulatory T cells. PMID:26619195

  10. Phosphate limitation promotes unsaturated fatty acids and arachidonic acid biosynthesis by microalgae Porphyridium purpureum.

    PubMed

    Su, Gaomin; Jiao, Kailin; Li, Zheng; Guo, Xiaoyi; Chang, Jingyu; Ndikubwimana, Theoneste; Sun, Yong; Zeng, Xianhai; Lu, Yinghua; Lin, Lu

    2016-07-01

    Polyunsaturated fatty acids (PUFAs) are highly appreciated on their nutritive value for human health and aquaculture. P. purpureum, one of the red microalgae acknowledged as a promising accumulator of ARA, was chosen as the target algae in the present research. Effects of sodium bicarbonate (0.04-1.2 g/L), temperature (25, 30 and 33 °C) and phosphate (0.00-0.14 g/L) on biomass yield, total fatty acids (TFA) and arachidonic acid (ARA) accumulation were investigated systemically. NaHCO3 dose of 0.8 g/L and moderate temperature of 30 °C were preferred. In addition, TFA and ARA production were significantly enhanced by an appropriate concentration of phosphate, and the highest TFA yield of 666.38 mg/L and ARA yield of 159.74 mg/L were obtained at a phosphate concentration of 0.035 g/L. Interestingly, with phosphate concentration continuing to fall, UFA/TFA and ARA/EPA ratios were increased accordingly, suggesting that phosphate limitation promoted unsaturated fatty acids and arachidonic acid biosynthesis. Low concentration of phosphate may be favored to increase the enzymatic activities of ∆6-desaturase, which played a key role in catalyzing the conversion of C16:0 to C18:2, and thus the selectivity of UFA increased. Meanwhile, the increase of ARA selectivity could be attributed to ω6 pathway promotion and ∆17-desaturase activity inhibition with phosphate limitation. Phosphate limitation strategy enhanced unsaturated fatty acids and ARA biosynthesis in P. purpureum, and can be applied in commercial scale manufacturing and commercialization of ARA. PMID:27004948

  11. Phosphate limitation promotes unsaturated fatty acids and arachidonic acid biosynthesis by microalgae Porphyridium purpureum.

    PubMed

    Su, Gaomin; Jiao, Kailin; Li, Zheng; Guo, Xiaoyi; Chang, Jingyu; Ndikubwimana, Theoneste; Sun, Yong; Zeng, Xianhai; Lu, Yinghua; Lin, Lu

    2016-07-01

    Polyunsaturated fatty acids (PUFAs) are highly appreciated on their nutritive value for human health and aquaculture. P. purpureum, one of the red microalgae acknowledged as a promising accumulator of ARA, was chosen as the target algae in the present research. Effects of sodium bicarbonate (0.04-1.2 g/L), temperature (25, 30 and 33 °C) and phosphate (0.00-0.14 g/L) on biomass yield, total fatty acids (TFA) and arachidonic acid (ARA) accumulation were investigated systemically. NaHCO3 dose of 0.8 g/L and moderate temperature of 30 °C were preferred. In addition, TFA and ARA production were significantly enhanced by an appropriate concentration of phosphate, and the highest TFA yield of 666.38 mg/L and ARA yield of 159.74 mg/L were obtained at a phosphate concentration of 0.035 g/L. Interestingly, with phosphate concentration continuing to fall, UFA/TFA and ARA/EPA ratios were increased accordingly, suggesting that phosphate limitation promoted unsaturated fatty acids and arachidonic acid biosynthesis. Low concentration of phosphate may be favored to increase the enzymatic activities of ∆6-desaturase, which played a key role in catalyzing the conversion of C16:0 to C18:2, and thus the selectivity of UFA increased. Meanwhile, the increase of ARA selectivity could be attributed to ω6 pathway promotion and ∆17-desaturase activity inhibition with phosphate limitation. Phosphate limitation strategy enhanced unsaturated fatty acids and ARA biosynthesis in P. purpureum, and can be applied in commercial scale manufacturing and commercialization of ARA.

  12. An Alternative Retinoic Acid-responsive Stra6 Promoter Regulated in Response to Retinol Deficiency*

    PubMed Central

    Laursen, Kristian B.; Kashyap, Vasundhra; Scandura, Joseph; Gudas, Lorraine J.

    2015-01-01

    Cellular uptake of vitamin A (retinol) is essential for many biological functions. The Stra6 protein binds the serum retinol-binding protein, RBP4, and acts in conjunction with the enzyme lecithin:retinol acyltransferase to facilitate retinol uptake in some cell types. We show that in embryonic stem (ES) cells and in some tissues, the Stra6 gene encodes two distinct mRNAs transcribed from two different promoters. Whereas both are all-trans-retinoic acid (RA)-responsive in ES cells, the downstream promoter contains a half-site RA response element (RARE) and drives an ∼13-fold, RA-associated increase in luciferase reporter activity. We employed CRISPR-Cas9 genome editing to show that the endogenous RARE is required for RA-induced transcription of both Stra6 isoforms. We further demonstrate that in ES cells, 1) both RARγ and RXRα are present at the Stra6 RARE; 2) RA increases co-activator p300 (KAT3B) binding and histone H3 Lys-27 acetylation at both promoters; 3) RA decreases Suz12 levels and histone H3 Lys-27 trimethylation epigenetic marks at both promoters; and 4) these epigenetic changes are diminished in the absence of RARγ. In the brains of WT mice, both the longer and the shorter Stra6 transcript (Stra6L and Stra6S, respectively) are highly expressed, whereas these transcripts are found only at low levels in RARγ−/− mice. In the brains of vitamin A-deficient mice, both Stra6L and Stra6S levels are decreased. In contrast, in the vitamin A-deficient kidneys, the Stra6L levels are greatly increased, whereas Stra6S levels are decreased. Our data show that kidneys respond to retinol deficiency by differential Stra6 promoter usage, which may play a role in the retention of retinol when vitamin A is low. PMID:25544292

  13. Ascorbic acid transport into cultured pituitary cells

    SciTech Connect

    Cullen, E.I.; May, V.; Eipper, R.A.

    1986-05-01

    An amidating enzyme designated peptidyl-glycine ..cap alpha..-amidating monooxygenase (PAM) has been studied in a variety of tissues and is dependent on molecular oxygen and stimulated by copper and ascorbic acid. To continue investigating the relationship among cellular ascorbic acid concentrations, amidating ability, and PAM activity, the authors studied ascorbic acid transport in three cell preparations that contain PAM and produce amidated peptides: primary cultures of rat anterior and intermediate pituitary and mouse AtT-20 tumor cells. When incubated in 50 ..mu..M (/sup 14/C)ascorbic acid all three cell preparations concentrated ascorbic acid 20- to 40-fold, producing intracellular ascorbate concentrations of 1 to 2 mM, based on experimentally determined cell volumes. All three cell preparations displayed saturable ascorbic acid uptake with half-maximal initial rates occurring between 9 and 18 ..mu..M ascorbate. Replacing NaCl in the uptake buffer with choline chloride significantly diminished ascorbate uptake in all three preparations. Ascorbic acid efflux from these cells was slow, displaying half-lives of 7 hours. Unlike systems that transport dehydroascorbic acid, the transport system for ascorbic acid in these cells was not inhibited by glucose. Thus, ascorbate is transported into pituitary cells by a sodium-dependent, active transport system.

  14. Fetal liver stromal cells promote hematopoietic cell expansion

    SciTech Connect

    Zhou, Kun; Hu, Caihong; Zhou, Zhigang; Huang, Lifang; Liu, Wenli; Sun, Hanying

    2009-09-25

    Future application of hematopoietic stem and progenitor cells (HSPCs) in clinical therapies largely depends on their successful expansion in vitro. Fetal liver (FL) is a unique hematopoietic organ in which hematopoietic cells markedly expand in number, but the mechanisms involved remain unclear. Stromal cells (StroCs) have been suggested to provide a suitable cellular environment for in vitro expansion of HSPCs. In this study, murine StroCs derived from FL at E14.5, with a high level of Sonic hedgehog (Shh) and Wnt expression, were found to have an increased ability to support the proliferation of HSPCs. This effect was inhibited by blocking Shh signaling. Supplementation with soluble Shh-N promoted the proliferation of hematopoietic cells by activating Wnt signaling. Our findings suggest that FL-derived StroCs support proliferation of HSPCs via Shh inducing an autocrine Wnt signaling loop. The use of FL-derived StroCs and regulation of the Shh pathway might further enhance HPSC expansion.

  15. Stabilizing platinum in phosphoric acid fuel cells

    NASA Technical Reports Server (NTRS)

    Remick, R. J.

    1982-01-01

    Platinum sintering on phosphoric acid fuel cell cathodes is discussed. The cathode of the phosphoric acid fuel cell uses a high surface area platinum catalyst dispersed on a conductive carbon support to minimize both cathode polarization and fabrication costs. During operation, however, the active surface area of these electrodes decreases, which in turn leads to decreased cell performance. This loss of active surface area is a major factor in the degradation of fuel cell performance over time.

  16. Recombinant sialidase NanA (rNanA) cleaves α2-3 linked sialic acid of host cell surface N-linked glycoprotein to promote Edwardsiella tarda infection.

    PubMed

    Chigwechokha, Petros Kingstone; Tabata, Mutsumi; Shinyoshi, Sayaka; Oishi, Kazuki; Araki, Kyosuke; Komatsu, Masaharu; Itakura, Takao; Shiozaki, Kazuhiro

    2015-11-01

    Edwardsiella tarda is one of the major pathogenic bacteria affecting both marine and freshwater fish species. Sialidase NanA expressed endogenously in E. tarda is glycosidase removing sialic acids from glycoconjugates. Recently, the relationship of NanA sialidase activity to E. tarda infection has been reported, however, the mechanism with which sialidase NanA aids the pathogenicity of E. tarda remained unclear. Here, we comprehensively determined the biochemical properties of NanA towards various substrates in vitro to provide novel insights on the potential NanA target molecule at the host cell. GAKS cell pretreated with recombinant NanA showed increased susceptibility to E. tarda infection. Moreover, sialidase inhibitor treated E. tarda showed a significantly reduced ability to infect GAKS cells. These results indicate that NanA-induced desialylation of cell surface glycoconjugates is essential for the initial step of E. tarda infection. Among the natural substrates, NanA exhibited the highest activity towards 3-sialyllactose, α2-3 linked sialic acid carrying sialoglycoconjugates. Supporting this finding, intact GAKS cell membrane exposed to recombinant NanA showed changes of glycoconjugates only in α2-3 sialo-linked glycoproteins, but not in glycolipids and α2-6 sialo-linked glycoproteins. Lectin staining of cell surface glycoprotein provided further evidence that α2-3 sialo-linkage of the N-linked glycoproteins was the most plausible target of NanA sialidase. To confirm the significance of α2-3 sialo-linkage desialylation for E. tarda infection, HeLa cells which possessed lower amount of α2-3 sialo-linkage glycoprotein were used for infection experiment along with GAKS cells. As a result, infection of HeLa cells by E. tarda was significantly reduced when compared to GAKS cells. Furthermore, E. tarda infection was significantly inhibited by mannose pretreatment suggesting that the bacterium potentially recognizes and binds to mannose or mannose containing

  17. 5-azacytidine promotes terminal differentiation of hepatic progenitor cells.

    PubMed

    He, Yun; Cui, Jiejie; He, Tongchuan; Bi, Yang

    2015-08-01

    5-azacytidine (5-azaC) is known to induce cardiomyocyte differentiation. However, its function in hepatocyte differentiation is unclear. The present study investigated the in vitro capability of 5-azaC to promote maturation and differentiation of mouse embryonic hepatic progenitor cells, with the aim of developing an approach for improving hepatic differentiation. Mouse embryonic hepatic progenitor cells (HP14.5 cells) were treated with 5-azaC at concentrations from 0 to 20 μmol/l, in addition to hepatocyte induction culture medium. Hepatocyte induction medium induces HP14.5 cell differentiation. 5-azaC may enhance the albumin promotor-driven Gaussia luciferase (ALB-GLuc) activity in induced HP14.5 cells. In the present study 2 μmol/l was found to be the optimum concentration with which to achieve this. The expression of hepatocyte-associated factors was not significantly different between the group treated with 5-azaC alone and the control group. The mRNA levels of ALB; cytokeratin 18 (CK18); tyrosine aminotransferase (TAT); and cytochrome p450, family 1, member A1 (CYP1A1); in addition to the protein levels of ALB, CK18 and uridine diphosphate glucuronyltransferase 1A (UGT1A) in the induced group with 5-azaC, were higher than those in the induced group without 5-azaC, although no significant differences were detected in expression of the hepatic stem cell markers, DLK and α-fetoprotein, between the two groups. Treatment with 5-azaC alone did not affect glycogen synthesis or indocyanine green (ICG) metabolic function in HP14.5 cells, although it significantly increased ICG uptake and periodic acid-Schiff-positive cell numbers amongst HP14.5 cells. Therefore, the present study demonstrated that treatment with 5-azaC alone exerted no effects on the maturation and differentiation of HP14.5 cells. However, 5-azaC exhibited a synergistic effect on the terminal differentiation of induced hepatic progenitor cells in association with a hepatic induction medium. PMID

  18. Lysophosphatidic acid activates Arf6 to promote the mesenchymal malignancy of renal cancer.

    PubMed

    Hashimoto, Shigeru; Mikami, Shuji; Sugino, Hirokazu; Yoshikawa, Ayumu; Hashimoto, Ari; Onodera, Yasuhito; Furukawa, Shotaro; Handa, Haruka; Oikawa, Tsukasa; Okada, Yasunori; Oya, Mototsugu; Sabe, Hisataka

    2016-01-01

    Acquisition of mesenchymal properties by cancer cells is critical for their malignant behaviour, but regulators of the mesenchymal molecular machinery and how it is activated remain elusive. Here we show that clear cell renal cell carcinomas (ccRCCs) frequently utilize the Arf6-based mesenchymal pathway to promote invasion and metastasis, similar to breast cancers. In breast cancer cells, ligand-activated receptor tyrosine kinases employ GEP100 to activate Arf6, which then recruits AMAP1; and AMAP1 then binds to the mesenchymal-specific protein EPB41L5, which promotes epithelial-mesenchymal transition and focal adhesion dynamics. In renal cancer cells, lysophosphatidic acid (LPA) activates Arf6 via its G-protein-coupled receptors, in which GTP-Gα12 binds to EFA6. The Arf6-based pathway may also contribute to drug resistance. Our results identify a specific mesenchymal molecular machinery of primary ccRCCs, which is triggered by a product of autotaxin and it is associated with poor outcome of patients. PMID:26854204

  19. Lysophosphatidic acid activates Arf6 to promote the mesenchymal malignancy of renal cancer

    PubMed Central

    Hashimoto, Shigeru; Mikami, Shuji; Sugino, Hirokazu; Yoshikawa, Ayumu; Hashimoto, Ari; Onodera, Yasuhito; Furukawa, Shotaro; Handa, Haruka; Oikawa, Tsukasa; Okada, Yasunori; Oya, Mototsugu; Sabe, Hisataka

    2016-01-01

    Acquisition of mesenchymal properties by cancer cells is critical for their malignant behaviour, but regulators of the mesenchymal molecular machinery and how it is activated remain elusive. Here we show that clear cell renal cell carcinomas (ccRCCs) frequently utilize the Arf6-based mesenchymal pathway to promote invasion and metastasis, similar to breast cancers. In breast cancer cells, ligand-activated receptor tyrosine kinases employ GEP100 to activate Arf6, which then recruits AMAP1; and AMAP1 then binds to the mesenchymal-specific protein EPB41L5, which promotes epithelial–mesenchymal transition and focal adhesion dynamics. In renal cancer cells, lysophosphatidic acid (LPA) activates Arf6 via its G-protein-coupled receptors, in which GTP-Gα12 binds to EFA6. The Arf6-based pathway may also contribute to drug resistance. Our results identify a specific mesenchymal molecular machinery of primary ccRCCs, which is triggered by a product of autotaxin and it is associated with poor outcome of patients. PMID:26854204

  20. Gallic Acid Promotes Wound Healing in Normal and Hyperglucidic Conditions.

    PubMed

    Yang, Dong Joo; Moh, Sang Hyun; Son, Dong Hwee; You, Seunghoon; Kinyua, Ann W; Ko, Chang Mann; Song, Miyoung; Yeo, Jinhee; Choi, Yun-Hee; Kim, Ki Woo

    2016-01-01

    Skin is the outermost layer of the human body that is constantly exposed to environmental stressors, such as UV radiation and toxic chemicals, and is susceptible to mechanical wounding and injury. The ability of the skin to repair injuries is paramount for survival and it is disrupted in a spectrum of disorders leading to skin pathologies. Diabetic patients often suffer from chronic, impaired wound healing, which facilitate bacterial infections and necessitate amputation. Here, we studied the effects of gallic acid (GA, 3,4,5-trihydroxybenzoic acid; a plant-derived polyphenolic compound) on would healing in normal and hyperglucidic conditions, to mimic diabetes, in human keratinocytes and fibroblasts. Our study reveals that GA is a potential antioxidant that directly upregulates the expression of antioxidant genes. In addition, GA accelerated cell migration of keratinocytes and fibroblasts in both normal and hyperglucidic conditions. Further, GA treatment activated factors known to be hallmarks of wound healing, such as focal adhesion kinases (FAK), c-Jun N-terminal kinases (JNK), and extracellular signal-regulated kinases (Erk), underpinning the beneficial role of GA in wound repair. Therefore, our results demonstrate that GA might be a viable wound healing agent and a potential intervention to treat wounds resulting from metabolic complications. PMID:27399667

  1. Bile acids induce hepatic differentiation of mesenchymal stem cells

    PubMed Central

    Sawitza, Iris; Kordes, Claus; Götze, Silke; Herebian, Diran; Häussinger, Dieter

    2015-01-01

    Mesenchymal stem cells (MSC) have the potential to differentiate into multiple cell lineages and their therapeutic potential has become obvious. In the liver, MSC are represented by stellate cells which have the potential to differentiate into hepatocytes after stimulation with growth factors. Since bile acids can promote liver regeneration, their influence on liver-resident and bone marrow-derived MSC was investigated. Physiological concentrations of bile acids such as tauroursodeoxycholic acid were able to initiate hepatic differentiation of MSC via the farnesoid X receptor and transmembrane G-protein-coupled bile acid receptor 5 as investigated with knockout mice. Notch, hedgehog, transforming growth factor-β/bone morphogenic protein family and non-canonical Wnt signalling were also essential for bile acid-mediated differentiation, whereas β-catenin-dependent Wnt signalling was able to attenuate this process. Our findings reveal bile acid-mediated signalling as an alternative way to induce hepatic differentiaion of stem cells and highlight bile acids as important signalling molecules during liver regeneration. PMID:26304833

  2. Opposing effects of bile acids deoxycholic acid and ursodeoxycholic acid on signal transduction pathways in oesophageal cancer cells.

    PubMed

    Abdel-Latif, Mohamed M; Inoue, Hiroyasu; Reynolds, John V

    2016-09-01

    Ursodeoxycholic acid (UDCA) was reported to reduce bile acid toxicity, but the mechanisms underlying its cytoprotective effects are not fully understood. The aim of the present study was to examine the effects of UDCA on the modulation of deoxycholic acid (DCA)-induced signal transduction in oesophageal cancer cells. Nuclear factor-κB (NF-κB) and activator protein-1 (AP-1) activity was assessed using a gel shift assay. NF-κB activation and translocation was performed using an ELISA-based assay and immunofluorescence analysis. COX-2 expression was analysed by western blotting and COX-2 promoter activity was assessed by luciferase assay. DCA induced NF-κB and AP-1 DNA-binding activities in SKGT-4 and OE33 cells. UDCA pretreatment inhibited DCA-induced NF-κB and AP-1 activation and NF-κB translocation. This inhibitory effect was coupled with a blockade of IκB-α degradation and inhibition of phosphorylation of IKK-α/β and ERK1/2. Moreover, UDCA pretreatment inhibited COX-2 upregulation. Using transient transfection of the COX-2 promoter, UDCA pretreatment abrogated DCA-induced COX-2 promoter activation. In addition, UDCA protected oesophageal cells from the apoptotic effects of deoxycholate. Our findings indicate that UDCA inhibits DCA-induced signalling pathways in oesophageal cancer cells. These data indicate a possible mechanistic role for the chemopreventive actions of UDCA in oesophageal carcinogenesis.

  3. Chimeric promoter mediates guard cell-specific gene expression in tobacco under water deficit.

    PubMed

    Na, Jong-Kuk; Metzger, James D

    2014-09-01

    The engineering of stomatal activity under water deficit through guard cell-specific gene regulation is an effective approach to improve drought tolerance of crops but it requires an appropriate promoter(s) inducible by water deficit in guard cells. We report that a chimeric promoter can induce guard cell-specific gene expression under water deficit. A chimeric promoter, p4xKST82-rd29B, was constructed using a tetramer of the 82 bp guard cell-specific regulatory region of potato KST1 promoter (4xKST82) and Arabidopsis dehydration-responsive rd29B promoter. Transgenic tobacco plants carrying p4xKST82-rd29B:mGFP-GUS exhibited GUS expression in response to water deficit. GUS enzyme activity of p4xKST82-rd29B:mGFP-GUS transgenic plants increased ~300 % by polyethylene glycol treatment compared to that of control plant but not by abscisic acid (ABA), indicating that the p4xKST82-rd29B chimeric promoter can be used to induce the guard cell-specific expression of genes of interest in response to water deficit in an ABA-independent manner.

  4. Retinol Promotes In Vitro Growth of Proximal Colon Organoids through a Retinoic Acid-Independent Mechanism

    PubMed Central

    Nibe, Yoichi; Akiyama, Shintaro; Matsumoto, Yuka; Nozaki, Kengo; Fukuda, Masayoshi; Hayashi, Ayumi; Mizutani, Tomohiro; Oshima, Shigeru; Watanabe, Mamoru; Nakamura, Tetsuya

    2016-01-01

    Retinol (ROL), the alcohol form of vitamin A, is known to control cell fate decision of various types of stem cells in the form of its active metabolite, retinoic acid (RA). However, little is known about whether ROL has regulatory effects on colonic stem cells. We examined in this study the effect of ROL on the growth of murine normal colonic cells cultured as organoids. As genes involved in RA synthesis from ROL were differentially expressed along the length of the colon, we tested the effect of ROL on proximal and distal colon organoids separately. We found that organoid forming efficiency and the expression level of Lgr5, a marker gene for colonic stem cells were significantly enhanced by ROL in the proximal colon organoids, but not in the distal ones. Interestingly, neither retinaldehyde (RAL), an intermediate product of the ROL-RA pathway, nor RA exhibited growth promoting effects on the proximal colon organoids, suggesting that ROL-dependent growth enhancement in organoids involves an RA-independent mechanism. This was confirmed by the observation that an inhibitor for RA-mediated gene transcription did not abrogate the effect of ROL on organoids. This novel role of ROL in stem cell maintenance in the proximal colon provides insights into the mechanism of region-specific regulation for colonic stem cell maintenance. PMID:27564706

  5. Diphenylarsinic acid promotes degradation of glutaminase C by mitochondrial Lon protease.

    PubMed

    Kita, Kayoko; Suzuki, Toshihide; Ochi, Takafumi

    2012-05-25

    Glutaminase C (GAC), a splicing variant of the kidney-type glutaminase (KGA) gene, is a vital mitochondrial enzyme protein that catalyzes glutamine to glutamate. Earlier studies have shown that GAC proteins in the human hepatocarcinoma cell line, HepG2, were down-regulated by diphenylarsinic acid (DPAA), but the mechanism by which DPAA induced GAC protein down-regulation remained poorly understood. Here, we showed that DPAA promoted GAC protein degradation without affecting GAC transcription and translation. Moreover, DPAA-induced GAC proteolysis was mediated by mitochondrial Lon protease. DPAA insolubilized 0.5% Triton X-100-soluble GAC protein and promoted the accumulation of insoluble GAC in Lon protease knockdown cells. DPAA destroyed the native tetrameric GAC conformation and promoted an increase in the unassembled form of GAC when DPAA was incubated with cell extracts. Decreases in the tetrameric form of GAC were observed in cells exposed to DPAA, and decreases occurred prior to a decrease in total GAC protein levels. In addition, decreases in the tetrameric form of GAC were observed independently with Lon protease. Mitochondrial heat shock protein 70 is known to be an indispensable protein that can bind to misfolded proteins, thereby supporting degradation of proteins sensitive to Lon protease. When cells were incubated with DPAA, GAC proteins that can bind with mtHsp70 increased. Interestingly, the association of mtHsp70 with GAC protein increased when the tetrameric form of GAC was reduced. These results suggest that degradation of native tetrameric GAC by DPAA may be a trigger in GAC protein degradation by Lon protease.

  6. Aminomethylphosphonic acid and methoxyacetic acid induce apoptosis in prostate cancer cells.

    PubMed

    Parajuli, Keshab R; Zhang, Qiuyang; Liu, Sen; You, Zongbing

    2015-01-01

    Aminomethylphosphonic acid (AMPA) and its parent compound herbicide glyphosate are analogs to glycine, which have been reported to inhibit proliferation and promote apoptosis of cancer cells, but not normal cells. Methoxyacetic acid (MAA) is the active metabolite of ester phthalates widely used in industry as gelling, viscosity and stabilizer; its exposure is associated with developmental and reproductive toxicities in both rodents and humans. MAA has been reported to suppress prostate cancer cell growth by inducing growth arrest and apoptosis. However, it is unknown whether AMPA and MAA can inhibit cancer cell growth. In this study, we found that AMPA and MAA inhibited cell growth in prostate cancer cell lines (LNCaP, C4-2B, PC-3 and DU-145) through induction of apoptosis and cell cycle arrest at the G1 phase. Importantly, the AMPA-induced apoptosis was potentiated with the addition of MAA, which was due to downregulation of the anti-apoptotic gene baculoviral inhibitor of apoptosis protein repeat containing 2 (BIRC2), leading to activation of caspases 7 and 3. These results demonstrate that the combination of AMPA and MAA can promote the apoptosis of prostate cancer cells, suggesting that they can be used as potential therapeutic drugs in the treatment of prostate cancer.

  7. Chlamydia pneumoniae Promotes Dysfunction of Pancreatic Beta Cells

    PubMed Central

    Rodriguez, Annette R.; Plascencia-Villa, Germán; Witt, Colleen M.; Yu, Jieh-Juen; José-Yacamán, Miguel; Chambers, James P.; Perry, George; Guentzel, M. Neal; Arulanandam, Bernard P.

    2015-01-01

    The human pathogen Chlamydia pneumoniae has been implicated in chronic inflammatory diseases including type 2 diabetes. Therefore, we designed a study to evaluate pancreatic beta cells and mast cells during chlamydial infection. Our study revealed that C. pneumoniae infected mast cells significantly (p< 0.005) decreased beta cell ATP and insulin production, in contrast to uninfected mast cells co-cultured with beta cells. Infected mast cells exhibited pyknotic nuclei and active caspase-3 and caspase-1 expression. Additionally, ex vivo analyses of tissues collected from C. pneumoniae infected mice showed increased interleukin-1β production in splenocytes and pancreatic tissues as was observed with in vitro mast cell-beta cell co-cultures during C. pneumoniae infection. Notably, infected mast cells promoted beta cell destruction. Our findings reveal the negative effect of C. pneumoniae on mast cells, and the consequential impact on pancreatic beta cell function and viability. PMID:25863744

  8. Resuscitation promoting factor (Rpf) from Tomitella biformata AHU 1821(T) promotes growth and resuscitates non-dividing cells.

    PubMed

    Dewi Puspita, Indun; Uehara, Moe; Katayama, Taiki; Kikuchi, Yoshitomo; Kitagawa, Wataru; Kamagata, Yoichi; Asano, Kozo; Nakatsu, Cindy H; Tanaka, Michiko

    2013-01-01

    Functional variation of Rpf, a growth factor found exclusively in Actinobacteria, is differentiated by its source and amino acid sequences. Only purified Rpf proteins from three species have been studied so far. To seek new Rpfs for use in future studies to understand their role in Actinobacteria, the objective of this study was to identify rpf gene homologs in Tomitella biformata AHU 1821(T), a novel Actinobacteria isolated from permafrost ice wedge. Amplification using degenerate primers targeting the essential Rpf domain led to the discovery of a new rpf gene in T. biformata. Gene structure and the deduced Rpf domain amino acid sequence indicated that this rpf gene was not identical to previously studied Rpf. Phylogenetic analysis placed T. biformata Rpf in a monophyletic branch in the RpfB subfamily. The deduced amino acid sequence was 44.9% identical to RpfB in Mycobacterium tuberculosis, the closest functionally tested Rpf. The gene was cloned and expressed in Escherichia coli; the recombinant Rpf protein (rRpf) promoted the growth of dividing cells and resuscitated non-dividing cells of T. biformata. Compared to other studies, this Rpf was required at higher concentrations to promote its growth and to resuscitate itself from a non-dividing state. The resuscitation function was likely due to the highly conserved Rpf domain. This study provides evidence that a genetically unique but functional Rpf can be found in novel members of Actinobacteria and can lead to a better understanding of bacterial cytokines in this phylum.

  9. Salicylic acid receptors activate jasmonic acid signalling through a non-canonical pathway to promote effector-triggered immunity

    PubMed Central

    Liu, Lijing; Sonbol, Fathi-Mohamed; Huot, Bethany; Gu, Yangnan; Withers, John; Mwimba, Musoki; Yao, Jian; He, Sheng Yang; Dong, Xinnian

    2016-01-01

    It is an apparent conundrum how plants evolved effector-triggered immunity (ETI), involving programmed cell death (PCD), as a major defence mechanism against biotrophic pathogens, because ETI-associated PCD could leave them vulnerable to necrotrophic pathogens that thrive on dead host cells. Interestingly, during ETI, the normally antagonistic defence hormones, salicylic acid (SA) and jasmonic acid (JA) associated with defence against biotrophs and necrotrophs respectively, both accumulate to high levels. In this study, we made the surprising finding that JA is a positive regulator of RPS2-mediated ETI. Early induction of JA-responsive genes and de novo JA synthesis following SA accumulation is activated through the SA receptors NPR3 and NPR4, instead of the JA receptor COI1. We provide evidence that NPR3 and NPR4 may mediate this effect by promoting degradation of the JA transcriptional repressor JAZs. This unique interplay between SA and JA offers a possible explanation of how plants can mount defence against a biotrophic pathogen without becoming vulnerable to necrotrophic pathogens. PMID:27725643

  10. TNFAIP3 promotes survival of CD4 T cells by restricting MTOR and promoting autophagy.

    PubMed

    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.

  11. TNFAIP3 promotes survival of CD4 T cells by restricting MTOR and promoting autophagy

    PubMed Central

    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. PMID:26043155

  12. Transcriptional activation by TFIIB mutants that are severely impaired in interaction with promoter DNA and acidic activation domains.

    PubMed Central

    Chou, S; Struhl, K

    1997-01-01

    Biochemical experiments indicate that the general transcription factor IIB (TFIIB) can interact directly with acidic activation domains and that activators can stimulate transcription by increasing recruitment of TFIIB to promoters. For promoters at which recruitment of TFIIB to promoters is limiting in vivo, one would predict that transcriptional activity should be particularly sensitive to TFIIB mutations that decrease the association of TFIIB with promoter DNA and/or with activation domains; i.e., such TFIIB mutations should exacerbate a limiting step that occurs in wild-type cells. Here, we describe mutations on the DNA-binding surface of TFIIB that severely affect both TATA-binding protein (TBP)-TFIIB-TATA complex formation and interaction with the VP16 activation domain in vitro. These TFIIB mutations affect the stability of the TBP-TFIIB-TATA complex in vivo because they are synthetically lethal in combination with TBP mutants impaired for TFIIB binding. Interestingly, these TFIIB derivatives support viability, and they efficiently respond to Gal4-VP16 and natural acidic activators in different promoter contexts. These results suggest that in vivo, recruitment of TFIIB is not generally a limiting step for acidic activators. However, one TFIIB derivative shows reduced transcription of GAL4, suggesting that TFIIB may be limiting at a subset of promoters in vivo. PMID:9372910

  13. Chemical modification of chitosan film via surface grafting of citric acid molecular to promote the biomineralization

    NASA Astrophysics Data System (ADS)

    Liu, Yang; Shen, Xin; Zhou, Huan; Wang, Yingjun; Deng, Linhong

    2016-05-01

    We develop a novel chitosan-citric acid film (abbreviated as CS-CA) suitable for biomedical applications in this study. In this CS-CA film, the citric acid, which is a harmless organic acid has been extensively investigated as a modifying agent on carbohydrate polymers, was cross-linked by 1-Ethyl-3-(3-dimethyl aminopropyl) carbodiimide (EDC) and N-hydroxysuccinimide (NHS) onto the surface of chitosan (CS) film. Fourier transform infrared spectroscopy (FTIR) and X-ray photoelectron spectroscopy (XPS) confirms the graft copolymerization of the modified chitosan film (CS-CA). Surface wettability, moisturizing performance, the capacity of mineralization in vitro and biocompatibility of the films were characterized. After modification, this CS-CA film has good hydrophilicity. It is very evident that the citric acid grafting treatment significantly promotes the biomineralization of the chitosan based substrates. Cell experiments show that the MC3T3-E1 osteoblasts can adhere and proliferate well on the surface of CS-CA film. This CS-CA film, which can be prepared in large quantities and at low cost, should have potential application in bone tissue engineering.

  14. Phosphoric acid fuel cell platinum use study

    NASA Technical Reports Server (NTRS)

    Lundblad, H. L.

    1983-01-01

    The U.S. Department of Energy is promoting the private development of phosphoric acid fuel cell (PAFC) power plants for terrestrial applications. Current PAFC technology utilizes platinum as catalysts in the power electrodes. The possible repercussions that the platinum demand of PAFC power plant commercialization will have on the worldwide supply and price of platinum from the outset of commercialization to the year 2000 are investigated. The platinum demand of PAFC commercialization is estimated by developing forecasts of platinum use per unit of generating capacity and penetration of PAFC power plants into the electric generation market. The ability of the platinum supply market to meet future demands is gauged by assessing the size of platinum reserves and the capability of platinum producers to extract, refine and market sufficient quantities of these reserves. The size and timing of platinum price shifts induced by the added demand of PAFC commercialization are investigated by several analytical methods. Estimates of these price shifts are then used to calculate the subsequent effects on PAFC power plant capital costs.

  15. Phosphoric acid fuel cell platinum use study

    NASA Astrophysics Data System (ADS)

    Lundblad, H. L.

    1983-05-01

    The U.S. Department of Energy is promoting the private development of phosphoric acid fuel cell (PAFC) power plants for terrestrial applications. Current PAFC technology utilizes platinum as catalysts in the power electrodes. The possible repercussions that the platinum demand of PAFC power plant commercialization will have on the worldwide supply and price of platinum from the outset of commercialization to the year 2000 are investigated. The platinum demand of PAFC commercialization is estimated by developing forecasts of platinum use per unit of generating capacity and penetration of PAFC power plants into the electric generation market. The ability of the platinum supply market to meet future demands is gauged by assessing the size of platinum reserves and the capability of platinum producers to extract, refine and market sufficient quantities of these reserves. The size and timing of platinum price shifts induced by the added demand of PAFC commercialization are investigated by several analytical methods. Estimates of these price shifts are then used to calculate the subsequent effects on PAFC power plant capital costs.

  16. Cryptococcal 3-Hydroxy Fatty Acids Protect Cells Against Amoebal Phagocytosis.

    PubMed

    Madu, Uju L; Ogundeji, Adepemi O; Mochochoko, Bonang M; Pohl, Carolina H; Albertyn, Jacobus; Swart, Chantel W; Allwood, J William; Southam, Andrew D; Dunn, Warwick B; May, Robin C; Sebolai, Olihile M

    2015-01-01

    We previously reported on a 3-hydroxy fatty acid that is secreted via cryptococcal capsular protuberances - possibly to promote pathogenesis and survival. Thus, we investigated the role of this molecule in mediating the fate of Cryptococcus (C.) neoformans and the related species C. gattii when predated upon by amoebae. We show that this molecule protects cells against the phagocytic effects of amoebae. C. neoformans UOFS Y-1378 (which produces 3-hydroxy fatty acids) was less sensitive toward amoebae compared to C. neoformans LMPE 046 and C. gattii R265 (both do not produce 3-hydroxy fatty acids) and addition of 3-hydroxy fatty acids to C. neoformans LMPE 046 and C. gattii R265 culture media, causes these strains to become more resistant to amoebal predation. Conversely, addition of aspirin (a 3-hydroxy fatty acid inhibitor) to C. neoformans UOFS Y-1378 culture media made cells more susceptible to amoebae. Our data suggest that this molecule is secreted at a high enough concentration to effect intracellular signaling within amoeba, which in turn, promotes fungal survival. PMID:26696972

  17. Cryptococcal 3-Hydroxy Fatty Acids Protect Cells Against Amoebal Phagocytosis

    PubMed Central

    Madu, Uju L.; Ogundeji, Adepemi O.; Mochochoko, Bonang M.; Pohl, Carolina H.; Albertyn, Jacobus; Swart, Chantel W.; Allwood, J. William; Southam, Andrew D.; Dunn, Warwick B.; May, Robin C.; Sebolai, Olihile M.

    2015-01-01

    We previously reported on a 3-hydroxy fatty acid that is secreted via cryptococcal capsular protuberances - possibly to promote pathogenesis and survival. Thus, we investigated the role of this molecule in mediating the fate of Cryptococcus (C.) neoformans and the related species C. gattii when predated upon by amoebae. We show that this molecule protects cells against the phagocytic effects of amoebae. C. neoformans UOFS Y-1378 (which produces 3-hydroxy fatty acids) was less sensitive toward amoebae compared to C. neoformans LMPE 046 and C. gattii R265 (both do not produce 3-hydroxy fatty acids) and addition of 3-hydroxy fatty acids to C. neoformans LMPE 046 and C. gattii R265 culture media, causes these strains to become more resistant to amoebal predation. Conversely, addition of aspirin (a 3-hydroxy fatty acid inhibitor) to C. neoformans UOFS Y-1378 culture media made cells more susceptible to amoebae. Our data suggest that this molecule is secreted at a high enough concentration to effect intracellular signaling within amoeba, which in turn, promotes fungal survival. PMID:26696972

  18. Cryptococcal 3-Hydroxy Fatty Acids Protect Cells Against Amoebal Phagocytosis.

    PubMed

    Madu, Uju L; Ogundeji, Adepemi O; Mochochoko, Bonang M; Pohl, Carolina H; Albertyn, Jacobus; Swart, Chantel W; Allwood, J William; Southam, Andrew D; Dunn, Warwick B; May, Robin C; Sebolai, Olihile M

    2015-01-01

    We previously reported on a 3-hydroxy fatty acid that is secreted via cryptococcal capsular protuberances - possibly to promote pathogenesis and survival. Thus, we investigated the role of this molecule in mediating the fate of Cryptococcus (C.) neoformans and the related species C. gattii when predated upon by amoebae. We show that this molecule protects cells against the phagocytic effects of amoebae. C. neoformans UOFS Y-1378 (which produces 3-hydroxy fatty acids) was less sensitive toward amoebae compared to C. neoformans LMPE 046 and C. gattii R265 (both do not produce 3-hydroxy fatty acids) and addition of 3-hydroxy fatty acids to C. neoformans LMPE 046 and C. gattii R265 culture media, causes these strains to become more resistant to amoebal predation. Conversely, addition of aspirin (a 3-hydroxy fatty acid inhibitor) to C. neoformans UOFS Y-1378 culture media made cells more susceptible to amoebae. Our data suggest that this molecule is secreted at a high enough concentration to effect intracellular signaling within amoeba, which in turn, promotes fungal survival.

  19. Dendritic Cells: A Spot on Sialic Acid

    PubMed Central

    Crespo, Hélio J.; Lau, Joseph T. Y.; Videira, Paula A.

    2013-01-01

    Glycans decorating cell surface and secreted proteins and lipids occupy the juncture where critical host–host and host-pathogen interactions occur. The role of glycan epitopes in cell–cell and cell-pathogen adhesive events is already well-established, and cell surface glycan structures change rapidly in response to stimulus and inflammatory cues. Despite the wide acceptance that glycans are centrally implicated in immunity, exactly how glycans and their changes contribute to the overall immune response remains poorly defined. Sialic acids are unique sugars that usually occupy the terminal position of the glycan chains and may be modified by external factors, such as pathogens, or upon specific physiological cellular events. At cell surface, sialic acid-modified structures form the key fundamental determinants for a number of receptors with known involvement in cellular adhesiveness and cell trafficking, such as the Selectins and the Siglec families of carbohydrate recognizing receptors. Dendritic cells (DCs) preside over the transition from innate to the adaptive immune repertoires, and no other cell has such relevant role in antigen screening, uptake, and its presentation to lymphocytes, ultimately triggering the adaptive immune response. Interestingly, sialic acid-modified structures are involved in all DC functions, such as antigen uptake, DC migration, and capacity to prime T cell responses. Sialic acid content changes along DC differentiation and activation and, while, not yet fully understood, these changes have important implications in DC functions. This review focuses on the developmental regulation of DC surface sialic acids and how manipulation of DC surface sialic acids can affect immune-critical DC functions by altering antigen endocytosis, pathogen and tumor cell recognition, cell recruitment, and capacity for T cell priming. The existing evidence points to a potential of DC surface sialylation as a therapeutic target to improve and diversify DC

  20. Osteoactivin Promotes Migration of Oral Squamous Cell Carcinomas.

    PubMed

    Arosarena, Oneida A; Dela Cadena, Raul A; Denny, Michael F; Bryant, Evan; Barr, Eric W; Thorpe, Ryan; Safadi, Fayez F

    2016-08-01

    Nearly 50% of patients with oral squamous cell carcinoma (OSCC) die of metastases or locoregional recurrence. Metastasis is mediated by cancer cell adhesion, migration, and invasion. Osteoactivin (OA) overexpression plays a role in metastases in several malignancies. The aims were to determine how integrin interactions modulate OA-induced OSCC cell migration; and to investigate OA effects on cell survival and proliferation. We confirmed OA mRNA and protein overexpression in OSCC cell lines. We assessed OA's interactions with integrins using adhesion inhibition assays, fluorescent immunocytochemistry and co-immunoprecipitation. We investigated OA-mediated activation of mitogen-activated protein kinases (MAPKs) and cell survival. Integrin inhibition effects on OA-mediated cell migration were determined. We assessed effects of OA knock-down on cell migration and proliferation. OA is overexpressed in OSCC cell lines, and serves as a migration-promoting adhesion molecule. OA co-localized with integrin subunits, and co-immunoprecipitated with the subunits. Integrin blocking antibodies, especially those directed against the β1 subunit, inhibited cell adhesion (P = 0.03 for SCC15 cells). Adhesion to OA activated MAPKs in UMSCC14a cells and OA treatment promoted survival of SCC15 cells. Integrin-neutralizing antibodies enhanced cell migration with OA in the extracellular matrix. OA knock-down resulted in decreased proliferation of SCC15 and SCC25 cells, but did not inhibit cell migration. OA in the extracellular matrix promotes OSCC cell adhesion and migration, and may be a novel target in the prevention of HNSCC spread. J. Cell. Physiol. 231: 1761-1770, 2016. © 2015 Wiley Periodicals, Inc. PMID:26636434

  1. Osteoactivin Promotes Migration of Oral Squamous Cell Carcinomas.

    PubMed

    Arosarena, Oneida A; Dela Cadena, Raul A; Denny, Michael F; Bryant, Evan; Barr, Eric W; Thorpe, Ryan; Safadi, Fayez F

    2016-08-01

    Nearly 50% of patients with oral squamous cell carcinoma (OSCC) die of metastases or locoregional recurrence. Metastasis is mediated by cancer cell adhesion, migration, and invasion. Osteoactivin (OA) overexpression plays a role in metastases in several malignancies. The aims were to determine how integrin interactions modulate OA-induced OSCC cell migration; and to investigate OA effects on cell survival and proliferation. We confirmed OA mRNA and protein overexpression in OSCC cell lines. We assessed OA's interactions with integrins using adhesion inhibition assays, fluorescent immunocytochemistry and co-immunoprecipitation. We investigated OA-mediated activation of mitogen-activated protein kinases (MAPKs) and cell survival. Integrin inhibition effects on OA-mediated cell migration were determined. We assessed effects of OA knock-down on cell migration and proliferation. OA is overexpressed in OSCC cell lines, and serves as a migration-promoting adhesion molecule. OA co-localized with integrin subunits, and co-immunoprecipitated with the subunits. Integrin blocking antibodies, especially those directed against the β1 subunit, inhibited cell adhesion (P = 0.03 for SCC15 cells). Adhesion to OA activated MAPKs in UMSCC14a cells and OA treatment promoted survival of SCC15 cells. Integrin-neutralizing antibodies enhanced cell migration with OA in the extracellular matrix. OA knock-down resulted in decreased proliferation of SCC15 and SCC25 cells, but did not inhibit cell migration. OA in the extracellular matrix promotes OSCC cell adhesion and migration, and may be a novel target in the prevention of HNSCC spread. J. Cell. Physiol. 231: 1761-1770, 2016. © 2015 Wiley Periodicals, Inc.

  2. Collective Cell Movement Promotes Synchronization of Coupled Genetic Oscillators

    PubMed Central

    Uriu, Koichiro; Morelli, Luis G.

    2014-01-01

    Collective cell movement is a crucial component of embryonic development. Intercellular interactions regulate collective cell movement by allowing cells to transfer information. A key question is how collective cell movement itself influences information flow produced in tissues by intercellular interactions. Here, we study the effect of collective cell movement on the synchronization of locally coupled genetic oscillators. This study is motivated by the segmentation clock in zebrafish somitogenesis, where short-range correlated movement of cells has been observed. We describe the segmentation clock tissue by a Voronoi diagram, cell movement by the force balance of self-propelled and repulsive forces between cells, the dynamics of the direction of self-propelled motion, and the synchronization of genetic oscillators by locally coupled phase oscillators. We find that movement with a correlation length of about 2 ∼ 3 cell diameters is optimal for the synchronization of coupled oscillators. Quantification of cell mixing reveals that this short-range correlation of cell movement allows cells to exchange neighbors most efficiently. Moreover, short-range correlated movement strongly destabilizes nonuniform spatial phase patterns, further promoting global synchronization. Our theoretical results suggest that collective cell movement may enhance the synchronization of the segmentation clock in zebrafish somitogenesis. More generally, collective cell movement may promote information flow in tissues by enhancing cell mixing and destabilizing spurious patterns. PMID:25028893

  3. Collective cell movement promotes synchronization of coupled genetic oscillators.

    PubMed

    Uriu, Koichiro; Morelli, Luis G

    2014-07-15

    Collective cell movement is a crucial component of embryonic development. Intercellular interactions regulate collective cell movement by allowing cells to transfer information. A key question is how collective cell movement itself influences information flow produced in tissues by intercellular interactions. Here, we study the effect of collective cell movement on the synchronization of locally coupled genetic oscillators. This study is motivated by the segmentation clock in zebrafish somitogenesis, where short-range correlated movement of cells has been observed. We describe the segmentation clock tissue by a Voronoi diagram, cell movement by the force balance of self-propelled and repulsive forces between cells, the dynamics of the direction of self-propelled motion, and the synchronization of genetic oscillators by locally coupled phase oscillators. We find that movement with a correlation length of about 2 ∼ 3 cell diameters is optimal for the synchronization of coupled oscillators. Quantification of cell mixing reveals that this short-range correlation of cell movement allows cells to exchange neighbors most efficiently. Moreover, short-range correlated movement strongly destabilizes nonuniform spatial phase patterns, further promoting global synchronization. Our theoretical results suggest that collective cell movement may enhance the synchronization of the segmentation clock in zebrafish somitogenesis. More generally, collective cell movement may promote information flow in tissues by enhancing cell mixing and destabilizing spurious patterns.

  4. Commensal bacteria promote migration of mast cells into the intestine.

    PubMed

    Kunii, Junichi; Takahashi, Kyoko; Kasakura, Kazumi; Tsuda, Masato; Nakano, Kou; Hosono, Akira; Kaminogawa, Shuichi

    2011-06-01

    Mast cells differentiate from hematopoietic stem cells in the bone marrow and migrate via the circulation to peripheral tissues, where they play a pivotal role in induction of both innate and adaptive immune responses. In this study, the effect of intestinal commensal bacteria on the migration of mast cells into the intestine was investigated. Histochemical analyses showed that germ-free (GF) mice had lower mast cell densities in the small intestine than normal mice. It was also shown that GF mice had lower mast cell proportion out of lamina propria leukocytes in the small intestine and higher mast cell percentages in the blood than normal mice by flow cytometry. These results indicate that migration of mast cells from the blood to the intestine is promoted by intestinal commensal bacteria. In addition, MyD88⁻/⁻ mice had lower densities of intestinal mast cells than CV mice, suggesting that the promotive effect of commensals is, at least in part, TLR-dependent. The ligands of CXC chemokine receptor 2 (CXCR2), which is critical for homing of mast cells to the intestine, were expressed higher in intestinal tissues and in intestinal epithelial cells (IECs) of normal mice than in those of GF or MyD88⁻/⁻ mice. Collectively, it is suggested that commensals promote migration of mast cells into the intestine through the induction of CXCR2 ligands from IECs in a TLR-dependent manner.

  5. Anacardic Acid, Salicylic Acid, and Oleic Acid Differentially Alter Cellular Bioenergetic Function in Breast Cancer Cells.

    PubMed

    Radde, Brandie N; Alizadeh-Rad, Negin; Price, Stephanie M; Schultz, David J; Klinge, Carolyn M

    2016-11-01

    Anacardic acid is a dietary and medicinal phytochemical that inhibits breast cancer cell proliferation and uncouples oxidative phosphorylation (OXPHOS) in isolated rat liver mitochondria. Since mitochondrial-targeted anticancer therapy (mitocans) may be useful in breast cancer, we examined the effect of anacardic acid on cellular bioenergetics and OXPHOS pathway proteins in breast cancer cells modeling progression to endocrine-independence: MCF-7 estrogen receptor α (ERα)+ endocrine-sensitive; LCC9 and LY2 ERα+, endocrine-resistant, and MDA-MB-231 triple negative breast cancer (TNBC) cells. At concentrations similar to cell proliferation IC50 s, anacardic acid reduced ATP-linked oxygen consumption rate (OCR), mitochondrial reserve capacity, and coupling efficiency while increasing proton leak, reflecting mitochondrial toxicity which was greater in MCF-7 compared to endocrine-resistant and TNBC cells. These results suggest tolerance in endocrine-resistant and TNBC cells to mitochondrial stress induced by anacardic acid. Since anacardic acid is an alkylated 2-hydroxybenzoic acid, the effects of salicylic acid (SA, 2-hydroxybenzoic acid moiety) and oleic acid (OA, monounsaturated alkyl moiety) were tested. SA inhibited whereas OA stimulated cell viability. In contrast to stimulation of basal OCR by anacardic acid (uncoupling effect), neither SA nor OA altered basal OCR- except OA inhibited basal and ATP-linked OCR, and increased ECAR, in MDA-MB-231 cells. Changes in OXPHOS proteins correlated with changes in OCR. Overall, neither the 2-hydroxybenzoic acid moiety nor the monounsaturated alky moiety of anacardic acid is solely responsible for the observed mitochondria-targeted anticancer activity in breast cancer cells and hence both moieties are required in the same molecule for the observed effects. J. Cell. Biochem. 117: 2521-2532, 2016. © 2016 Wiley Periodicals, Inc. PMID:26990649

  6. Nitrated fatty acids reverse pulmonary fibrosis by dedifferentiating myofibroblasts and promoting collagen uptake by alveolar macrophages

    PubMed Central

    Reddy, Aravind T.; Lakshmi, Sowmya P.; Zhang, Yingze; Reddy, Raju C.

    2014-01-01

    Idiopathic pulmonary fibrosis (IPF) is a progressive, fatal disease, thought to be largely transforming growth factor β (TGFβ) driven, for which there is no effective therapy. We assessed the potential benefits in IPF of nitrated fatty acids (NFAs), which are unique endogenous agonists of peroxisome proliferator-activated receptor γ (PPARγ), a nuclear hormone receptor that exhibits wound-healing and antifibrotic properties potentially useful for IPF therapy. We found that pulmonary PPARγ is down-regulated in patients with IPF. In vitro, knockdown or knockout of PPARγ expression in isolated human and mouse lung fibroblasts induced a profibrotic phenotype, whereas treating human fibroblasts with NFAs up-regulated PPARγ and blocked TGFβ signaling and actions. NFAs also converted TGFβ to inactive monomers in cell-free solution, suggesting an additional mechanism through which they may inhibit TGFβ. In vivo, treating mice bearing experimental pulmonary fibrosis with NFAs reduced disease severity. Also, NFAs up-regulated the collagen-targeting factor milk fat globule-EGF factor 8 (MFG-E8), stimulated collagen uptake and degradation by alveolar macrophages, and promoted myofibroblast dedifferentiation. Moreover, treating mice with established pulmonary fibrosis using NFAs reversed their existing myofibroblast differentiation and collagen deposition. These findings raise the prospect of treating IPF with NFAs to halt and perhaps even reverse the progress of IPF.—Reddy, A. T., Lakshmi, S. P., Zhang, Y., Reddy, R. C. Nitrated fatty acids reverse pulmonary fibrosis by dedifferentiating myofibroblasts and promoting collagen uptake by alveolar macrophages. PMID:25252739

  7. Interferon regulatory factor-1 binds c-Cbl, enhances mitogen activated protein kinase signaling and promotes retinoic acid-induced differentiation of HL-60 human myelo-monoblastic leukemia cells.

    PubMed

    Shen, Miaoqing; Bunaciu, Rodica P; Congleton, Johanna; Jensen, Holly A; Sayam, Lavanya G; Varner, Jeffrey D; Yen, Andrew

    2011-12-01

    All-trans retinoic acid (RA) and interferons (IFNs) have efficacy in treating certain leukemias and lymphomas, respectively, motivating interest in their mechanism of action to improve therapy. Both RA and IFNs induce interferon regulatory factor-1 (IRF-1). We find that in HL-60 myeloblastic leukemia cells which undergo mitogen activated protien kinase (MAPK)-dependent myeloid differentiation in response to RA, IRF-1 propels differentiation. RA induces MAPK-dependent expression of IRF-1. IRF-1 binds c-Cbl, a MAPK related adaptor. Ectopic IRF-1 expression causes CD38 expression and activation of the Raf/MEK/ERK axis, and enhances RA-induced differentiation by augmenting CD38, CD11b, respiratory burst and G0 arrest. Ectopic IRF-1 expression also decreases the activity of aldehyde dehydrogenase 1, a stem cell marker, and enhances RA-induced ALDH1 down-regulation. Interestingly, expression of aryl hydrocarbon receptor (AhR), which is RA-induced and known to down-regulate Oct4 and drive RA-induced differentiation, also enhances IRF-1 expression. The data are consistent with a model whereby IRF-1 acts downstream of RA and AhR to enhance Raf/MEK/ERK activation and propel differentiation.

  8. TP508 accelerates fracture repair by promoting cell growth over cell death

    SciTech Connect

    Li Xinmin; Wang Hali; Touma, Edward; Qi Yuchen; Rousseau, Emma; Quigg, Richard J.; Ryaby, James T.

    2007-12-07

    TP508 is a synthetic 23-amino acid peptide representing a receptor-binding domain of human thrombin. We have previously shown that a single injection of TP508 accelerates fracture healing in a rat femoral fracture model. To understand how TP508 acts at the protein level during fracture healing, we compared the translational profiles between saline-control and fractured femur at six time points after TP508 treatment using the second generation of BD Clontech{sup TM} Antibody Microarray. Here, we demonstrate that TP508 accelerates fracture healing by modulating expression levels of proteins primarily involved in the functional categories of cell cycle, cellular growth and proliferation, and cell death. The majority of those proteins are physically interrelated and functionally overlapped. The action of those proteins is highlighted by a central theme of promoting cell growth via balance of cell survival over cell death signals. This appears to occur through the stimulation of several bone healing pathways including cell cycle-G1/S checkpoint regulation, apoptosis, JAK/STAT, NF-{kappa}B, PDGF, PI3K/AKT, PTEN, and ERK/MAPK.

  9. Acid treatment of melanoma cells selects for invasive phenotypes.

    PubMed

    Moellering, Raymond E; Black, Kvar C; Krishnamurty, Chetan; Baggett, Brenda K; Stafford, Phillip; Rain, Matthew; Gatenby, Robert A; Gillies, Robert J

    2008-01-01

    Solid tumors become acidic due to hypoxia and upregulated glycolysis. We have hypothesized that this acidosis leads to more aggressive invasive behavior during carcinogenesis (Nature Reviews Cancer 4:891-899, 2004). Previous work on this subject has shown mixed results. While some have observed an induction of metastasis and invasion with acid treatments, others have not. To investigate this, human melanoma cells were acclimated to low pH growth conditions. Significant cell mortality occurred during acclimation, suggesting that acidosis selected for resistant phenotypes. Cells maintained under acidic conditions exhibited a greater range of motility, a reduced capacity to form flank tumors in SCID mice and did not invade more rapidly in vitro, compared to non-selected control cells. However, re-acclimation of these selected cells to physiological pH gave rise to stable populations with significantly higher in vitro invasion. These re-acclimated cells maintained higher invasion and higher motility for multiple generations. Transcriptomic analyses of these three phenotypes revealed significant differences, including upregulation of relevant pathways important for tissue remodeling, cell cycle control and proliferation. These results reinforce the hypothesis that acidosis promotes selection of stable, more invasive phenotypes, rather than inductive changes, which would be reversible.

  10. Interleukin-8 derived from local tissue-resident stromal cells promotes tumor cell invasion.

    PubMed

    Welte, Gabriel; Alt, Eckhard; Devarajan, Eswaran; Krishnappa, Srinivasalu; Jotzu, Constantin; Song, Yao-Hua

    2012-11-01

    The aim of this study is to evaluate the role of adipose tissue resident stromal cells on tumor cell invasion. Our data show that a subpopulation of adipose tissue derived stromal cells expressing Nestin, NG2, α-smooth muscle actin and PDGFR-α migrate toward the cancer cells. Microarray analysis revealed the upregulation of IL-8 in the migrated cells. We demonstrated that stromal cell derived IL-8 promote the invasion and the anchorage-independent growth of cancer cells. We conclude that human breast cancer cells attract a subpopulation of stromal cells that secrete IL-8 to promote tumor cell invasion in a paracrine fashion.

  11. Histone deacetylase inhibitors epigenetically promote reparative events in primary dental pulp cells

    SciTech Connect

    Duncan, Henry F.; Smith, Anthony J.; Fleming, Garry J.P.; Cooper, Paul R.

    2013-06-10

    Application of histone deacetylase inhibitors (HDACi) to cells epigenetically alters their chromatin structure and induces transcriptional and cellular reparative events. This study investigated the application of two HDACi, valproic acid (VPA) and trichostatin A (TSA) on the induction of repair-associated responses in primary dental pulp cell (DPC) cultures. Flow cytometry demonstrated that TSA (100 nM, 400 nM) significantly increased cell viability. Neither HDACi was cytotoxic, although cell growth analysis revealed significant anti-proliferative effects at higher concentrations for VPA (>0.5 mM) and TSA (>50 nM). While high-content-analysis demonstrated that HDACi did not significantly induce caspase-3 or p21 activity, p53-expression was increased by VPA (3 mM, 5 mM) at 48 h. HDACi-exposure induced mineralization per cell dose-dependently to a plateau level (VPA-0.125 mM and TSA-25 nM) with accompanying increases in mineralization/dentinogenic-associated gene expression at 5 days (DMP-1, BMP-2/-4, Nestin) and 10 days (DSPP, BMP-2/-4). Both HDACis, at a range of concentrations, significantly stimulated osteopontin and BMP-2 protein expression at 10 and 14 days further supporting the ability of HDACi to promote differentiation. HDACi exert different effects on primary compared with transformed DPCs and promote mineralization and differentiation events without cytotoxic effects. These novel data now highlight the potential in restorative dentistry for applying low concentrations of HDACi in vital pulp treatment. -- Highlights: • Valproic acid and trichostatin A promoted mineralization in primary pulp cells. • Cell viability, apoptosis, caspase-3, p21 unaltered; p53 increased by valproic acid. • Trichostatin A increased cell viability at 24 h at selected concentrations. • Altered cell toxicity and differentiation between primary and transformed cells. • HDACi-induced the differentiation marker proteins osteopontin and BMP-2.

  12. Acid Strength and Bifunctional Catalytic Behavior of Alloys Comprised of Noble Metals and Oxophilic Metal Promoters

    SciTech Connect

    Hibbitts, David D.; Tan, Qiaohua; Neurock, Matthew

    2014-06-01

    The promotion of metal catalysts with partially oxidized oxophilic MOx species, such as ReOx-promoted Rh, has been demonstrated to produce Brønsted acid sites that can promote hydrogenolysis of oxygenate intermediates such as those found in biomass-derived species. A wide variety of alloy compositions and structures are examined in this work to investigate strongly acidic promoters by using DFT-calculated deprotonation energies (DPE) as a measure of acid strength. Sites with the highest acid strength had DPE less than 1100 kJ mol-1, similar to DPE values of heteropolyacids or acid-containing zeolites, and were found on alloys composed of an oxophilic metal (such as Re or W) with a noble metal (such as Rh or Pt). NH3 adsorbs more strongly to sites with increasing acid strength and the activation barriers for acid-catalyzed ring opening of a furan ring decrease with increasing acid strength, which was also shown to be stronger for OH acid sites bound to multiple oxophilic metal atoms in a three-fold configuration rather than OH sites adsorbed in an atop configuration on one oxophilic metal, indicating that small MOx clusters may yield sites with the highest acid strength.

  13. Bile-acid-induced cell injury and protection

    PubMed Central

    Perez, Maria J; Briz, Oscar

    2009-01-01

    Several studies have characterized the cellular and molecular mechanisms of hepatocyte injury caused by the retention of hydrophobic bile acids (BAs) in cholestatic diseases. BAs may disrupt cell membranes through their detergent action on lipid components and can promote the generation of reactive oxygen species that, in turn, oxidatively modify lipids, proteins, and nucleic acids, and eventually cause hepatocyte necrosis and apoptosis. Several pathways are involved in triggering hepatocyte apoptosis. Toxic BAs can activate hepatocyte death receptors directly and induce oxidative damage, thereby causing mitochondrial dysfunction, and induce endoplasmic reticulum stress. When these compounds are taken up and accumulate inside biliary cells, they can also cause apoptosis. Regarding extrahepatic tissues, the accumulation of BAs in the systemic circulation may contribute to endothelial injury in the kidney and lungs. In gastrointestinal cells, BAs may behave as cancer promoters through an indirect mechanism involving oxidative stress and DNA damage, as well as acting as selection agents for apoptosis-resistant cells. The accumulation of BAs may have also deleterious effects on placental and fetal cells. However, other BAs, such as ursodeoxycholic acid, have been shown to modulate BA-induced injury in hepatocytes. The major beneficial effects of treatment with ursodeoxycholic acid are protection against cytotoxicity due to more toxic BAs; the stimulation of hepatobiliary secretion; antioxidant activity, due in part to an enhancement in glutathione levels; and the inhibition of liver cell apoptosis. Other natural BAs or their derivatives, such as cholyl-N-methylglycine or cholylsarcosine, have also aroused pharmacological interest owing to their protective properties. PMID:19360911

  14. Corrosion free phosphoric acid fuel cell

    DOEpatents

    Wright, Maynard K.

    1990-01-01

    A phosphoric acid fuel cell with an electrolyte fuel system which supplies electrolyte via a wick disposed adjacent a cathode to an absorbent matrix which transports the electrolyte to portions of the cathode and an anode which overlaps the cathode on all sides to prevent corrosion within the cell.

  15. Activated mast cells promote differentiation of B cells into effector cells

    PubMed Central

    Palm, Anna-Karin E.; Garcia-Faroldi, Gianni; Lundberg, Marcus; Pejler, Gunnar; Kleinau, Sandra

    2016-01-01

    Based on the known accumulation of mast cells (MCs) in B cell-dependent inflammatory diseases, including rheumatoid arthritis, we hypothesized that MCs directly modulate B cells. We show here that degranulated, and to a lesser extent naïve or IgE-sensitized, MCs activate both naïve and B cell receptor-activated B cells. This was shown by increased proliferation, blast formation, and expression of CD19, MHC class II and CD86 in the B cells. Further, MCs stimulated the secretion of IgM and IgG in IgM+ B cells, indicating that MCs can induce class-switch recombination in B cells. We also show that coculture of MCs with B cells promotes surface expression of L-selectin, a homing receptor, on the B cells. The effects of MCs on B cells were partly dependent on cell-cell contact and both follicular and marginal zone B cells could be activated by MCs. Our findings suggest that degranulated MCs support optimal activation of B cells, a finding that is in line with in vivo studies showing that MCs frequently degranulate in the context of B-cell driven pathologies such as arthritis. Together, our findings show that MCs have the capacity to differentiate B cells to effector cells. PMID:26847186

  16. Maternal bile acid transporter deficiency promotes neonatal demise

    PubMed Central

    Zhang, Yuanyuan; Li, Fei; Wang, Yao; Pitre, Aaron; Fang, Zhong-ze; Frank, Matthew W.; Calabrese, Christopher; Krausz, Kristopher W.; Neale, Geoffrey; Frase, Sharon; Vogel, Peter; Rock, Charles O.; Gonzalez, Frank J.; Schuetz, John D.

    2015-01-01

    Intrahepatic cholestasis of pregnancy (ICP) is associated with adverse neonatal survival and is estimated to impact between 0.4 and 5% of pregnancies worldwide. Here we show that maternal cholestasis (due to Abcb11 deficiency) produces neonatal death among all offspring within 24 h of birth due to atelectasis-producing pulmonary hypoxia, which recapitulates the neonatal respiratory distress of human ICP. Neonates of Abcb11-deficient mothers have elevated pulmonary bile acids and altered pulmonary surfactant structure. Maternal absence of Nr1i2 superimposed on Abcb11 deficiency strongly reduces maternal serum bile acid concentrations and increases neonatal survival. We identify pulmonary bile acids as a key factor in the disruption of the structure of pulmonary surfactant in neonates of ICP. These findings have important implications for neonatal respiratory failure, especially when maternal bile acids are elevated during pregnancy, and highlight potential pathways and targets amenable to therapeutic intervention to ameliorate this condition. PMID:26416771

  17. Maternal bile acid transporter deficiency promotes neonatal demise.

    PubMed

    Zhang, Yuanyuan; Li, Fei; Wang, Yao; Pitre, Aaron; Fang, Zhong-Ze; Frank, Matthew W; Calabrese, Christopher; Krausz, Kristopher W; Neale, Geoffrey; Frase, Sharon; Vogel, Peter; Rock, Charles O; Gonzalez, Frank J; Schuetz, John D

    2015-09-29

    Intrahepatic cholestasis of pregnancy (ICP) is associated with adverse neonatal survival and is estimated to impact between 0.4 and 5% of pregnancies worldwide. Here we show that maternal cholestasis (due to Abcb11 deficiency) produces neonatal death among all offspring within 24 h of birth due to atelectasis-producing pulmonary hypoxia, which recapitulates the neonatal respiratory distress of human ICP. Neonates of Abcb11-deficient mothers have elevated pulmonary bile acids and altered pulmonary surfactant structure. Maternal absence of Nr1i2 superimposed on Abcb11 deficiency strongly reduces maternal serum bile acid concentrations and increases neonatal survival. We identify pulmonary bile acids as a key factor in the disruption of the structure of pulmonary surfactant in neonates of ICP. These findings have important implications for neonatal respiratory failure, especially when maternal bile acids are elevated during pregnancy, and highlight potential pathways and targets amenable to therapeutic intervention to ameliorate this condition.

  18. Purinergic signaling promotes proliferation of adult mouse subventricular zone cells.

    PubMed

    Suyama, Satoshi; Sunabori, Takehiko; Kanki, Hiroaki; Sawamoto, Kazunobu; Gachet, Christian; Koizumi, Schuichi; Okano, Hideyuki

    2012-07-01

    In adult mammalian brains, neural stem cells (NSCs) exist in the subventricular zone (SVZ), where persistent neurogenesis continues throughout life. Those NSCs produce neuroblasts that migrate into the olfactory bulb via formation of transit-amplifying cells, which are committed precursor cells of the neuronal lineage. In this SVZ niche, cell-cell communications conducted by diffusible factors as well as physical cell-cell contacts are important for the regulation of the proliferation and fate determination of NSCs. Previous studies have suggested that extracellular purinergic signaling, which is mediated by purine compounds such as ATP, plays important roles in cell-cell communication in the CNS. Purinergic signaling also promotes the proliferation of adult NSCs in vitro. However, the in vivo roles of purinergic signaling in the neurogenic niche still remain unknown. In this study, ATP infusion into the lateral ventricle of the mouse brain resulted in an increase in the numbers of rapidly dividing cells and Mash1-positive transit-amplifying cells (Type C cells) in the SVZ. Mash1-positive cells express the P2Y1 purinergic signaling receptor and infusion of the P2Y1 receptor-specific antagonist MRS2179 decreased the number of rapidly dividing bromodeoxyuridine (BrdU)-positive cells and Type C cells. Moreover, a 17% reduction of rapidly dividing BrdU-positive cells and a 19% reduction of Mash1-positive cells were observed in P2Y1 knock-out mice. Together, these results suggest that purinergic signaling promotes the proliferation of rapidly dividing cells and transit-amplifying cells, in the SVZ niche through the P2Y1 receptor. PMID:22764232

  19. Hospicells (ascites-derived stromal cells) promote tumorigenicity and angiogenesis.

    PubMed

    Pasquet, Marlene; Golzio, Muriel; Mery, Eliane; Rafii, Arash; Benabbou, Nadia; Mirshahi, Pezhman; Hennebelle, Isabelle; Bourin, Philippe; Allal, Ben; Teissie, Justin; Mirshahi, Massoud; Couderc, Bettina

    2010-05-01

    The microenvironment is known to play a dominant role in cancer progression. Cells closely associated with tumoral cells, named hospicells, have been recently isolated from the ascites of ovarian cancer patients. Whilst these cells present no specific markers from known cell lineages, they do share some homology with bone marrow-derived or adipose tissue-derived human mesenchymal stem cells (CD9, CD10, CD29, CD146, CD166, HLA-1). We studied the role of hospicells in ovarian carcinoma progression. In vitro, these cells had no effect on the growth of human ovarian carcinoma cell lines OVCAR-3, SKOV-1 and IGROV-1. In vivo, their co-injection with adenocarcinoma cells enhanced tumor growth whatever the tumor model used (subcutaneous and intraperitoneally established xenografts in athymic mice). In addition, their injection increased the development of ascites in tumor-bearing mice. Fluorescent macroscopy revealed an association between hospicells and ovarian adenocarcinoma cells within the tumor mass. Tumors obtained by coinjection of hospicells and human ovarian adenocarcinoma cells presented an increased microvascularization indicating that the hospicells could promote tumorigenicity of ovarian tumor cells in vivovia their action on angiogenesis. This effect on angiogenesis could be attributed to the increased HIF1alpha and VEGF expression associated with the presence of the hospicells. Collectively, these data indicate a role for these ascite-derived stromal cells in promoting tumor growth by increasing angiogenesis.

  20. HIF-1α Promotes A Hypoxia-Independent Cell Migration.

    PubMed

    Li, Liyuan; Madu, Chikezie O; Lu, Andrew; Lu, Yi

    2010-01-01

    Hypoxia-inducible factor-1α (HIF-1α) is known as a transactivator for VEGF gene promoter. It can be induced by hypoxia. However, no study has been done so far to dissect HIF-1α-mediated effects from hypoxia or VEGF-mediated effects. By using a HIF-1α knockout (HIF-1α KO) cell system in mouse embryonic fibroblast (MEF) cells, this study analyzes cell migration and HIF-1α, hypoxia and VEGF activation. A hypoxia-mediated HIF-1α induction and VEGF transactivation were observed: both HIF-1α WT lines had significantly increased VEGF transactivation, as an indicator for HIF-1α induction, in hypoxia compared to normoxia; in contrast, HIF-1α KO line had no increased VEGF transactivation under hypoxia. HIF-1α promotes cell migration: HIF-1α-KO cells had a significantly reduced migration compared to that of the HIF-1α WT cells under both normoxia and hypoxia. The significantly reduced cell migration in HIF-1α KO cells can be partially rescued by the restoration of WT HIF-1α expression mediated by adenoviral-mediated gene transfer. Interestingly, hypoxia has no effect on cell migration: the cells had a similar cell migration rate under hypoxic and normoxic conditions for both HIF-1α WT and HIF-1α KO lines, respectively. Collectively, these data suggest that HIF-1α plays a role in MEF cell migration that is independent from hypoxia-mediated effects.

  1. HIF-1α Promotes A Hypoxia-Independent Cell Migration

    PubMed Central

    Li, Liyuan; Madu, Chikezie O.; Lu, Andrew; Lu, Yi

    2010-01-01

    Hypoxia-inducible factor-1α (HIF-1α) is known as a transactivator for VEGF gene promoter. It can be induced by hypoxia. However, no study has been done so far to dissect HIF-1α-mediated effects from hypoxia or VEGF-mediated effects. By using a HIF-1α knockout (HIF-1α KO) cell system in mouse embryonic fibroblast (MEF) cells, this study analyzes cell migration and HIF-1α, hypoxia and VEGF activation. A hypoxia-mediated HIF-1α induction and VEGF transactivation were observed: both HIF-1α WT lines had significantly increased VEGF transactivation, as an indicator for HIF-1α induction, in hypoxia compared to normoxia; in contrast, HIF-1α KO line had no increased VEGF transactivation under hypoxia. HIF-1α promotes cell migration: HIF-1α-KO cells had a significantly reduced migration compared to that of the HIF-1α WT cells under both normoxia and hypoxia. The significantly reduced cell migration in HIF-1α KO cells can be partially rescued by the restoration of WT HIF-1α expression mediated by adenoviral-mediated gene transfer. Interestingly, hypoxia has no effect on cell migration: the cells had a similar cell migration rate under hypoxic and normoxic conditions for both HIF-1α WT and HIF-1α KO lines, respectively. Collectively, these data suggest that HIF-1α plays a role in MEF cell migration that is independent from hypoxia-mediated effects. PMID:20882121

  2. Promoting justice in stem cell intellectual property.

    PubMed

    Regenberg, Alan; Mathews, Debra J H

    2011-11-01

    According to the World Trade Organization, intellectual property rights are "rights given to persons over the creations of their minds. They usually give the creator an exclusive right over the use of his/her creation for a certain period of time." The rationale behind intellectual property rights is to offer a quid pro quo, between creators and the public, intended to spur innovation. Inventors gain exclusivity (and an opportunity for profits) in exchange for publicly disclosing details about their creations. The public gains free access to information - information that can then be used to support further innovation. Innovation is seen as an inherent good in this context, as it can lead to the development of things people need (e.g., treatments for disease, green energy technologies or a better mousetrap). Exclusive rights to intellectual property are managed via patents and licenses, with patenting being primarily regulated at the national level. Intellectual property rights are the dominant mechanism used in innovation policy, particularly in science. However, myriad modifications and alternatives to intellectual property rights have been proposed and utilized, including patent pooling, intellectual property exchanges and clearing houses, innovation prizes and open-source licenses. The challenges related to competing models of innovation policy present in a fairly consistent manner across most fields of science. However, this paper will focus exclusively on intellectual property rights and models of innovation policy in the context of stem cell science. It is not that the issues themselves are unique in this context, but rather that there are a series of factors that make a discussion of intellectual property rights and models of innovation policy particularly important in the context of stem cell science. PMID:21999266

  3. Promoting justice in stem cell intellectual property.

    PubMed

    Regenberg, Alan; Mathews, Debra J H

    2011-11-01

    According to the World Trade Organization, intellectual property rights are "rights given to persons over the creations of their minds. They usually give the creator an exclusive right over the use of his/her creation for a certain period of time." The rationale behind intellectual property rights is to offer a quid pro quo, between creators and the public, intended to spur innovation. Inventors gain exclusivity (and an opportunity for profits) in exchange for publicly disclosing details about their creations. The public gains free access to information - information that can then be used to support further innovation. Innovation is seen as an inherent good in this context, as it can lead to the development of things people need (e.g., treatments for disease, green energy technologies or a better mousetrap). Exclusive rights to intellectual property are managed via patents and licenses, with patenting being primarily regulated at the national level. Intellectual property rights are the dominant mechanism used in innovation policy, particularly in science. However, myriad modifications and alternatives to intellectual property rights have been proposed and utilized, including patent pooling, intellectual property exchanges and clearing houses, innovation prizes and open-source licenses. The challenges related to competing models of innovation policy present in a fairly consistent manner across most fields of science. However, this paper will focus exclusively on intellectual property rights and models of innovation policy in the context of stem cell science. It is not that the issues themselves are unique in this context, but rather that there are a series of factors that make a discussion of intellectual property rights and models of innovation policy particularly important in the context of stem cell science.

  4. Promoter DNA Hypermethylation and Gene Repression in Undifferentiated Arabidopsis Cells

    PubMed Central

    Berdasco, María; Alcázar, Rubén; García-Ortiz, María Victoria; Ballestar, Esteban; Fernández, Agustín F.; Roldán-Arjona, Teresa; Tiburcio, Antonio F.; Altabella, Teresa; Buisine, Nicolas; Quesneville, Hadi; Baudry, Antoine; Lepiniec, Loïc; Alaminos, Miguel; Rodríguez, Roberto; Lloyd, Alan; Colot, Vincent; Bender, Judith; Canal, María Jesús; Esteller, Manel; Fraga, Mario F.

    2008-01-01

    Maintaining and acquiring the pluripotent cell state in plants is critical to tissue regeneration and vegetative multiplication. Histone-based epigenetic mechanisms are important for regulating this undifferentiated state. Here we report the use of genetic and pharmacological experimental approaches to show that Arabidopsis cell suspensions and calluses specifically repress some genes as a result of promoter DNA hypermethylation. We found that promoters of the MAPK12, GSTU10 and BXL1 genes become hypermethylated in callus cells and that hypermethylation also affects the TTG1, GSTF5, SUVH8, fimbrin and CCD7 genes in cell suspensions. Promoter hypermethylation in undifferentiated cells was associated with histone hypoacetylation and primarily occurred at CpG sites. Accordingly, we found that the process specifically depends on MET1 and DRM2 methyltransferases, as demonstrated with DNA methyltransferase mutants. Our results suggest that promoter DNA methylation may be another important epigenetic mechanism for the establishment and/or maintenance of the undifferentiated state in plant cells. PMID:18827894

  5. Harzianic acid, an antifungal and plant growth promoting metabolite from Trichoderma harzianum.

    PubMed

    Vinale, Francesco; Flematti, Gavin; Sivasithamparam, Krishnapillai; Lorito, Matteo; Marra, Roberta; Skelton, Brian W; Ghisalberti, Emilio L

    2009-11-01

    A Trichoderma harzianum strain, isolated from composted hardwood bark in Western Australia, was found to produce a metabolite with antifungal and plant growth promoting activity. The structure and absolute configuration of the fungal compound, harzianic acid (1), were determined by X-ray diffraction studies. Harzianic acid showed antibiotic activity against Pythium irregulare, Sclerotinia sclerotiorum, and Rhizoctonia solani. A plant growth promotion effect was observed at low concentrations of 1.

  6. Differential effects of eicosapentaenoic acid and docosahexaenoic acid in promoting the differentiation of 3T3-L1 preadipocytes.

    PubMed

    Murali, Ganesan; Desouza, Cyrus V; Clevenger, Michelle E; Ramalingam, Ramesh; Saraswathi, Viswanathan

    2014-01-01

    The objective of this study was to determine the effects of enrichment with n-3 fatty acids, eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA), on the differentiation of 3T3-L1 preadipocytes. Enrichment with DHA but not EPA significantly increased the differentiation markers compared to control differentiated cells. DHA compared to EPA treatment led to a greater increase in adiponectin secretion and, conditioned media collected from DHA treated cells inhibited monocyte migration. Moreover, DHA treatment resulted in inhibition of pro-inflammatory signaling pathways. DHA treated cells predominantly accumulated DHA in phospholipids whereas EPA treatment led to accumulation of both EPA and its elongation product docosapentaenoic acid (DPA), an n-3 fatty acid. Of note, adding DPA to DHA inhibited DHA-induced differentiation. The differential effects of EPA and DHA on preadipocyte differentiation may be due, in part, to differences in their intracellular modification which could impact the type of n-3 fatty acids incorporated into the cells.

  7. Mesothelial cells promote early ovarian cancer metastasis through fibronectin secretion

    PubMed Central

    Kenny, Hilary A.; Chiang, Chun-Yi; White, Erin A.; Schryver, Elizabeth M.; Habis, Mohammed; Romero, Iris L.; Ladanyi, Andras; Penicka, Carla V.; George, Joshy; Matlin, Karl; Montag, Anthony; Wroblewski, Kristen; Yamada, S. Diane; Mazar, Andrew P.; Bowtell, David; Lengyel, Ernst

    2014-01-01

    Ovarian cancer (OvCa) metastasizes to organs in the abdominal cavity, such as the omentum, which are covered by a single layer of mesothelial cells. Mesothelial cells are generally thought to be “bystanders” to the metastatic process and simply displaced by OvCa cells to access the submesothelial extracellular matrix. Here, using organotypic 3D cultures, we found that primary human mesothelial cells secrete fibronectin in the presence of OvCa cells. Moreover, we evaluated the tumor stroma of 108 human omental metastases and determined that fibronectin was consistently overexpressed in these patients. Blocking fibronectin production in primary mesothelial cells in vitro or in murine models, either genetically (fibronectin 1 floxed mouse model) or via siRNA, decreased adhesion, invasion, proliferation, and metastasis of OvCa cells. Using a coculture model, we determined that OvCa cells secrete TGF-β1, which in turn activates a TGF-β receptor/RAC1/SMAD-dependent signaling pathway in the mesothelial cells that promotes a mesenchymal phenotype and transcriptional upregulation of fibronectin. Additionally, blocking α5 or β1 integrin function with antibodies reduced metastasis in an orthotopic preclinical model of OvCa metastasis. These findings indicate that cancer-associated mesothelial cells promote colonization during the initial steps of OvCa metastasis and suggest that mesothelial cells actively contribute to metastasis. PMID:25202979

  8. Effect of Ultrasonic Frequency on Lactic Acid Fermentation Promotion by Ultrasonic Irradiation

    NASA Astrophysics Data System (ADS)

    Shimada, Tadayuki; Ohdaira, Etsuzo; Masuzawa, Nobuyoshi

    2004-05-01

    The authors have been researching the promotion of lactic acid fermentation by ultrasonic irradiation. In the past research, it was proven that ultrasonic irradiation is effective in the process of fermentation, and the production of yoghurt and kefir was promoted. In this study, the effect of the ultrasonic frequency in this fermentation process was examined. In the frequency range of this study, it was found that the action of fermentation promotion was exponentially proportionate to the irradiated ultrasonic frequency.

  9. Scaffold architecture and fibrin gels promote meniscal cell proliferation

    SciTech Connect

    Pawelec, K. M. E-mail: jw626@cam.ac.uk; Best, S. M.; Cameron, R. E.; Wardale, R. J. E-mail: jw626@cam.ac.uk

    2015-01-01

    Stability of the knee relies on the meniscus, a complex connective tissue with poor healing ability. Current meniscal tissue engineering is inadequate, as the signals for increasing meniscal cell proliferation have not been established. In this study, collagen scaffold structure, isotropic or aligned, and fibrin gel addition were tested. Metabolic activity was promoted by fibrin addition. Cellular proliferation, however, was significantly increased by both aligned architectures and fibrin addition. None of the constructs impaired collagen type I production or triggered adverse inflammatory responses. It was demonstrated that both fibrin gel addition and optimized scaffold architecture effectively promote meniscal cell proliferation.

  10. Progesterone promotes propagation and viability of mouse embryonic stem cells.

    PubMed

    Shen, Shan-Wei; Song, Hou-Yan

    2009-10-25

    It has been known that estrogen-17beta stimulates proliferation of mouse embryonic stem (mES) cells. To explore the function of another steroid hormone progesterone, we used MTT method and BrdU incorporation assay to obtain growth curves, clone forming assay to detect the propagation and viability of individual mES cells, Western blot to test the expression of ES cell marker gene Oct-4, fluorescence activated cell sorter (FACS) to test cell cycle, and real-time PCR to detect the expressions of cyclins, cyclin-dependent kinases and proto-oncogenes. The results showed that progesterone promoted proliferation of mES cells. The number of clones was more in progesterone-treated group than that in the control group. The expression of pluripotency-associated transcriptional factor Oct-4 changed little after progesterone treatment as shown by Western blot, indicating that most of mES cells were in undifferentiated state. The results of FACS proved that progesterone promoted DNA synthesis in mES cells. The proportion of mES cells in S+G(2)/M phase was higher in progesterone-treated group than that in the control group. Cyclins and cyclin-dependent kinases, as well as proto-oncogenes (c-myc, c-fos) were up-regulated when cells were treated with progesterone. The results obtained indicate that progesterone promotes propagation and viability of mES cells. The up-regulation of cell cycle-related factors might contribute to the function of progesterone.

  11. Nucleic Acid Aptamers for Living Cell Analysis

    NASA Astrophysics Data System (ADS)

    Xiong, Xiangling; Lv, Yifan; Chen, Tao; Zhang, Xiaobing; Wang, Kemin; Tan, Weihong

    2014-06-01

    Cells as the building blocks of life determine the basic functions and properties of a living organism. Understanding the structure and components of a cell aids in the elucidation of its biological functions. Moreover, knowledge of the similarities and differences between diseased and healthy cells is essential to understanding pathological mechanisms, identifying diagnostic markers, and designing therapeutic molecules. However, monitoring the structures and activities of a living cell remains a challenging task in bioanalytical and life science research. To meet the requirements of this task, aptamers, as “chemical antibodies,” have become increasingly powerful tools for cellular analysis. This article reviews recent advances in the development of nucleic acid aptamers in the areas of cell membrane analysis, cell detection and isolation, real-time monitoring of cell secretion, and intracellular delivery and analysis with living cell models. Limitations of aptamers and possible solutions are also discussed.

  12. Acidic Chitinase Limits Allergic Inflammation and Promotes Intestinal Nematode Expulsion

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Acidic mammalian chitinase (AMCase) is stereotypically induced during mammalian immune responses to helminths and allergens—yet, its precise role in immunity and inflammation is unclear. Here we show that in the lung, genetic ablation of AMCase failed to diminish type 2 inflammation against helmint...

  13. Overexpression of Mucin 13 due to Promoter Methylation Promotes Aggressive Behavior in Ovarian Cancer Cells

    PubMed Central

    Sung, Hye Youn; Park, Ae Kyung

    2014-01-01

    Purpose Recent discoveries suggest that aberrant DNA methylation provides cancer cells with advanced metastatic properties. However, the precise regulatory mechanisms controlling metastasis genes and their role in metastatic transformation are largely unknown. To address epigenetically-regulated gene products involved in ovarian cancer metastasis, we examined the mechanisms regulating mucin 13 (MUC13) expression and its influence on aggressive behaviors of ovarian malignancies. Materials and Methods We injected SK-OV-3 ovarian cancer cells peritoneally into nude mice to mimic human ovarian tumor metastasis. Overexpression of MUC13 mRNA was detected in metastatic implants from the xenografts by expression microarray analysis and quantitative reverse-transcription polymerase chain reaction (qRT-PCR). The DNA methylation status within the MUC13 promoter region was determined using bisulfite sequencing PCR and quantitative methylation-specific PCR. We evaluated the effects of exogenous MUC13 on cell invasion and migration using in vitro transwell assays. Results MUC13 mRNA expression was up-regulated, and methylation of specific CpG sites within the promoter was reduced in the metastatic implants relative to those in wild-type SK-OV-3 cells. Addition of a DNA methyltransferase inhibitor to SK-OV-3 cells induced MUC13 expression, thereby implying epigenetic regulation of MUC13 by promoter methylation. MUC13 overexpression increased migration and invasiveness, compared to control cells, suggesting aberrant up-regulation of MUC13 is strongly associated with progression of aggressive behaviors in ovarian cancer. Conclusion We provide novel evidence for epigenetic regulation of MUC13 in ovarian cancer. We suggest that the DNA methylation status within the MUC13 promoter region may be a potential biomarker of aggressive behavior in ovarian cancer. PMID:25048476

  14. Micro-electro-mechanical systems phosphoric acid fuel cell

    DOEpatents

    Sopchak, David A.; Morse, Jeffrey D.; Upadhye, Ravindra S.; Kotovsky, Jack; Graff, Robert T.

    2010-08-17

    A phosphoric acid fuel cell system comprising a porous electrolyte support, a phosphoric acid electrolyte in the porous electrolyte support, a cathode electrode contacting the phosphoric acid electrolyte, and an anode electrode contacting the phosphoric acid electrolyte.

  15. Micro-electro-mechanical systems phosphoric acid fuel cell

    DOEpatents

    Sopchak, David A.; Morse, Jeffrey D.; Upadhye, Ravindra S.; Kotovsky, Jack; Graff, Robert T.

    2010-12-21

    A phosphoric acid fuel cell system comprising a porous electrolyte support, a phosphoric acid electrolyte in the porous electrolyte support, a cathode electrode contacting the phosphoric acid electrolyte, and an anode electrode contacting the phosphoric acid electrolyte.

  16. An epigenetically distinct breast cancer cell subpopulation promotes collective invasion.

    PubMed

    Westcott, Jill M; Prechtl, Amanda M; Maine, Erin A; Dang, Tuyen T; Esparza, Matthew A; Sun, Han; Zhou, Yunyun; Xie, Yang; Pearson, Gray W

    2015-05-01

    Tumor cells can engage in a process called collective invasion, in which cohesive groups of cells invade through interstitial tissue. Here, we identified an epigenetically distinct subpopulation of breast tumor cells that have an enhanced capacity to collectively invade. Analysis of spheroid invasion in an organotypic culture system revealed that these "trailblazer" cells are capable of initiating collective invasion and promote non-trailblazer cell invasion, indicating a commensal relationship among subpopulations within heterogenous tumors. Canonical mesenchymal markers were not sufficient to distinguish trailblazer cells from non-trailblazer cells, suggesting that defining the molecular underpinnings of the trailblazer phenotype could reveal collective invasion-specific mechanisms. Functional analysis determined that DOCK10, ITGA11, DAB2, PDFGRA, VASN, PPAP2B, and LPAR1 are highly expressed in trailblazer cells and required to initiate collective invasion, with DOCK10 essential for metastasis. In patients with triple-negative breast cancer, expression of these 7 genes correlated with poor outcome. Together, our results indicate that spontaneous conversion of the epigenetic state in a subpopulation of cells can promote a transition from in situ to invasive growth through induction of a cooperative form of collective invasion and suggest that therapeutic inhibition of trailblazer cell invasion may help prevent metastasis.

  17. An epigenetically distinct breast cancer cell subpopulation promotes collective invasion

    PubMed Central

    Westcott, Jill M.; Prechtl, Amanda M.; Maine, Erin A.; Dang, Tuyen T.; Esparza, Matthew A.; Sun, Han; Zhou, Yunyun; Xie, Yang; Pearson, Gray W.

    2015-01-01

    Tumor cells can engage in a process called collective invasion, in which cohesive groups of cells invade through interstitial tissue. Here, we identified an epigenetically distinct subpopulation of breast tumor cells that have an enhanced capacity to collectively invade. Analysis of spheroid invasion in an organotypic culture system revealed that these “trailblazer” cells are capable of initiating collective invasion and promote non-trailblazer cell invasion, indicating a commensal relationship among subpopulations within heterogenous tumors. Canonical mesenchymal markers were not sufficient to distinguish trailblazer cells from non-trailblazer cells, suggesting that defining the molecular underpinnings of the trailblazer phenotype could reveal collective invasion-specific mechanisms. Functional analysis determined that DOCK10, ITGA11, DAB2, PDFGRA, VASN, PPAP2B, and LPAR1 are highly expressed in trailblazer cells and required to initiate collective invasion, with DOCK10 essential for metastasis. In patients with triple-negative breast cancer, expression of these 7 genes correlated with poor outcome. Together, our results indicate that spontaneous conversion of the epigenetic state in a subpopulation of cells can promote a transition from in situ to invasive growth through induction of a cooperative form of collective invasion and suggest that therapeutic inhibition of trailblazer cell invasion may help prevent metastasis. PMID:25844900

  18. ANKHD1 represses p21 (WAF1/CIP1) promoter and promotes multiple myeloma cell growth.

    PubMed

    Dhyani, Anamika; Machado-Neto, João A; Favaro, Patricia; Saad, Sara T Olalla

    2015-01-01

    ANKHD1 (Ankyrin repeat and KH domain-containing protein 1) is highly expressed and plays an important role in the proliferation and cell cycle progression of multiple myeloma (MM) cells. ANKHD1 downregulation modulates cell cycle gene expression and upregulates p21 irrespective of the TP53 mutational status of MM cell lines. The present study was aimed to investigate the role of ANKHD1 in MM in vitro clonogenicity and in vivo tumourigenicity, as well as the role of ANKHD1 in p21 transcriptional regulation. ANKHD1 silencing in MM cells resulted in significantly low no. of colonies formed and in slow migration as compared to control cells (p < 0.05). Furthermore, in xenograft MM mice models, tumour growth was visibly suppressed in mice injected with ANKHD1 silenced cells compared to the control group. There was a significant decrease in tumour volume (p = 0.006) as well as in weight (p = 0.02) in the group injected with silenced cells compared to those of the control group. Co-immunoprecipitation and chromatin immunoprecipitation (ChIP) assays confirmed the interaction between p21 and ANKHD1. Moreover, overexpression of ANKHD1 downregulated the activity of a p21 promoter in luciferase assays. Decrease in luciferase activity suggests a direct role of ANKHD1 in p21 transcriptional regulation. In addition confocal analysis after U266 cells were treated with Leptomycin B (LMB) for 24 h showed accumulation of ANKHD1 inside the nucleus as compared to untreated cells where ANKHD1 was found to be predominantly in cytoplasm. This suggests ANKHD1 might be shuttling between cytoplasm and nucleus. In conclusion, ANKHD1 promotes MM growth by repressing p21 a potent cell cycle regulator.

  19. AtWRKY22 promotes susceptibility to aphids and modulates salicylic acid and jasmonic acid signalling

    PubMed Central

    Kloth, Karen J.; Wiegers, Gerrie L.; Busscher-Lange, Jacqueline; van Haarst, Jan C.; Kruijer, Willem; Bouwmeester, Harro J.; Dicke, Marcel; Jongsma, Maarten A.

    2016-01-01

    Aphids induce many transcriptional perturbations in their host plants, but the signalling cascades responsible and the effects on plant resistance are largely unknown. Through a genome-wide association (GWA) mapping study in Arabidopsis thaliana, we identified WRKY22 as a candidate gene associated with feeding behaviour of the green peach aphid, Myzus persicae. The transcription factor WRKY22 is known to be involved in pathogen-triggered immunity, and WRKY22 gene expression has been shown to be induced by aphids. Assessment of aphid population development and feeding behaviour on knockout mutants and overexpression lines showed that WRKY22 increases susceptibility to M. persicae via a mesophyll-located mechanism. mRNA sequencing analysis of aphid-infested wrky22 knockout plants revealed the up-regulation of genes involved in salicylic acid (SA) signalling and down-regulation of genes involved in plant growth and cell-wall loosening. In addition, mechanostimulation of knockout plants by clip cages up-regulated jasmonic acid (JA)-responsive genes, resulting in substantial negative JA–SA crosstalk. Based on this and previous studies, WRKY22 is considered to modulate the interplay between the SA and JA pathways in response to a wide range of biotic and abiotic stimuli. Its induction by aphids and its role in suppressing SA and JA signalling make WRKY22 a potential target for aphids to manipulate host plant defences. PMID:27107291

  20. AtWRKY22 promotes susceptibility to aphids and modulates salicylic acid and jasmonic acid signalling.

    PubMed

    Kloth, Karen J; Wiegers, Gerrie L; Busscher-Lange, Jacqueline; van Haarst, Jan C; Kruijer, Willem; Bouwmeester, Harro J; Dicke, Marcel; Jongsma, Maarten A

    2016-05-01

    Aphids induce many transcriptional perturbations in their host plants, but the signalling cascades responsible and the effects on plant resistance are largely unknown. Through a genome-wide association (GWA) mapping study in Arabidopsis thaliana, we identified WRKY22 as a candidate gene associated with feeding behaviour of the green peach aphid, Myzus persicae The transcription factor WRKY22 is known to be involved in pathogen-triggered immunity, and WRKY22 gene expression has been shown to be induced by aphids. Assessment of aphid population development and feeding behaviour on knockout mutants and overexpression lines showed that WRKY22 increases susceptibility to M. persicae via a mesophyll-located mechanism. mRNA sequencing analysis of aphid-infested wrky22 knockout plants revealed the up-regulation of genes involved in salicylic acid (SA) signalling and down-regulation of genes involved in plant growth and cell-wall loosening. In addition, mechanostimulation of knockout plants by clip cages up-regulated jasmonic acid (JA)-responsive genes, resulting in substantial negative JA-SA crosstalk. Based on this and previous studies, WRKY22 is considered to modulate the interplay between the SA and JA pathways in response to a wide range of biotic and abiotic stimuli. Its induction by aphids and its role in suppressing SA and JA signalling make WRKY22 a potential target for aphids to manipulate host plant defences. PMID:27107291

  1. AtWRKY22 promotes susceptibility to aphids and modulates salicylic acid and jasmonic acid signalling.

    PubMed

    Kloth, Karen J; Wiegers, Gerrie L; Busscher-Lange, Jacqueline; van Haarst, Jan C; Kruijer, Willem; Bouwmeester, Harro J; Dicke, Marcel; Jongsma, Maarten A

    2016-05-01

    Aphids induce many transcriptional perturbations in their host plants, but the signalling cascades responsible and the effects on plant resistance are largely unknown. Through a genome-wide association (GWA) mapping study in Arabidopsis thaliana, we identified WRKY22 as a candidate gene associated with feeding behaviour of the green peach aphid, Myzus persicae The transcription factor WRKY22 is known to be involved in pathogen-triggered immunity, and WRKY22 gene expression has been shown to be induced by aphids. Assessment of aphid population development and feeding behaviour on knockout mutants and overexpression lines showed that WRKY22 increases susceptibility to M. persicae via a mesophyll-located mechanism. mRNA sequencing analysis of aphid-infested wrky22 knockout plants revealed the up-regulation of genes involved in salicylic acid (SA) signalling and down-regulation of genes involved in plant growth and cell-wall loosening. In addition, mechanostimulation of knockout plants by clip cages up-regulated jasmonic acid (JA)-responsive genes, resulting in substantial negative JA-SA crosstalk. Based on this and previous studies, WRKY22 is considered to modulate the interplay between the SA and JA pathways in response to a wide range of biotic and abiotic stimuli. Its induction by aphids and its role in suppressing SA and JA signalling make WRKY22 a potential target for aphids to manipulate host plant defences.

  2. Method for promoting specific alignment of short oligonucleotides on nucleic acids

    DOEpatents

    Studier, F. William; Kieleczawa, Jan; Dunn, John J.

    1996-01-01

    Disclosed is a method for promoting specific alignment of short oligonucleotides on a nucleic acid polymer. The nucleic acid polymer is incubated in a solution containing a single-stranded DNA-binding protein and a plurality of oligonucleotides which are perfectly complementary to distinct but adjacent regions of a predetermined contiguous nucleotide sequence in the nucleic acid polymer. The plurality of oligonucleotides anneal to the nucleic acid polymer to form a contiguous region of double stranded nucleic acid. Specific application of the methods disclosed include priming DNA synthesis and template-directed ligation.

  3. Metabolic pathways promoting cancer cell survival and growth.

    PubMed

    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.

  4. Formic acid fuel cells and catalysts

    DOEpatents

    Masel, Richard I.; Larsen, Robert; Ha, Su Yun

    2010-06-22

    An exemplary fuel cell of the invention includes a formic acid fuel solution in communication with an anode (12, 134), an oxidizer in communication with a cathode (16, 135) electrically linked to the anode, and an anode catalyst that includes Pd. An exemplary formic acid fuel cell membrane electrode assembly (130) includes a proton-conducting membrane (131) having opposing first (132) and second surfaces (133), a cathode catalyst on the second membrane surface, and an anode catalyst including Pd on the first surface.

  5. Global view of enhancer–promoter interactome in human cells

    PubMed Central

    He, Bing; Chen, Changya; Teng, Li; Tan, Kai

    2014-01-01

    Enhancer mapping has been greatly facilitated by various genomic marks associated with it. However, little is available in our toolbox to link enhancers with their target promoters, hampering mechanistic understanding of enhancer–promoter (EP) interaction. We develop and characterize multiple genomic features for distinguishing true EP pairs from noninteracting pairs. We integrate these features into a probabilistic predictor for EP interactions. Multiple validation experiments demonstrate a significant improvement over state-of-the-art approaches. Systematic analyses of EP interactions across 12 cell types reveal several global features of EP interactions: (i) a larger fraction of EP interactions are cell type specific than enhancers; (ii) promoters controlled by multiple enhancers have higher tissue specificity, but the regulating enhancers are less conserved; (iii) cohesin plays a role in mediating tissue-specific EP interactions via chromatin looping in a CTCF-independent manner. Our approach presents a systematic and effective strategy to decipher the mechanisms underlying EP communication. PMID:24821768

  6. Identification of a novel temperature sensitive promoter in cho cells

    PubMed Central

    2011-01-01

    Background The Chinese hamster ovary (CHO) expression system is the leading production platform for manufacturing biopharmaceuticals for the treatment of numerous human diseases. Efforts to optimize the production process also include the genetic construct encoding the therapeutic gene. Here we report about the successful identification of an endogenous highly active gene promoter obtained from CHO cells which shows conditionally inducible gene expression at reduced temperature. Results Based on CHO microarray expression data abundantly transcribed genes were selected as potential promoter candidates. The S100a6 (calcyclin) and its flanking regions were identified from a genomic CHO-K1 lambda-phage library. Computational analyses showed a predicted TSS, a TATA-box and several TFBSs within the 1.5 kb region upstream the ATG start signal. Various constructs were investigated for promoter activity at 37°C and 33°C in transient luciferase reporter gene assays. Most constructs showed expression levels even higher than the SV40 control and on average a more than two-fold increase at lower temperature. We identified the core promoter sequence (222 bp) comprising two SP1 sites and could show a further increase in activity by duplication of this minimal sequence. Conclusions This novel CHO promoter permits conditionally high-level gene expression. Upon a shift to 33°C, a two to three-fold increase of basal productivity (already higher than SV40 promoter) is achieved. This property is of particular advantage for a process with reduced expression during initial cell growth followed by the production phase at low temperature with a boost in expression. Additionally, production of toxic proteins becomes feasible, since cell metabolism and gene expression do not directly interfere. The CHO S100a6 promoter can be characterized as cold-shock responsive with the potential for improving process performance of mammalian expression systems. PMID:21569433

  7. Sorafenib-induced defective autophagy promotes cell death by necroptosis.

    PubMed

    Kharaziha, Pedram; Chioureas, Dimitris; Baltatzis, George; Fonseca, Pedro; Rodriguez, Patricia; Gogvadze, Vladimir; Lennartsson, Lena; Björklund, Ann-Charlotte; Zhivotovsky, Boris; Grandér, Dan; Egevad, Lars; Nilsson, Sten; Panaretakis, Theocharis

    2015-11-10

    Autophagy is one of the main cytoprotective mechanisms that cancer cells deploy to withstand the cytotoxic stress and survive the lethal damage induced by anti-cancer drugs. However, under specific conditions, autophagy may, directly or indirectly, induce cell death. In our study, treatment of the Atg5-deficient DU145 prostate cancer cells, with the multi-tyrosine kinase inhibitor, sorafenib, induces mitochondrial damage, autophagy and cell death. Molecular inhibition of autophagy by silencing ULK1 and Beclin1 rescues DU145 cells from cell death indicating that, in this setting, autophagy promotes cell death. Re-expression of Atg5 restores the lipidation of LC3 and rescues DU145 and MEF atg5-/- cells from sorafenib-induced cell death. Despite the lack of Atg5 expression and LC3 lipidation, DU145 cells form autophagosomes as demonstrated by transmission and immuno-electron microscopy, and the formation of LC3 positive foci. However, the lack of cellular content in the autophagosomes, the accumulation of long-lived proteins, the presence of GFP-RFP-LC3 positive foci and the accumulated p62 protein levels indicate that these autophagosomes may not be fully functional. DU145 cells treated with sorafenib undergo a caspase-independent cell death that is inhibited by the RIPK1 inhibitor, necrostatin-1. Furthermore, treatment with sorafenib induces the interaction of RIPK1 with p62, as demonstrated by immunoprecipitation and a proximity ligation assay. Silencing of p62 decreases the RIPK1 protein levels and renders necrostatin-1 ineffective in blocking sorafenib-induced cell death. In summary, the formation of Atg5-deficient autophagosomes in response to sorafenib promotes the interaction of p62 with RIPK leading to cell death by necroptosis. PMID:26416459

  8. Sorafenib-induced defective autophagy promotes cell death by necroptosis

    PubMed Central

    Kharaziha, Pedram; Chioureas, Dimitris; Baltatzis, George; Fonseca, Pedro; Rodriguez, Patricia; Gogvadze, Vladimir; Lennartsson, Lena; Björklund, Ann-Charlotte; Zhivotovsky, Boris; Grandér, Dan; Egevad, Lars; Nilsson, Sten; Panaretakis, Theocharis

    2015-01-01

    Autophagy is one of the main cytoprotective mechanisms that cancer cells deploy to withstand the cytotoxic stress and survive the lethal damage induced by anti-cancer drugs. However, under specific conditions, autophagy may, directly or indirectly, induce cell death. In our study, treatment of the Atg5-deficient DU145 prostate cancer cells, with the multi-tyrosine kinase inhibitor, sorafenib, induces mitochondrial damage, autophagy and cell death. Molecular inhibition of autophagy by silencing ULK1 and Beclin1 rescues DU145 cells from cell death indicating that, in this setting, autophagy promotes cell death. Re-expression of Atg5 restores the lipidation of LC3 and rescues DU145 and MEF atg5−/− cells from sorafenib-induced cell death. Despite the lack of Atg5 expression and LC3 lipidation, DU145 cells form autophagosomes as demonstrated by transmission and immuno-electron microscopy, and the formation of LC3 positive foci. However, the lack of cellular content in the autophagosomes, the accumulation of long-lived proteins, the presence of GFP-RFP-LC3 positive foci and the accumulated p62 protein levels indicate that these autophagosomes may not be fully functional. DU145 cells treated with sorafenib undergo a caspase-independent cell death that is inhibited by the RIPK1 inhibitor, necrostatin-1. Furthermore, treatment with sorafenib induces the interaction of RIPK1 with p62, as demonstrated by immunoprecipitation and a proximity ligation assay. Silencing of p62 decreases the RIPK1 protein levels and renders necrostatin-1 ineffective in blocking sorafenib-induced cell death. In summary, the formation of Atg5-deficient autophagosomes in response to sorafenib promotes the interaction of p62 with RIPK leading to cell death by necroptosis. PMID:26416459

  9. Pancreatic Tumor Cell Secreted CCN1/Cyr61 Promotes Endothelial cell migration and Aberrant Neovascularization

    PubMed Central

    Maity, Gargi; Mehta, Smita; Haque, Inamul; Dhar, Kakali; Sarkar, Sandipto; Banerjee, Sushanta K.; Banerjee, Snigdha

    2014-01-01

    The complex signaling networks between cancer cells and adjacent endothelial cells make it challenging to unravel how cancer cells send extracellular messages to promote aberrant vascularization or tumor angiogenesis. Here, in vitro and in vivo models show that pancreatic cancer cell generated unique microenvironments can underlie endothelial cell migration and tumor angiogenesis. Mechanistically, we find that pancreatic cancer cell secreted CCN1/Cyr61 matricellular protein rewires the microenvironment to promote endothelial cell migration and tumor angiogenesis. This event can be overcome by Sonic Hedgehog (SHh) antibody treatment. Collectively, these studies identify a novel CCN1 signaling program in pancreatic cancer cells which activates SHh through autocrine-paracrine circuits to promote endothelial cell migration and tumor angiogenesis and suggests that CCN1 signaling of pancreatic cancer cells is vital for the regulation of tumor angiogenesis. Thus CCN1 signaling could be an ideal target for tumor vascular disruption in pancreatic cancer. PMID:24833309

  10. Algae as an electron donor promoting sulfate reduction for the bioremediation of acid rock drainage.

    PubMed

    Ayala-Parra, Pedro; Sierra-Alvarez, Reyes; Field, Jim A

    2016-11-01

    This study assessed bioremediation of acid rock drainage in simulated permeable reactive barriers (PRB) using algae, Chlorella sorokiniana, as the sole electron donor for sulfate-reducing bacteria. Lipid extracted algae (LEA), the residues of biodiesel production, were compared with whole cell algae (WCA) as an electron donor to promote sulfate-reducing activity. Inoculated columns containing anaerobic granular sludge were fed a synthetic medium containing H2SO4 and Cu(2+). Sulfate, sulfide, Cu(2+) and pH were monitored throughout the experiment of 123d. Cu recovered in the column packing at the end of the experiment was evaluated using sequential extraction. Both WCA and LEA promoted 80% of sulfate removal (12.7mg SO4(2-) d(-1)) enabling near complete Cu removal (>99.5%) and alkalinity generation raising the effluent pH to 6.5. No noteworthy sulfate reduction, alkalinity formation and Cu(2+) removal were observed in the endogenous control. In algae amended-columns, Cu(2+) was precipitated with biogenic H2S produced by sulfate reduction. Formation of CuS was evidenced by sequential extraction and X-ray diffraction. LEA and WCA provided similar levels of electron donor based on the COD balance. The results demonstrate an innovative passive remediation system using residual algae biomass from the biodiesel industry.

  11. Algae as an electron donor promoting sulfate reduction for the bioremediation of acid rock drainage.

    PubMed

    Ayala-Parra, Pedro; Sierra-Alvarez, Reyes; Field, Jim A

    2016-11-01

    This study assessed bioremediation of acid rock drainage in simulated permeable reactive barriers (PRB) using algae, Chlorella sorokiniana, as the sole electron donor for sulfate-reducing bacteria. Lipid extracted algae (LEA), the residues of biodiesel production, were compared with whole cell algae (WCA) as an electron donor to promote sulfate-reducing activity. Inoculated columns containing anaerobic granular sludge were fed a synthetic medium containing H2SO4 and Cu(2+). Sulfate, sulfide, Cu(2+) and pH were monitored throughout the experiment of 123d. Cu recovered in the column packing at the end of the experiment was evaluated using sequential extraction. Both WCA and LEA promoted 80% of sulfate removal (12.7mg SO4(2-) d(-1)) enabling near complete Cu removal (>99.5%) and alkalinity generation raising the effluent pH to 6.5. No noteworthy sulfate reduction, alkalinity formation and Cu(2+) removal were observed in the endogenous control. In algae amended-columns, Cu(2+) was precipitated with biogenic H2S produced by sulfate reduction. Formation of CuS was evidenced by sequential extraction and X-ray diffraction. LEA and WCA provided similar levels of electron donor based on the COD balance. The results demonstrate an innovative passive remediation system using residual algae biomass from the biodiesel industry. PMID:27318730

  12. Ethanol promotes T cell apoptosis through the mitochondrial pathway

    PubMed Central

    Kapasi, Aditi A; Patel, Geeta; Goenka, Anuj; Nahar, Nilay; Modi, Neeraj; Bhaskaran, Madhu; Reddy, Krishna; Franki, Nicholas; Patel, Jaimita; Singhal, Pravin C

    2003-01-01

    Clinical reports suggest that acute ethanol intoxication is often associated with lymphopenia. Previously, ethanol was reported to invoke thymocyte apoptosis. We studied the effect of ethanol on T cell apoptosis. In addition, we evaluated the molecular mechanism of ethanol-induced T cell apoptosis. Human T cells harvested from healthy subjects after an alcohol drinking binge showed enhanced T cell apoptosis (before, 0·4 ± 0·2% versus after, 19·6 ± 2·5% apoptotic lymphocytes/field; P < 0·001). In in vitro studies, ethanol in a concentration of 50 mm and higher enhanced the apoptosis of Jurkat cells. DNA isolated from ethanol-treated Jurkat cells displayed integer multiples of 180 base pairs. Ethanol decreased Jurkat cell expression of Bcl-2, whereas ethanol increased Jurkat cell expression of Bax. Jurkat cells treated with ethanol also showed translocation of cytochrome C into cytosol. Moreover, a caspase-9 inhibitor partially inhibited ethanol-induced Jurkat cell apoptosis. In in vivo studies, after binge drinking, T cell expression of Bcl-2 also decreased. In addition, binge drinking induced the cleavage of caspase-3, suggesting activation of caspase-3 in T cells. These results suggest that ethanol promotes T cell apoptosis through the activation of intrinsic or mitochondrial pathway. PMID:12603597

  13. Phosphoric Acid Fuel Cell Technology Status

    NASA Technical Reports Server (NTRS)

    Simons, S. N.; King, R. B.; Prokopius, P. R.

    1981-01-01

    A review of the current phosphoric acid fuel cell system technology development efforts is presented both for multimegawatt systems for electric utility applications and for multikilowatt systems for on-site integrated energy system applications. Improving fuel cell performance, reducing cost, and increasing durability are the technology drivers at this time. Electrodes, matrices, intercell cooling, bipolar/separator plates, electrolyte management, and fuel selection are discussed.

  14. Solid acids as fuel cell electrolytes.

    PubMed

    Haile, S M; Boysen, D A; Chisholm, C R; Merle, R B

    2001-04-19

    Fuel cells are attractive alternatives to combustion engines for electrical power generation because of their very high efficiencies and low pollution levels. Polymer electrolyte membrane fuel cells are generally considered to be the most viable approach for mobile applications. However, these membranes require humid operating conditions, which limit the temperature of operation to less than 100 degrees C; they are also permeable to methanol and hydrogen, which lowers fuel efficiency. Solid, inorganic, acid compounds (or simply, solid acids) such as CsHSO4 and Rb3H(SeO4)2 have been widely studied because of their high proton conductivities and phase-transition behaviour. For fuel-cell applications they offer the advantages of anhydrous proton transport and high-temperature stability (up to 250 degrees C). Until now, however, solid acids have not been considered viable fuel-cell electrolyte alternatives owing to their solubility in water and extreme ductility at raised temperatures (above approximately 125 degrees C). Here we show that a cell made of a CsHSO4 electrolyte membrane (about 1.5 mm thick) operating at 150-160 degrees C in a H2/O2 configuration exhibits promising electrochemical performances: open circuit voltages of 1.11 V and current densities of 44 mA cm-2 at short circuit. Moreover, the solid-acid properties were not affected by exposure to humid atmospheres. Although these initial results show promise for applications, the use of solid acids in fuel cells will require the development of fabrication techniques to reduce electrolyte thickness, and an assessment of possible sulphur reduction following prolonged exposure to hydrogen.

  15. Eliminating aluminum toxicity in an acid sulfate soil for rice cultivation using plant growth promoting bacteria.

    PubMed

    Panhwar, Qurban Ali; Naher, Umme Aminun; Radziah, Othman; Shamshuddin, Jusop; Razi, Ismail Mohd

    2015-01-01

    Aluminum toxicity is widely considered as the most important limiting factor for plants growing in acid sulfate soils. A study was conducted in laboratory and in field to ameliorate Al toxicity using plant growth promoting bacteria (PGPB), ground magnesium limestone (GML) and ground basalt. Five-day-old rice seedlings were inoculated by Bacillus sp., Stenotrophomonas maltophila, Burkholderia thailandensis and Burkholderia seminalis and grown for 21 days in Hoagland solution (pH 4.0) at various Al concentrations (0, 50 and 100 μM). Toxicity symptoms in root and leaf were studied using scanning electron microscope. In the field, biofertilizer (PGPB), GML and basalt were applied (4 t·ha-1 each). Results showed that Al severely affected the growth of rice. At high concentrations, the root surface was ruptured, leading to cell collapse; however, no damages were observed in the PGPB inoculated seedlings. After 21 days of inoculation, solution pH increased to >6.0, while the control treatment remained same. Field study showed that the highest rice growth and yield were obtained in the bio-fertilizer and GML treatments. This study showed that Al toxicity was reduced by PGPB via production of organic acids that were able to chelate the Al and the production of polysaccharides that increased solution pH. The release of phytohormones further enhanced rice growth that resulted in yield increase. PMID:25710843

  16. Selective Retinoic Acid Receptor γ Agonists Promote Repair of Injured Skeletal Muscle in Mouse

    PubMed Central

    Di Rocco, Agnese; Uchibe, Kenta; Larmour, Colleen; Berger, Rebecca; Liu, Min; Barton, Elisabeth R.; Iwamoto, Masahiro

    2016-01-01

    Retinoic acid signaling regulates several biological events, including myogenesis. We previously found that retinoic acid receptor γ (RARγ) agonist blocks heterotopic ossification, a pathological bone formation that mostly occurs in the skeletal muscle. Interestingly, RARγ agonist also weakened deterioration of muscle architecture adjacent to the heterotopic ossification lesion, suggesting that RARγ agonist may oppose skeletal muscle damage. To test this hypothesis, we generated a critical defect in the tibialis anterior muscle of 7-week-old mice with a cautery, treated them with RARγ agonist or vehicle corn oil, and examined the effects of RARγ agonist on muscle repair. The muscle defects were partially repaired with newly regenerating muscle cells, but also filled with adipose and fibrous scar tissue in both RARγ-treated and control groups. The fibrous or adipose area was smaller in RARγ agonist–treated mice than in the control. In addition, muscle repair was remarkably delayed in RARγ-null mice in both critical defect and cardiotoxin injury models. Furthermore, we found a rapid increase in retinoid signaling in lacerated muscle, as monitored by retinoid signaling reporter mice. Together, our results indicate that endogenous RARγ signaling is involved in muscle repair and that selective RARγ agonists may be beneficial to promote repair in various types of muscle injuries. PMID:26205250

  17. Impaired DNA replication within progenitor cell pools promotes leukemogenesis.

    PubMed

    Bilousova, Ganna; Marusyk, Andriy; Porter, Christopher C; Cardiff, Robert D; DeGregori, James

    2005-12-01

    Impaired cell cycle progression can be paradoxically associated with increased rates of malignancies. Using retroviral transduction of bone marrow progenitors followed by transplantation into mice, we demonstrate that inhibition of hematopoietic progenitor cell proliferation impairs competition, promoting the expansion of progenitors that acquire oncogenic mutations which restore cell cycle progression. Conditions that impair DNA replication dramatically enhance the proliferative advantage provided by the expression of Bcr-Abl or mutant p53, which provide no apparent competitive advantage under conditions of healthy replication. Furthermore, for the Bcr-Abl oncogene the competitive advantage in contexts of impaired DNA replication dramatically increases leukemogenesis. Impaired replication within hematopoietic progenitor cell pools can select for oncogenic events and thereby promote leukemia, demonstrating the importance of replicative competence in the prevention of tumorigenesis. The demonstration that replication-impaired, poorly competitive progenitor cell pools can promote tumorigenesis provides a new rationale for links between tumorigenesis and common human conditions of impaired DNA replication such as dietary folate deficiency, chemotherapeutics targeting dNTP synthesis, and polymorphisms in genes important for DNA metabolism. PMID:16277552

  18. Ginseng Berry Extract Promotes Maturation of Mouse Dendritic Cells

    PubMed Central

    Zhang, Wei; Cho, Si-Young; Xiang, Gao; Min, Kyung-Jin; Yu, Qing; Jin, Jun-O

    2015-01-01

    Ginseng extract has been shown to possess certain anti-virus, anti-tumor and immune-activating effects. However, the immunostimulatory effect of ginseng berry extract (GB) has been less well characterized. In this study, we investigated the effect of GB on the activation of mouse dendritic cells (DCs) in vitro and in vivo. GB treatment induced up-regulation of co-stimulatory molecules in bone marrow-derived DCs (BMDCs). Interestingly, GB induced a higher degree of co-stimulatory molecule up-regulation than ginseng root extract (GR) at the same concentrations. Moreover, in vivo administration of GB promoted up-regulation of CD86, MHC class I and MHC class II and production of IL-6, IL-12 and TNF-α in spleen DCs. GB also promoted the generation of Th1 and Tc1 cells. Furthermore, Toll like receptor 4 (TLR4) and myeloid differentiation primary response 88 (MyD88) signaling pathway were essential for DC activation induced by GB. In addition, GB strongly prompted the proliferation of ovalbumin (OVA)-specific CD4 and CD8 T cells. Finally, GB induced DC activation in tumor-bearing mice and the combination of OVA and GB treatment inhibited B16-OVA tumor cell growth in C57BL/6 mice. These results demonstrate that GB is a novel tumor therapeutic vaccine adjuvant by promoting DC and T cell activation. PMID:26090808

  19. MRG15 activates the cdc2 promoter via histone acetylation in human cells

    SciTech Connect

    Pena, AndreAna N.; Tominaga, Kaoru; Pereira-Smith, Olivia M.

    2011-07-01

    Chromatin remodeling is required for transcriptional activation and repression. MRG15 (MORF4L1), a chromatin modulator, is a highly conserved protein and is present in complexes containing histone acetyltransferases (HATs) as well as histone deacetylases (HDACs). Loss of expression of MRG15 in mice and Drosophila results in embryonic lethality and fibroblast and neural stem/progenitor cells cultured from Mrg15 null mouse embryos exhibit marked proliferative defects when compared with wild type cells. To determine the role of MRG15 in cell cycle progression we performed chromatin immunoprecipitation with an antibody to MRG15 on normal human fibroblasts as they entered the cell cycle from a quiescent state, and analyzed various cell cycle gene promoters. The results demonstrated a 3-fold increase in MRG15 occupancy at the cdc2 promoter during S phase of the cell cycle and a concomitant increase in acetylated histone H4. H4 lysine 12 was acetylated at 24 h post-serum stimulation while there was no change in acetylation of lysine 16. HDAC1 and 2 were decreased at this promoter during cell cycle progression. Over-expression of MRG15 in HeLa cells activated a cdc2 promoter-reporter construct in a dose-dependent manner, whereas knockdown of MRG15 resulted in decreased promoter activity. In order to implicate HAT activity, we treated cells with the HAT inhibitor anacardic acid and determined that HAT inhibition results in loss of expression of cdc2 mRNA. Further, chromatin immunoprecipitation with Tip60 localizes the protein to the same 110 bp stretch of the cdc2 promoter pulled down by MRG15. Additionally, we determined that cotransfection of MRG15 with the known associated HAT Tip60 had a cooperative effect in activating the cdc2 promoter. These results suggest that MRG15 is acting in a HAT complex involving Tip60 to modify chromatin via acetylation of histone H4 at the cdc2 promoter to activate transcription.

  20. Radiation induced cell inactivation as cause for cancer promotion

    NASA Astrophysics Data System (ADS)

    Heidenreich, W. F.; Paretzke, H. G.

    In space research, estimates of health risks from high-LET radiation are needed. Features of several applications of the biologically based two step clonal expansion (TSCE) model on data with the high-LET radiation, normally alpha particles, from radon and from Thorotrast are reviewed. One conclusion is that radiation might not only influence the initiating event in carcinogenesis, but may also act as a promoter. A possible mechanism which gives a promoting action from cell inactivation is presented for the organs "lung", with a two-dimensional arrangement of the cells at risk, and for "liver" where the sensitive cells are distributed in all three dimensions. Inferences for dose-response curves at low doses and dose rates are drawn.

  1. Overexpression of ankyrin1 promotes pancreatic cancer cell growth

    PubMed Central

    Omura, Noriyuki; Mizuma, Masamichi; MacGregor, Anne; Hong, Seung-Mo; Ayars, Michael; Almario, Jose Alejandro; Borges, Michael; Kanda, Mitsuro; Li, Ang; Vincent, Audrey; Maitra, Anirban; Goggins, Michael

    2016-01-01

    The methylation status of a promoter influences gene expression and aberrant methylation during tumor development has important functional consequences for pancreatic and other cancers. Using methylated CpG island amplification and promoter microarrays, we identified ANK1 as hypomethylated in pancreatic cancers. Expression analysis determined ANK1 as commonly overexpressed in pancreatic cancers relative to normal pancreas. ANK1 was co-expressed with miR-486 in pancreatic cancer cells. Stable knockdown of ANK1 in the pancreatic cancer cell line AsPC1 led to changes in cell morphology, and decreases in colony formation. Stable knockdown of ANK1 also marked reduced the growth of tumors in athymic nude mice. Among patients undergoing pancreaticoduodenectomy, those with pancreatic cancers expressing ANK1 had a poorer prognosis than those without ANK1 expression. These findings indicate a role for ANK1 overexpression in mediating pancreatic cancer tumorigenicity. PMID:27144336

  2. The reorientation of cell nucleus promotes the establishment of front-rear polarity in migrating fibroblasts.

    PubMed

    Maninová, Miloslava; Klímová, Zuzana; Parsons, J Thomas; Weber, Michael J; Iwanicki, Marcin P; Vomastek, Tomáš

    2013-06-12

    The establishment of cell polarity is an essential step in the process of cell migration. This process requires precise spatiotemporal coordination of signaling pathways that in most cells create the typical asymmetrical profile of a polarized cell with nucleus located at the cell rear and the microtubule organizing center (MTOC) positioned between the nucleus and the leading edge. During cell polarization, nucleus rearward positioning promotes correct microtubule organizing center localization and thus the establishment of front-rear polarity and directional migration. We found that cell polarization and directional migration require also the reorientation of the nucleus. Nuclear reorientation is manifested as temporally restricted nuclear rotation that aligns the nuclear axis with the axis of cell migration. We also found that nuclear reorientation requires physical connection between the nucleus and cytoskeleton mediated by the LINC (linker of nucleoskeleton and cytoskeleton) complex. Nuclear reorientation is controlled by coordinated activity of lysophosphatidic acid (LPA)-mediated activation of GTPase Rho and the activation of integrin, FAK (focal adhesion kinase), Src, and p190RhoGAP signaling pathway. Integrin signaling is spatially induced at the leading edge as FAK and p190RhoGAP are predominantly activated or localized at this location. We suggest that integrin activation within lamellipodia defines cell front, and subsequent FAK, Src, and p190RhoGAP signaling represents the polarity signal that induces reorientation of the nucleus and thus promotes the establishment of front-rear polarity.

  3. Noncytotoxic and Antitumour-Promoting Activities of Garcinia Acid Esters from Garcinia atroviridis Griff. ex T. Anders (Guttiferae)

    PubMed Central

    Mackeen, Mukram M.; Mooi, Lim Y.; Amran, Mohidin; Mat, Nashriyah; Lajis, Nordin H.; Ali, Abdul M.

    2012-01-01

    The in vitro antitumour-promoting, cytotoxic, and antioxidant activities of two ester derivatives of garcinia acid, that is, 2-(butoxycarbonylmethyl)-3-butoxycarbonyl-2-hydroxy-3-propanolide (1) and 1′,1′′-dibutyl methyl hydroxycitrate (2), that had been previously isolated from the fruits of Garcinia atroviridis Griff. ex T. Anders (Guttiferae), were examined. Based on the inhibition of Epstein-Barr virus early antigen (EBV-EA) activation, compound 1 (IC50: 70 μM) showed much higher (8-fold) antitumour-promoting activity than compound 2 (IC50: 560 μM). In addition, both compounds were nontoxic towards CEM-SS (human T-lymphoblastic leukemia) cells (CD50: >100 μM), Raji (human B-lymphoblastoid) cells (CD50: >600 μM), and brine shrimp (LD50: >300 μM). Although the antitumour-promoting activity of compound 1 is moderate compared with the known antitumour promoter genistein, its non-toxicity suggests the potential of compound 1 and related structures as chemopreventive agents. The weak antioxidant activity displayed by both compounds also suggested that the primary antitumour-promoting mechanism of compound 1 did not involve oxidative-stress quenching. PMID:22685487

  4. TBX2 blocks myogenesis and promotes proliferation in rhabdomyosarcoma cells

    PubMed Central

    Zhu, Bo; Zhang, Meiling; Byrum, Stephanie D.; Tackett, Alan J.; Davie, Judith K.

    2014-01-01

    Rhabdomyosarcomas (RMS) are the most frequent soft tissue sarcomas in children that share many features of developing skeletal muscle. We have discovered that a T-box family member, TBX2, is highly up regulated in tumor cells of both major RMS subtypes. TBX2 is a repressor that is often over expressed in cancer cells and is thought to function in bypassing cell growth control, including repression of p14 and p21. The cell cycle regulator p21 is required for the terminal differentiation of skeletal muscle cells and is silenced in RMS cells. We have found that TBX2 interacts with the myogenic regulatory factors MyoD and myogenin and inhibits the activity of these factors. TBX2 is expressed in primary myoblasts and C2C12 cells, but is strongly down regulated upon differentiation. TBX2 recruits the histone deacetylase HDAC1 and is a potent inhibitor of the expression of muscle specific genes and the cell cycle regulators, p21 and p14. TBX2 promotes the proliferation of RMS cells and either depletions of TBX2 or dominant negative TBX2 up regulate p21 and muscle specific genes. Significantly, depletion or interference with TBX2 completely inhibits tumor growth in a xenograft assay, highlighting the oncogenic role of TBX2 in RMS cells. Thus, the data demonstrate that elevated expression of TBX2 contributes to the pathology of RMS cells by promoting proliferation and repressing differentiation specific gene expression. These results show that deregulated TBX2 serves as an oncogene in RMS, suggesting that TBX2 may serve as a new diagnostic marker or therapeutic target for RMS tumors. PMID:24470334

  5. Multi-input regulation and logic with T7 promoters in cells and cell free systems

    SciTech Connect

    Iyer, Sukanya; Karig, David K; Norred, Sarah E; Simpson, Michael L; Doktycz, Mitchel John

    2014-01-01

    Engineered gene circuits offer an opportunity to harness biological systems for biotechnological and biomedical applications. However, reliance on host E. coli promoters for the construction of circuit elements, such as logic gates, makes implementation of predictable, independently functioning circuits difficult. In contrast, T7 promoters offer a simple orthogonal expression system for use in a variety of cellular backgrounds and even in cell free systems. Here we develop a T7 promoter system that can be regulated by two different transcriptional repressors for the construction of a logic gate that functions in cells and in cell free systems. We first present LacI repressible T7lacO promoters that are regulated from a distal lac operator site for repression. We next explore the positioning of a tet operator site within the T7lacO framework to create T7 promoters that respond to tet and lac repressors and realize an IMPLIES gate. Finally, we demonstrate that these dual input sensitive promoters function in a commercially available E. coli cell-free protein expression system. Together, our results contribute to the first demonstration of multi-input regulation of T7 promoters and expand the utility of T7 promoters in cell based as well as cell-free gene circuits.

  6. Ligand-Promoted, Boron-Mediated Chemoselective Carboxylic Acid Aldol Reaction.

    PubMed

    Nagai, Hideoki; Morita, Yuya; Shimizu, Yohei; Kanai, Motomu

    2016-05-01

    The first carboxylic acid selective aldol reaction mediated by boron compounds and a mild organic base (DBU) was developed. Inclusion of electron-withdrawing groups in the amino acid derivative ligands reacted with BH3·SMe2 forms a boron promoter with increased Lewis acidity at the boron atom and facilitated the carboxylic acid selective enolate formation, even in the presence of other carbonyl groups such as amides, esters, ketones, or aliphatic aldehydes. The remarkable ligand effect led to the broad substrate scope including biologically relevant compounds. PMID:27104352

  7. Genistein promotes DNA demethylation of the steroidogenic factor 1 (SF-1) promoter in endometrial stromal cells

    SciTech Connect

    Matsukura, Hiroshi; Aisaki, Ken-ichi; Igarashi, Katsuhide; Matsushima, Yuko; Kanno, Jun; Muramatsu, Masaaki; Sudo, Katsuko; Sato, Noriko

    2011-08-26

    Highlights: {yields} Genistein (GEN) is a phytoestrogen found in soy products. {yields} GEN demethylated/unsilenced the steroidogenic factor 1 gene in endometrial tissue. {yields} GEN thus altered mRNA expression in uteri of ovariectomized (OVX) mice. {yields} A high-resolution melting assay was used to screen for epigenetic change. {yields} We isolated an endometrial cell clone that was epigenetically modulated by GEN. -- Abstract: It has recently been demonstrated that genistein (GEN), a phytoestrogen in soy products, is an epigenetic modulator in various types of cells; but its effect on endometrium has not yet been determined. We investigated the effects of GEN on mouse uterine cells, in vivo and in vitro. Oral administration of GEN for 1 week induced mild proliferation of the endometrium in ovariectomized (OVX) mice, which was accompanied by the induction of steroidogenic factor 1 (SF-1) gene expression. GEN administration induced demethylation of multiple CpG sites in the SF-1 promoter; these sites are extensively methylated and thus silenced in normal endometrium. The GEN-mediated promoter demethylation occurred predominantly on the luminal side, as opposed to myometrium side, indicating that the epigenetic change was mainly shown in regenerated cells. Primary cultures of endometrial stromal cell colonies were screened for GEN-mediated alterations of DNA methylation by a high-resolution melting (HRM) method. One out of 20 colony-forming cell clones showed GEN-induced demethylation of SF-1. This clone exhibited a high proliferation capacity with continuous colony formation activity through multiple serial clonings. We propose that only a portion of endometrial cells are capable of receiving epigenetic modulation by GEN.

  8. Mesd extrinsically promotes phagocytosis by retinal pigment epithelial cells.

    PubMed

    Chen, Xiuping; Guo, Feiye; LeBlanc, Michelle E; Ding, Ying; Zhang, Chenming; Shakya, Akhalesh; Li, Wei

    2016-08-01

    Phagocytosis is a critical process to maintain tissue homeostasis. In the retina, photoreceptor cells renew their photoexcitability by shedding photoreceptor outer segments (POSs) in a diurnal rhythm. Shed POSs are phagocytosed by retinal pigment epithelial (RPE) cells to prevent debris accumulation, retinal degeneration, and blindness. Phagocytosis ligands are the key to understanding how RPE recognizes shed POSs. Here, we characterized mesoderm development candidate 2 (Mesd or Mesdc2), an endoplasmic reticulum (ER) chaperon for low-density lipoprotein receptor-related proteins (LRPs), to extrinsically promote RPE phagocytosis. The results showed that Mesd stimulated phagocytosis of fluorescence-labeled POS vesicles by D407 RPE cells. Ingested POSs were partially degraded within 3 h in some RPE cells to dispense undegradable fluorophore throughout the cytoplasm. Internalized POSs were colocalized with phagosome biomarker Rab7, suggesting that Mesd-mediated engulfment is involved in a phagocytosis pathway. Mesd also facilitated phagocytosis of POSs by primary RPE cells. Mesd bound to unknown phagocytic receptor(s) on RPE cells. Mesd was detected in the cytoplasm, but not nuclei, of different retinal layers and is predominantly expressed in the ER-free cellular compartment of POSs. Mesd was not secreted into medium from healthy cells but passively released from apoptotic cells with increased membrane permeability. Released Mesd selectively bound to the surface of POS vesicles and apoptotic cells, but not healthy cells. These results suggest that Mesd may be released from and bind to shed POSs to facilitate their phagocytic clearance.

  9. Lycium barbarum polysaccharides promotes in vivo proliferation of adult rat retinal progenitor cells

    PubMed Central

    Wang, Hua; Lau, Benson Wui-Man; Wang, Ning-li; Wang, Si-ying; Lu, Qing-jun; Chang, Raymond Chuen-Chung; So, Kwok-fai

    2015-01-01

    Lycium barbarum is a widely used Chinese herbal medicine prescription for protection of optic nerve. However, it remains unclear regarding the effects of Lycium barbarum polysaccharides, the main component of Lycium barbarum, on in vivo proliferation of adult ciliary body cells. In this study, adult rats were intragastrically administered low- and high-dose Lycium barbarum polysaccharides (1 and 10 mg/kg) for 35 days and those intragastrically administered phosphate buffered saline served as controls. The number of Ki-67-positive cells in rat ciliary body in the Lycium barbarum polysaccharides groups, in particular low-dose Lycium barbarum polysaccharides group, was significantly greater than that in the phosphate buffered saline group. Ki-67-positive rat ciliary body cells expressed nestin but they did not express glial fibrillary acidic protein. These findings suggest that Lycium barbarum polysaccharides can promote the proliferation of adult rat retinal progenitor cells and the proliferated cells present with neuronal phenotype. PMID:26889185

  10. Hydroxycitric acid does not promote inflammation or liver toxicity.

    PubMed

    Clouatre, Dallas L; Preuss, Harry G

    2013-11-28

    Garcinia cambogia extract (GC) with its active component consisting of hydroxycitric acid (HCA) is widely utilized for weight loss. Various HCA salts are available, including calcium, magnesium, potassium and mixtures of these. Experimentally, these salts exhibit different properties with some, but not all, improving glucose tolerance and blood pressure. Recently, obesity-prone C57BL/6J mice were fed a high-fat diet (HFD, 45 kcal% fat) with or without GC (1%, w/w) for 16 wk. The active arm reduced visceral fat, adipocyte size and serum glucose, yet purportedly also exhibited hepatic collagen accumulation, lipid peroxidation and increased mRNA levels of genes related to oxidative stress. The latter findings are at odds with a large body of animal and human studies that have been conducted on the safety and efficacy of HCA. This literature shows HCA to be protective against the liver toxicity associated with ethanol and dexamethasone administration, and to maintain serum aspartate aminotransferase, alanine aminotransferase and alkaline phosphatase at near normal levels. In both animal and clinical literature, elevated intakes of HCA per se have not led to signs of inflammation or hepatotoxicity. The compound has been found to reduce markers of inflammation in brain, intestines, kidney and serum. PMID:24307814

  11. Hydroxycitric acid does not promote inflammation or liver toxicity

    PubMed Central

    Clouatre, Dallas L; Preuss, Harry G

    2013-01-01

    Garcinia cambogia extract (GC) with its active component consisting of hydroxycitric acid (HCA) is widely utilized for weight loss. Various HCA salts are available, including calcium, magnesium, potassium and mixtures of these. Experimentally, these salts exhibit different properties with some, but not all, improving glucose tolerance and blood pressure. Recently, obesity-prone C57BL/6J mice were fed a high-fat diet (HFD, 45 kcal% fat) with or without GC (1%, w/w) for 16 wk. The active arm reduced visceral fat, adipocyte size and serum glucose, yet purportedly also exhibited hepatic collagen accumulation, lipid peroxidation and increased mRNA levels of genes related to oxidative stress. The latter findings are at odds with a large body of animal and human studies that have been conducted on the safety and efficacy of HCA. This literature shows HCA to be protective against the liver toxicity associated with ethanol and dexamethasone administration, and to maintain serum aspartate aminotransferase, alanine aminotransferase and alkaline phosphatase at near normal levels. In both animal and clinical literature, elevated intakes of HCA per se have not led to signs of inflammation or hepatotoxicity. The compound has been found to reduce markers of inflammation in brain, intestines, kidney and serum. PMID:24307814

  12. High folic acid increases cell turnover and lowers differentiation and iron content in human HT29 colon cancer cells.

    PubMed

    Pellis, Linette; Dommels, Yvonne; Venema, Dini; Polanen, Ab van; Lips, Esther; Baykus, Hakan; Kok, Frans; Kampman, Ellen; Keijer, Jaap

    2008-04-01

    Folate, a water-soluble B vitamin, is a cofactor in one-carbon metabolism and is essential for DNA synthesis, amino acid interconversion, methylation and, consequently, normal cell growth. In animals with existing pre-neoplastic and neoplastic lesions, folic acid supplementation increases the tumour burden. To identify processes that are affected by increased folic acid levels, we compared HT29 human colon cancer cells exposed to a chronic supplemental (100 ng/ml) level of folic acid to cells exposed to a normal (10 ng/ml) level of folic acid, in the presence of vitamin B12 and other micronutrients involved in the folate-methionine cycle. In addition to higher intracellular folate levels, HT29 cells at 100 ng folic acid/ml displayed faster growth and higher metabolic activity. cDNA microarray analysis indicated an effect on cell turnover and Fe metabolism. We fully confirmed these effects at the physiological level. At 100 ng/ml, cell assays showed higher proliferation and apoptosis, while gene expression analysis and a lower E-cadherin protein expression indicated decreased differentiation. These results are in agreement with the promoting effect of folic acid supplementation on established colorectal neoplasms. The lower expression of genes related to Fe metabolism at 100 ng folic acid/ml was confirmed by lower intracellular Fe levels in the cells exposed to folic acid at 100 ng/ml. This suggests an effect of folate on Fe metabolism.

  13. β-Catenin promotes colitis and colon cancer through imprinting of proinflammatory properties in T cells.

    PubMed

    Keerthivasan, Shilpa; Aghajani, Katayoun; Dose, Marei; Molinero, Luciana; Khan, Mohammad W; Venkateswaran, Vysak; Weber, Christopher; Emmanuel, Akinola Olumide; Sun, Tianjao; Bentrem, David J; Mulcahy, Mary; Keshavarzian, Ali; Ramos, Elena M; Blatner, Nichole; Khazaie, Khashayarsha; Gounari, Fotini

    2014-02-26

    The density and type of lymphocytes that infiltrate colon tumors are predictive of the clinical outcome of colon cancer. High densities of T helper 17 (T(H)17) cells and inflammation predict poor outcome, whereas infiltration by T regulatory cells (Tregs) that naturally suppress inflammation is associated with longer patient survival. However, the role of Tregs in cancer remains controversial. We recently reported that Tregs in colon cancer patients can become proinflammatory and tumor-promoting. These properties were directly linked with their expression of RORγt (retinoic acid-related orphan receptor-γt), the signature transcription factor of T(H)17 cells. We report that Wnt/β-catenin signaling in T cells promotes expression of RORγt. Expression of β-catenin was elevated in T cells, including Tregs, of patients with colon cancer. Genetically engineered activation of β-catenin in mouse T cells resulted in enhanced chromatin accessibility in the proximity of T cell factor-1 (Tcf-1) binding sites genome-wide, induced expression of T(H)17 signature genes including RORγt, and promoted T(H)17-mediated inflammation. Strikingly, the mice had inflammation of small intestine and colon and developed lesions indistinguishable from colitis-induced cancer. Activation of β-catenin only in Tregs was sufficient to produce inflammation and initiate cancer. On the basis of these findings, we conclude that activation of Wnt/β-catenin signaling in effector T cells and/or Tregs is causatively linked with the imprinting of proinflammatory properties and the promotion of colon cancer.

  14. β-Catenin promotes colitis and colon cancer through imprinting of proinflammatory properties in T cells.

    PubMed

    Keerthivasan, Shilpa; Aghajani, Katayoun; Dose, Marei; Molinero, Luciana; Khan, Mohammad W; Venkateswaran, Vysak; Weber, Christopher; Emmanuel, Akinola Olumide; Sun, Tianjao; Bentrem, David J; Mulcahy, Mary; Keshavarzian, Ali; Ramos, Elena M; Blatner, Nichole; Khazaie, Khashayarsha; Gounari, Fotini

    2014-02-26

    The density and type of lymphocytes that infiltrate colon tumors are predictive of the clinical outcome of colon cancer. High densities of T helper 17 (T(H)17) cells and inflammation predict poor outcome, whereas infiltration by T regulatory cells (Tregs) that naturally suppress inflammation is associated with longer patient survival. However, the role of Tregs in cancer remains controversial. We recently reported that Tregs in colon cancer patients can become proinflammatory and tumor-promoting. These properties were directly linked with their expression of RORγt (retinoic acid-related orphan receptor-γt), the signature transcription factor of T(H)17 cells. We report that Wnt/β-catenin signaling in T cells promotes expression of RORγt. Expression of β-catenin was elevated in T cells, including Tregs, of patients with colon cancer. Genetically engineered activation of β-catenin in mouse T cells resulted in enhanced chromatin accessibility in the proximity of T cell factor-1 (Tcf-1) binding sites genome-wide, induced expression of T(H)17 signature genes including RORγt, and promoted T(H)17-mediated inflammation. Strikingly, the mice had inflammation of small intestine and colon and developed lesions indistinguishable from colitis-induced cancer. Activation of β-catenin only in Tregs was sufficient to produce inflammation and initiate cancer. On the basis of these findings, we conclude that activation of Wnt/β-catenin signaling in effector T cells and/or Tregs is causatively linked with the imprinting of proinflammatory properties and the promotion of colon cancer. PMID:24574339

  15. Plasmacytoid dendritic cells sense skin injury and promote wound healing through type I interferons.

    PubMed

    Gregorio, Josh; Meller, Stephan; Conrad, Curdin; Di Nardo, Anna; Homey, Bernhard; Lauerma, Antti; Arai, Naoko; Gallo, Richard L; Digiovanni, John; Gilliet, Michel

    2010-12-20

    Plasmacytoid dendritic cells (pDCs) are specialized type I interferon (IFN-α/β)-producing cells that express intracellular toll-like receptor (TLR) 7 and TLR9 and recognize viral nucleic acids in the context of infections. We show that pDCs also have the ability to sense host-derived nucleic acids released in common skin wounds. pDCs were found to rapidly infiltrate both murine and human skin wounds and to transiently produce type I IFNs via TLR7- and TLR9-dependent recognition of nucleic acids. This process was critical for the induction of early inflammatory responses and reepithelization of injured skin. Cathelicidin peptides, which facilitate immune recognition of released nucleic acids by promoting their access to intracellular TLR compartments, were rapidly induced in skin wounds and were sufficient but not necessary to stimulate pDC activation and type I IFN production. These data uncover a new role of pDCs in sensing tissue damage and promoting wound repair at skin surfaces.

  16. Adipose-derived mesenchymal stem cells promote cell proliferation and invasion of epithelial ovarian cancer

    SciTech Connect

    Chu, Yijing; Tang, Huijuan; Guo, Yan; Guo, Jing; Huang, Bangxing; Fang, Fang; Cai, Jing Wang, Zehua

    2015-09-10

    Adipose-derived mesenchymal stem cell (ADSC) is an important component of tumor microenvironment. However, whether ADSCs have a hand in ovarian cancer progression remains unclear. In this study, we investigated the impact of human ADSCs derived from the omentum of normal donors on human epithelial ovarian cancer (EOC) cells in vitro and in vivo. Direct and indirect co-culture models including ADSCs and human EOC cell lines were established and the effects of ADSCs on EOC cell proliferation were evaluated by EdU incorporation and flow cytometry. Transwell migration assays and detection of MMPs were performed to assess the invasion activity of EOC cells in vitro. Mouse models were established by intraperitoneal injection of EOC cells with or without concomitant ADSCs to investigate the role of ADSCs in tumor progression in vivo. We found that ADSCs significantly promoted proliferation and invasion of EOC cells in both direct and indirect co-culture assays. In addition, after co-culture with ADSCs, EOC cells secreted higher levels of matrix metalloproteinases (MMPs), and inhibition of MMP2 and MMP9 partially relieved the tumor-promoting effects of ADSCs in vitro. In mouse xenograft models, we confirmed that ADSCs promoted EOC growth and metastasis and elevated the expression of MMP2 and MMP9. Our findings indicate that omental ADSCs play a promotive role during ovarian cancer progression. - Highlights: • Omental adipose derived stem cells enhanced growth and invasion properties of ovarian cancer cells. • Adipose derived stem cells promoted the growth and metastasis of ovarian cancer in mice models. • Adipose derived stem cells promoted MMPs expression and secretion of ovarian cancer cells. • Elevated MMPs mediated the tumor promoting effects of ADSCs.

  17. EBI2 augments Tfh cell fate by promoting interaction with IL-2-quenching dendritic cells.

    PubMed

    Li, Jianhua; Lu, Erick; Yi, Tangsheng; Cyster, Jason G

    2016-05-01

    T follicular helper (Tfh) cells are a subset of T cells carrying the CD4 antigen; they are important in supporting plasma cell and germinal centre responses. The initial induction of Tfh cell properties occurs within the first few days after activation by antigen recognition on dendritic cells, although how dendritic cells promote this cell-fate decision is not fully understood. Moreover, although Tfh cells are uniquely defined by expression of the follicle-homing receptor CXCR5 (refs 1, 2), the guidance receptor promoting the earlier localization of activated T cells at the interface of the B-cell follicle and T zone has been unclear. Here we show that the G-protein-coupled receptor EBI2 (GPR183) and its ligand 7α,25-dihydroxycholesterol mediate positioning of activated CD4 T cells at the interface of the follicle and T zone. In this location they interact with activated dendritic cells and are exposed to Tfh-cell-promoting inducible co-stimulator (ICOS) ligand. Interleukin-2 (IL-2) is a cytokine that has multiple influences on T-cell fate, including negative regulation of Tfh cell differentiation. We demonstrate that activated dendritic cells in the outer T zone further augment Tfh cell differentiation by producing membrane and soluble forms of CD25, the IL-2 receptor α-chain, and quenching T-cell-derived IL-2. Mice lacking EBI2 in T cells or CD25 in dendritic cells have reduced Tfh cells and mount defective T-cell-dependent plasma cell and germinal centre responses. These findings demonstrate that distinct niches within the lymphoid organ T zone support distinct cell fate decisions, and they establish a function for dendritic-cell-derived CD25 in controlling IL-2 availability and T-cell differentiation.

  18. Probiotics promote endocytic allergen degradation in gut epithelial cells

    SciTech Connect

    Song, Chun-Hua; Liu, Zhi-Qiang; Huang, Shelly; Zheng, Peng-Yuan; Yang, Ping-Chang

    2012-09-14

    Highlights: Black-Right-Pointing-Pointer Knockdown of A20 compromised the epithelial barrier function. Black-Right-Pointing-Pointer The fusion of endosome/lysosome was disturbed in the A20-deficient HT-29 cells. Black-Right-Pointing-Pointer Antigens transported across A20-deficient HT-29 monolayers conserved antigenicity. Black-Right-Pointing-Pointer Probiotic proteins increased the expression of A20 in HT-29 cells. -- Abstract: Background and aims: Epithelial barrier dysfunction plays a critical role in the pathogenesis of allergic diseases; the mechanism is to be further understood. The ubiquitin E3 ligase A20 (A20) plays a role in the endocytic protein degradation in the cells. This study aims to elucidate the role of A20 in the maintenance of gut epithelial barrier function. Methods: Gut epithelial cell line, HT-29 cell, was cultured into monolayers to evaluate the barrier function in transwells. RNA interference was employed to knock down the A20 gene in HT-29 cells to test the role of A20 in the maintenance of epithelial barrier function. Probiotic derived proteins were extracted from the culture supernatants using to enhance the expression of A20 in HT-29 cells. Results: The results showed that the knockdown of A20 compromised the epithelial barrier function in HT-29 monolayers, mainly increased the intracellular permeability. The fusion of endosome/lysosome was disturbed in the A20-deficient HT-29 cells. Allergens collected from the transwell basal chambers of A20-deficient HT-29 monolayers still conserved functional antigenicity. Treating with probiotic derived proteins increased the expression of A20 in HT-29 cells and promote the barrier function. Conclusion: A20 plays an important role in the maintenance of epithelial barrier function as shown by HT-29 monolayer. Probiotic derived protein increases the expression of A20 and promote the HT-29 monolayer barrier function.

  19. Daucosterol promotes the proliferation of neural stem cells.

    PubMed

    Jiang, Li-hua; Yang, Nian-yun; Yuan, Xiao-lin; Zou, Yi-jie; Zhao, Feng-ming; Chen, Jian-ping; Wang, Ming-yan; Lu, Da-xiang

    2014-03-01

    Neural stem cells (NSCs) are self-regenerating cells, but their regenerative capacity is limited. The present study was conducted to investigate the effect of daucosterol (a sterolin) on the promotion of NSC proliferation and determine the corresponding molecular mechanism. Results of cell counting kit-8 (CCK-8) assay showed that daucosterol significantly increased the quantity of viable cells and the effectiveness of daucosterol was similar to that of basic fibroblast growth factor (bFGF) and epidermal growth factor (EGF). Flow cytometry detection of CFSE-labeled (CFSE, carboxyfluorescein diacetate succinimidyl ester) NSCs showed that Div Index (or the average number of cell divisions) and % Divided (or the percentage of cells that divided at least once) of the cells were increased, indicating that daucosterol increased the percentage of NSCs re-entering the cell cycle. mRNA microarray analysis showed that 333 genes that are mostly involved in the mitotic cell cycle were up-regulated. By contrast, 627 genes that are mostly involved in differentiation were down-regulated. In particular, insulin-like growth factor I (IGF1) was considered as an important regulatory gene that functionally promoted NSC proliferation, and the increased expression of IGF1 protein was validated by ELISA. In addition, the phosphorylation of AKT was increased, indicating that the proliferation-enhancing activity of daucosterol may be involved in IGF1-AKT pathway. Our study provided information about daucosterol as an efficient and inexpensive growth factor alternative that could be used in clinical medicine and research applications. PMID:24333794

  20. Glial fibrillary acidic protein promoters direct adenovirus early 1A gene and human telomerase reverse transcriptase promoters direct sodium iodide symporter expression for malignant glioma radioiodine therapy.

    PubMed

    Li, Wei; Tan, Jian; Wang, Peng; Li, Ning; Li, Chengxia

    2015-01-01

    Malignant glioma can be treated with radioiodine following transfection with human sodium iodide symporter (hNIS) gene. Ad-Tp-E1A-Gp-NIS is engineered with human telomerase reverse transcriptase (hTERT) and glial fibrillary acidic protein (GFAP) promoters to express early region 1A (E1A) and hNIS genes, which may be useful in targeted gene therapy. The Ad-Tp-E1A-Gp-NIS was constructed and purified using the E1A and hNIS genes regulated by the hTERT and GFAP promoters, respectively. Glioma cells were infected by Ad-Tp-E1A-Gp-NIS. Selective replication ability of Ad-Tp-E1A-Gp-NIS was then evaluated by plaque forming assay, transgene expression by Western blot, (125)I-iodide uptake and efflux, clonogenicity following (131)I-iodide treatment in the tumor cells, and radioiodine therapy using nude mouse model. The Ad-Tp-E1A-Gp-NIS could selectively replicate; the hNIS gene was successfully expressed under the GFAP promoter. Western blot analyses using E1A- and hNIS-specific antibodies revealed two bands of approximately 40 and 70 kDa. In addition, the cells showed about 93.4 and 107.1 times higher (125)I uptake in U251 and U87 cells than in the control cells, respectively. Clonogenic assay indicated that >90% of cells transfected with Ad-Tp-E1A-Gp-NIS were killed. The Ad-Tp-E1A-Gp-NIS-transfected and 2 mCi (131)I-injected U87 xenograft nude mice survived the longest among the three groups. Ad-Tp-E1A-Gp-NIS has a good ability of selective replication and strong antitumor selectivity. An effective therapy of (131)I was achieved activity in malignant glioma cells after induction of tumor-specific iodide uptake activity by GFAP promoter-directed hNIS gene expression in vitro and in vivo.

  1. Medium-chain fatty acids inhibit mitochondrial metabolism in astrocytes promoting astrocyte-neuron lactate and ketone body shuttle systems.

    PubMed

    Thevenet, Jonathan; De Marchi, Umberto; Domingo, Jaime Santo; Christinat, Nicolas; Bultot, Laurent; Lefebvre, Gregory; Sakamoto, Kei; Descombes, Patrick; Masoodi, Mojgan; Wiederkehr, Andreas

    2016-05-01

    Medium-chain triglycerides have been used as part of a ketogenic diet effective in reducing epileptic episodes. The health benefits of the derived medium-chain fatty acids (MCFAs) are thought to result from the stimulation of liver ketogenesis providing fuel for the brain. We tested whether MCFAs have direct effects on energy metabolism in induced pluripotent stem cell-derived human astrocytes and neurons. Using single-cell imaging, we observed an acute pronounced reduction of the mitochondrial electrical potential and a concomitant drop of the NAD(P)H signal in astrocytes, but not in neurons. Despite the observed effects on mitochondrial function, MCFAs did not lower intracellular ATP levels or activate the energy sensor AMP-activated protein kinase. ATP concentrations in astrocytes were unaltered, even when blocking the respiratory chain, suggesting compensation through accelerated glycolysis. The MCFA decanoic acid (300 μM) promoted glycolysis and augmented lactate formation by 49.6%. The shorter fatty acid octanoic acid (300 μM) did not affect glycolysis but increased the rates of astrocyte ketogenesis 2.17-fold compared with that of control cells. MCFAs may have brain health benefits through the modulation of astrocyte metabolism leading to activation of shuttle systems that provide fuel to neighboring neurons in the form of lactate and ketone bodies.-Thevenet, J., De Marchi, U., Santo Domingo, J., Christinat, N., Bultot, L., Lefebvre, G., Sakamoto, K., Descombes, P., Masoodi, M., Wiederkehr, A. Medium-chain fatty acids inhibit mitochondrial metabolism in astrocytes promoting astrocyte-neuron lactate and ketone body shuttle systems.

  2. Medium-chain fatty acids inhibit mitochondrial metabolism in astrocytes promoting astrocyte-neuron lactate and ketone body shuttle systems.

    PubMed

    Thevenet, Jonathan; De Marchi, Umberto; Domingo, Jaime Santo; Christinat, Nicolas; Bultot, Laurent; Lefebvre, Gregory; Sakamoto, Kei; Descombes, Patrick; Masoodi, Mojgan; Wiederkehr, Andreas

    2016-05-01

    Medium-chain triglycerides have been used as part of a ketogenic diet effective in reducing epileptic episodes. The health benefits of the derived medium-chain fatty acids (MCFAs) are thought to result from the stimulation of liver ketogenesis providing fuel for the brain. We tested whether MCFAs have direct effects on energy metabolism in induced pluripotent stem cell-derived human astrocytes and neurons. Using single-cell imaging, we observed an acute pronounced reduction of the mitochondrial electrical potential and a concomitant drop of the NAD(P)H signal in astrocytes, but not in neurons. Despite the observed effects on mitochondrial function, MCFAs did not lower intracellular ATP levels or activate the energy sensor AMP-activated protein kinase. ATP concentrations in astrocytes were unaltered, even when blocking the respiratory chain, suggesting compensation through accelerated glycolysis. The MCFA decanoic acid (300 μM) promoted glycolysis and augmented lactate formation by 49.6%. The shorter fatty acid octanoic acid (300 μM) did not affect glycolysis but increased the rates of astrocyte ketogenesis 2.17-fold compared with that of control cells. MCFAs may have brain health benefits through the modulation of astrocyte metabolism leading to activation of shuttle systems that provide fuel to neighboring neurons in the form of lactate and ketone bodies.-Thevenet, J., De Marchi, U., Santo Domingo, J., Christinat, N., Bultot, L., Lefebvre, G., Sakamoto, K., Descombes, P., Masoodi, M., Wiederkehr, A. Medium-chain fatty acids inhibit mitochondrial metabolism in astrocytes promoting astrocyte-neuron lactate and ketone body shuttle systems. PMID:26839375

  3. Binding sequence-dependent regulation of the human proliferating cell nuclear antigen promoter by p53

    SciTech Connect

    Shan Bin . E-mail: gmorris2@tulane.edu

    2005-04-15

    Exposure of a lung epithelial cell line to ionizing radiation (IR) arrests cell cycle progression through 48 h post-exposure. Coincidentally, IR differentially activates expression of the cell cycle inhibitor, p21/WAF1, and the DNA replication protein, proliferating cell nuclear antigen (PCNA). p21/WAF1 mRNA levels remain elevated through 48 h post-exposure to IR, while PCNA mRNA levels increase transiently at early times. Since p21/WAF1 inhibits DNA replication by directly binding PCNA, the relative levels of the two proteins can determine cell cycle progression. The PCNA p53-binding site displayed reduced p53 binding affinity in vitro relative to the distal p21/WAF1 p53-binding site. Substitution of the p21/WAF1 site for the resident p53-binding site in the PCNA promoter altered the responses to increasing amounts of p53 or IR in transient expression assays. The p21/WAF1 p53-binding site sustained activation of the chimeric PCNA promoter under conditions (high p53 levels or high dose IR) that the PCNA p53-binding site did not. Binding site-specific regulation by wild-type p53 was not observed with mutant p53 harboring a serine to alanine change at amino acid 46. Limited activation of the PCNA promoter by p53 and its modified forms would restrict the amount of PCNA made available for DNA repair.

  4. Acetylsalicylic acid induces programmed cell death in Arabidopsis cell cultures.

    PubMed

    García-Heredia, José M; Hervás, Manuel; De la Rosa, Miguel A; Navarro, José A

    2008-06-01

    Acetylsalicylic acid (ASA), a derivative from the plant hormone salicylic acid (SA), is a commonly used drug that has a dual role in animal organisms as an anti-inflammatory and anticancer agent. It acts as an inhibitor of cyclooxygenases (COXs), which catalyze prostaglandins production. It is known that ASA serves as an apoptotic agent on cancer cells through the inhibition of the COX-2 enzyme. Here, we provide evidences that ASA also behaves as an agent inducing programmed cell death (PCD) in cell cultures of the model plant Arabidopsis thaliana, in a similar way than the well-established PCD-inducing agent H(2)O(2), although the induction of PCD by ASA requires much lower inducer concentrations. Moreover, ASA is herein shown to be a more efficient PCD-inducing agent than salicylic acid. ASA treatment of Arabidopsis cells induces typical PCD-linked morphological and biochemical changes, namely cell shrinkage, nuclear DNA degradation, loss of mitochondrial membrane potential, cytochrome c release from mitochondria and induction of caspase-like activity. However, the ASA effect can be partially reverted by jasmonic acid. Taking together, these results reveal the existence of common features in ASA-induced animal apoptosis and plant PCD, and also suggest that there are similarities between the pathways of synthesis and function of prostanoid-like lipid mediators in animal and plant organisms.

  5. Polysialic Acid Directs Tumor Cell Growth by Controlling Heterophilic Neural Cell Adhesion Molecule Interactions

    PubMed Central

    Seidenfaden, Ralph; Krauter, Andrea; Schertzinger, Frank; Gerardy-Schahn, Rita; Hildebrandt, Herbert

    2003-01-01

    Polysialic acid (PSA), a carbohydrate polymer attached to the neural cell adhesion molecule (NCAM), promotes neural plasticity and tumor malignancy, but its mode of action is controversial. Here we establish that PSA controls tumor cell growth and differentiation by interfering with NCAM signaling at cell-cell contacts. Interactions between cells with different PSA and NCAM expression profiles were initiated by enzymatic removal of PSA and by ectopic expression of NCAM or PSA-NCAM. Removal of PSA from the cell surface led to reduced proliferation and activated extracellular signal-regulated kinase (ERK), inducing enhanced survival and neuronal differentiation of neuroblastoma cells. Blocking with an NCAM-specific peptide prevented these effects. Combinatorial transinteraction studies with cells and membranes with different PSA and NCAM phenotypes revealed that heterophilic NCAM binding mimics the cellular responses to PSA removal. In conclusion, our data demonstrate that PSA masks heterophilic NCAM signals, having a direct impact on tumor cell growth. This provides a mechanism for how PSA may promote the genesis and progression of highly aggressive PSA-NCAM-positive tumors. PMID:12897159

  6. Estrogen promotes Leydig cell engulfment by macrophages in male infertility.

    PubMed

    Yu, Wanpeng; Zheng, Han; Lin, Wei; Tajima, Astushi; Zhang, Yong; Zhang, Xiaoyan; Zhang, Hongwen; Wu, Jihua; Han, Daishu; Rahman, Nafis A; Korach, Kenneth S; Gao, George Fu; Inoue, Ituro; Li, Xiangdong

    2014-06-01

    Male infertility accounts for almost half of infertility cases worldwide. A subset of infertile men exhibit reduced testosterone and enhanced levels of estradiol (E2), though it is unclear how increased E2 promotes deterioration of male fertility. Here, we utilized a transgenic mouse strain that overexpresses human CYP19, which encodes aromatase (AROM+ mice), and mice with knockout of Esr1, encoding estrogen receptor α (ERαKO mice), to analyze interactions between viable Leydig cells (LCs) and testicular macrophages that may lead to male infertility. In AROM+ males, enhanced E2 promoted LC hyperplasia and macrophage activation via ERα signaling. E2 stimulated LCs to produce growth arrest-specific 6 (GAS6), which mediates phagocytosis of apoptotic cells by bridging cells with surface exposed phosphatidylserine (PS) to macrophage receptors, including the tyrosine kinases TYRO3, AXL, and MER. Overproduction of E2 increased apoptosis-independent extrusion of PS on LCs, which in turn promoted engulfment by E2/ERα-activated macrophages that was mediated by AXL-GAS6-PS interaction. We further confirmed E2-dependant engulfment of LCs by real-time 3D imaging. Furthermore, evaluation of molecular markers in the testes of patients with nonobstructive azoospermia (NOA) revealed enhanced expression of CYP19, GAS6, and AXL, which suggests that the AROM+ mouse model reflects human infertility. Together, these results suggest that GAS6 has a potential as a clinical biomarker and therapeutic target for male infertility.

  7. Epigenetic modulators promote mesenchymal stem cell phenotype switches.

    PubMed

    Alexanian, Arshak R

    2015-07-01

    Discoveries in recent years have suggested that some tissue specific adult stem cells in mammals might have the ability to differentiate into cell types from different germ layers. This phenomenon has been referred to as stem cell transdifferentiation or plasticity. Despite controversy, the current consensus holds that transdifferentiation does occur in mammals, but only within a limited range. Understanding the mechanisms that underlie the switches in phenotype and development of the methods that will promote such type of conversions can open up endless possibilities for regenerative medicine. Epigenetic control contributes to various processes that lead to cellular plasticity and DNA and histone covalent modifications play a key role in these processes. Recently, we have been able to convert human mesenchymal stem cells (hMSCs) into neural-like cells by exposing cells to epigenetic modifiers and neural inducing factors. The goal of this study was to investigate the stability and plasticity of these transdifferentiated cells. To this end, neurally induced MSCs (NI-hMSCs) were exposed to adipocyte inducing factors. Grown for 24-48 h in fat induction media NI-hMSCs reversed their morphology into fibroblast-like cells and regained their proliferative properties. After 3 weeks approximately 6% of hMSCs differentiated into multilocular or plurivacuolar adipocyte cells that demonstrated by Oil Red O staining. Re-exposure of these cultures or the purified adipocytes to neural induction medium induced the cells to re-differentiate into neuronal-like cells. These data suggest that cell plasticity can be manipulated by the combination of small molecule modulators of chromatin modifying enzymes and specific cell signaling pathways.

  8. Epigenetic modulators promote mesenchymal stem cell phenotype switches.

    PubMed

    Alexanian, Arshak R

    2015-07-01

    Discoveries in recent years have suggested that some tissue specific adult stem cells in mammals might have the ability to differentiate into cell types from different germ layers. This phenomenon has been referred to as stem cell transdifferentiation or plasticity. Despite controversy, the current consensus holds that transdifferentiation does occur in mammals, but only within a limited range. Understanding the mechanisms that underlie the switches in phenotype and development of the methods that will promote such type of conversions can open up endless possibilities for regenerative medicine. Epigenetic control contributes to various processes that lead to cellular plasticity and DNA and histone covalent modifications play a key role in these processes. Recently, we have been able to convert human mesenchymal stem cells (hMSCs) into neural-like cells by exposing cells to epigenetic modifiers and neural inducing factors. The goal of this study was to investigate the stability and plasticity of these transdifferentiated cells. To this end, neurally induced MSCs (NI-hMSCs) were exposed to adipocyte inducing factors. Grown for 24-48 h in fat induction media NI-hMSCs reversed their morphology into fibroblast-like cells and regained their proliferative properties. After 3 weeks approximately 6% of hMSCs differentiated into multilocular or plurivacuolar adipocyte cells that demonstrated by Oil Red O staining. Re-exposure of these cultures or the purified adipocytes to neural induction medium induced the cells to re-differentiate into neuronal-like cells. These data suggest that cell plasticity can be manipulated by the combination of small molecule modulators of chromatin modifying enzymes and specific cell signaling pathways. PMID:25936755

  9. Fabp3 inhibits proliferation and promotes apoptosis of embryonic myocardial cells.

    PubMed

    Zhu, C; Hu, D L; Liu, Y Q; Zhang, Q J; Chen, F K; Kong, X Q; Cao, K J; Zhang, J S; Qian, L M

    2011-07-01

    Fatty acid binding protein 3 (FABP3) is a member of a family of binding proteins. The protein is mainly expressed in cardiac and skeletal muscle cells, and it has been linked to fatty acid metabolism, trafficking, and signaling. Using suppression subtractive hybridization, we previously found that FABP3 is highly regulated in ventricular septal defect (VSD) patients and may play a significant role in the development of human VSD. We therefore aimed to identify the biological characteristics of the FABP3 gene in embryonic myocardial cells. On the basis of RT-PCR and western blotting analyses, we demonstrated that the expression levels of FABP3 mRNA and protein were up-regulated initially and then gradually decreased with P19 cell differentiation. MTT assays and cell cycle analysis showed that FABP3 inhibits P19 cell proliferation, and data from annexin V-FITC assays revealed that FABP3 can promote apoptosis of P19 cells. Further data from quantitative real-time RT-PCR revealed lower expression levels of cardiac muscle-specific molecular markers (cTnT, alpha-MHC, GATA4, and MEF2c) in FABP3-overexpressing cell lines than in the control cells during differentiation. Our results demonstrate that FABP3 may be involved in the differentiation of cardiac myocytes. PMID:21293949

  10. The endocannabinoid system promotes astroglial differentiation by acting on neural progenitor cells.

    PubMed

    Aguado, Tania; Palazuelos, Javier; Monory, Krisztina; Stella, Nephi; Cravatt, Benjamin; Lutz, Beat; Marsicano, Giovanni; Kokaia, Zaal; Guzmán, Manuel; Galve-Roperh, Ismael

    2006-02-01

    Endocannabinoids exert an important neuromodulatory role via presynaptic cannabinoid CB1 receptors and may also participate in the control of neural cell death and survival. The function of the endocannabinoid system has been extensively studied in differentiated neurons, but its potential role in neural progenitor cells remains to be elucidated. Here we show that the CB1 receptor and the endocannabinoid-inactivating enzyme fatty acid amide hydrolase are expressed, both in vitro and in vivo, in postnatal radial glia (RC2+ cells) and in adult nestin type I (nestin(+)GFAP+) neural progenitor cells. Cell culture experiments show that CB1 receptor activation increases progenitor proliferation and differentiation into astroglial cells in vitro. In vivo analysis evidences that, in postnatal CB1(-/-) mouse brain, progenitor proliferation and astrogliogenesis are impaired. Likewise, in adult CB1-deficient mice, neural progenitor proliferation is decreased but is increased in fatty acid amide hydrolase-deficient mice. In addition, endocannabinoid signaling controls neural progenitor differentiation in the adult brain by promoting astroglial differentiation of newly born cells. These results show a novel physiological role of endocannabinoids, which constitute a new family of signaling cues involved in the regulation of neural progenitor cell function.

  11. Obesity promotes aerobic glycolysis in prostate cancer cells.

    PubMed

    Cavazos, David A; deGraffenried, Matthew J; Apte, Shruti A; Bowers, Laura W; Whelan, Kaitlin A; deGraffenried, Linda A

    2014-01-01

    Obesity is the leading preventable comorbidity associated with increased prostate cancer-related recurrence and mortality. Epidemiological and clinical studies indicate that a body mass index >30 is associated with increased oxidative DNA damage within the prostate gland and increased prostate cancer-related mortality. Here we provide evidence that obesity promotes worse clinical outcome through induction of metabolic abnormalities known to promote genotoxic stress. We have previously reported that blood serum derived from obese mice may enhance the proliferative and invasive potential of human prostate cancer cell lines ex vivo. Here we show that a 1-h exposure of LNCaP or PacMetUT1 prostate cancer cell lines and nonmalignant RWPE-1 prostate epithelial cells to 2% serum from obese mice induces markers of aerobic glycolysis relative to those exposed to serum from nonobese mice. This metabolic change was correlated with accumulation of reactive oxygen species (ROS) and increased frequency of DNA double-strand breaks. Interestingly, N-tert-Butylhydroxylamine, an antioxidant, significantly suppressed markers of aerobic glycolysis in the cells exposed to the blood serum of obese mice, suggesting that ROS contributes to a metabolic shift toward aerobic glycolysis. Here we describe obesity-induced changes in key metabolic markers that impact prostate cancer cell progression and explore the role of antioxidants in ameliorating these effects. PMID:25264717

  12. Decreased Ferroportin Promotes Myeloma Cell Growth and Osteoclast Differentiation

    PubMed Central

    Gu, Zhimin; Wang, He; Xia, Jiliang; Yang, Ye; Jin, Zhendong; Xu, Hongwei; Shi, Jumei; De Domenico, Ivana; Tricot, Guido; Zhan, Fenghuang

    2016-01-01

    Iron homeostasis is disrupted in multiple myeloma, a difficult-to-cure plasma cell malignancy with lytic bone lesions. Here, we systematically analyzed iron gene expression signature and demonstrated that mRNA expression of iron exporter ferroportin (FPN1) is significantly downregulated in myeloma cells and correlates negatively with clinic outcome. Restoring expression of FPN1 reduces intracellular liable iron pool, inhibits STAT3-MCL-1 signaling, and suppresses myeloma cells growth. Furthermore, we demonstrated that mRNA of FPN1 is also downregulated at the initial stages of osteoclast differentiation and suppresses myeloma cell–induced osteoclast differentiation through regulating iron regulator TFRC, NF-κB, and JNK pathways. Altogether, we demonstrated that downregulation of FPN1 plays critical roles in promoting myeloma cell growth and bone resorption in multiple myeloma. PMID:25855377

  13. Relative efficacy of organic acids and antibiotics as growth promoters in broiler chicken

    PubMed Central

    Bagal, Vikrant Laxman; Khatta, Vinod Kumar; Tewatia, Bachu Singh; Sangwan, Sandeep Kumar; Raut, Subhash Shamrao

    2016-01-01

    Aim: The objective of this study was to evaluate the effect of organic acids as replacer to antibiotics in their various combinations on feed consumption, body weight gain, and feed conversion ratio (FCR) in broiler chicks during different phases of growth. Materials and Methods: Antibiotics and organic acids were incorporated into boiler feed in different combinations to form 10 maize based test diets (T1 to T10). Each test diet was offered to four replicates of 10 birds each constituting a total of 400 birds kept for 45 days. Results: Significantly better effect in terms of body weight gain from supplementation of 1% citric acid and 1% citric acid along with antibiotic was observed throughout the entire study, whereas the effect of tartaric acid supplementation was similar to control group. Citric acid (1%) along with antibiotic supplementation showed highest feed intake during the experimental period. Significantly better FCR was observed in groups supplemented with 1% citric acid and 1% citric acid along with antibiotic followed by antibiotic along with organic acids supplemented group. Conclusion: Growth performance of birds in terms of body weight, body weight gain, and FCR improved significantly in 1% citric acid which was significantly higher than antibiotic supplemented group. 1% citric acid can effectively replace antibiotic growth promoter (chlortetracycline) without affecting growth performance of birds. PMID:27182133

  14. Stabilizing platinum in phosphoric acid fuel cells

    NASA Technical Reports Server (NTRS)

    Remick, R. J.

    1981-01-01

    The cathode of the phosphoric acid fuel cell uses a high surface area platinum catalyst supported on a carbon substrate. During operation, the small platinum crystallites sinter, causing loss in cell performance. A support was developed that stabilizes platinum in the high surface area condition by retarding or preventing the sintering process. The approach is to form etch pits in the carbon by oxidizing the carbon in the presence of a metal oxide catalyst, remove the metal oxide by an acid wash, and then deposit platinum in these pits. Results confirm the formation of etch pits in each of the three supports chosen for investigation: Vulcan XC-72R, Vulcan XC-72 that was graphized at 2500 C, and Shawinigan Acetylene Black.

  15. Okadaic acid: An additional non-phorbol-12-tetradecanoate-13-acetate-type tumor promoter

    SciTech Connect

    Suganuma, Masami; Fujiki, Hirota; Suguri, Hiroko; Yoshizawa, Shigeru; Hirota, Mitsuru; Nakayasu, Michie ); Ojika, Makoto; Wakamatsu, Kazumasa; Yamada, Kiyoyuki ); Sugimura, Takashi )

    1988-03-01

    Okadaic acid is a polyether compound of a C{sub 38} fatty acid, isolated from a black sponge, Halichondria okadai. Previous studies showed that okadaic acid is a skin irritant and induces ornithine decarboxylase in mouse skin 4 hr after its application to the skin. This induction was strongly inhibited by pretreatment of the skin with 13-cis-retinoic acid. A two-stage carcinogenesis experiment in mouse skin initiated by a single application of 100 {mu}g of 7,12-dimethylbenz(a)anthracene (DMBA) and followed by application of 10 {mu}g of okadaic acid twice a week revealed that okadaic acid is a potent additional tumor promoter: tumors developed in 93% of the mice treated with DMBA and okadaic acid by week 16. In contrast, tumors were found in only one mouse each in the groups treated with DMBA alone or okadaic acid alone. An average of 2.6 tumors per mouse was found in week 30 in the group treated with DMBA and okadaic acid. Unlike phorbol 12-tetradecanoate 13-acetate (TPA), teleocidin, and aplysiatoxin, okadaic acid did not inhibit the specific binding of ({sup 3}H)TPA to a mouse skin particulate fraction when added up to 100 {mu}M or activate calcium-activated, phospholipid-dependent protein kinase (protein kinase C) in vitro when added up to 1.2 {mu}M. Therefore, the actions of okadaic acid and phorbol ester may be mediated in different ways. These results show that okadaic acid is a non-TPA-type tumor promoter in mouse skin carcinogenesis.

  16. Dynamic mast cell-stromal cell interactions promote growth of pancreatic cancer

    PubMed Central

    Ma, Ying; Hwang, Rosa F.; Logsdon, Craig D.; Ullrich, Stephen E.

    2013-01-01

    Pancreatic ductal adenocarcinoma (PDAC) exists in a complex desmoplastic microenvironment, which includes cancer-associated fibroblasts (also known as pancreatic stellate cells, PSCs) and immune cells that provide a fibrotic niche that impedes successful cancer therapy. We have found that mast cells are essential for PDAC tumorigenesis. Whether mast cells contribute to the growth of PDAC and/or PSCs is unknown. Here we tested the hypothesis that mast cells contribute to the growth of PSCs and tumor cells, thus contributing to PDAC development. Tumor cells promoted mast cell migration. Both tumor cells and PSCs stimulated mast cell activation. Conversely, mast cell-derived IL-13 and tryptase stimulated PSC proliferation. Treating tumor-bearing mice with agents that block mast cell migration and function depressed PDAC growth. Our findings suggest that mast cells exacerbate the cellular and extracellular dynamics of the tumor microenvironment found in PDAC. Therefore, targeting mast cells may inhibit stromal formation and improve therapy. PMID:23633481

  17. Transgelin promotes migration and invasion of cancer stem cells.

    PubMed

    Lee, Eun-Kyung; Han, Gi-Yeon; Park, Hye Won; Song, Yeo-Ju; Kim, Chan-Wha

    2010-10-01

    Recent studies have suggested the existence of a small subset of cancer cells called cancer stem cells (CSCs), which possess the ability to initiate malignancies, promote tumor formation, drive metastasis, and evade conventional chemotherapies. Elucidation of the specific signaling pathway and mechanism underlying the action of CSCs might improve the efficacy of cancer treatments. In this study, we analyzed differentially expressed proteins between tumerigenic and nontumorigenic cells isolated from the human hepatocellular carcinoma (HCC) cell line, Huh7, via proteomic analysis to identify proteins correlated with specific features of CSCs. The expression level of Transgelin was 25-fold higher in tumorigenic cells than nontumorigenic cells. Similar results were also observed in tumorigenic cells derived from colorectal adenocarcinoma and prostate carcinoma. More importantly, the elevated levels of Transgelin significantly increased the invasiveness of tumorigenic cells, whereas reduced levels decreased the invasive potential. Moreover, in tumors derived from Huh7-induced xenografts, Transgelin was also co-expressed with CXCR4, which is responsible for tumor invasion. Taken together, these results indicate that the metastatic potential of CSCs arises from highly expressed Transgelin.

  18. Nolz1 promotes striatal neurogenesis through the regulation of retinoic acid signaling

    PubMed Central

    2010-01-01

    Background Nolz1 is a zinc finger transcription factor whose expression is enriched in the lateral ganglionic eminence (LGE), although its function is still unknown. Results Here we analyze the role of Nolz1 during LGE development. We show that Nolz1 expression is high in proliferating neural progenitor cells (NPCs) of the LGE subventricular zone. In addition, low levels of Nolz1 are detected in the mantle zone, as well as in the adult striatum. Similarly, Nolz1 is highly expressed in proliferating LGE-derived NPC cultures, but its levels rapidly decrease upon cell differentiation, pointing to a role of Nolz1 in the control of NPC proliferation and/or differentiation. In agreement with this hypothesis, we find that Nolz1 over-expression promotes cell cycle exit of NPCs in neurosphere cultures and negatively regulates proliferation in telencephalic organotypic cultures. Within LGE primary cultures, Nolz1 over-expression promotes the acquisition of a neuronal phenotype, since it increases the number of β-III tubulin (Tuj1)- and microtubule-associated protein (MAP)2-positive neurons, and inhibits astrocyte generation and/or differentiation. Retinoic acid (RA) is one of the most important morphogens involved in striatal neurogenesis, and regulates Nolz1 expression in different systems. Here we show that Nolz1 also responds to this morphogen in E12.5 LGE-derived cell cultures. However, Nolz1 expression is not regulated by RA in E14.5 LGE-derived cell cultures, nor is it affected during LGE development in mouse models that present decreased RA levels. Interestingly, we find that Gsx2, which is necessary for normal RA signaling during LGE development, is also required for Nolz1 expression, which is lost in Gsx2 knockout mice. These findings suggest that Nolz1 might act downstream of Gsx2 to regulate RA-induced neurogenesis. Keeping with this hypothesis, we show that Nolz1 induces the selective expression of the RA receptor (RAR)β without altering RARα or RARγ. In

  19. Expression of Beta-Defensin 131 Promotes an Innate Immune Response in Human Prostate Epithelial Cells

    PubMed Central

    Kim, Hae Jong; Lee, Jaehyouk; Myung, Soon Chul

    2015-01-01

    Previously, using the Illumina HumanHT-12 microarray we found that β-defensin 131 (DEFB131), an antimicrobial peptide, is upregulated in the human prostate epithelial cell line RWPE-1 upon stimulation with lipoteichoic acid (LTA; a gram-positive bacterial component), than that in the untreated RWPE-1 cells. In the current study, we aimed to investigate the role of DEFB131 in RWPE-1 cells during bacterial infection. We examined the intracellular signaling pathways and nuclear responses in RWPE-1 cells that contribute to DEFB131 gene induction upon stimulation with LTA. Chromatin immunoprecipitation was performed to determine whether NF-κB directly binds to the DEFB131 promoter after LTA stimulation in RWPE-1 cells. We found that DEFB131 expression was induced by LTA stimulation through TLR2 and p38MAPK/NF-κB activation, which was evident in the phosphorylation of both p38MAPK and IκBα. We also found that SB203580 and Bay11-7082, inhibitors of p38MAPK and NF-κB, respectively, suppressed LTA-induced DEFB131 expression. The chromatin immunoprecipitation assay showed that NF-κB directly binds to the DEFB131 promoter, suggesting that NF-κB is a direct regulator, and is necessary for LTA-induced DEFB131 expression in RWPE-1 cells. Interestingly, with DEFB131 overexpression in RWPE-1 cells, the accumulation of mRNA and protein secretion of cytokines (IL-1α, IL-1β, IL-6, and IL-12α) and chemokines (CCL20, CCL22, and CXCL8) were significantly enhanced. In addition, DEFB131-transfected RWPE-1 cells markedly induced chemotactic activity in THP-1 monocytes. We concluded that DEFB131 induces cytokine and chemokine upregulation through the TLR2/NF-κB signaling pathway in RWPE-1 cells during bacterial infection and promotes an innate immune response. PMID:26649771

  20. Oxygen-18 study of the mechanism of promoter action of thiocyanate ions in the electrosynthesis of persulfuric acid and ammonium persulfate at platinum anodes

    SciTech Connect

    Kasatkin, E.V.; Larchenko, L.I.; Potapova, G.F.

    1987-02-01

    The authors use labelled oxygen to study the involvement of water and sulfate ions in molecular oxygen evolution during the anodic synthesis of persulfuric acid and ammonium persulfate at a platinum anode in an electrolytic cell with and without thiocyanate as a promoter for the electrocatalytic reaction.

  1. World wide IFC phosphoric acid fuel cell implementation

    SciTech Connect

    King, J.M. Jr

    1996-04-01

    International Fuel Cells, a subsidary of United technologies Corporation, is engaged in research and development of all types of fuel cell technologies and currently manufactures alkaline fuel cell power plants for the U.S. manned space flight program and natural gas fueled stationary power plants using phosphoric acid fuel cells. This paper describes the phosphoric acid fuel cell power plants.

  2. Lewis acid promoted highly diastereoselective Petasis Borono-Mannich reaction: efficient synthesis of optically active β,γ-unsaturated α-amino acids.

    PubMed

    Li, Yi; Xu, Ming-Hua

    2012-04-20

    An efficient and straightforward method for the preparation of highly enantiomerically enriched β,γ-unsaturated α-amino acid derivatives by a Lewis acid promoted diastereoselective Petasis reaction of vinylboronic acid, N-tert-butanesulfinamide, and glyoxylic acid has been developed. The synthetic utilities of the approach were demonstrated by the rapid and convenient construction of challenging cyclopenta[c]proline derivatives.

  3. Nifedipine Promotes the Proliferation and Migration of Breast Cancer Cells

    PubMed Central

    Guo, Dong-Qing; Zhang, Hao; Tan, Sheng-Jiang; Gu, Yu-Chun

    2014-01-01

    Nifedipine is widely used as a calcium channel blocker (CCB) to treat angina and hypertension,but it is controversial with respect the risk of stimulation of cancers. In this study, we demonstrated that nifedipine promoted the proliferation and migration of breast cancer cells both invivo and invitro. However, verapamil, another calcium channel blocker, didn’t exert the similar effects. Nifedipine and high concentration KCl failed to alter the [Ca2+]i in MDA-MB-231 cells, suggesting that such nifedipine effect was not related with calcium channel. Moreover, nifedipine decreased miRNA-524-5p, resulting in the up-regulation of brain protein I3 (BRI3). Erk pathway was consequently activated and led to the proliferation and migration of breast cancer cells. Silencing BRI3 reversed the promoting effect of nifedipine on the breast cancer. In a summary, nifedipine stimulated the proliferation and migration of breast cancer cells via the axis of miRNA-524-5p-BRI3–Erk pathway independently of its calcium channel-blocking activity. Our findings highlight that nifedipine but not verapamil is conducive for breast cancer growth and metastasis, urging that the caution should be taken in clinic to prescribe nifedipine to women who suffering both hypertension and breast cancer, and hypertension with a tendency in breast cancers. PMID:25436889

  4. Neural crest cell-derived VEGF promotes embryonic jaw extension

    PubMed Central

    Wiszniak, Sophie; Mackenzie, Francesca E.; Anderson, Peter; Kabbara, Samuela; Ruhrberg, Christiana; Schwarz, Quenten

    2015-01-01

    Jaw morphogenesis depends on the growth of Meckel’s cartilage during embryogenesis. However, the cell types and signals that promote chondrocyte proliferation for Meckel’s cartilage growth are poorly defined. Here we show that neural crest cells (NCCs) and their derivatives provide an essential source of the vascular endothelial growth factor (VEGF) to enhance jaw vascularization and stabilize the major mandibular artery. We further show in two independent mouse models that blood vessels promote Meckel’s cartilage extension. Coculture experiments of arterial tissue with NCCs or chondrocytes demonstrated that NCC-derived VEGF promotes blood vessel growth and that blood vessels secrete factors to instruct chondrocyte proliferation. Computed tomography and X-ray scans of patients with hemifacial microsomia also showed that jaw hypoplasia correlates with mandibular artery dysgenesis. We conclude that cranial NCCs and their derivatives provide an essential source of VEGF to support blood vessel growth in the developing jaw, which in turn is essential for normal chondrocyte proliferation, and therefore jaw extension. PMID:25922531

  5. Farnesoid X receptor activation promotes cell proliferation via PDK4-controlled metabolic reprogramming

    PubMed Central

    Xie, Yang; Wang, Hong; Cheng, Xuefang; Wu, Yuzheng; Cao, Lijuan; Wu, Mengqiu; Xie, Wen; Wang, Guangji; Hao, Haiping

    2016-01-01

    Farnesoid X receptor (FXR) plays a pivotal role in the regulation of various metabolic pathways as well as liver regeneration. However, the casual link between cell proliferative effects during liver regeneration and metabolic regulation of FXR was elusive. In this study, we found that FXR activation significantly promotes HepG2 cell proliferation accompanied with metabolic switch towards the excessive accumulation of aerobic glycolytic intermediates including lactic acid, pyruvate and the subsequently increased biosynthesis of glycine. This FXR-induced metabolic switch was found dependent on an up-regulation of pyruvate dehydrogenate kinase 4 (PDK4), a FXR target gene. FXR agonists were found to promote liver regeneration in the murine model of APAP induced liver injury, which was associated with a metabolic switch favoring the accumulation of glycolytic intermediates as precursors for generation of biomass. However, FXR activation has little effect on the glycolytic metabolism in healthy primary hepatocytes in vitro and the liver of healthy mice in vivo. Therefore, we conclude that FXR may promote the proliferation of tumor cells and the hepatocytes in the process of liver regeneration by activating the PDK4-mediated metabolic reprogramming to generate glycolytic intermediates essential for rapid biomass generation, establishing a mechanistic link between cell proliferation and metabolic switch. PMID:26728993

  6. Ellagic acid promotes A{beta}42 fibrillization and inhibits A{beta}42-induced neurotoxicity

    SciTech Connect

    Feng, Ying; Yang, Shi-gao; Du, Xue-ting; Zhang, Xi; Sun, Xiao-xia; Zhao, Min; Sun, Gui-yuan; Liu, Rui-tian

    2009-12-25

    Smaller, soluble oligomers of {beta}-amyloid (A{beta}) play a critical role in the pathogenesis of Alzheimer's disease (AD). Selective inhibition of A{beta} oligomer formation provides an optimum target for AD therapy. Some polyphenols have potent anti-amyloidogenic activities and protect against A{beta} neurotoxicity. Here, we tested the effects of ellagic acid (EA), a polyphenolic compound, on A{beta}42 aggregation and neurotoxicity in vitro. EA promoted A{beta} fibril formation and significant oligomer loss, contrary to previous results that polyphenols inhibited A{beta} aggregation. The results of transmission electron microscopy (TEM) and Western blot displayed more fibrils in A{beta}42 samples co-incubated with EA in earlier phases of aggregation. Consistent with the hypothesis that plaque formation may represent a protective mechanism in which the body sequesters toxic A{beta} aggregates to render them harmless, our MTT results showed that EA could significantly reduce A{beta}42-induced neurotoxicity toward SH-SY5Y cells. Taken together, our results suggest that EA, an active ingredient in many fruits and nuts, may have therapeutic potential in AD.

  7. Vascular Mural Cells Promote Noradrenergic Differentiation of Embryonic Sympathetic Neurons.

    PubMed

    Fortuna, Vitor; Pardanaud, Luc; Brunet, Isabelle; Ola, Roxana; Ristori, Emma; Santoro, Massimo M; Nicoli, Stefania; Eichmann, Anne

    2015-06-23

    The sympathetic nervous system controls smooth muscle tone and heart rate in the cardiovascular system. Postganglionic sympathetic neurons (SNs) develop in close proximity to the dorsal aorta (DA) and innervate visceral smooth muscle targets. Here, we use the zebrafish embryo to ask whether the DA is required for SN development. We show that noradrenergic (NA) differentiation of SN precursors temporally coincides with vascular mural cell (VMC) recruitment to the DA and vascular maturation. Blocking vascular maturation inhibits VMC recruitment and blocks NA differentiation of SN precursors. Inhibition of platelet-derived growth factor receptor (PDGFR) signaling prevents VMC differentiation and also blocks NA differentiation of SN precursors. NA differentiation is normal in cloche mutants that are devoid of endothelial cells but have VMCs. Thus, PDGFR-mediated mural cell recruitment mediates neurovascular interactions between the aorta and sympathetic precursors and promotes their noradrenergic differentiation.

  8. Promoting folic acid to Spanish-speaking Hispanic women: evaluating existing campaigns to guide new development.

    PubMed

    Mackert, Michael; Kahlor, Leeann; Silva, Kristi; Padilla, Yolanda

    2010-06-01

    Hispanic women are 1.5-3 times as likely as non-Hispanic white women to have a child affected by neural tube defects. This disparity exists in spite of varied interventions designed to address the problem. The purpose of this research was to investigate Hispanic women's knowledge of folic acid, perceptions of existing education campaigns, and provide guidance for future promotion efforts. Three focus groups with Hispanic mothers (N = 18) were conducted to garner insights on these issues. Results suggested that these women understood the benefits of folic acid, did not see major cultural barriers to consuming folic acid-rich foods, and did not perceive insurmountable challenges to consuming a multivitamin with folic acid. For many women, an initial pregnancy served as their initial cue to action, suggesting a need for the continued development of education strategies that communicate the benefits of folic acid supplementation prior to pregnancy. Such strategies may necessitate targeting younger audiences, including teenagers.

  9. Bidirectional Promoter Engineering for Single Cell MicroRNA Sensors in Embryonic Stem Cells

    PubMed Central

    Sladitschek, Hanna L.

    2016-01-01

    MicroRNAs have emerged as important markers and regulators of cell identity. Precise measurements of cellular miRNA levels rely traditionally on RNA extraction and thus do not allow to follow miRNA expression dynamics at the level of single cells. Non-invasive miRNA sensors present an ideal solution but they critically depend on the performance of suitable ubiquitous promoters that reliably drive expression both in pluripotent and differentiated cell types. Here we describe the engineering of bidirectional promoters that drive the expression of precise ratiometric fluorescent miRNA sensors in single mouse embryonic stem cells (mESCs) and their differentiated derivatives. These promoters are based on combinations of the widely used CAG, EF1α and PGK promoters as well as the CMV and PGK enhancers. miR-142-3p, which is known to be bimodally expressed in mESCs, served as a model miRNA to gauge the precision of the sensors. The performance of the resulting miRNA sensors was assessed by flow cytometry in single stable transgenic mESCs undergoing self-renewal or differentiation. EF1α promoters arranged back-to-back failed to drive the robustly correlated expression of two transgenes. Back-to-back PGK promoters were shut down during mESC differentiation. However, we found that a back-to-back arrangement of CAG promoters with four CMV enhancers provided both robust expression in mESCs undergoing differentiation and the best signal-to-noise for measurement of miRNA activity in single cells among all the sensors we tested. Such a bidirectional promoter is therefore particularly well suited to study the dynamics of miRNA expression during cell fate transitions at the single cell level. PMID:27152616

  10. Bidirectional Promoter Engineering for Single Cell MicroRNA Sensors in Embryonic Stem Cells.

    PubMed

    Sladitschek, Hanna L; Neveu, Pierre A

    2016-01-01

    MicroRNAs have emerged as important markers and regulators of cell identity. Precise measurements of cellular miRNA levels rely traditionally on RNA extraction and thus do not allow to follow miRNA expression dynamics at the level of single cells. Non-invasive miRNA sensors present an ideal solution but they critically depend on the performance of suitable ubiquitous promoters that reliably drive expression both in pluripotent and differentiated cell types. Here we describe the engineering of bidirectional promoters that drive the expression of precise ratiometric fluorescent miRNA sensors in single mouse embryonic stem cells (mESCs) and their differentiated derivatives. These promoters are based on combinations of the widely used CAG, EF1α and PGK promoters as well as the CMV and PGK enhancers. miR-142-3p, which is known to be bimodally expressed in mESCs, served as a model miRNA to gauge the precision of the sensors. The performance of the resulting miRNA sensors was assessed by flow cytometry in single stable transgenic mESCs undergoing self-renewal or differentiation. EF1α promoters arranged back-to-back failed to drive the robustly correlated expression of two transgenes. Back-to-back PGK promoters were shut down during mESC differentiation. However, we found that a back-to-back arrangement of CAG promoters with four CMV enhancers provided both robust expression in mESCs undergoing differentiation and the best signal-to-noise for measurement of miRNA activity in single cells among all the sensors we tested. Such a bidirectional promoter is therefore particularly well suited to study the dynamics of miRNA expression during cell fate transitions at the single cell level.

  11. Bidirectional Promoter Engineering for Single Cell MicroRNA Sensors in Embryonic Stem Cells.

    PubMed

    Sladitschek, Hanna L; Neveu, Pierre A

    2016-01-01

    MicroRNAs have emerged as important markers and regulators of cell identity. Precise measurements of cellular miRNA levels rely traditionally on RNA extraction and thus do not allow to follow miRNA expression dynamics at the level of single cells. Non-invasive miRNA sensors present an ideal solution but they critically depend on the performance of suitable ubiquitous promoters that reliably drive expression both in pluripotent and differentiated cell types. Here we describe the engineering of bidirectional promoters that drive the expression of precise ratiometric fluorescent miRNA sensors in single mouse embryonic stem cells (mESCs) and their differentiated derivatives. These promoters are based on combinations of the widely used CAG, EF1α and PGK promoters as well as the CMV and PGK enhancers. miR-142-3p, which is known to be bimodally expressed in mESCs, served as a model miRNA to gauge the precision of the sensors. The performance of the resulting miRNA sensors was assessed by flow cytometry in single stable transgenic mESCs undergoing self-renewal or differentiation. EF1α promoters arranged back-to-back failed to drive the robustly correlated expression of two transgenes. Back-to-back PGK promoters were shut down during mESC differentiation. However, we found that a back-to-back arrangement of CAG promoters with four CMV enhancers provided both robust expression in mESCs undergoing differentiation and the best signal-to-noise for measurement of miRNA activity in single cells among all the sensors we tested. Such a bidirectional promoter is therefore particularly well suited to study the dynamics of miRNA expression during cell fate transitions at the single cell level. PMID:27152616

  12. Platelet microparticles promote neural stem cell proliferation, survival and differentiation.

    PubMed

    Hayon, Yael; Dashevsky, Olga; Shai, Ela; Varon, David; Leker, Ronen R

    2012-07-01

    Platelet microparticles (PMP) are small subcellular fragments, shed upon platelet activation. PMP host a variety of cytokines and growth factor that were previously shown to affect angiogenesis and postischemic tissue regeneration. This study attempted to explore the effect of PMP on neural stem cell (NSC) proliferation, survival and differentiation. Cells were grown as neurospheres and treated with PMP, or relevant growth factors, sphere size and cell fates were evaluated. PMP treatment led to larger neurospheres with increased cell survival. PMP treatment was comparable with the effect of acceptable single growth factors such as fibroblastic growth factor (FGF), vascular endothelial growth factor (VEGF) and platelet derived growth factor (PDGF). PMP treatment also increased the differentiation potential of NSC to glia and neurons. Specific growth factor inhibitors only partly blocked these effects, which were associated with increments in ERK and Akt phosphorylation. In this study, we show that various growth factors contained within the PMP promote neuronal cell proliferation, survival and differentiation. The results suggest a role for platelet microparticles in augmenting endogenous neural progenitor and stem cells angiogenesis and neurogenesis that might be utilized for treatment following brain injury.

  13. Hedgehog signaling in myofibroblasts directly promotes prostate tumor cell growth†

    PubMed Central

    Domenech, Maribella; Bjerregaard, Robert; Bushman, Wade; Beebe, David J.

    2012-01-01

    Despite strong evidence for the involvement of the stroma in Hedgehog signaling, little is known about the identity of the stromal cells and the signaling mechanisms that mediate the growth promoting effect of Hh signaling. We developed an in vitro co-culture model using microchannel technology to examine the effect of paracrine Hh signaling on proliferation of prostate cancer cells. We show here that activation of Hh signaling in myofibroblasts is sufficient to accelerate tumor cell growth. This effect was independent of any direct effect of Hh ligand on tumor cells or other cellular components of the tumor stroma. Further, the trophic effect of Hh pathway activation in myofibroblasts does not require collaboration of other elements of the stroma or direct physical interaction with the cancer cells. By isolating the tropic effect of Hh pathway activation in prostate stroma, we have taken the first step toward identifying cell-specific mechanisms that mediate the effect of paracrine Hh signaling on tumor growth. PMID:22234342

  14. SNAILs promote G1 phase in selected cancer cells.

    PubMed

    Wu, Ya-Lan; Xue, Jian-Xin; Zhou, Lin; Deng, Lei; Shang, Yan-Na; Liu, Fang; Mo, Xian-Ming; Lu, You

    2015-11-01

    Cells can acquire a stem-like cell phenotype through epithelial-mesenchymal transition (EMT). However, it is not known which of the stem-like cancer cells are generated by these phenotype transitions. We studied the EMT-inducing roles of SNAILs (the key inducers for the onset of EMT) in selected cancer cells (lung cancer cell line with relatively stable genome), in order to provide more implications for the investigation of EMT-related phenotype transitions in cancer. However, SNAILs fail to induce completed EMT. In addition, we proved that Snail accelerates the early G1 phase whereas Slug accelerates the late G1 phase. Blocking G1 phase is one of the basic conditions for the onset of EMT-related phenotype transitions (e.g., metastasis, acquiring stemness). The discovery of this unexpected phenomenon (promoting G1 phase) typically reveals the heterogeneity of cancer cells. The onset of EMT-related phenotype transitions in cancer needs not only the induction and activation of SNAILs, but also some particular heredity alterations (genetic or epigenetic alterations, which cause heterogeneity). The new connection between heredity alteration (heterogeneity) and phenotype transition suggests a novel treatment strategy, the heredity alteration-directed specific target therapy. Further investigations need to be conducted to study the relevant heredity alterations.

  15. TLR9 ligation in pancreatic stellate cells promotes tumorigenesis

    PubMed Central

    Zambirinis, Constantinos P.; Levie, Elliot; Nguy, Susanna; Avanzi, Antonina; Barilla, Rocky; Xu, Yijie; Seifert, Lena; Daley, Donnele; Greco, Stephanie H.; Deutsch, Michael; Jonnadula, Saikiran; Torres-Hernandez, Alejandro; Tippens, Daniel; Pushalkar, Smruti; Eisenthal, Andrew; Saxena, Deepak; Ahn, Jiyoung; Hajdu, Cristina; Engle, Dannielle D.; Tuveson, David

    2015-01-01

    Modulation of Toll-like receptor (TLR) signaling can have protective or protumorigenic effects on oncogenesis depending on the cancer subtype and on specific inflammatory elements within the tumor milieu. We found that TLR9 is widely expressed early during the course of pancreatic transformation and that TLR9 ligands are ubiquitous within the tumor microenvironment. TLR9 ligation markedly accelerates oncogenesis, whereas TLR9 deletion is protective. We show that TLR9 activation has distinct effects on the epithelial, inflammatory, and fibrogenic cellular subsets in pancreatic carcinoma and plays a central role in cross talk between these compartments. Specifically, TLR9 activation can induce proinflammatory signaling in transformed epithelial cells, but does not elicit oncogene expression or cancer cell proliferation. Conversely, TLR9 ligation induces pancreatic stellate cells (PSCs) to become fibrogenic and secrete chemokines that promote epithelial cell proliferation. TLR9-activated PSCs mediate their protumorigenic effects on the epithelial compartment via CCL11. Additionally, TLR9 has immune-suppressive effects in the tumor microenvironment (TME) via induction of regulatory T cell recruitment and myeloid-derived suppressor cell proliferation. Collectively, our work shows that TLR9 has protumorigenic effects in pancreatic carcinoma which are distinct from its influence in extrapancreatic malignancies and from the mechanistic effects of other TLRs on pancreatic oncogenesis. PMID:26481685

  16. Tumour-cell-induced endothelial cell necroptosis via death receptor 6 promotes metastasis.

    PubMed

    Strilic, Boris; Yang, Lida; Albarrán-Juárez, Julián; Wachsmuth, Laurens; Han, Kang; Müller, Ulrike C; Pasparakis, Manolis; Offermanns, Stefan

    2016-08-11

    Metastasis is the leading cause of cancer-related death in humans. It is a complex multistep process during which individual tumour cells spread primarily through the circulatory system to colonize distant organs. Once in the circulation, tumour cells remain vulnerable, and their metastatic potential largely depends on a rapid and efficient way to escape from the blood stream by passing the endothelial barrier. Evidence has been provided that tumour cell extravasation resembles leukocyte transendothelial migration. However, it remains unclear how tumour cells interact with endothelial cells during extravasation and how these processes are regulated on a molecular level. Here we show that human and murine tumour cells induce programmed necrosis (necroptosis) of endothelial cells, which promotes tumour cell extravasation and metastasis. Treatment of mice with the receptor-interacting serine/threonine-protein kinase 1 (RIPK1)-inhibitor necrostatin-1 or endothelial-cell-specific deletion of RIPK3 reduced tumour-cell-induced endothelial necroptosis, tumour cell extravasation and metastasis. In contrast, pharmacological caspase inhibition or endothelial-cell-specific loss of caspase-8 promoted these processes. We furthermore show in vitro and in vivo that tumour-cell-induced endothelial necroptosis leading to extravasation and metastasis requires amyloid precursor protein expressed by tumour cells and its receptor, death receptor 6 (DR6), on endothelial cells as the primary mediators of these effects. Our data identify a new mechanism underlying tumour cell extravasation and metastasis, and suggest endothelial DR6-mediated necroptotic signalling pathways as targets for anti-metastatic therapies. PMID:27487218

  17. Interleukin-22 Promotes Intestinal Stem Cell-Mediated Epithelial Regeneration

    PubMed Central

    Dudakov, Jarrod A.; Jenq, Robert R.; Velardi, Enrico; Young, Lauren F.; Smith, Odette M.; Lawrence, Gillian; Ivanov, Juliet A.; Fu, Ya-Yuan; Takashima, Shuichiro; Hua, Guoqiang; Martin, Maria L.; O'Rourke, Kevin P.; Lo, Yuan-Hung; Mokry, Michal; Romera-Hernandez, Monica; Cupedo, Tom; Dow, Lukas; Nieuwenhuis, Edward E.; Shroyer, Noah F.; Liu, Chen; Kolesnick, Richard

    2015-01-01

    Epithelial regeneration is critical for barrier maintenance and organ function after intestinal injury. The intestinal stem cell (ISC) niche provides Wnt, Notch, and epidermal growth factor (EGF) signals supporting Lgr5+ crypt base columnar ISCs for normal epithelial maintenance1,2. However, little is known about the regulation of the ISC compartment after tissue damage. Utilizing ex vivo organoid cultures, we provide evidence that innate lymphoid cells (ILCs), potent producers of Interleukin-22 (IL-22) after intestinal injury3,4, increased the growth of murine small intestine (SI) organoids in an IL-22-dependent fashion. Recombinant IL-22 directly targeted ISCs, augmenting the growth of both murine and human intestinal organoids, increasing proliferation, and promoting ISC expansion. IL-22 induced Stat3 phosphorylation in Lgr5+ ISCs, and Stat3 was critical for both organoid formation and IL-22-mediated regeneration. Treatment with IL-22 in vivo after murine allogeneic bone marrow transplantation (BMT) enhanced recovery of ISCs, increased epithelial regeneration, and reduced intestinal pathology and mortality from graft vs. host disease (GVHD). Atoh1-deficient organoid culture demonstrated that IL-22 induced epithelial regeneration independent of the Paneth cell niche. Our findings reveal a fundamental mechanism by which the immune system is able to support intestinal epithelium, activating ISCs to promote regeneration. PMID:26649819

  18. CONJUGATED LINOLEIC ACID PROMOTES HUMAN ADIPOCYTE INSULIN RESISTANCE THROUGH NFκB-DEPENDENT CYTOKINE PRODUCTION

    PubMed Central

    Chung1, Soonkyu; Brown2, J. Mark; Provo1, J. Nathan; Hopkins1, Robin; McIntosh1, Michael K.

    2005-01-01

    We previously demonstrated that trans-10, cis-12 conjugated linoleic acid (CLA) reduced the triglyceride (TG) content of human adipocytes by activating mitogen-activated protein kinase kinase/extracellular signal-related kinase (MEK/ERK) signaling via interleukins-6 (IL-6) and 8 (IL-8). However, the upstream mechanism is unknown. Here we show that CLA increased (≥ 6 h) the secretion of IL-6 and IL-8 in cultures containing both differentiated adipocytes and stromal vascular (SV) cells, non-differentiated SV cells, and adipose tissue explants. CLA’s isomer-specific induction of IL-6 and tumor necrosis factor-α (TNF-α) was associated with the activation of nuclear factor κB (NFκB) as evidenced by: 1) phosphorylation of IκBα, IκBα kinase (IKK), and NFκB p65; 2) IκBα degradation; and 3) nuclear translocation of NFκB. Pretreatment with selective NFκB inhibitors and the MEK/ERK inhibitor U0126 blocked CLA-mediated IL-6 gene expression. Trans-10, cis-12 CLA’s suppression of insulin-stimulated glucose uptake at 24 h was associated with decreased total and plasma membrane glucose transporter 4 (Glut4) proteins. Inhibition of NFκB activation or depletion of NFκB by RNA interference using siNFκB p65 attenuated CLA’s suppression of Glut4 and peroxisome proliferator activated receptor gamma (PPARγ) proteins and glucose uptake. Collectively, these data demonstrate for the first time that trans-10, cis-12 CLA promotes NFκB activation and subsequent induction of IL-6 which are, at least in part, responsible for trans-10, cis-12 CLA-mediated suppression of PPARγ target gene expression and insulin sensitivity in mature human adipocytes. PMID:16155293

  19. Ratite oils promote keratinocyte cell growth and inhibit leukocyte activation

    PubMed Central

    Bennett, Darin C.; Leung, Gigi; Wang, Eddy; Ma, Sam; Lo, Blanche K. K.; McElwee, Kevin J.; Cheng, Kimberly M.

    2015-01-01

    Traditionally, native Australian aborigines have used emu oil for the treatment of inflammation and to accelerate wound healing. Studies on mice suggest that topically applied emu oil may have anti-inflammatory properties and may promote wound healing. We investigated the effects of ratite oils (6 emu, 3 ostrich, 1 rhea) on immortalized human keratinocytes (HaCaT cells) in vitro by culturing the cells in media with oil concentrations of 0%, 0.5%, and 1.0%. Peking duck, tea tree, and olive oils were used as comparative controls. The same oils at 0.5% concentration were evaluated for their influence on peripheral blood mononuclear cell (PBMC) survival over 48 hr and their ability to inhibit IFNγ production in PBMCs activated by phytohemagglutinin (PHA) in ELISpot assays. Compared to no oil control, significantly shorter population doubling time durations were observed for HaCaT cells cultured in emu oil (1.51 × faster), ostrich oil (1.46 × faster), and rhea oil (1.64 × faster). Tea tree oil demonstrated significant antiproliferative activity and olive oil significantly prolonged (1.35 × slower) cell population doubling time. In contrast, almost all oils, particularly tea tree oil, significantly reduced PBMC viability. Different oils had different levels of inhibitory effect on IFNγ production with individual emu, ostrich, rhea, and duck oil samples conferring full inhibition. This preliminary investigation suggests that emu oil might promote wound healing by accelerating the growth rate of keratinocytes. Combined with anti-inflammatory properties, ratite oil may serve as a useful component in bandages and ointments for the treatment of wounds and inflammatory skin conditions. PMID:26217022

  20. Ratite oils promote keratinocyte cell growth and inhibit leukocyte activation.

    PubMed

    Bennett, Darin C; Leung, Gigi; Wang, Eddy; Ma, Sam; Lo, Blanche K K; McElwee, Kevin J; Cheng, Kimberly M

    2015-09-01

    Traditionally, native Australian aborigines have used emu oil for the treatment of inflammation and to accelerate wound healing. Studies on mice suggest that topically applied emu oil may have anti-inflammatory properties and may promote wound healing. We investigated the effects of ratite oils (6 emu, 3 ostrich, 1 rhea) on immortalized human keratinocytes (HaCaT cells) in vitro by culturing the cells in media with oil concentrations of 0%, 0.5%, and 1.0%. Peking duck, tea tree, and olive oils were used as comparative controls. The same oils at 0.5% concentration were evaluated for their influence on peripheral blood mononuclear cell (PBMC) survival over 48 hr and their ability to inhibit IFNγ production in PBMCs activated by phytohemagglutinin (PHA) in ELISpot assays. Compared to no oil control, significantly shorter population doubling time durations were observed for HaCaT cells cultured in emu oil (1.51×faster), ostrich oil (1.46×faster), and rhea oil (1.64×faster). Tea tree oil demonstrated significant antiproliferative activity and olive oil significantly prolonged (1.35×slower) cell population doubling time. In contrast, almost all oils, particularly tea tree oil, significantly reduced PBMC viability. Different oils had different levels of inhibitory effect on IFNγ production with individual emu, ostrich, rhea, and duck oil samples conferring full inhibition. This preliminary investigation suggests that emu oil might promote wound healing by accelerating the growth rate of keratinocytes. Combined with anti-inflammatory properties, ratite oil may serve as a useful component in bandages and ointments for the treatment of wounds and inflammatory skin conditions. PMID:26217022

  1. Modeling keratinocyte wound healing dynamics: Cell-cell adhesion promotes sustained collective migration.

    PubMed

    Nardini, John T; Chapnick, Douglas A; Liu, Xuedong; Bortz, David M

    2016-07-01

    The in vitro migration of keratinocyte cell sheets displays behavioral and biochemical similarities to the in vivo wound healing response of keratinocytes in animal model systems. In both cases, ligand-dependent Epidermal Growth Factor Receptor (EGFR) activation is sufficient to elicit collective cell migration into the wound. Previous mathematical modeling studies of in vitro wound healing assays assume that physical connections between cells have a hindering effect on cell migration, but biological literature suggests a more complicated story. By combining mathematical modeling and experimental observations of collectively migrating sheets of keratinocytes, we investigate the role of cell-cell adhesion during in vitro keratinocyte wound healing assays. We develop and compare two nonlinear diffusion models of the wound healing process in which cell-cell adhesion either hinders or promotes migration. Both models can accurately fit the leading edge propagation of cell sheets during wound healing when using a time-dependent rate of cell-cell adhesion strength. The model that assumes a positive role of cell-cell adhesion on migration, however, is robust to changes in the leading edge definition and yields a qualitatively accurate density profile. Using RNAi for the critical adherens junction protein, α-catenin, we demonstrate that cell sheets with wild type cell-cell adhesion expression maintain migration into the wound longer than cell sheets with decreased cell-cell adhesion expression, which fails to exhibit collective migration. Our modeling and experimental data thus suggest that cell-cell adhesion promotes sustained migration as cells pull neighboring cells into the wound during wound healing.

  2. Modeling keratinocyte wound healing dynamics: Cell-cell adhesion promotes sustained collective migration.

    PubMed

    Nardini, John T; Chapnick, Douglas A; Liu, Xuedong; Bortz, David M

    2016-07-01

    The in vitro migration of keratinocyte cell sheets displays behavioral and biochemical similarities to the in vivo wound healing response of keratinocytes in animal model systems. In both cases, ligand-dependent Epidermal Growth Factor Receptor (EGFR) activation is sufficient to elicit collective cell migration into the wound. Previous mathematical modeling studies of in vitro wound healing assays assume that physical connections between cells have a hindering effect on cell migration, but biological literature suggests a more complicated story. By combining mathematical modeling and experimental observations of collectively migrating sheets of keratinocytes, we investigate the role of cell-cell adhesion during in vitro keratinocyte wound healing assays. We develop and compare two nonlinear diffusion models of the wound healing process in which cell-cell adhesion either hinders or promotes migration. Both models can accurately fit the leading edge propagation of cell sheets during wound healing when using a time-dependent rate of cell-cell adhesion strength. The model that assumes a positive role of cell-cell adhesion on migration, however, is robust to changes in the leading edge definition and yields a qualitatively accurate density profile. Using RNAi for the critical adherens junction protein, α-catenin, we demonstrate that cell sheets with wild type cell-cell adhesion expression maintain migration into the wound longer than cell sheets with decreased cell-cell adhesion expression, which fails to exhibit collective migration. Our modeling and experimental data thus suggest that cell-cell adhesion promotes sustained migration as cells pull neighboring cells into the wound during wound healing. PMID:27105673

  3. [(-)-Epigallocatechin gallate, the main constituent of Japanese green tea, inhibits tumor promotion of okadaic acid].

    PubMed

    Yoshizawa, S

    1996-10-01

    (-)-Epigallocatechin gallate (EGCG), the main constituent of green tea, inhibited a tumor promoting activity of okadaic acid in a two-stage carcinogenesis experiment on mouse skin. The group treated with a single application of 100 micrograms 7, 12-dimethylbenz (a) anthracene followed by repeated applications of 1 microgram okadaic acid resulted in 80% of tumor-bearing mice and 4.7 of average numbers of tumors per mouse in week 20. Repeated applications of 5 mg EGCG, prior to okadaic acid, completely inhibited the tumor formation in mice up to week 20. The inhibitory effects of EGCG with two different doses of each application, 1 mg and 5 mg, were dose-dependent. A topical application of 5 mg EGCG immediately reduced the specific binding of [3H]okadaic acid to a particulate fraction of mouse skin to as low as 30% of control. According to the Scatchard analysis, the reduction of specific [3H]okadaic acid binding was mainly due to the reduction of the binding sites, not due to the change of the affinity. The reduction of the specific binding was closely related to the inhibitory effct of EGCG on tumor promotion of okadaic acid. Since EGCG is a non-toxic compound, ingested in green tea in daily life in Japan, EGCG is one of the candidates for practical cancer chemopreventive agents.

  4. The folate-coupled enzyme MTHFD2 is a nuclear protein and promotes cell proliferation.

    PubMed

    Gustafsson Sheppard, Nina; Jarl, Lisa; Mahadessian, Diana; Strittmatter, Laura; Schmidt, Angelika; Madhusudan, Nikhil; Tegnér, Jesper; Lundberg, Emma K; Asplund, Anna; Jain, Mohit; Nilsson, Roland

    2015-10-13

    Folate metabolism is central to cell proliferation and a target of commonly used cancer chemotherapeutics. In particular, the mitochondrial folate-coupled metabolism is thought to be important for proliferating cancer cells. The enzyme MTHFD2 in this pathway is highly expressed in human tumors and broadly required for survival of cancer cells. Although the enzymatic activity of the MTHFD2 protein is well understood, little is known about its larger role in cancer cell biology. We here report that MTHFD2 is co-expressed with two distinct gene sets, representing amino acid metabolism and cell proliferation, respectively. Consistent with a role for MTHFD2 in cell proliferation, MTHFD2 expression was repressed in cells rendered quiescent by deprivation of growth signals (serum) and rapidly re-induced by serum stimulation. Overexpression of MTHFD2 alone was sufficient to promote cell proliferation independent of its dehydrogenase activity, even during growth restriction. In addition to its known mitochondrial localization, we found MTHFD2 to have a nuclear localization and co-localize with DNA replication sites. These findings suggest a previously unknown role for MTHFD2 in cancer cell proliferation, adding to its known function in mitochondrial folate metabolism.

  5. LPA Promotes T Cell Recruitment through Synthesis of CXCL13

    PubMed Central

    Hui, Weili; Zhao, Chenqi; Bourgoin, Sylvain G.

    2015-01-01

    Lysophosphatidic acid (LPA) is a bioactive phospholipid playing an important role in various inflammatory diseases by inducing expression and secretion of many inflammatory cytokines/chemokines. Here we report in a murine air pouch model of inflammation that LPA induced CXCL13 secretion in a time-dependent manner and with exacerbation of the response when LPA was administered after a pretreatment with TNF-α, a key inflammatory cytokine. LPA mediates recruitment of leukocytes, including that of CD3+ cells into unprimed and TNF-α-primed air pouches. CXCL13 neutralization using a blocking antibody injected into air pouches prior to administration of LPA into TNF-α-primed air pouches decreased CD3+ cell influx. Our data highlight that LPA-mediated CXCL13 secretion plays a role in T cell recruitment and participates in regulation of the inflammatory response. PMID:26339130

  6. Human neural stem cells promote proliferation of endogenous neural stem cells and enhance angiogenesis in ischemic rat brain

    PubMed Central

    Ryu, Sun; Lee, Seung-Hoon; Kim, Seung U.; Yoon, Byung-Woo

    2016-01-01

    Transplantation of human neural stem cells into the dentate gyrus or ventricle of rodents has been reportedly to enhance neurogenesis. In this study, we examined endogenous stem cell proliferation and angiogenesis in the ischemic rat brain after the transplantation of human neural stem cells. Focal cerebral ischemia in the rat brain was induced by middle cerebral artery occlusion. Human neural stem cells were transplanted into the subventricular zone. The behavioral performance of human neural stem cells-treated ischemic rats was significantly improved and cerebral infarct volumes were reduced compared to those in untreated animals. Numerous transplanted human neural stem cells were alive and preferentially localized to the ipsilateral ischemic hemisphere. Furthermore, 5-bromo-2′-deoxyuridine-labeled endogenous neural stem cells were observed in the subventricular zone and hippocampus, where they differentiated into cells immunoreactive for the neural markers doublecortin, neuronal nuclear antigen NeuN, and astrocyte marker glial fibrillary acidic protein in human neural stem cells-treated rats, but not in the untreated ischemic animals. The number of 5-bromo-2′-deoxyuridine-positive ⁄ anti-von Willebrand factor-positive proliferating endothelial cells was higher in the ischemic boundary zone of human neural stem cells-treated rats than in controls. Finally, transplantation of human neural stem cells in the brains of rats with focal cerebral ischemia promoted the proliferation of endogenous neural stem cells and their differentiation into mature neural-like cells, and enhanced angiogenesis. This study provides valuable insights into the effect of human neural stem cell transplantation on focal cerebral ischemia, which can be applied to the development of an effective therapy for stroke. PMID:27073384

  7. Live-cell Imaging of Pol II Promoter Activity to Monitor Gene expression with RNA IMAGEtag reporters

    SciTech Connect

    Shin, Ilchung; Ray, Judhajeet; Gupta, Vinayak; Ilgu, Muslum; Beasley, Jonathan; Bendickson, Lee; Mehanovic, Samir; Kraus, George A.; Nilsen-Hamilton, Marit

    2014-04-20

    We describe a ribonucleic acid (RNA) reporter system for live-cell imaging of gene expression to detect changes in polymerase II activity on individual promoters in individual cells. The reporters use strings of RNA aptamers that constitute IMAGEtags (Intracellular MultiAptamer GEnetic tags) that can be expressed from a promoter of choice. For imaging, the cells are incubated with their ligands that are separately conjugated with one of the FRET pair, Cy3 and Cy5. The IMAGEtags were expressed in yeast from the GAL1, ADH1 or ACT1 promoters. Transcription from all three promoters was imaged in live cells and transcriptional increases from the GAL1 promoter were observed with time after adding galactose. Expression of the IMAGEtags did not affect cell proliferation or endogenous gene expression. Advantages of this method are that no foreign proteins are produced in the cells that could be toxic or otherwise influence the cellular response as they accumulate, the IMAGEtags are short lived and oxygen is not required to generate their signals. The IMAGEtag RNA reporter system provides a means of tracking changes in transcriptional activity in live cells and in real time.

  8. Promotion of cell migration by neural cell adhesion molecule (NCAM) is enhanced by PSA in a polysialyltransferase-specific manner.

    PubMed

    Guan, Feng; Wang, Xin; He, Fa

    2015-01-01

    Neural cell adhesion molecule 140 (NCAM-140) is a glycoprotein and always highly polysialylated in cancer. Functions of polysialic acid (PSA) that binds to N-glycan termini on NCAM remain unclear. ldlD-14 cells, a CHO cell mutant deficient in UDP-Gal 4-epimerase, are useful for structural and functional studies of Gal-containing glycoproteins because their abnormal glycosylation can be converted to normal status by exogenous addition of galactose (Gal). We cloned the genes for NCAM-140 and for polysialyltransferases STX and PST (responsible for PSA synthesis) from normal murine mammary gland epithelial (NMuMG) cells and transfected them into ldlD-14 and human breast cancer cells MCF-7. The effect of PSA on NCAM-mediated cell proliferation, motility, migration and adhesion was studied. We found that NCAM-140 significantly promoted cell proliferation, motility and migration, while polysialylation of NCAM-140 catalyzed by STX, but not by PST, enhanced NCAM-mediated cell migration, but not cell proliferation or motility. In addition, PSA catalyzed by different polysialyltransferases affected the adhesion of NCAM to different extracellular matrix (ECM) components. PMID:25885924

  9. Targeted Lipidomics Studies Reveal that Linolenic Acid Promotes Cotton Fiber Elongation by Activating Phosphatidylinositol and Phosphatidylinositol Monophosphate Biosynthesis.

    PubMed

    Liu, Gao-Jun; Xiao, Guang-Hui; Liu, Ning-Jing; Liu, Dan; Chen, Pei-Shuang; Qin, Yong-Mei; Zhu, Yu-Xian

    2015-06-01

    The membrane lipids from fast-elongating wild-type cotton (Gossypium hirsutum) fibers at 10 days post-anthesis, wild-type ovules with fiber cells removed, and ovules from the fuzzless-lintless mutant harvested at the same age, were extracted, separated, and quantified. Fiber cells contained significantly higher amounts of phosphatidylinositol (PI) than both ovule samples with PI 34:3 being the most predominant species. The genes encoding fatty acid desaturases (Δ(15)GhFAD), PI synthase (PIS) and PI kinase (PIK) were expressed in a fiber-preferential manner. Further analysis of phosphatidylinositol monophosphate (PIP) indicated that elongating fibers contained four- to five-fold higher amounts of PIP 34:3 than the ovules. Exogenously applied linolenic acid (C18:3), soybean L-α-PI, and PIPs containing PIP 34:3 promoted significant fiber growth, whereas a liver PI lacking the C18:3 moiety, linoleic acid, and PIP 36:2 were completely ineffective. The growth inhibitory effects of carbenoxolone, 5-hydroxytryptamine, and wortmannin were reverted by C18:3, PI, or PIP, respectively, suggesting that PIP signaling is essential for fiber cell growth. Furthermore, cotton plants expressing virus-induced gene-silencing constructs that specifically suppressed GhΔ(15)FAD, GhPIS, or GhPIK expression, resulted in significantly short-fibered phenotypes. Our data provide the basis for in-depth studies on the roles of PI and PIP in mediating cotton fiber growth.

  10. Specificity of a retinoic acid response element in the phosphoenolpyruvate carboxykinase gene promoter: consequences of both retinoic acid and thyroid hormone receptor binding.

    PubMed Central

    Lucas, P C; Forman, B M; Samuels, H H; Granner, D K

    1991-01-01

    The ability of a retinoic acid (RA) response element (RARE) in the phosphoenolpyruvate carboxykinase (PEPCK) gene promoter to mediate effects of either RA or thyroid hormone (T3) on gene expression was studied. Fusion gene constructs consisting of PEPCK promoter sequences ligated to the chloramphenicol acetyltransferase (CAT) reporter gene were used for this analysis. While T3 induced CAT expression to a small degree (about twofold) when such constructs were transiently transfected into H4IIE rat hepatoma cells, along with an expression vector encoding the alpha subtype of the T3 receptor (TR), this effect was mediated by promoter sequences distinct from the PEPCK RARE. Although TRs were capable of binding the PEPCK RARE in the form of putative monomers, dimers, and heterodimers with RA receptors (RARs), this element failed to mediate any positive effect of T3 on gene expression. In contrast, the PEPCK RARE mediated six- to eightfold induction of CAT expression by RA. When TRs were coexpressed along with RARs in transfected H4IIE cells, this RA induction was substantially blunted in a T3-independent manner. This inhibitory effect may be due to the binding of nonfunctional TRs or TR-RAR heterodimers to the PEPCK RARE. A model is proposed to explain the previously observed in vivo effects of T3 on PEPCK gene expression. Images PMID:1656224

  11. Restricted Transgene Expression in the Brain with Cell-Type Specific Neuronal Promoters

    PubMed Central

    Delzor, Aurélie; Dufour, Noelle; Petit, Fanny; Guillermier, Martine; Houitte, Diane; Auregan, Gwennaelle; Brouillet, Emmanuel; Hantraye, Philippe

    2012-01-01

    Abstract Tissue-targeted expression is of major interest for studying the contribution of cellular subpopulations to neurodegenerative diseases. However, in vivo methods to investigate this issue are limited. Here, we report an analysis of the cell specificity of expression of fluorescent reporter genes driven by six neuronal promoters, with the ubiquitous phosphoglycerate kinase 1 (PGK) promoter used as a reference. Quantitative analysis of AcGFPnuc expression in the striatum and hippocampus of rodents showed that all lentiviral vectors (LV) exhibited a neuronal tropism; however, there was substantial diversity of transcriptional activity and cell-type specificity of expression. The promoters with the highest activity were those of the 67 kDa glutamic acid decarboxylase (GAD67), homeobox Dlx5/6, glutamate receptor 1 (GluR1), and preprotachykinin 1 (Tac1) genes. Neuron-specific enolase (NSE) and dopaminergic receptor 1 (Drd1a) promoters showed weak activity, but the integration of an amplification system into the LV overcame this limitation. In the striatum, the expression profiles of Tac1 and Drd1a were not limited to the striatonigral pathway, whereas in the hippocampus, Drd1a and Dlx5/6 showed the expected restricted pattern of expression. Regulation of the Dlx5/6 promoter was observed in a disease condition, whereas Tac1 activity was unaffected. These vectors provide safe tools that are more selective than others available, for the administration of therapeutic molecules in the central nervous system (CNS). Nevertheless, additional characterization of regulatory elements in neuronal promoters is still required. PMID:22934828

  12. Effect of palmitic acid and linoleic acid on expression of ICAM-1 and VCAM-1 in human bone marrow endothelial cells (HBMECs)

    PubMed Central

    Sanadgol, Nima; Mansouri, Kamran; Bahrami, Gholamreza

    2012-01-01

    Introduction The amount and type of fatty acids (FAs) in the diet influence the risk of atherosclerosis. Palmitic acid and linoleic acid exist at high levels in Iranian edible oils. In this study, we investigated the effect of palmitic acid and linoleic acid on expression of soluble and cell-associated forms of intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) in human bone marrow endothelial cells (HBMECs). Material and methods The endothelial cells were induced with bacterial lipopolysaccharide (LPS) or tumor necrosis factor α (TNF-α), and thereafter incubated with palmitic or linoleic acid. The level of soluble and cell-associated VCAM-1 and ICAM-1 were analyzed using ELISA and western blot. Results Our findings indicated that palmitic acid up-regulates the expression of ICAM-1 and VCAM-1 in HBMECs when these cells are induced with TNF-α or LPS. In addition, the results suggest that linoleic acid could sustain up-regulated ICAM-1 and VCAM-1 in activated endothelial cells. Conclusions Chronic activation of endothelial cells in the presence of palmitic and linoleic may account for pathogenesis of cardiovascular events. These findings provide further support for the detrimental effects of these fatty acids, especially palmitic acid, in promotion and induction of cardiovascular diseases which are prevalent in the Iranian population. PMID:22661989

  13. Lactic acid bacterial cell factories for gamma-aminobutyric acid.

    PubMed

    Li, Haixing; Cao, Yusheng

    2010-11-01

    Gamma-aminobutyric acid is a non-protein amino acid that is widely present in organisms. Several important physiological functions of gamma-aminobutyric acid have been characterized, such as neurotransmission, induction of hypotension, diuretic effects, and tranquilizer effects. Many microorganisms can produce gamma-aminobutyric acid including bacteria, fungi and yeasts. Among them, gamma-aminobutyric acid-producing lactic acid bacteria have been a focus of research in recent years, because lactic acid bacteria possess special physiological activities and are generally regarded as safe. They have been extensively used in food industry. The production of lactic acid bacterial gamma-aminobutyric acid is safe and eco-friendly, and this provides the possibility of production of new naturally fermented health-oriented products enriched in gamma-aminobutyric acid. The gamma-aminobutyric acid-producing species of lactic acid bacteria and their isolation sources, the methods for screening of the strains and increasing their production, the enzymatic properties of glutamate decarboxylases and the relative fundamental research are reviewed in this article. And the potential applications of gamma-aminobutyric acid-producing lactic acid bacteria were also referred to.

  14. TGFβ Signaling Promotes Juvenile Granulosa Cell Tumorigenesis by Suppressing Apoptosis

    PubMed Central

    Mansouri-Attia, Nadéra; Tripurani, Swamy K.; Gokul, Nisha; Piard, Hermann; Anderson, Matthew L.; Eldin, Karen

    2014-01-01

    Molecular changes that give rise to granulosa cell tumors of the ovary are not well understood. Previously, we showed that deletion in granulosa cells of the bone morphogenetic protein receptor-signaling transcription factors, Smad1 and Smad5, causes development of metastatic granulosa cell tumors that phenocopy the juvenile form of granulosa cell tumors (JGCTs) in humans. The TGFβ-SMAD2/3 pathway is active in JGCTs, but its role is unknown. We tested the in vivo contribution of TGFβ-SMAD signaling to JGCT development by genetically deleting the common Smad4 from Smad1/5 double knockout mice. Smad1/5/4 triple knockout mice were sterile and had significantly increased survival and delayed tumor development compared to those for the Smad1/5 double knockout mice. The few tumors that did develop were smaller, showed no evidence of metastasis, and had increased apoptosis. In the human JGCT cell line COV434, TGFβ1 increased viability by inhibiting apoptosis through a TGFβ type I receptor–dependent repression of caspase activity and inhibition of poly(ADP-ribose) polymerase cleavage. These data support a tumor-promoting function of TGFβ in JGCTs through its ability to repress apoptosis. PMID:25243859

  15. PARP activation promotes nuclear AID accumulation in lymphoma cells.

    PubMed

    Tepper, Sandra; Jeschke, Julia; Böttcher, Katrin; Schmidt, Angelika; Davari, Kathrin; Müller, Peter; Kremmer, Elisabeth; Hemmerich, Peter; Pfeil, Ines; Jungnickel, Berit

    2016-03-15

    Activation-induced cytidine deaminase (AID) initiates immunoglobulin diversification in germinal center B cells by targeted introduction of DNA damage. As aberrant nuclear AID action contributes to the generation of B cell lymphoma, the protein's activity is tightly regulated, e.g. by nuclear/cytoplasmic shuttling and nuclear degradation. In the present study, we asked whether DNA damage may affect regulation of the AID protein. We show that exogenous DNA damage that mainly activates base excision repair leads to prevention of proteasomal degradation of AID and hence its nuclear accumulation. Inhibitor as well as knockout studies indicate that activation of poly (ADP-ribose) polymerase (PARP) by DNA damaging agents promotes both phenomena. These findings suggest that PARP inhibitors influence DNA damage dependent AID regulation, with interesting implications for the regulation of AID function and chemotherapy of lymphoma.

  16. PARP activation promotes nuclear AID accumulation in lymphoma cells

    PubMed Central

    Böttcher, Katrin; Schmidt, Angelika; Davari, Kathrin; Müller, Peter; Kremmer, Elisabeth; Hemmerich, Peter; Pfeil, Ines; Jungnickel, Berit

    2016-01-01

    Activation-induced cytidine deaminase (AID) initiates immunoglobulin diversification in germinal center B cells by targeted introduction of DNA damage. As aberrant nuclear AID action contributes to the generation of B cell lymphoma, the protein's activity is tightly regulated, e.g. by nuclear/cytoplasmic shuttling and nuclear degradation. In the present study, we asked whether DNA damage may affect regulation of the AID protein. We show that exogenous DNA damage that mainly activates base excision repair leads to prevention of proteasomal degradation of AID and hence its nuclear accumulation. Inhibitor as well as knockout studies indicate that activation of poly (ADP-ribose) polymerase (PARP) by DNA damaging agents promotes both phenomena. These findings suggest that PARP inhibitors influence DNA damage dependent AID regulation, with interesting implications for the regulation of AID function and chemotherapy of lymphoma. PMID:26921193

  17. Zfp423 promotes adipogenic differentiation of bovine stromal vascular cells.

    PubMed

    Huang, Yan; Das, Arun Kr; Yang, Qi-Yuan; Zhu, Mei-Jun; Du, Min

    2012-01-01

    Intramuscular fat or marbling is critical for the palatability of beef. In mice, very recent studies show that adipocytes and fibroblasts share a common pool of progenitor cells, with Zinc finger protein 423 (Zfp423) as a key initiator of adipogenic differentiation. To evaluate the role of Zfp423 in intramuscular adipogenesis and marbling in beef cattle, we sampled beef muscle for separation of stromal vascular cells. These cells were immortalized with pCI neo-hEST2 and individual clones were selected by G418. A total of 288 clones (3×96 well plates) were isolated and induced to adipogenesis. The presence of adipocytes was assessed by Oil-Red-O staining. Three clones with high and low adipogenic potential respectively were selected for further analyses. In addition, fibro/adipogenic progenitor cells were selected using a surface marker, platelet derived growth factor receptor (PDGFR) α. The expression of Zfp423 was much higher (307.4±61.9%, P<0.05) in high adipogenic cells, while transforming growth factor (TGF)-β was higher (156.1±48.7%, P<0.05) in low adipogenic cells. Following adipogenic differentiation, the expression of peroxisome proliferator-activated receptor γ (PPARγ) and CCAAT/enhancer binding protein α (C/EBPα) were much higher (239.4±84.1% and 310.7±138.4%, respectively, P<0.05) in high adipogenic cells. Over-expression of Zfp423 in stromal vascular cells and cloned low adipogenic cells dramatically increased their adipogenic differentiation, accompanied with the inhibition of TGF-β expression. Zfp423 knockdown by shRNA in high adipogenic cells largely prevented their adipogenic differentiation. The differential regulation of Zfp423 and TGF-β between low and high adipogenic cells is associated with the DNA methylation in their promoters. In conclusion, data show that Zfp423 is a critical regulator of adipogenesis in stromal vascular cells of bovine muscle, and Zfp423 may provide a molecular target for enhancing intramuscular adipogenesis

  18. Sertoli cells promote proliferation of bone marrow-derived mesenchymal stem cells in co-culture.

    PubMed

    Zhang, Fenxi; Lu, Ming; Liu, Hengxing; Ren, Tongming; Miao, Yingying; Wang, Jingjing

    2016-05-01

    Bone marrow-derived mesenchymal stem cells (BMSCs) are a major source for cell transplantation. The proliferative ability of BMSCs is an important determinant of the efficiency of transplant therapy. Sertoli cells are "nurse" cells for development of sperm cells. Our recent study showed that Sertoli cells promoted proliferation of human umbilical cord mesenchymal stem cells (hUCMSCs) in co-culture. Studies by other groups also showed that Sertoli cells promoted growth of endothelial cells and neural stem cells. In this study, we investigated the effect of Sertoli cells on proliferation of BMSCs. Our results showed that Sertoli cells in co-culture significantly enhanced proliferation of BMSCs (P < 0.01). Moreover, co-culture with Sertoli cells also markedly increased mRNA and/or protein expressions of Mdm2, p-Akt and Cyclin D1, and decreased p53 expression in BMSCs (P < 0.01 or < 0.05). These findings indicate that Sertoli cells have the potential to enhance proliferation of BMSCs. PMID:27319049

  19. Salicylic acid induces apoptosis in colon carcinoma cells grown in-vitro: Influence of oxygen and salicylic acid concentration

    SciTech Connect

    Zitta, Karina; Meybohm, Patrick; Bein, Berthold; Huang, Ying; Heinrich, Christin; Scholz, Jens; Steinfath, Markus; Albrecht, Martin

    2012-04-15

    In solid tumors the hypoxic environment can promote tumor progression and resistance to therapy. Recently, acetylsalicylic acid a major component of analgesic drugs and its metabolite salicylic acid (SA) have been shown to reduce the risk of colon cancer, but the mechanisms of action remain still unclear. Here we elucidate the effects of physiologically relevant concentrations of SA on colon carcinoma cells (CaCo-2) grown under normoxic and hypoxic conditions. Western blotting, caspase-3/7 apoptosis assays, MTS cell-proliferation assays, LDH cytotoxicity assays and hydrogen peroxide measurements were performed to investigate the effects of 1 and 10 {mu}M SA on CaCo-2 cells grown under normoxic conditions and cells exposed to hypoxia. Under normoxic conditions, SA did not influence cell proliferation or LDH release of CaCo-2 cells. However, caspase-3/7 activity was significantly increased. Under hypoxia, cell proliferation was reduced and LDH release and caspase-3/7 activities were increased. None of these parameters was altered by the addition of SA under hypoxic conditions. Hypoxia increased hydrogen peroxide concentrations 300-fold and SA significantly augmented the release of hydrogen peroxide under normoxic, but not under hypoxic conditions. Phosphorylation of the pro-survival kinases akt and erk1/2 was not changed by SA under hypoxic conditions, whereas under normoxia SA reduced phosphorylation of erk1/2 after 2 hours. We conclude that in colon carcinoma cells effects of SA on apoptosis and cellular signaling are dependent on the availability of oxygen. -- Highlights: Black-Right-Pointing-Pointer Effects of salicylic acid on colon carcinoma cells grown under normoxic and hypoxic conditions Black-Right-Pointing-Pointer Salicylic acid increases caspase-3/7 activity and hydrogen peroxide release under normoxia Black-Right-Pointing-Pointer Salicylic acid decreases pro-survival erk-1/2 phosphorylation under normoxia Black-Right-Pointing-Pointer Salicylic acid does

  20. Cholesteatoma Fibroblasts Promote Epithelial Cell Proliferation through Overexpression of Epiregulin

    PubMed Central

    Yoshikawa, Mamoru; Kojima, Hiromi; Yaguchi, Yuichiro; Okada, Naoko; Saito, Hirohisa; Moriyama, Hiroshi

    2013-01-01

    To investigate whether keratinocytes proliferate in response to epiregulin produced by subepithelial fibroblasts derived from middle ear cholesteatoma. Tissue samples were obtained from patients undergoing tympanoplasty. The quantitative polymerase chain reaction and immunohistochemistry were performed to examine epiregulin expression and localization in cholesteatoma tissues and retroauricular skin tissues. Fibroblasts were cultured from cholesteatoma tissues and from normal retroauricular skin. These fibroblasts were used as feeder cells for culture with a human keratinocyte cell line (PHK16-0b). To investigate the role of epiregulin in colony formation by PHK16-0b cells, epiregulin mRNA expression was knocked down in fibroblasts by using short interfering RNA and epiregulin protein was blocked with a neutralizing antibody. Epiregulin mRNA expression was significantly elevated in cholesteatoma tissues compared with that in normal retroauricular skin. Staining for epiregulin was more intense in the epithelial cells and subepithelial fibroblasts of cholesteatoma tissues than in retroauricular skin. When PHK16-0b cells were cultured with cholesteatoma fibroblasts, their colony-forming efficiency was 50% higher than when these cells were cultured with normal skin fibroblasts. Also, knockdown of epiregulin mRNA in cholesteatoma fibroblasts led to greater suppression of colony formation than knockdown in skin fibroblasts. Furthermore, the colony-forming efficiency of PHK16-0b cells was significantly reduced after treatment with an epiregulin neutralizing antibody in co-culture with cholesteatoma fibroblasts, but not in co-culture with skin fibroblasts. These results suggest that keratinocyte hyperproliferation in cholesteatoma is promoted through overexpression of epiregulin by subepithelial fibroblasts via epithelial–mesenchymal interactions, which may play a crucial role in the pathogenesis of middle ear cholesteatoma. PMID:23826119

  1. Exploitation of the health-promoting and sensory properties of organic pomegranate (Punica granatum L.) juice through lactic acid fermentation.

    PubMed

    Filannino, Pasquale; Azzi, Loubna; Cavoski, Ivana; Vincentini, Olimpia; Rizzello, Carlo G; Gobbetti, Marco; Di Cagno, Raffaella

    2013-05-15

    Two strains (POM1 and C2) or LP09 of Lactobacillus plantarum, which were previously isolated from tomatoes and carrots, and another commercial strain of L. plantarum (LP09), were selected to singly ferment (30 °C for 120 h) pomegranate juice (PJ) under standardized protocol. PJs were further stored at 4 °C for 30 days. Filtered PJ, not added of starters (unstarted PJ), was used as the control. After fermentation, all starters grew to ca. 9.0 Log CFU/mL. Viable cells of strain LP09 sharply decreased during storage. The other two strains survived to ca. 7.0 and 8.0 Log CFU/mL. Lactic acid bacteria consumed glucose, fructose, malic acid, and branched chain and aromatic amino acids. The concentration of free fatty acids increased for all started PJs. Compared to unstarted PJ, color and browning indexes of fermented PJs were preferable. The concentration of total polyphenolic compounds and antioxidant activity were the highest for started PJs, with some differences that depended on the starter used. Fermentation increased the concentration of ellagic acid, and enhanced the antimicrobial activity. Fermented PJs scavenged the reactive oxygen species generated by H2O2 and modulated the synthesis of immune-mediators from peripheral blood mononuclear cells (PBMC). Unstarted and fermented PJs inhibited the growth of K562 tumor cells. The sensory attributes of fermented PJs were preferred. The fermentation of pomegranate juice would represent a novel technology option, which joins health-promoting, sensory and preservative features to exploit the potential of pomegranate fruits.

  2. Arabidopsis ENHANCED DISEASE SUSCEPTIBILITY1 promotes systemic acquired resistance via azelaic acid and its precursor 9-oxo nonanoic acid.

    PubMed

    Wittek, Finni; Hoffmann, Thomas; Kanawati, Basem; Bichlmeier, Marlies; Knappe, Claudia; Wenig, Marion; Schmitt-Kopplin, Philippe; Parker, Jane E; Schwab, Wilfried; Vlot, A Corina

    2014-11-01

    Systemic acquired resistance (SAR) is a form of inducible disease resistance that depends on salicylic acid and its upstream regulator ENHANCED DISEASE SUSCEPTIBILITY1 (EDS1). Although local Arabidopsis thaliana defence responses activated by the Pseudomonas syringae effector protein AvrRpm1 are intact in eds1 mutant plants, SAR signal generation is abolished. Here, the SAR-specific phenotype of the eds1 mutant is utilized to identify metabolites that contribute to SAR. To this end, SAR bioassay-assisted fractionation of extracts from the wild type compared with eds1 mutant plants that conditionally express AvrRpm1 was performed. Using high-performance liquid chromatography followed by mass spectrometry, systemic immunity was associated with the accumulation of 60 metabolites, including the putative SAR signal azelaic acid (AzA) and its precursors 9-hydroperoxy octadecadienoic acid (9-HPOD) and 9-oxo nonanoic acid (ONA). Exogenous ONA induced SAR in systemic untreated leaves when applied at a 4-fold lower concentration than AzA. The data suggest that in planta oxidation of ONA to AzA might be partially responsible for this response and provide further evidence that AzA mobilizes Arabidopsis immunity in a concentration-dependent manner. The AzA fragmentation product pimelic acid did not induce SAR. The results link the C9 lipid peroxidation products ONA and AzA with systemic rather than local resistance and suggest that EDS1 directly or indirectly promotes the accumulation of ONA, AzA, or one or more of their common precursors possibly by activating one or more pathways that either result in the release of these compounds from galactolipids or promote lipid peroxidation.

  3. Uric acid promotes left ventricular diastolic dysfunction in mice fed a Western diet.

    PubMed

    Jia, Guanghong; Habibi, Javad; Bostick, Brian P; Ma, Lixin; DeMarco, Vincent G; Aroor, Annayya R; Hayden, Melvin R; Whaley-Connell, Adam T; Sowers, James R

    2015-03-01

    The rising obesity rates parallel increased consumption of a Western diet, high in fat and fructose, which is associated with increased uric acid. Population-based data support that elevated serum uric acids are associated with left ventricular hypertrophy and diastolic dysfunction. However, the mechanism by which excess uric acid promotes these maladaptive cardiac effects has not been explored. In assessing the role of Western diet-induced increases in uric acid, we hypothesized that reductions in uric acid would prevent Western diet-induced development of cardiomyocyte hypertrophy, cardiac stiffness, and impaired diastolic relaxation by reducing growth and profibrotic signaling pathways. Four-weeks-old C57BL6/J male mice were fed excess fat (46%) and fructose (17.5%) with or without allopurinol (125 mg/L), a xanthine oxidase inhibitor, for 16 weeks. The Western diet-induced increases in serum uric acid along with increases in cardiac tissue xanthine oxidase activity temporally related to increases in body weight, fat mass, and insulin resistance without changes in blood pressure. The Western diet induced cardiomyocte hypertrophy, myocardial oxidative stress, interstitial fibrosis, and impaired diastolic relaxation. Further, the Western diet enhanced activation of the S6 kinase-1 growth pathway and the profibrotic transforming growth factor-β1/Smad2/3 signaling pathway and macrophage proinflammatory polarization. All results improved with allopurinol treatment, which lowered cardiac xanthine oxidase as well as serum uric acid levels. These findings support the notion that increased production of uric acid with intake of a Western diet promotes cardiomyocyte hypertrophy, inflammation, and oxidative stress that lead to myocardial fibrosis and associated impaired diastolic relaxation.

  4. Laminin 411 and 511 promote the cholangiocyte differentiation of human induced pluripotent stem cells.

    PubMed

    Takayama, Kazuo; Mitani, Seiji; Nagamoto, Yasuhito; Sakurai, Fuminori; Tachibana, Masashi; Taniguchi, Yukimasa; Sekiguchi, Kiyotoshi; Mizuguchi, Hiroyuki

    2016-05-20

    The drug discovery research for cholestatic liver diseases has been hampered by the lack of a well-established human cholangiocyte model. Functional cholangiocyte-like cells differentiated from human induced pluripotent stem (iPS) cells are expected to be a promising candidate for such research, but there remains no well-established method for differentiating cholangiocytes from human iPS cells. In this study, we searched for a suitable extracellular matrix to promote cholangiocyte differentiation from human iPS cells, and found that both laminin 411 and laminin 511 were suitable for this purpose. The gene expression levels of the cholangiocyte markers, aquaporin 1 (AQP1), SRY-box 9 (SOX9), cystic fibrosis transmembrane conductance regulator (CFTR), G protein-coupled bile acid receptor 1 (GPBAR1), Jagged 1 (JAG1), secretin receptor (SCTR), and γ-glutamyl transferase (GGT1) were increased by using laminin 411 or laminin 511 as a matrix. In addition, the percentage of AQP1-positive cells was increased from 61.8% to 92.5% by using laminin 411 or laminin 511. Furthermore, the diameter and number of cysts consisted of cholangiocyte-like cells were increased when using either matrix. We believe that the human iPS cell-derived cholangiocyte-like cells, which were generated by using our differentiation technology, would be useful for the drug discovery research of cholestatic liver diseases. PMID:27103433

  5. Ferulic acid esters and weight-loss promoting effects in rats.

    PubMed

    Totani, Nagao; Tateishi, Sayuri; Takimoto, Tatsuya; Shinohara, Risa; Sasaki, Hideaki

    2012-01-01

    We previously fed rats with an ester (90 ppm in a powdered AIN93G diet) synthesized from gallic acid and 1,2-dioleoyl glycerol and found that it promoted weight loss more effectively than either octyl gallate or gallic acid. Here, we esterified ferulic acid (FerA) with oleic acid, monooleoyl glycerol and dioleoyl glycerol to obtain oleoyl ferulic acid (FO), feruloyl monooleoyl glycerol (FMO) and feruloyl dioleoyl glycerol (FDO) esters, respectively. A mixture of AIN93G and 90 ppm of each ester and FerA was fed to 10-week-old male Wistar rats for 12 weeks. The FMO and FDO groups weighed less than the control group starting from approximately 16 weeks of age. At 21 and 22 weeks of age, weight significantly differed between the FMO and both groups, respectively, and controls. The FO, FerA and control groups did not significantly differ in terms of body, liver, kidney and retroperitoneal fat tissue weights and serum biochemical findings. We concluded that the hydroxyl group of FerA is essential for promoting weight loss and that the carboxyl group should be esterified with alcohol. In addition, monooleoyl glycerol and dioleoyl glycerol did not show any difference as the alcohol moiety of the ester in the weight loss effect. PMID:22687778

  6. Ferulic acid esters and weight-loss promoting effects in rats.

    PubMed

    Totani, Nagao; Tateishi, Sayuri; Takimoto, Tatsuya; Shinohara, Risa; Sasaki, Hideaki

    2012-01-01

    We previously fed rats with an ester (90 ppm in a powdered AIN93G diet) synthesized from gallic acid and 1,2-dioleoyl glycerol and found that it promoted weight loss more effectively than either octyl gallate or gallic acid. Here, we esterified ferulic acid (FerA) with oleic acid, monooleoyl glycerol and dioleoyl glycerol to obtain oleoyl ferulic acid (FO), feruloyl monooleoyl glycerol (FMO) and feruloyl dioleoyl glycerol (FDO) esters, respectively. A mixture of AIN93G and 90 ppm of each ester and FerA was fed to 10-week-old male Wistar rats for 12 weeks. The FMO and FDO groups weighed less than the control group starting from approximately 16 weeks of age. At 21 and 22 weeks of age, weight significantly differed between the FMO and both groups, respectively, and controls. The FO, FerA and control groups did not significantly differ in terms of body, liver, kidney and retroperitoneal fat tissue weights and serum biochemical findings. We concluded that the hydroxyl group of FerA is essential for promoting weight loss and that the carboxyl group should be esterified with alcohol. In addition, monooleoyl glycerol and dioleoyl glycerol did not show any difference as the alcohol moiety of the ester in the weight loss effect.

  7. Differentiation of axon-related Schwann cells in vitro. I. Ascorbic acid regulates basal lamina assembly and myelin formation

    PubMed Central

    1987-01-01

    Rat Schwann cells cultured with dorsal root ganglion neurons in a serum- free defined medium fail to ensheathe or myelinate axons or assemble basal laminae. Replacement of defined medium with medium that contains human placental serum (HPS) and chick embryo extract (EE) results in both basal lamina and myelin formation. In the present study, the individual effects of HPS and EE on basal lamina assembly and on myelin formation by Schwann cells cultured with neurons have been examined. Some batches of HPS were unable to promote myelin formation in the absence of EE, as assessed by quantitative evaluation of cultures stained with Sudan black; such HPS also failed to promote basal lamina assembly, as assessed by immunofluorescence using antibodies against laminin, type IV collagen, and heparan sulfate proteoglycan. The addition of EE or L-ascorbic acid with such HPS led to the formation of large quantities of myelin and to the assembly of basal laminae. Pretreatment of EE with ascorbic acid oxidase abolished the EE activity, whereas trypsin did not. Other batches of HPS were found to promote both basal lamina and myelin formation in the absence of either EE or ascorbic acid. Ascorbic acid oxidase treatment or dialysis of these batches of HPS abolished their ability to promote Schwann cell differentiation, whereas the subsequent addition of ascorbic acid restored that ability. Ascorbic acid in the absence of serum was relatively ineffective in promoting either basal lamina or myelin formation. Fetal bovine serum was as effective as HPS in allowing ascorbic acid (and several analogs but not other reducing agents) to manifest its ability to promote Schwann cell differentiation. We suggest that ascorbic acid promotes Schwann cell myelin formation by enabling the Schwann cell to assemble a basal lamina, which is required for complete differentiation. PMID:3624305

  8. New applications for phosphoric acid fuel cells

    NASA Technical Reports Server (NTRS)

    Stickles, R. P.; Breuer, C. T.

    1983-01-01

    New applications for phosphoric acid fuel cells were identified and evaluated. Candidates considered included all possibilities except grid connected electric utility applications, on site total energy systems, industrial cogeneration, opportunistic use of waste hydrogen, space and military applications, and applications smaller than 10 kW. Applications identified were screened, with the most promising subjected to technical and economic evaluation using a fuel cell and conventional power system data base developed in the study. The most promising applications appear to be the underground mine locomotive and the railroad locomotive. Also interesting are power for robotic submersibles and Arctic villages. The mine locomotive is particularly attractive since it is expected that the fuel cell could command a very high price and still be competitive with the conventionally used battery system. The railroad locomotive's attractiveness results from the (smaller) premium price which the fuel cell could command over the conventional diesel electric system based on its superior fuel efficiency, and on the large size of this market and the accompanying opportunities for manufacturing economy.

  9. New applications for phosphoric acid fuel cells

    SciTech Connect

    Stickles, R.P.; Breuer, C.T.

    1983-11-01

    New applications for phosphoric acid fuel cells were identified and evaluated. Candidates considered included all possibilities except grid connected electric utility applications, on-site total energy systems, industrial co-generation, opportunistic use of waste hydrogen, space and military applications, and applications smaller than 10 kW. Applications identified were screened, with the most promising subjected to technical and economic evaluation using a fuel cell and conventional power system data base developed in the study. The most promising applications appear to be the underground mine locomotive and the railroad locomotive. Also interesting is power for robotic submersibles and Arctic villages. The mine locomotive is particularly attractive since it is expected that the fuel cell could command a very high price and still be competitive with the conventionally used battery system. The railroad locomotive's attractiveness results from the (smaller) premium price which the fuel cell could command over the conventional diesel electric system based on its superior fuel efficiency, and on the large size of this market and the accompanying opportunities for manufacturing economy.

  10. New charge-bearing amino acid residues that promote β-sheet secondary structure.

    PubMed

    Maynard, Stacy J; Almeida, Aaron M; Yoshimi, Yasuharu; Gellman, Samuel H

    2014-11-26

    Proteinogenic amino acid residues that promote β-sheet secondary structure are hydrophobic (e.g., Ile or Val) or only moderately polar (e.g., Thr). The design of peptides intended to display β-sheet secondary structure in water typically requires one set of residues to ensure conformational stability and an orthogonal set, with charged side chains, to ensure aqueous solubility and discourage self-association. Here we describe new amino acids that manifest substantial β-sheet propensity, by virtue of β-branching, and also bear an ionizable group in the side chain. PMID:25393077

  11. Exposure to Brefeldin A promotes initiation of meiosis in murine female germ cells.

    PubMed

    Zhang, Lian-Jun; Chen, Bo; Feng, Xin-Lei; Ma, Hua-Gang; Sun, Li-Lan; Feng, Yan-Min; Liang, Gui-Jin; Cheng, Shun-Feng; Li, Lan; Shen, Wei

    2015-01-01

    In mammals, ontogenesis starts from a fusion of spermatozoon and oocyte, which are produced by reductive nuclear division of a diploid germ cell in a specialised but complex biological process known as meiosis. However, little is known about the mechanism of meiotic initiation in germ cells, although many factors may be responsible for meiosis both in male and female gonads. In this study, 11.5 days post coitum (dpc) female fetal mouse genital ridges were cultured in vitro with exposure to Brefeldin A (BFA) for 6h, and the changes in meiosis were detected. Synaptonemal-complex analysis implied that BFA played a positive role in meiosis initiation and this hypothesis was confirmed by quantitative PCR of meiosis-specific genes: stimulated by retinoic acid gene 8 (Stra8) and deleted in a zoospermia-like (DAZL). At the same time, mRNA expression of retinoic acid synthetase (Raldh2) and retinoic acid (RA) receptors increased in female gonads with in vitro exposure to BFA. Transplanting genital ridges treated with BFA into the kidney capsule of immunodeficient mice demonstrated that the development capacity of female germ cells was normal, while formation of primordial follicles was seen to be a result of accelerated meiosis after exposure to BFA. In conclusion, the study indicated that BFA stimulated meiosis initiation partly by RA signalling and then promoted the development of follicles.

  12. Lactobionic and cellobionic acid production profiles of the resting cells of acetic acid bacteria.

    PubMed

    Kiryu, Takaaki; Kiso, Taro; Nakano, Hirofumi; Murakami, Hiromi

    2015-01-01

    Lactobionic acid was produced by acetic acid bacteria to oxidize lactose. Gluconobacter spp. and Gluconacetobacter spp. showed higher lactose-oxidizing activities than Acetobacter spp. Gluconobacter frateurii NBRC3285 produced the highest amount of lactobionic acid per cell, among the strains tested. This bacterium assimilated neither lactose nor lactobionic acid. At high lactose concentration (30%), resting cells of the bacterium showed sufficient oxidizing activity for efficient production of lactobionic acid. These properties may contribute to industrial production of lactobionic acid by the bacterium. The bacterium showed higher oxidizing activity on cellobiose than that on lactose and produced cellobionic acid. PMID:25965080

  13. Induced accumulation of oleanolic acid and ursolic acid in cell suspension cultures of Uncaria tomentosa.

    PubMed

    Feria-Romero, Iris; Lazo, Elizabeth; Ponce-Noyola, Teresa; Cerda-García-Rojas, Carlos M; Ramos-Valdivia, Ana C

    2005-06-01

    Increasing sucrose from 20 to 50 g l(-1) in Uncaria tomentosa cell suspension cultures enhanced ursolic acid and oleanolic acid production from 129 +/- 61 to 553 +/- 193 microg g(-1) cell dry wt. The maximal concentration of both triterpenes (1680 +/- 39 microg g(-1) cell dry wt) was 8 days after elicitation by jasmonic acid, while yeast extract or citrus pectin treatments produced 1189 +/- 20 or 1120 +/- 26 microg g(-1) cell dry wt, respectively. The ratio of ursolic acid:oleanolic acid was constant at 70:30.

  14. Liver cyst cytokines promote endothelial cell proliferation and development.

    PubMed

    Brodsky, Kelley S; McWilliams, Ryan R; Amura, Claudia R; Barry, Nicholas P; Doctor, R Brian

    2009-10-01

    Autosomal dominant polycystic kidney (ADPKD) is highly prevalent genetic disease. Liver cyst disease is the most common extrarenal manifestation in ADPKD and accounts for up to 10% of ADPKD morbidity and mortality. The clinical features of ADPKD liver disease arise from dramatic increases in liver cyst volumes. To identify mechanisms that promote liver cyst growth, the present study characterized the degree of vascularization of liver cyst walls and determined that cyst-specific cytokines and growth factors can drive endothelial cell proliferation and development. Microscopic techniques demonstrated liver cyst walls are well vascularized. A comparative analysis found the vascular density in free liver cyst walls was greater in mice than in humans. Treatment of human micro-vascular endothelial cells (HMEC-1) with human liver cyst fluid (huLCF) induced a rapid increase in vascular endothelium growth factor receptor 2 (VEGFR2) phosphorylation that persisted for 45-60 min and was blocked by 20 microM SU5416, a VEGFR tyrosine kinase inhibitor. Similarly, huLCF treatment of HMEC-1 cells induced an increase in the cell proliferation rate (131 +/- 6% of control levels; P > 0.05) and the degree of vascular development ('tube' diameter assay: 92 +/- 14 microm for huLCF vs. 12 +/- 7 microm for vehicle); P > 0.05). Both cell proliferation and vascular development were sensitive to SU5416. These studies indicate that factors secreted by liver cyst epithelia can activate VEGF signaling pathways and induce endothelial cell proliferation and differentiation. The present studies suggest that targeting VEGFR2-dependent angiogenesis may be an effective therapeutic strategy in blocking ADPKD liver cyst vascularization and growth. PMID:19596832

  15. Molecular mechanisms promoting the pathogenesis of Schwann cell neoplasms.

    PubMed

    Carroll, Steven L

    2012-03-01

    Neurofibromas, schwannomas and malignant peripheral nerve sheath tumors (MPNSTs) all arise from the Schwann cell lineage. Despite their common origin, these tumor types have distinct pathologies and clinical behaviors; a growing body of evidence indicates that they also arise via distinct pathogenic mechanisms. Identification of the genes that are mutated in genetic diseases characterized by the development of either neurofibromas and MPNSTs [neurofibromatosis type 1 (NF1)] or schwannomas [neurofibromatosis type 2 (NF2), schwannomatosis and Carney complex type 1] has greatly advanced our understanding of these mechanisms. The development of genetically engineered mice with ablation of NF1, NF2, SMARCB1/INI1 or PRKAR1A has confirmed the key role these genes play in peripheral nerve sheath tumorigenesis. Establishing the functions of the NF1, NF2, SMARCB1/INI1 and PRKAR1A gene products has led to the identification of key cytoplasmic signaling pathways promoting Schwann cell neoplasia and identified new therapeutic targets. Analyses of human neoplasms and genetically engineered mouse models have established that interactions with other tumor suppressors such as TP53 and CDKN2A promote neurofibroma-MPNST progression and indicate that intratumoral interactions between neoplastic and non-neoplastic cell types play an essential role in peripheral nerve sheath tumorigenesis. Recent advances have also provided new insights into the identity of the neural crest-derived populations that give rise to different types of peripheral nerve sheath tumors. Based on these findings, we now have an initial outline of the molecular mechanisms driving the pathogenesis of neurofibromas, MPNSTs and schwannomas. However, this improved understanding in turn raises a host of intriguing new questions. PMID:22160322

  16. Ubc9 promotes invasion and metastasis of lung cancer cells.

    PubMed

    Li, Hui; Niu, Huiyan; Peng, Yang; Wang, Jiahe; He, Ping

    2013-04-01

    Lung cancer is the leading cause of cancer-related mortality worldwide. The mortality is high mainly due to the lack of known effective screening procedures; there is a high tendency for early spread and systemic therapies do not cure metastatic disease. Thus, it is important to investigate the molecular mechanism(s) of lung cancer development and, specifically, to identify an effective method by which to inhibit the invasion and metastasis of lung cancer. Ubiquitin-conjugating enzyme 9 (Ubc9), the sole conjugating enzyme for sumoylation, regulates protein function and plays a key role in tumorigenesis. Whether Ubc9 is involved in the invasion and metastasis of lung cancer remains unknown. Herein, we report that Ubc9 exhibits an important role in lung cancer invasion and metastasis. We first investigated the biological effect of Ubc9 on lung cancer by cloning the Ubc9 gene into a eukaryotic expression plasmid and stably expressing it in the human small cell lung cancer cell line NCI-H446 in order to observe any biological changes. We further analyzed the effect of Ubc9 in an in vivo experiment, injecting NCI-H446 cells stably overexpressing Ubc9 into nude mice and analyzing their metastatic ability. Our results demonstrated that Ubc9 is expressed at higher levels in primary lung cancer tissue and metastatic nodules as compared to premalignant and/or normal tissue. Furthermore, we demonstrated that upregulation of Ubc9 expression promotes migration and invasion. Ubc9 likely plays an important role in cancer progression by promoting invasion and metastasis in lung cancer. PMID:23381475

  17. Functional analysis of Drosophila HSP70 promoter with different HSE numbers in human cells.

    PubMed

    Kust, Nadezda; Rybalkina, Ekaterina; Mertsalov, Ilya; Savchenko, Ekaterina; Revishchin, Alexander; Pavlova, Gali

    2014-01-01

    The activation of genetic constructs including the Drosophila hsp70 promoter with four and eight HSE sequences in the regulatory region has been described in human cells. The promoter was shown to be induced at lower temperatures compared to the human hsp70 promoter. The promoter activity increased after a 60-min heat shock already at 38 °C in human cells. The promoter activation was observed 24 h after heat shock for the constructs with eight HSEs, while those with four HSEs required 48 h. After transplantation of in vitro heat-shocked transfected cells, the promoter activity could be maintained for 3 days with a gradual decline. The promoter activation was confirmed in vivo without preliminary heat shock in mouse ischemic brain foci. Controlled expression of the Gdnf gene under a Drosophila hsp70 promoter was demonstrated. This promoter with four and eight HSE sequences in the regulatory region can be proposed as a regulated promoter in genetic therapeutic systems.

  18. Design of stereoelectronically promoted super lewis acids and unprecedented chemistry of their complexes.

    PubMed

    Foroutan-Nejad, Cina; Vicha, Jan; Marek, Radek

    2014-09-01

    A new family of stereoelectronically promoted aluminum and scandium super Lewis acids is introduced on the basis of state-of-the-art computations. Structures of these molecules are designed to minimize resonance electron donation to central metal atoms in the Lewis acids. Acidity of these species is evaluated on the basis of their fluoride-ion affinities relative to the antimony pentafluoride reference system. It is demonstrated that introduced changes in the stereochemistry of the designed ligands increase acidity considerably relative to Al and Sc complexes with analogous monodentate ligands. The high stability of fluoride complexes of these species makes them ideal candidates to be used as weakly coordinating anions in combination with highly reactive cations instead of conventional Lewis acid-fluoride complexes. Further, the interaction of all designed molecules with methane is investigated. All studied acids form stable pentavalent-carbon complexes with methane. In addition, interactions of the strongest acid of this family with very weak bases, namely, H2, N2, carbon oxides, and noble gases were investigated; it is demonstrated that this compound can form considerably stable complexes with the aforementioned molecules. To the best of our knowledge, carbonyl and nitrogen complexes of this species are the first hypothetical four-coordinated carbonyl and nitrogen complexes of aluminum. The nature of bonding in these systems is studied in detail by various bonding analysis approaches.

  19. Lactobacillus casei combats acid stress by maintaining cell membrane functionality.

    PubMed

    Wu, Chongde; Zhang, Juan; Wang, Miao; Du, Guocheng; Chen, Jian

    2012-07-01

    Lactobacillus casei strains have traditionally been recognized as probiotics and frequently used as adjunct culture in fermented dairy products where lactic acid stress is a frequently encountered environmental condition. We have investigated the effect of lactic acid stress on the cell membrane of L. casei Zhang [wild type (WT)] and its acid-resistant mutant Lbz-2. Both strains were grown under glucose-limiting conditions in chemostats; following challenge by low pH, the cell membrane stress responses were investigated. In response to acid stress, cell membrane fluidity decreased and its fatty acid composition changed to reduce the damage caused by lactic acid. Compared with the WT, the acid-resistant mutant exhibited numerous survival advantages, such as higher membrane fluidity, higher proportions of unsaturated fatty acids, and higher mean chain length. In addition, cell integrity analysis showed that the mutant maintained a more intact cellular structure and lower membrane permeability after environmental acidification. These results indicate that alteration in membrane fluidity, fatty acid distribution, and cell integrity are common mechanisms utilized by L. casei to withstand severe acidification and to reduce the deleterious effect of lactic acid on the cell membrane. This detailed comparison of cell membrane responses between the WT and mutant add to our knowledge of the acid stress adaptation and thus enable new strategies to be developed aimed at improving the industrial performance of this species under acid stress. PMID:22366811

  20. GABA A receptor π subunit promotes apoptosis of HTR-8/SVneo trophoblastic cells: Implications in preeclampsia

    PubMed Central

    LU, JUNJIE; ZHANG, QIAN; TAN, DONGMEI; LUO, WENPING; ZHAO, HAI; MA, JING; LIANG, HAO; TAN, YI

    2016-01-01

    Gamma-aminobutyric acid (GABA) functions primarily as an inhibitory neurotransmitter through its receptors in the mature central nervous system. The GABA type A receptor π subunit (GABRP) has been identified in the tissues of the reproductive system, particularly in the uterus. In addition, we have previously detected GABRP expression in both human and mouse placentas. To examine the role of GABRP in trophoblastic cell invasion, we constructed a pIRES2-GABRP-EGFP plasmid which was used for the transfection of a human placental cell line derived from first trimester extravillous trophoblasts (HTR-8/SVneo). The number of invaded cells was decreased by GABRP overexpression. Notably, the decrease in the invasive cell number may be due to the increased apoptosis of the HTR-8/SVneo cells following GABRP transfection, which was further confirmed by flow cytometry, reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and western blot analysis. Based on the increased apoptosis of trophoblastic cells in pregnancies complicated by preeclampsia (PE) and the fact that GABRP promotes the apoptosis of trophoblastic cells, we hypothesized that GABRP expression is increased in the placental tissues from patients with PE compared with that in the normal groups and this hypothesis was confirmed by RT-qPCR and immunohistochemical analysis. Taken together, these findings imply that GABRP plays an important role in placentation and this pathway may be a promising molecular target for the development of novel therapeutic strategies for PE. PMID:27221053

  1. CXCR4 engagement promotes dendritic cell survival and maturation

    SciTech Connect

    Kabashima, Kenji Sugita, Kazunari; Shiraishi, Noriko; Tamamura, Hirokazu; Fujii, Nobutaka; Tokura, Yoshiki

    2007-10-05

    It has been reported that human monocyte derived-dendritic cells (DCs) express CXCR4, responsible for chemotaxis to CXCL12. However, it remains unknown whether CXCR4 is involved in other functions of DCs. Initially, we found that CXCR4 was expressed on bone marrow-derived DCs (BMDCs). The addition of specific CXCR4 antagonist, 4-F-Benzoyl-TN14003, to the culture of mouse BMDCs decreased their number, especially the mature subset of them. The similar effect was found on the number of Langerhans cells (LCs) but not keratinocytes among epidermal cell suspensions. Since LCs are incapable of proliferating in vitro, these results indicate that CXCR4 engagement is important for not only maturation but also survival of DCs. Consistently, the dinitrobenzene sulfonic acid-induced, antigen-specific in vitro proliferation of previously sensitized lymph node cells was enhanced by CXCL12, and suppressed by CXCR4 antagonist. These findings suggest that CXCL12-CXCR4 engagement enhances DC maturation and survival to initiate acquired immune response.

  2. Pancreatic stellate cells promote epithelial-mesenchymal transition in pancreatic cancer cells

    SciTech Connect

    Kikuta, Kazuhiro; Masamune, Atsushi; Watanabe, Takashi; Ariga, Hiroyuki; Itoh, Hiromichi; Hamada, Shin; Satoh, Kennichi; Egawa, Shinichi; Unno, Michiaki; Shimosegawa, Tooru

    2010-12-17

    Research highlights: {yields} Recent studies have shown that pancreatic stellate cells (PSCs) promote the progression of pancreatic cancer. {yields} Pancreatic cancer cells co-cultured with PSCs showed loose cell contacts and scattered, fibroblast-like appearance. {yields} PSCs decreased the expression of epithelial markers but increased that of mesenchymal markers, along with increased migration. {yields} This study suggests epithelial-mesenchymal transition as a novel mechanism by which PSCs contribute to the aggressive behavior of pancreatic cancer cells. -- Abstract: The interaction between pancreatic cancer cells and pancreatic stellate cells (PSCs), a major profibrogenic cell type in the pancreas, is receiving increasing attention. There is accumulating evidence that PSCs promote the progression of pancreatic cancer by increasing cancer cell proliferation and invasion as well as by protecting them from radiation- and gemcitabine-induced apoptosis. Because epithelial-mesenchymal transition (EMT) plays a critical role in the progression of pancreatic cancer, we hypothesized that PSCs promote EMT in pancreatic cancer cells. Panc-1 and SUIT-2 pancreatic cancer cells were indirectly co-cultured with human PSCs isolated from patients undergoing operation for pancreatic cancer. The expression of epithelial and mesenchymal markers was examined by real-time PCR and immunofluorescent staining. The migration of pancreatic cancer cells was examined by scratch and two-chamber assays. Pancreatic cancer cells co-cultured with PSCs showed loose cell contacts and a scattered, fibroblast-like appearance. The expression of E-cadherin, cytokeratin 19, and membrane-associated {beta}-catenin was decreased, whereas vimentin and Snail (Snai-1) expression was increased more in cancer cells co-cultured with PSCs than in mono-cultured cells. The migration of pancreatic cancer cells was increased by co-culture with PSCs. The PSC-induced decrease of E-cadherin expression was not altered

  3. Promotion of Growth of Tumour Cells in Acutely Inflamed Tissues

    PubMed Central

    van den Brenk, H. A. S.; Stone, M.; Kelly, H.; Orton, C.; Sharpington, C.

    1974-01-01

    Acute inflammatory reactions were induced in rats by the intravenous injection of cellulose sulphate (CS) or an extract of normal rat lung homogenate (LH), or by intraperitoneal injections of Compound 48/80. These treatments greatly increased survival and clonogenic growth in the lungs of rats of intravenously injected allogeneic W-256 and Y-P388 tumour cells. Increase in the dose of intravenously injected CS caused a logarithmic increase in colony forming efficiency (CFE) of tumour cells in the lungs. CFE was not stimulated by the intravenous injection of rats with pharmacological mediators of inflammation (histamine, 5-hydroxytryptamine, bradykinin and prostaglandins PGE1 and PGF2α) which are released from tissues by agents which induce inflammation. Stimulation of CFE by CS occurred in adrenalectomized rats but was inhibited by treatment of rats with an anti-inflammatory steroid, dexamethasone. CFE was stimulated by CS in tumour immunized rats; the inflammatory state did not prevent the expression of immunity but “rescued” a proportion (approximately 20%) of the injected tumour cells from immunodestruction in the lungs. A higher proportion of tumours grew in the paws of rats when a small number of W-256 cells were injected interdigitally into the acute inflammatory swellings produced by the local injection of paws with LH or CS. CS is a “synthetic heparin” which causes marked prolongation of blood clotting time and also increases fibrinolytic activity of the blood. Anticoagulant treatment of rats with heparin did not affect CFE. Thus, there was no direct correlation between blood clotting time and CFE of blood borne tumour cells in the rat. The mechanisms which may be responsible for the nonspecific growth promoting effects of inflammatory reactions induced by various types of tissue injury on tumour induction and growth are discussed. ImagesFig. 2 PMID:4451630

  4. Increasing sucrose concentrations promote phenylpropanoid biosynthesis in grapevine cell cultures.

    PubMed

    Ferri, Maura; Righetti, Laura; Tassoni, Annalisa

    2011-02-15

    Vitis vinifera cell suspensions are a suitable model system to study the metabolic regulation of a large range of high valuable polyphenols that are important in understanding the physiology of the plant and for nutraceutical, pharmaceutical and medical purposes. Increasing sucrose concentrations were found to promote cell growth and phenylpropanoid biosynthesis in grape cell cultures obtained from cv. Barbera immature berries. This led to an intracellular accumulation and/or release into the media of specific polyphenol families (in particular, anthocyanins, catechins and stilbenes). This effect was partially correlated with a sucrose modulation of the transcription of some key biosynthetic enzymes, such as phenylalanine ammonia lyase, chalcone synthase, chalcone-flavanone isomerase and stilbene synthase. Total catechin amounts, both endogenous and released in the media, were increased in proportion to the sugar concentration, as were anthocyanin and stilbene production. Sugar treatment notably improved the endogenous accumulation and release in the culture media of resveratroloside, a resveratrol mono-glycoside, which is the most abundant stilbene found in grape cultures, especially in cv. Barbera. We hypothesize that high sucrose concentrations (exceeding those naturally-occurring in ripe berries) could play a role in plant defense via the induction of secondary metabolites, such as stilbenes.

  5. 18β-glycyrrhetinic acid suppresses experimental autoimmune encephalomyelitis through inhibition of microglia activation and promotion of remyelination.

    PubMed

    Zhou, Jieru; Cai, Wei; Jin, Min; Xu, Jingwei; Wang, Yanan; Xiao, Yichuan; Hao, Li; Wang, Bei; Zhang, Yanyun; Han, Jie; Huang, Rui

    2015-01-01

    Microglia are intrinsic immune cells in the central nervous system (CNS). The under controlled microglia activation plays important roles in inflammatory demyelination diseases, such as multiple sclerosis (MS). However, the means to modulate microglia activation as a therapeutic modality and the underlying mechanisms remain elusive. Here we show that administration of 18β-glycyrrhetinic acid (GRA), by using both preventive and therapeutic treatment protocols, significantly suppresses disease severity of experimental autoimmune encephalomyelitis (EAE) in C57BL/6 mice. The treatment effect of GRA on EAE is attributed to its regulatory effect on microglia. GRA-modulated microglia significantly decreased pro-inflammatory profile in the CNS through suppression of MAPK signal pathway. The ameliorated CNS pro-inflammatory profile prevented the recruitment of encephalitogenic T cells into the CNS, which alleviated inflammation-induced demyelination. In addition, GRA treatment promoted remyelination in the CNS of EAE mice. The induced remyelination can be mediated by the overcome of inflammation-induced blockade of brain-derived neurotrophic factor expression in microglia, as well as enhancing oligodendrocyte precursor cell proliferation. Collectively, our results demonstrate that GRA-modulated microglia suppresses EAE through inhibiting microglia activation-mediated CNS inflammation, and promoting neuroprotective effect of microglia, which represents a potential therapeutic strategy for MS and maybe other neuroinflammatory diseases associated with microglia activation.

  6. [Advances in the mechanism of mesenchymal stem cells in promoting wound healing].

    PubMed

    Zhu, Wenjing; Sun, Haobo; Lyu, Guozhong

    2015-12-01

    Mesenchymal stem cells possess the ability of self-renewal and multiple differentiation potential, thus exert immunomodulatory effect during tissue repair. Mesenchymal stem cells can stimulate angiogenesis and promote tissue repair through transdifferentiation and secreting a variety of growth factors and cytokines. This review outlines the advances in the mechanism of mesenchymal stem cells in promoting wound healing, including alleviation of inflammatory response, induction of angiogenesis, and promotion of migration of mesenchymal stem cells to the site of tissue injury.

  7. Gluconic acid production and phosphate solubilization by the plant growth-promoting bacterium Azospirillum spp.

    NASA Astrophysics Data System (ADS)

    Rodriguez, Hilda; Gonzalez, Tania; Goire, Isabel; Bashan, Yoav

    2004-11-01

    In vitro gluconic acid formation and phosphate solubilization from sparingly soluble phosphorus sources by two strains of the plant growth-promoting bacteria A. brasilense (Cd and 8-I) and one strain of A. lipoferum JA4 were studied. Strains of A. brasilense were capable of producing gluconic acid when grown in sparingly soluble calcium phosphate medium when their usual fructose carbon source is amended with glucose. At the same time, there is a reduction in pH of the medium and release of soluble phosphate. To a greater extent, gluconic acid production and pH reduction were observed for A. lipoferum JA4. For the three strains, clearing halos were detected on solid medium plates with calcium phosphate. This is the first report of in vitro gluconic acid production and direct phosphate solubilization by A. brasilense and the first report of P solubilization by A. lipoferum. This adds to the very broad spectrum of plant growth-promoting abilities of this genus.

  8. Stromal uptake and transmission of acid is a pathway for venting cancer cell-generated acid.

    PubMed

    Hulikova, Alzbeta; Black, Nicholas; Hsia, Lin-Ting; Wilding, Jennifer; Bodmer, Walter F; Swietach, Pawel

    2016-09-01

    Proliferation and invasion of cancer cells require favorable pH, yet potentially toxic quantities of acid are produced metabolically. Membrane-bound transporters extrude acid from cancer cells, but little is known about the mechanisms that handle acid once it is released into the poorly perfused extracellular space. Here, we studied acid handling by myofibroblasts (colon cancer-derived Hs675.T, intestinal InMyoFib, embryonic colon-derived CCD-112-CoN), skin fibroblasts (NHDF-Ad), and colorectal cancer (CRC) cells (HCT116, HT29) grown in monoculture or coculture. Expression of the acid-loading transporter anion exchanger 2 (AE2) (SLC4A2 product) was detected in myofibroblasts and fibroblasts, but not in CRC cells. Compared with CRC cells, Hs675.T and InMyoFib myofibroblasts had very high capacity to absorb extracellular acid. Acid uptake into CCD-112-CoN and NHDF-Ad cells was slower and comparable to levels in CRC cells, but increased alongside SLC4A2 expression under stimulation with transforming growth factor β1 (TGFβ1), a cytokine involved in cancer-stroma interplay. Myofibroblasts and fibroblasts are connected by gap junctions formed by proteins such as connexin-43, which allows the absorbed acid load to be transmitted across the stromal syncytium. To match the stimulatory effect on acid uptake, cell-to-cell coupling in NHDF-Ad and CCD-112-CoN cells was strengthened with TGFβ1. In contrast, acid transmission was absent between CRC cells, even after treatment with TGFβ1. Thus, stromal cells have the necessary molecular apparatus for assembling an acid-venting route that can improve the flow of metabolic acid through tumors. Importantly, the activities of stromal AE2 and connexin-43 do not place an energetic burden on cancer cells, allowing resources to be diverted for other activities. PMID:27543333

  9. Stromal uptake and transmission of acid is a pathway for venting cancer cell-generated acid

    PubMed Central

    Hulikova, Alzbeta; Black, Nicholas; Hsia, Lin-Ting; Wilding, Jennifer; Bodmer, Walter F.; Swietach, Pawel

    2016-01-01

    Proliferation and invasion of cancer cells require favorable pH, yet potentially toxic quantities of acid are produced metabolically. Membrane-bound transporters extrude acid from cancer cells, but little is known about the mechanisms that handle acid once it is released into the poorly perfused extracellular space. Here, we studied acid handling by myofibroblasts (colon cancer-derived Hs675.T, intestinal InMyoFib, embryonic colon-derived CCD-112-CoN), skin fibroblasts (NHDF-Ad), and colorectal cancer (CRC) cells (HCT116, HT29) grown in monoculture or coculture. Expression of the acid-loading transporter anion exchanger 2 (AE2) (SLC4A2 product) was detected in myofibroblasts and fibroblasts, but not in CRC cells. Compared with CRC cells, Hs675.T and InMyoFib myofibroblasts had very high capacity to absorb extracellular acid. Acid uptake into CCD-112-CoN and NHDF-Ad cells was slower and comparable to levels in CRC cells, but increased alongside SLC4A2 expression under stimulation with transforming growth factor β1 (TGFβ1), a cytokine involved in cancer–stroma interplay. Myofibroblasts and fibroblasts are connected by gap junctions formed by proteins such as connexin-43, which allows the absorbed acid load to be transmitted across the stromal syncytium. To match the stimulatory effect on acid uptake, cell-to-cell coupling in NHDF-Ad and CCD-112-CoN cells was strengthened with TGFβ1. In contrast, acid transmission was absent between CRC cells, even after treatment with TGFβ1. Thus, stromal cells have the necessary molecular apparatus for assembling an acid-venting route that can improve the flow of metabolic acid through tumors. Importantly, the activities of stromal AE2 and connexin-43 do not place an energetic burden on cancer cells, allowing resources to be diverted for other activities. PMID:27543333

  10. Fascinating and challenging role of tungstate promoted vanadium phosphate towards solvent free esterification of oleic acid.

    PubMed

    Behera, Gobinda Chandra; Parida, K M

    2012-01-28

    A novel solid acid catalyst has been extensively used for the esterification reaction. Herein, tungstate promoted vanadium phosphate material is fabricated from its precursor, VOHPO(4)·0.5H(2)O and its catalytic activities and structure are investigated in detail. This kind of catalyst is, for the first time, applied for the effective production of biodiesel from fatty acids. Although vanadium phosphate has been extensively used in gas phase oxidation reactions, it has not drawn much attention for its application in liquid phase reactions. Our recent results indicate that vanadium phosphate is an effective, minimally polluting and re-usable catalyst that is highly suited to the production of biodiesel from fatty acids. This work extends the possibility of using VPO in other liquid phase reactions.

  11. Lewis acid promoted ruthenium(II)-catalyzed etherifications by selective hydrogenation of carboxylic acids/esters.

    PubMed

    Li, Yuehui; Topf, Christoph; Cui, Xinjiang; Junge, Kathrin; Beller, Matthias

    2015-04-20

    Ethers are of fundamental importance in organic chemistry and they are an integral part of valuable flavors, fragrances, and numerous bioactive compounds. In general, the reduction of esters constitutes the most straightforward preparation of ethers. Unfortunately, this transformation requires large amounts of metal hydrides. Presented herein is a bifunctional catalyst system, consisting of Ru/phosphine complex and aluminum triflate, which allows selective synthesis of ethers by hydrogenation of esters or carboxylic acids. Different lactones were reduced in good yields to the desired products. Even challenging aromatic and aliphatic esters were reduced to the desired products. Notably, the in situ formed catalyst can be reused several times without any significant loss of activity.

  12. Lewis acid promoted ruthenium(II)-catalyzed etherifications by selective hydrogenation of carboxylic acids/esters.

    PubMed

    Li, Yuehui; Topf, Christoph; Cui, Xinjiang; Junge, Kathrin; Beller, Matthias

    2015-04-20

    Ethers are of fundamental importance in organic chemistry and they are an integral part of valuable flavors, fragrances, and numerous bioactive compounds. In general, the reduction of esters constitutes the most straightforward preparation of ethers. Unfortunately, this transformation requires large amounts of metal hydrides. Presented herein is a bifunctional catalyst system, consisting of Ru/phosphine complex and aluminum triflate, which allows selective synthesis of ethers by hydrogenation of esters or carboxylic acids. Different lactones were reduced in good yields to the desired products. Even challenging aromatic and aliphatic esters were reduced to the desired products. Notably, the in situ formed catalyst can be reused several times without any significant loss of activity. PMID:25728921

  13. Aspirin promotes TFF2 gene activation in human gastric cancer cell lines.

    PubMed

    Azarschab, P; Al-Azzeh, E; Kornberger, W; Gött, P

    2001-01-19

    Trefoil factor family (TFF) peptides promote cell migration, heal the mucosa and may suppress tumor growth. In reporter gene assays we show that aspirin (1-12 mM) evokes a six-fold up-regulation of TFF2, but not TFF1 and TFF3 transcription in human gastrointestinal cell lines. 6 h after application up-regulation of endogenous TFF2 mRNA was observed. TFF2 transcription was enhanced by indomethacin and arachidonic acid but repressed by staurosporine, suggesting mediation via protein kinase C. We mapped an aspirin responding element -546 to -758 bp upstream of TFF2. Up-regulation of TFF2 by aspirin may partially explain the chemopreventive potential of low dose aspirin in gastrointestinal carcinogenesis.

  14. Group X Secreted Phospholipase A2 Releases ω3 Polyunsaturated Fatty Acids, Suppresses Colitis, and Promotes Sperm Fertility.

    PubMed

    Murase, Remi; Sato, Hiroyasu; Yamamoto, Kei; Ushida, Ayako; Nishito, Yasumasa; Ikeda, Kazutaka; Kobayashi, Tetsuyuki; Yamamoto, Toshinori; Taketomi, Yoshitaka; Murakami, Makoto

    2016-03-25

    Within the secreted phospholipase A2(sPLA2) family, group X sPLA2(sPLA2-X) has the highest capacity to hydrolyze cellular membranes and has long been thought to promote inflammation by releasing arachidonic acid, a precursor of pro-inflammatory eicosanoids. Unexpectedly, we found that transgenic mice globally overexpressing human sPLA2-X (PLA2G10-Tg) displayed striking immunosuppressive and lean phenotypes with lymphopenia and increased M2-like macrophages, accompanied by marked elevation of free ω3 polyunsaturated fatty acids (PUFAs) and their metabolites. Studies usingPla2g10-deficient mice revealed that endogenous sPLA2-X, which is highly expressed in the colon epithelium and spermatozoa, mobilized ω3 PUFAs or their metabolites to protect against dextran sulfate-induced colitis and to promote fertilization, respectively. In colitis, sPLA2-X deficiency increased colorectal expression of Th17 cytokines, and ω3 PUFAs attenuated their production by lamina propria cells partly through the fatty acid receptor GPR120. In comparison, cytosolic phospholipase A2(cPLA2α) protects from colitis by mobilizing ω6 arachidonic acid metabolites, including prostaglandin E2 Thus, our results underscore a previously unrecognized role of sPLA2-X as an ω3 PUFA mobilizerin vivo, segregated mobilization of ω3 and ω6 PUFA metabolites by sPLA2-X and cPLA2α, respectively, in protection against colitis, and the novel role of a particular sPLA2-X-driven PUFA in fertilization.

  15. Downregulation of KDM4A Suppresses the Survival of Glioma Cells by Promoting Autophagy.

    PubMed

    Wang, Bo; Fan, Xinxin; Ma, Chong; Lei, Hui; Long, Qianfa; Chai, Yuan

    2016-10-01

    Glioma is the most common type of primary intracranial tumor and has a poor prognosis. It has been reported that lysine-specific demethylase 4A (KDM4A) can promote tumor progression; however, its role in human glioma remains unclear. Western blot and qRT-PCR analyses showed that KDM4A was highly expressed in U87MG and T98G cells. 48 h after transfection with siKDM4A, the protein level of KDM4A was significantly downregulated. The silenced expression of KDM4A in T98G or U87MG cells inhibited cell viability and invasion, and aggravated cell apoptosis. We found that the siKDM4A led to a significant increase in acidic vesicular organelles (AVOs) and upregulated the expression of autophagy-related proteins, including LC3B-phosphatidylethanolamine conjugate, a cytosolic form of LC3B (LC3B-II/LC3B-I) and Beclin 1 in T98G and U87MG cells. Further studies demonstrated that after pretreatment with 3-MA (3 mmol/L) for 48 h, siKDM4A-transfected cells showed a prominent decrease in LC3B-II/LC3B-I and Beclin 1, accompanied by increased viability and invasion and decreased apoptosis. Our results suggest that the inhibition of KDM4A expression might efficiently suppress glioma cell survival by promoting autophagy, providing a promising agent for treating malignant gliomas. PMID:27514525

  16. Adenovirus-mediated GDF-5 promotes the extracellular matrix expression in degenerative nucleus pulposus cells*

    PubMed Central

    Luo, Xu-wei; Liu, Kang; Chen, Zhu; Zhao, Ming; Han, Xiao-wei; Bai, Yi-guang; Feng, Gang

    2016-01-01

    Objective: To construct a recombinant adenovirus vector-carrying human growth and differentiation factor-5 (GDF-5) gene, investigate the biological effects of adenovirus-mediated GDF-5 (Ad-GDF-5) on extracellular matrix (ECM) expression in human degenerative disc nucleus pulposus (NP) cells, and explore a candidate gene therapy method for intervertebral disc degeneration (IDD). Methods: Human NP cells of a degenerative disc were isolated, cultured, and infected with Ad-GDF-5 using the AdEasy-1 adenovirus vector system. On Days 3, 7, 14, and 21, the contents of the sulfated glycosaminoglycan (sGAG), deoxyribonucleic acid (DNA) and hydroxyproline (Hyp), synthesis of proteoglycan and collagen II, gene expression of collagen II and aggrecan, and NP cell proliferation were assessed. Results: The adenovirus was an effective vehicle for gene delivery with prolonged expression of GDF-5. Biochemical analysis revealed increased sGAG and Hyp contents in human NP cells infected by Ad-GDF-5 whereas there was no conspicuous change in basal medium (BM) or Ad-green fluorescent protein (GFP) groups. Only cells in the Ad-GDF-5 group promoted the production of ECM, as demonstrated by the secretion of proteoglycan and up-regulation of collagen II and aggrecan at both protein and mRNA levels. The NP cell proliferation was significantly promoted. Conclusions: The data suggest that Ad-GDF-5 gene therapy is a potential treatment for IDD, which restores the functions of degenerative intervertebral disc through enhancing the ECM production of human NP cells. PMID:26739524

  17. Valproic Acid Increases the Hepatic Differentiation Potential of Salivary Gland Cells.

    PubMed

    Petrakova, O S; Ashapkin, V V; Shtratnikova, V Y; Kutueva, L I; Vorotelyak, E A; Borisov, M A; Terskikh, V V; Gvazava, I G; Vasiliev, A V

    2015-01-01

    The studies of cell plasticity and differentiation abilities are important problems in modern cellular biology. The use of histone deacetylase inhibitor - valproic acid is a promising approach to increasing the differentiation efficiency of various cell types. In this paper we investigate the ability of mouse submandibular salivary gland cells to differentiate into the hepatic direction and the effect of valproic acid on the efficiency of this differentiation. It was shown that the gene expression levels of hepatocyte markers (Aat, Afp, G6p, Pepck, Tat, Cyp3a13) and liver-enriched transcription factors (Hnf-3α, Hnf-3β, Hnf-4α, Hnf-6) were increased after differentiation in salivary gland cells. Valproic acid increases the specificity of hepatic differentiation, reducing the expression levels of the ductal (Krt19, Hhex1, Cyp7a1) and acinar (Ptf1a) markers. After valproic acid exposure, the efficiency of hepatic differentiation also increases, as evidenced by the increase in the gene expression level of Alb and Tdo, and increase in urea production by differentiated cells. No change was found in DNA methylation of the promoter regions of the genes; however, valproic acid treatment and subsequent hepatic differentiation largely affected the histone H3 methylation of liver-enriched genes. Thus, mouse submandibular salivary gland cells are capable of effective differentiation in the hepatic direction. Valproic acid increases the specificity and efficiency of the hepatic differentiation of these cells.

  18. Cell cycle nucleic acids, polypeptides and uses thereof

    DOEpatents

    Gordon-Kamm, William J.; Lowe, Keith S.; Larkins, Brian A.; Dilkes, Brian R.; Sun, Yuejin

    2007-08-14

    The invention provides isolated nucleic acids and their encoded proteins that are involved in cell cycle regulation. The invention further provides recombinant expression cassettes, host cells, transgenic plants, and antibody compositions. The present invention provides methods and compositions relating to altering cell cycle protein content, cell cycle progression, cell number and/or composition of plants.

  19. Co-culture with Sertoli cells promotes proliferation and migration of umbilical cord mesenchymal stem cells

    SciTech Connect

    Zhang, Fenxi; Hong, Yan; Liang, Wenmei; Ren, Tongming; Jing, Suhua; Lin, Juntang

    2012-10-12

    Highlights: Black-Right-Pointing-Pointer Co-culture of Sertoli cells (SCs) with human umbilical cord mesenchymal stem cells (UCMSCs). Black-Right-Pointing-Pointer Presence of SCs dramatically increased proliferation and migration of UCMSCs. Black-Right-Pointing-Pointer Presence of SCs stimulated expression of Mdm2, Akt, CDC2, Cyclin D, CXCR4, MAPKs. -- Abstract: Human umbilical cord mesenchymal stem cells (hUCMSCs) have been recently used in transplant therapy. The proliferation and migration of MSCs are the determinants of the efficiency of MSC transplant therapy. Sertoli cells are a kind of 'nurse' cells that support the development of sperm cells. Recent studies show that Sertoli cells promote proliferation of endothelial cells and neural stem cells in co-culture. We hypothesized that co-culture of UCMSCs with Sertoli cells may also promote proliferation and migration of UCMSCs. To examine this hypothesis, we isolated UCMSCs from human cords and Sertoli cells from mouse testes, and co-cultured them using a Transwell system. We found that UCMSCs exhibited strong proliferation ability and potential to differentiate to other cell lineages such as osteocytes and adipocytes. The presence of Sertoli cells in co-culture significantly enhanced the proliferation and migration potential of UCMSCs (P < 0.01). Moreover, these phenotypic changes were accompanied with upregulation of multiple genes involved in cell proliferation and migration including phospho-Akt, Mdm2, phospho-CDC2, Cyclin D1, Cyclin D3 as well as CXCR4, phospho-p44 MAPK and phospho-p38 MAPK. These findings indicate that Sertoli cells boost UCMSC proliferation and migration potential.

  20. SV40 enhancer activation during retinoic acid-induced differentiation of F9 embryonal carcinoma cells.

    PubMed Central

    Sleigh, M J; Lockett, T J

    1985-01-01

    The transient expression vector pSV2CAT, which carries the bacterial chloramphenicol acetyl transferase (CAT) gene under the control of the SV40 early promoter, was used to transfect the murine embryonal carcinoma cell line F9 at various times during the retinoic acid-induced differentiation of these cells. Expression of the CAT gene under SV40 promoter control was found to increase markedly on F9 cell differentiation, measured relative to expression from the thymidine kinase promoter in the same cells. A series of constructs was prepared to identify the features of the SV40 early promoter required for transcription in differentiated and undifferentiated cells, as well as the factors limiting transcription in each case. The increased transcription seen on F9 cell differentiation was not observed when cells were transfected with molecules lacking a functional enhancer. It appears that as embryonal carcinoma cells differentiate, increased SV40 transcription results from enhancer sequence activation. In both differentiated and undifferentiated cell types the level of transcription was found to be limited by the availability and/or activity of cellular factors necessary for enhancer function. Images Fig. 1. PMID:3004973

  1. Uric acid: a modulator of prostate cells and activin sensitivity.

    PubMed

    Sangkop, Febbie; Singh, Geeta; Rodrigues, Ely; Gold, Elspeth; Bahn, Andrew

    2016-03-01

    Elevated serum uric acid (SUA) or urate is associated with inflammation and gout. Recent evidence has linked urate to cancers, but little is known about urate effects in prostate cancer. Activins are inflammatory cytokines and negative growth regulators in the prostate. A hallmark of prostate cancer progression is activin insensitivity; however, mechanisms underlying this are unclear. We propose that elevated SUA is associated with prostate cancer counteracting the growth inhibitory effects of activins. The expression of activins A and B, urate transporter GLUT9 and tissue urate levels were examined in human prostate disease. Intracellular and secreted urate and GLUT9 expression were assessed in human prostate cancer cell lines. Furthermore, the effects of urate and probenecid, a known urate transport inhibitor, were determined in combination with activin A. Activin A expression was increased in low-grade prostate cancer, whereas activin B expression was reduced in high-grade prostate cancer. Intracellular urate levels decreased in all prostate pathologies, while GLUT9 expression decreased in benign prostatic hyperplasia, prostatitis and high-grade prostate cancer. Activin responsive LNCaP cells had higher intracellular and lower secreted urate levels than activin-insensitive PC3 cells. GLUT9 expression in prostate cancer cells was progressively lower than in prostate epithelial cells. Elevated extracellular urate was growth promoting in vitro, which was abolished by the gout medication probenecid, and it antagonized the growth inhibitory effects of activins. This study shows for the first time that a change in plasma or intracellular urate levels, possibly involving GLUT9 and a urate efflux transporter, has an impact on prostate cancer cell growth, and that lowering SUA levels in prostate cancer is likely to be therapeutically beneficial. PMID:26910779

  2. 9-Cis Retinoic Acid Promotes Lymphangiogenesis and Enhances Lymphatic Vessel Regeneration: Therapeutic Implications of 9-Cis Retinoic Acid for Secondary Lymphedema

    PubMed Central

    Choi, Inho; Lee, Sunju; Chung, Hee Kyoung; Lee, Yong Suk; Kim, Kyu Eui; Choi, Dongwon; Park, Eun Kyung; Yang, Dongyun; Ecoiffier, Tatiana; Monahan, John; Chen, Wen; Aguilar, Berenice; Lee, Ha Neul; Yoo, Jaehyuk; Koh, Chester J.; Chen, Lu; Wong, Alex K.; Hong, Young-Kwon

    2012-01-01

    Background The lymphatic system plays a key role in tissue fluid homeostasis and lymphatic dysfunction due to genetic defects or lymphatic vessel obstruction can cause lymphedema, disfiguring tissue swellings often associated with fibrosis and recurrent infections without available cures to date. In this study, retinoic acids (RAs) were determined to be a potent therapeutic agent that is immediately applicable to reduce secondary lymphedema. Methods and Results We report that RAs promote proliferation, migration and tube formation of cultured lymphatic endothelial cells (LECs) by activating FGF-receptor signaling. Moreover, RAs control the expression of cell-cycle checkpoint regulators such as p27Kip1, p57Kip2 and the aurora kinases through both an Akt-mediated non-genomic action and a transcription-dependent genomic action that is mediated by Prox1, a master regulator of lymphatic development. Moreover, 9-cisRA was found to activate in vivo lymphangiogenesis in animals based on mouse trachea, matrigel plug and cornea pocket assays. Finally, we demonstrate that 9-cisRA can provide a strong therapeutic efficacy in ameliorating the experimental mouse tail lymphedema by enhancing lymphatic vessel regeneration. Conclusions These in vitro and animal studies demonstrate that 9-cisRA potently activates lymphangiogenesis and promotes lymphatic regeneration in an experimental lymphedema model, presenting it as a promising novel therapeutic agent to treat human lymphedema patients. PMID:22275501

  3. Electrochemical characteristics of acid electrolytes for fuel cells

    NASA Astrophysics Data System (ADS)

    Adzic, R.; Gervasio, D.; Kanamura, K.; Razaq, A.; Razaq, M.; Yeager, Ernest B.

    1990-01-01

    Five topics investigated by the Gas Research Institute (GRI) contractors at Case Western Reserve University (CWRU) during the past year included: (1) electrochemical evaluation of perfluorinated electrolyte, (2) the Nafion solid polymer electrolyte (SPE) fuel cell, (3) electrochemistry of single crystal Pt electrodes in acid solution, (4) catalytic effects of adatoms entrapped on electrode surfaces by bipolar or monopolar ion exchange membrane layers, (5) investigations of the Fleischmann-Pons phenomenon. The principal objective of the project is to evaluate new acid electrolytes. Electrochemical evaluation was made for two bisphosphonic acids as a replacement for phosphoric acid as a fuel cell electrolyte, and also a bis-sulfonyl carbon acid as an additive to concentrated phosphoric acid electrolyte for acid H2-O2 fuel cells. Electrochemical characteristics were found for these new perfluorinated acids.

  4. Survival of activated human T lymphocytes is promoted by retinoic acid via induction of IL-2.

    PubMed

    Engedal, Nikolai; Ertesvag, Aase; Blomhoff, Heidi Kiil

    2004-03-01

    At the end of an immune response, most activated T cells spontaneously undergo programmed cell death (apoptosis). In the present study we show that all-trans retinoic acid (atRA), a major vitamin A metabolite, can inhibit the spontaneous apoptosis of activated human T lymphocytes in vitro. Isolated peripheral blood T lymphocytes were activated by 12-O-tetradecanoyl phorbol 13-acetate and cultured for up to 11 days without any further stimuli. With time, a gradual increase in cell death was observed. This spontaneous death of activated T cells was apoptotic, as demonstrated by cell shrinkage, DNA fragmentation and depolarization of the mitochondrial membrane. In the presence of physiological concentrations of atRA, the percentage of T cells exhibiting these apoptotic features was significantly reduced. After 5 days of stimulation, the percentage of TUNEL+ T cells decreased from 28 to 12% in the presence of atRA. The anti-apoptotic effect of atRA was mimicked by the retinoic acid receptor (RAR)-selective agonists 4-[(E)-2-(5,6,7,8-tetrahydro-5,5,8,8-tetramethyl-2-naphthalenyl)-1-propenyl]benzoic acid and AM-580, and totally abrogated by the RAR-selective antagonist Ro 41-5253. Cytokines of the IL-2 family have been shown to improve the survival of activated T cells. Strikingly, we found that the ability of atRA to inhibit apoptosis was significantly correlated with its ability to increase the production of IL-2. Furthermore, a blocking anti-IL-2 receptor antibody completely abrogated the anti-apoptotic effect of atRA. Together, these results suggest that retinoic acid inhibits spontaneous apoptosis of activated T lymphocytes through a RAR-dependent increase in IL-2 production.

  5. Growth modulating effects of chlorinated oleic acid in cell cultures.

    PubMed

    Høstmark, A T; Lystad, E; Jebens, E; Skramstad, J; Frøyen, P

    1998-07-01

    Chlorinated fatty acids represent a major fraction of extractable, organically bound chlorine in fish. After dietary intake such fatty acids may be transferred from the mother to the foetus through the placenta, and via breast milk to the child. In the present work we have studied the effect of chlorinated oleic acid on the growth of three widely differing types of cells in culture. Chlorinated oleic acid inhibited growth of Human Microvascular Endothelial Cells (HMVEC), Immortilized Human Kidney Epithelial (IHKE) cells, and human Hepatoma cells (HepG2). The order of potency was: HMVEC > IHKE > HepG2. Vitamin E counteracted the inhibitory effect of chlorinated oleic acid on HepG2 cells, but did not significantly affect the fatty acid effect on HMVEC or IHKE. Defatted serum albumin stimulated the growth of HMVEC and IHKE. With HMVEC there was no major interaction between the effect of albumin and chlorinated oleic acid on cell growth. In contrast, with IHKE albumin at low concentration abolished the growth inhibiting effect of chlorinated oleic acid and appreciably counteracted growth inhibition by the fatty acid of HepG2. We conclude that the growth modulation by chlorinated oleic acid and its interaction with vitamin E and albumin are cell specific.

  6. Acetylsalicylic acid and salicylic acid decrease tumor cell viability and glucose metabolism modulating 6-phosphofructo-1-kinase structure and activity.

    PubMed

    Spitz, Guilherme A; Furtado, Cristiane M; Sola-Penna, Mauro; Zancan, Patricia

    2009-01-01

    The common observation that cancer cells present higher glycolytic rates when compared to control cells leads to the proposal of glycolysis as a potential target for the development of anti-tumoral agents. Anti-inflammatory drugs, such as acetylsalicylic acid (ASA) and salicylic acid (SA), present anti-tumoral properties, inducing apoptosis and altering tumor glucose utilization. The present work aims at evaluating whether ASA could directly decrease cell glycolysis through inhibition of the major regulatory enzyme within this pathway, 6-phosphofructo-1-kinase (PFK). We show that ASA and SA inhibit purified PFK in a dose-dependent manner, and that this inhibition occurs due to the modulation of the enzyme quaternary structure. ASA and SA promote the dissociation of the enzyme active tetramers into quite inactive dimers, a common regulatory mechanism of this enzyme. The inhibitory effects of ASA and SA on PFK are fully reversible and can be prevented or reverted by the binding of the enzyme to the actin filaments. Both drugs are also able to decrease glucose consumption by human breast cancer cell line MCF-7, as well as its viability, which decrease parallelly to the inhibition of PFK on these cells. In the end, we demonstrate the ability of ASA and SA to directly modulate an important regulatory intracellular enzyme, and propose that this is one of their mechanisms promoting anti-tumoral effects.

  7. The Influence of 13-cis Retinoic Acid on Human Meibomian Gland Epithelial Cells

    PubMed Central

    Ding, Juan; Kam, Wendy R.; Dieckow, Julia; Sullivan, David A.

    2013-01-01

    Purpose. Meibomian gland dysfunction (MGD) is a primary cause of dry eye disease. One of the risk factors for MGD is exposure to 13-cis retinoic acid (13-cis RA), a metabolite of vitamin A. However, the mechanism is not well understood. We hypothesize that 13-cis RA inhibits cell proliferation, promotes cell death, alters gene and protein expressions, and attenuates cell survival pathways in human meibomian gland epithelial cells. Methods. To test our hypotheses, immortalized human meibomian gland epithelial cells were cultured with or without 13-cis RA for varying doses and time. Cell proliferation, cell death, gene expression, and proteins involved in proliferation/survival and inflammation were evaluated. Results. We found that 13-cis RA inhibited cell proliferation, induced cell death, and significantly altered the expression of 6726 genes, including those involved in cell proliferation, cell death, differentiation, keratinization, and inflammation, in human meibomian gland epithelial cells. Further, 13-cis RA also reduced the phosphorylation of Akt and increased the generation of interleukin-1β and matrix metallopeptidase 9. Conclusions. Exposure to 13-cis RA inhibits cell proliferation, increases cell death, alters gene expression, changes signaling pathways, and promotes inflammatory mediator and protease expression in meibomian gland epithelial cells. These effects may be responsible, at least in part, for the 13-cis RA–related induction of MGD. PMID:23722388

  8. Amino acids in the cultivation of mammalian cells.

    PubMed

    Salazar, Andrew; Keusgen, Michael; von Hagen, Jörg

    2016-05-01

    Amino acids are crucial for the cultivation of mammalian cells. This importance of amino acids was realized soon after the development of the first cell lines, and a solution of a mixture of amino acids has been supplied to cultured cells ever since. The importance of amino acids is further pronounced in chemically defined mammalian cell culture media, making the consideration of their biological and chemical properties necessary. Amino acids concentrations have been traditionally adjusted to their cellular consumption rates. However, since changes in the metabolic equilibrium of amino acids can be caused by changes in extracellular concentrations, metabolomics in conjunction with flux balance analysis is being used in the development of culture media. The study of amino acid transporters is also gaining importance since they control the intracellular concentrations of these molecules and are influenced by conditions in cell culture media. A better understanding of the solubility, stability, dissolution kinetics, and interactions of these molecules is needed for an exploitation of these properties in the development of dry powdered chemically defined media for mammalian cells. Due to the complexity of these mixtures however, this has proven to be challenging. Studying amino acids in mammalian cell culture media will help provide a better understanding of how mammalian cells in culture interact with their environment. It would also provide insight into the chemical behavior of these molecules in solutions of complex mixtures, which is important in the understanding of the contribution of individual amino acids to protein structure. PMID:26832172

  9. Amino acids in the cultivation of mammalian cells.

    PubMed

    Salazar, Andrew; Keusgen, Michael; von Hagen, Jörg

    2016-05-01

    Amino acids are crucial for the cultivation of mammalian cells. This importance of amino acids was realized soon after the development of the first cell lines, and a solution of a mixture of amino acids has been supplied to cultured cells ever since. The importance of amino acids is further pronounced in chemically defined mammalian cell culture media, making the consideration of their biological and chemical properties necessary. Amino acids concentrations have been traditionally adjusted to their cellular consumption rates. However, since changes in the metabolic equilibrium of amino acids can be caused by changes in extracellular concentrations, metabolomics in conjunction with flux balance analysis is being used in the development of culture media. The study of amino acid transporters is also gaining importance since they control the intracellular concentrations of these molecules and are influenced by conditions in cell culture media. A better understanding of the solubility, stability, dissolution kinetics, and interactions of these molecules is needed for an exploitation of these properties in the development of dry powdered chemically defined media for mammalian cells. Due to the complexity of these mixtures however, this has proven to be challenging. Studying amino acids in mammalian cell culture media will help provide a better understanding of how mammalian cells in culture interact with their environment. It would also provide insight into the chemical behavior of these molecules in solutions of complex mixtures, which is important in the understanding of the contribution of individual amino acids to protein structure.

  10. Transepithelial transport of ferulic acid by monocarboxylic acid transporter in Caco-2 cell monolayers.

    PubMed

    Konishi, Yutaka; Shimizu, Makoto

    2003-04-01

    Our previous study (Biosci. Biotechnol. Biochem., 66, 2449-2457 (2002)), suggested that ferulic acid was transported via a monocarboxylic acid transporter (MCT). Transepithelial transport of ferulic acid was examined in this study by directly measuring the rate of its transport across Caco-2 cell monolayers. Ferulic acid transport was dependent on pH, and in a vectorical way in the apical-basolateral direction. The permeation of ferulic acid was concentration-dependent and saturable; the Michaelis constant was 16.2 mM and the maximum velocity was 220.4 nmol min-1 (mg protein)-1. Various substrates for MCTs, such as benzoic acid and acetic acid, strongly inhibited the permeation of ferulic acid, demonstrating that ferulic acid is obviously transported by MCT. Antioxidative phenolic acid compounds from dietary sources like ferulic acid would be recognized and transported by MCT by intestinal absorption.

  11. Valproic acid protects neurons and promotes neuronal regeneration after brachial plexus avulsion

    PubMed Central

    Li, Qiang; Wu, Dianxiu; Li, Rui; Zhu, Xiaojuan; Cui, Shusen

    2013-01-01

    Valproic acid has been shown to exert neuroprotective effects and promote neurite outgrowth in several peripheral nerve injury models. However, whether valproic acid can exert its beneficial effect on neurons after brachial plexus avulsion injury is currently unknown. In this study, brachial plexus root avulsion models, established in Wistar rats, were administered daily with valproic acid dissolved in drinking water (300 mg/kg) or normal water. On days 1, 2, 3, 7, 14 and 28 after avulsion injury, tissues of the C5–T1 spinal cord segments of the avulsion injured side were harvested to investigate the expression of Bcl-2, c-Jun and growth associated protein 43 by real-time PCR and western blot assay. Results showed that valproic acid significantly increased the expression of Bcl-2 and growth associated protein 43, and reduced the c-Jun expression after brachial plexus avulsion. Our findings indicate that valproic acid can protect neurons in the spinal cord and enhance neuronal regeneration following brachial plexus root avulsion. PMID:25206605

  12. Dendritic Cells Promote Pancreatic Viability in Mice with Acute Pancreatitis

    PubMed Central

    Bedrosian, Andrea S.; Nguyen, Andrew H.; Hackman, Michael; Connolly, Michael K.; Malhotra, Ashim; Ibrahim, Junaid; Cieza-Rubio, Napoleon E.; Henning, Justin R.; Barilla, Rocky; Rehman, Adeel; Pachter, H. Leon; Medina-Zea, Marco V.; Cohen, Steven M.; Frey, Alan B.; Acehan, Devrim; Miller, George

    2011-01-01

    Background & Aims Acute pancreatitis increases morbidity and mortality from organ necrosis by mechanisms that are incompletely understood. Dendritic cells (DCs) can promote or suppress inflammation, depending on their subtype and context. We investigated the roles of DC in development of acute pancreatitis. Methods Acute pancreatitis was induced in CD11c.DTR mice using caerulein or L-arginine; DCs were depleted by administration of diphtheria toxin. Survival was analyzed using Kaplan-Meier analysis. Results Numbers of MHC II+CD11c+DC increased 100-fold in pancreas of mice with acute pancreatitis, to account for nearly 15% of intra-pancreatic leukocytes. Intra-pancreatic DC acquired an immune phenotype in mice with acute pancreatitis; they expressed higher levels of MHC II and CD86 and increased production of interleukin-6, membrane cofactor protein (MCP)-1, and tumor necrosis factor (TNF)-α. However, rather than inducing an organ-destructive inflammatory process, DC were required for pancreatic viability; the exocrine pancreas died in mice that were depleted of DC and challenged with caerulein or L-arginine. All mice with pancreatitis that were depleted of DC died from acinar cell death within 4 days. Depletion of DC from mice with pancreatitis resulted in neutrophil infiltration and increased levels of systemic markers of inflammation. However, the organ necrosis associated with depletion of DC did not require infiltrating neutrophils, activation of NF-κB, or signaling by mitogen-activated protein kinase or TNF-α. Conclusions DC are required for pancreatic viability in mice with acute pancreatitis and might protect organs against cell stress. PMID:21801698

  13. Electrochemical technique and copper-promoted transformations: selective hydroxylation and amination of arylboronic acids.

    PubMed

    Qi, Hong-Lin; Chen, Dong-Song; Ye, Jian-Shan; Huang, Jing-Mei

    2013-08-01

    An efficient and selective electrosynthesis of phenols and anilines from arylboronic acids in aqueous ammonia is achieved in an undivided cell. By simply changing the concentration of aqueous ammonia and the anode potential, good yields of phenols and anilines can be obtained chemoselectively with high reaction rates. We propose that anodic oxidation could have played an important role in these transformations. PMID:23808633

  14. Human hair follicle pluripotent stem (hfPS) cells promote regeneration of peripheral-nerve injury: an advantageous alternative to ES and iPS cells.

    PubMed

    Amoh, Yasuyuki; Kanoh, Maho; Niiyama, Shiro; Hamada, Yuko; Kawahara, Katsumasa; Sato, Yuichi; Hoffman, Robert M; Katsuoka, Kensei

    2009-08-01

    The optimal source of stem cells for regenerative medicine is a major question. Embryonic stem (ES) cells have shown promise for pluripotency but have ethical issues and potential to form teratomas. Pluripotent stem cells have been produced from skin cells by either viral-, plasmid- or transposon-mediated gene transfer. These stem cells have been termed induced pluripotent stem cells or iPS cells. iPS cells may also have malignant potential and are inefficiently produced. Embryonic stem cells may not be suited for individualized therapy, since they can undergo immunologic rejection. To address these fundamental problems, our group is developing hair follicle pluripotent stem (hfPS) cells. Our previous studies have shown that mouse hfPS cells can differentiate to neurons, glial cells in vitro, and other cell types, and can promote nerve and spinal cord regeneration in vivo. hfPS cells are located above the hair follicle bulge in what we have termed the hfPS cell area (hfPSA) and are nestin positive and keratin 15 (K-15) negative. Human hfPS cells can also differentiate into neurons, glia, keratinocytes, smooth muscle cells, and melanocytes in vitro. In the present study, human hfPS cells were transplanted in the severed sciatic nerve of the mouse where they differentiated into glial fibrillary-acidic-protein (GFAP)-positive Schwann cells and promoted the recovery of pre-existing axons, leading to nerve generation. The regenerated nerve recovered function and, upon electrical stimulation, contracted the gastrocnemius muscle. The hfPS cells can be readily isolated from the human scalp, thereby providing an accessible, autologous and safe source of stem cells for regenerative medicine that have important advantages over ES or iPS cells.

  15. Reconstitution of bile acid transport in the rat hepatoma McArdle RH-7777 cell line.

    PubMed

    Torchia, E C; Shapiro, R J; Agellon, L B

    1996-07-01

    The liver recovers bile acids from the portal circulation primarily via an active process that is dependent on sodium ions. Hepatocytes lose the ability to transport bile acids in culture, and, in liver-derived permanent cell lines, this ability is severely reduced or absent. To study the importance of bile acids in regulating liver-specific functions (e.g., cellular bile acid and cholesterol metabolism), we have re-established active bile acid transport in cultured cells. The complementary DNA (cDNA) encoding the rat sodium/taurocholate cotransporting polypeptide (ntcp) was placed under the control of a cytomegalovirus promoter and transfected into the rat hepatoma cell line, McArdle RH-7777. Transfected cells were screened for the ability to take up [3H]-taurocholate. Clones that displayed the ability to take up taurocholate were expanded (designated McNtcp) and further characterized. The apparent Michaelis constant (Km) for taurocholate uptake was similar among the different clones. The observed maximum velocity (Vmax), however, differed and was positively correlated with the abundance of recombinant ntcp messenger RNA (mRNA). The highest level of taurocholate uptake activity observed in McNtcp cells was comparable with that of freshly isolated hepatocytes. Efflux of accumulated taurocholate from McNtcp cells proceeded in a manner similar to primary hepatocytes, indicating that McArdle RH-7777 cells have retained the ability to secrete bile acids. Moreover, taurocholate uptake in McNtcp cells was inhibited by other bile acid species. Based on the observed kinetic parameters, the reconstituted McArdle RH-7777 cells mimic the ability of primary hepatocytes to transport bile acids.

  16. Omega 3 but not omega 6 fatty acids inhibit AP-1 activity and cell transformation in JB6 cells

    PubMed Central

    Liu, Guangming; Bibus, Douglas M.; Bode, Ann M.; Ma, Wei-Ya; Holman, Ralph T.; Dong, Zigang

    2001-01-01

    Epidemiological and animal-based investigations have indicated that the development of skin cancer is in part associated with poor dietary practices. Lipid content and subsequently the derived fatty acid composition of the diet are believed to play a major role in the development of tumorigenesis. Omega 3 (ω3) fatty acids, including docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA), can effectively reduce the risk of skin cancer whereas omega 6 (ω6) fatty acids such as arachidonic acid (AA) reportedly promote risk. To investigate the effects of fatty acids on tumorigenesis, we performed experiments to examine the effects of the ω3 fatty acids EPA and DHA and of the ω6 fatty acid AA on phorbol 12-tetradecanoate 13-acetate (TPA)-induced or epidermal growth factor (EGF)-induced transcription activator protein 1 (AP-1) transactivation and on the subsequent cellular transformation in a mouse epidermal JB6 cell model. DHA treatment resulted in marked inhibition of TPA- and EGF-induced cell transformation by inhibiting AP-1 transactivation. EPA treatment also inhibited TPA-induced AP-1 transactivation and cell transformation but had no effect on EGF-induced transformation. AA treatment had no effect on either TPA- or EGF-induced AP-1 transactivation or transformation, but did abrogate the inhibitory effects of DHA on TPA- or EGF-induced AP-1 transactivation and cell transformation in a dose-dependent manner. The results of this study demonstrate that the inhibitory effects of ω3 fatty acids on tumorigenesis are more significant for DHA than for EPA and are related to an inhibition of AP-1. Similarly, because AA abrogates the beneficial effects of DHA, the dietary ratio of ω6 to ω3 fatty acids may be a significant factor in mediating tumor development. PMID:11416221

  17. Parasitic worms stimulate host NADPH oxidases to produce reactive oxygen species that limit plant cell death and promote infection.

    PubMed

    Siddique, Shahid; Matera, Christiane; Radakovic, Zoran S; Hasan, M Shamim; Gutbrod, Philipp; Rozanska, Elzbieta; Sobczak, Miroslaw; Torres, Miguel Angel; Grundler, Florian M W

    2014-04-01

    Plants and animals produce reactive oxygen species (ROS) in response to infection. In plants, ROS not only activate defense responses and promote cell death to limit the spread of pathogens but also restrict the amount of cell death in response to pathogen recognition. Plants also use hormones, such as salicylic acid, to mediate immune responses to infection. However, there are long-lasting biotrophic plant-pathogen interactions, such as the interaction between parasitic nematodes and plant roots during which defense responses are suppressed and root cells are reorganized to specific nurse cell systems. In plants, ROS are primarily generated by plasma membrane-localized NADPH (reduced form of nicotinamide adenine dinucleotide phosphate) oxidases, and loss of NADPH oxidase activity compromises immune responses and cell death. We found that infection of Arabidopsis thaliana by the parasitic nematode Heterodera schachtii activated the NADPH oxidases RbohD and RbohF to produce ROS, which was necessary to restrict infected plant cell death and promote nurse cell formation. RbohD- and RbohF-deficient plants exhibited larger regions of cell death in response to nematode infection, and nurse cell formation was greatly reduced. Genetic disruption of SID2, which is required for salicylic acid accumulation and immune activation in nematode-infected plants, led to the increased size of nematodes in RbohD- and RbohF-deficient plants, but did not decrease plant cell death. Thus, by stimulating NADPH oxidase-generated ROS, parasitic nematodes fine-tune the pattern of plant cell death during the destructive root invasion and may antagonize salicylic acid-induced defense responses during biotrophic life stages.

  18. Endothelial cells mitigate DNA damage and promote the regeneration of hematopoietic stem cells after radiation injury

    PubMed Central

    Zachman, Derek K.; Leon, Ronald P.; Das, Prerna; Goldman, Devorah C.; Hamlin, Kimberly L.; Guha, Chandan; Fleming, William H.

    2014-01-01

    Endothelial cells (ECs) are an essential component of the hematopoietic microenvironment, which maintains and regulates hematopoietic stem cells (HSCs). Although ECs can support the regeneration of otherwise lethally-irradiated HSCs, the mechanisms are not well understood. To further understand this phenomenon, we studied HSC regeneration from irradiated bone marrow using co-culture with human aortic endothelial cells (HAECs). Co-culture with HAECs induced a 24-fold expansion of long-term HSCs (CD150+, lineagelo, Sca-1+, c-Kit+; CD150+LSK cells) in vitro. These cells gave rise to functional hematopoietic stem and progenitor cells (HSPCs) with colony-forming activity, multilineage reconstitution and serial transplantation potential. Furthermore, HAECs significantly reduced DNA damage in irradiated LSK cells within 24 hours. Remarkably, we were able to delay the exposure of irradiated bone marrow to the regenerative, HAEC-derived signals for up to 48 hours and still rescue functional HSCs. G-CSF is the gold standard for promoting hematopoietic regeneration in vivo. However, when compared to HAECs, in vitro G-CSF treatment promoted lineage differentiation and regenerated 5-fold fewer CD150+LSK cells. Together, our results show that HAECs are powerful, direct mitigators of HSC injury and DNA damage. Identification of the HAEC-derived factors that rescue HSCs may lead to improved therapies for hematopoietic regeneration after radiation injury. PMID:23939266

  19. Coniferyl Aldehyde Attenuates Radiation Enteropathy by Inhibiting Cell Death and Promoting Endothelial Cell Function

    PubMed Central

    Son, Yeonghoon; Jang, Jun-Ho; Lee, Yoon-Jin; Kim, Sung-Ho; Ko, Young-Gyo; Lee, Yun-Sil; Lee, Hae-June

    2015-01-01

    Radiation enteropathy is a common complication in cancer patients. The aim of this study was to investigate whether radiation-induced intestinal injury could be alleviated by coniferyl aldehyde (CA), an HSF1-inducing agent that increases cellular HSP70 expression. We systemically administered CA to mice with radiation enteropathy following abdominal irradiation (IR) to demonstrate the protective effects of CA against radiation-induced gastrointestinal injury. CA clearly alleviated acute radiation-induced intestinal damage, as reflected by the histopathological data and it also attenuated sub-acute enteritis. CA prevented intestinal crypt cell death and protected the microvasculature in the lamina propria during the acute and sub-acute phases of damage. CA induced HSF1 and HSP70 expression in both intestinal epithelial cells and endothelial cells in vitro. Additionally, CA protected against not only the apoptotic cell death of both endothelial and epithelial cells but also the loss of endothelial cell function following IR, indicating that CA has beneficial effects on the intestine. Our results provide novel insight into the effects of CA and suggest its role as a therapeutic candidate for radiation-induced enteropathy due to its ability to promote rapid re-proliferation of the intestinal epithelium by the synergic effects of the inhibition of cell death and the promotion of endothelial cell function. PMID:26029925

  20. Ethylene Biosynthesis Is Promoted by Very-Long-Chain Fatty Acids during Lysigenous Aerenchyma Formation in Rice Roots.

    PubMed

    Yamauchi, Takaki; Shiono, Katsuhiro; Nagano, Minoru; Fukazawa, Aya; Ando, Miho; Takamure, Itsuro; Mori, Hitoshi; Nishizawa, Naoko K; Kawai-Yamada, Maki; Tsutsumi, Nobuhiro; Kato, Kiyoaki; Nakazono, Mikio

    2015-09-01

    In rice (Oryza sativa) roots, lysigenous aerenchyma, which is created by programmed cell death and lysis of cortical cells, is constitutively formed under aerobic conditions, and its formation is further induced under oxygen-deficient conditions. Ethylene is involved in the induction of aerenchyma formation. reduced culm number1 (rcn1) is a rice mutant in which the gene encoding the ATP-binding cassette transporter RCN1/OsABCG5 is defective. Here, we report that the induction of aerenchyma formation was reduced in roots of rcn1 grown in stagnant deoxygenated nutrient solution (i.e. under stagnant conditions, which mimic oxygen-deficient conditions in waterlogged soils). 1-Aminocyclopropane-1-carboxylic acid synthase (ACS) is a key enzyme in ethylene biosynthesis. Stagnant conditions hardly induced the expression of ACS1 in rcn1 roots, resulting in low ethylene production in the roots. Accumulation of saturated very-long-chain fatty acids (VLCFAs) of 24, 26, and 28 carbons was reduced in rcn1 roots. Exogenously supplied VLCFA (26 carbons) increased the expression level of ACS1 and induced aerenchyma formation in rcn1 roots. Moreover, in rice lines in which the gene encoding a fatty acid elongase, CUT1-LIKE (CUT1L; a homolog of the gene encoding Arabidopsis CUT1, which is required for cuticular wax production), was silenced, both ACS1 expression and aerenchyma formation were reduced. Interestingly, the expression of ACS1, CUT1L, and RCN1/OsABCG5 was induced predominantly in the outer part of roots under stagnant conditions. These results suggest that, in rice under oxygen-deficient conditions, VLCFAs increase ethylene production by promoting 1-aminocyclopropane-1-carboxylic acid biosynthesis in the outer part of roots, which, in turn, induces aerenchyma formation in the root cortex.

  1. Ethylene Biosynthesis Is Promoted by Very-Long-Chain Fatty Acids during Lysigenous Aerenchyma Formation in Rice Roots1

    PubMed Central

    Yamauchi, Takaki; Shiono, Katsuhiro; Nagano, Minoru; Fukazawa, Aya; Ando, Miho; Takamure, Itsuro; Mori, Hitoshi; Nishizawa, Naoko K.; Kawai-Yamada, Maki; Tsutsumi, Nobuhiro; Kato, Kiyoaki; Nakazono, Mikio

    2015-01-01

    In rice (Oryza sativa) roots, lysigenous aerenchyma, which is created by programmed cell death and lysis of cortical cells, is constitutively formed under aerobic conditions, and its formation is further induced under oxygen-deficient conditions. Ethylene is involved in the induction of aerenchyma formation. reduced culm number1 (rcn1) is a rice mutant in which the gene encoding the ATP-binding cassette transporter RCN1/OsABCG5 is defective. Here, we report that the induction of aerenchyma formation was reduced in roots of rcn1 grown in stagnant deoxygenated nutrient solution (i.e. under stagnant conditions, which mimic oxygen-deficient conditions in waterlogged soils). 1-Aminocyclopropane-1-carboxylic acid synthase (ACS) is a key enzyme in ethylene biosynthesis. Stagnant conditions hardly induced the expression of ACS1 in rcn1 roots, resulting in low ethylene production in the roots. Accumulation of saturated very-long-chain fatty acids (VLCFAs) of 24, 26, and 28 carbons was reduced in rcn1 roots. Exogenously supplied VLCFA (26 carbons) increased the expression level of ACS1 and induced aerenchyma formation in rcn1 roots. Moreover, in rice lines in which the gene encoding a fatty acid elongase, CUT1-LIKE (CUT1L; a homolog of the gene encoding Arabidopsis CUT1, which is required for cuticular wax production), was silenced, both ACS1 expression and aerenchyma formation were reduced. Interestingly, the expression of ACS1, CUT1L, and RCN1/OsABCG5 was induced predominantly in the outer part of roots under stagnant conditions. These results suggest that, in rice under oxygen-deficient conditions, VLCFAs increase ethylene production by promoting 1-aminocyclopropane-1-carboxylic acid biosynthesis in the outer part of roots, which, in turn, induces aerenchyma formation in the root cortex. PMID:26036614

  2. Formic acid and acetic acid induce a programmed cell death in pathogenic Candida species.

    PubMed

    Lastauskienė, Eglė; Zinkevičienė, Auksė; Girkontaitė, Irutė; Kaunietis, Arnoldas; Kvedarienė, Violeta

    2014-09-01

    Cutaneous fungal infections are common and widespread. Antifungal agents used for the treatment of these infections often have undesirable side effects. Furthermore, increased resistance of the microorganisms to the antifungal drugs becomes the growing problem. Accordingly, the search for natural antifungal compounds continues to receive attention. Apoptosis is highly regulated programmed cell death. During yeast cell apoptosis, amino acids and peptides are released and can stimulate regeneration of human epithelium cells. Thus, detection of chemical compounds inducing apoptosis in yeast and nontoxic for humans is of great medical relevance. The aim of this study was to detect chemical compound inducing apoptosis in pathogenic Candida species with the lowest toxicity to the mammalian cells. Five chemical compounds--acetic acid, sodium bicarbonate, potassium carbonate, lithium acetate, and formic acid--were tested for evaluation of antifungal activity on C. albicans, C. guilliermondii, and C. lusitaniae. The results showed that acetic acid and formic acid at the lowest concentrations induced yeast cells death. Apoptosis analysis revealed that cells death was accompanied by activation of caspase. Minimal inhibitory concentrations of potassium carbonate and sodium bicarbonate induced Candida cells necrosis. Toxicity test with mammalian cell cultures showed that formic acid has the lowest effect on the growth of Jurkat and NIH 3T3 cells. In conclusion, our results show that a low concentration of formic acid induces apoptosis-like programmed cell death in the Candida yeast and has a minimal effect on the survivability of mammalian cells, suggesting potential applications in the treatment of these infections. PMID:24752490

  3. White-to-brite conversion in human adipocytes promotes metabolic reprogramming towards fatty acid anabolic and catabolic pathways

    PubMed Central

    Barquissau, V.; Beuzelin, D.; Pisani, D.F.; Beranger, G.E.; Mairal, A.; Montagner, A.; Roussel, B.; Tavernier, G.; Marques, M.-A.; Moro, C.; Guillou, H.; Amri, E.-Z.; Langin, D.

    2016-01-01

    Objective Fat depots with thermogenic activity have been identified in humans. In mice, the appearance of thermogenic adipocytes within white adipose depots (so-called brown-in-white i.e., brite or beige adipocytes) protects from obesity and insulin resistance. Brite adipocytes may originate from direct conversion of white adipocytes. The purpose of this work was to characterize the metabolism of human brite adipocytes. Methods Human multipotent adipose-derived stem cells were differentiated into white adipocytes and then treated with peroxisome proliferator-activated receptor (PPAR)γ or PPARα agonists between day 14 and day 18. Gene expression profiling was determined using DNA microarrays and RT-qPCR. Variations of mRNA levels were confirmed in differentiated human preadipocytes from primary cultures. Fatty acid and glucose metabolism was investigated using radiolabelled tracers, Western blot analyses and assessment of oxygen consumption. Pyruvate dehydrogenase kinase 4 (PDK4) knockdown was achieved using siRNA. In vivo, wild type and PPARα-null mice were treated with a β3-adrenergic receptor agonist (CL316,243) to induce appearance of brite adipocytes in white fat depot. Determination of mRNA and protein levels was performed on inguinal white adipose tissue. Results PPAR agonists promote a conversion of white adipocytes into cells displaying a brite molecular pattern. This conversion is associated with transcriptional changes leading to major metabolic adaptations. Fatty acid anabolism i.e., fatty acid esterification into triglycerides, and catabolism i.e., lipolysis and fatty acid oxidation, are increased. Glucose utilization is redirected from oxidation towards glycerol-3-phophate production for triglyceride synthesis. This metabolic shift is dependent on the activation of PDK4 through inactivation of the pyruvate dehydrogenase complex. In vivo, PDK4 expression is markedly induced in wild-type mice in response to CL316,243, while this increase is blunted

  4. A strategy for promoting astaxanthin accumulation in Haematococcus pluvialis by 1-aminocyclopropane-1-carboxylic acid application.

    PubMed

    Lee, Changsu; Choi, Yoon-E; Yun, Yeoung-Sang

    2016-10-20

    The green algae Haematococcus pluvialis is a freshwater unicellular microalga belonging to Chlorophyceae. It is one of the best natural sources of astaxanthin, a secondary metabolite commonly used as an antioxidant and anti-inflammatory agent. Due to the importance of astaxanthin, various efforts have been made to increase its production. In this study, we attempted to develop a strategy for promoting astaxanthin accumulation in H. pluvialis using 1-aminocyclopropane-1-carboxylic acid (ACC), a precursor of ethylene (normally known as an aging hormone in plants). Our results demonstrated that ACC could enhance the growth of H. pluvialis, thereby promoting astaxanthin accumulation. Therefore, ACC has an indirect influence on astaxanthin production. We further verified the effect of ACC with a direct treatment of ethylene originated from banana peels. These results indicate that ethylene could be applied as an indirect method for enhancing growth and astaxanthin biosynthesis in H. pluvialis. PMID:27544287

  5. Dentin Matrix Protein-1 Isoforms Promote Differential Cell Attachment and Migration*S⃞

    PubMed Central

    von Marschall, Zofia; Fisher, Larry W.

    2008-01-01

    Dentin matrix protein-1 (DMP1), bone sialoprotein (BSP), and osteopontin (OPN) are three SIBLINGs (small integrin-binding ligand, N-linked glycoproteins) co-expressed/secreted by skeletal and active ductal epithelial cells. Although etiological mechanisms remain unclear, DMP1 is the only one of these three genes currently known to have mutations resulting in human disease, and yet it remains the least studied. All three contain the highly conserved integrin-binding tripeptide, RGD, and experiments comparing the cell attachment and haptotactic migration-enhancing properties of DMP1 to BSP and OPN were performed using human skeletal (MG63 and primary dental pulp cells) and salivary gland (HSG) cells. Mutation of any SIBLING's RGD destroyed all attachment and migration activity. Using itsαVβ5 integrin, HSG cells attached to BSP but not to DMP1 or OPN. However, HSG cells could not migrate onto BSP in a modified Boyden chamber assay. Expression of αVβ3 integrin enhanced HSG attachment to DMP1 and OPN and promoted haptotactic migration onto all three proteins. Interchanging the first four coding exons or the conserved amino acids adjacent to the RGD of DMP1 with corresponding sequences of BSP did not enhance the ability of DMP1 to bindαVβ5. For αVβ3-expressing cells, intact DMP1, its BMP1-cleaved C-terminal fragment, and exon six lacking all post-translational modifications worked equally well but the proteoglycan isoform of DMP1 had greatly reduced ability for cell attachment and migration. The sequence specificity of the proposed BMP1-cleavage site of DMP1 was verified by mutation analysis. Direct comparison of the three proteins showed that cells discriminate among these SIBLINGs and among DMP1 isoforms. PMID:18819913

  6. Vectofusin-1, a new viral entry enhancer, strongly promotes lentiviral transduction of human hematopoietic stem cells.

    PubMed

    Fenard, David; Ingrao, Dina; Seye, Ababacar; Buisset, Julien; Genries, Sandrine; Martin, Samia; Kichler, Antoine; Galy, Anne

    2013-05-07

    Gene transfer into hCD34(+) hematopoietic stem/progenitor cells (HSCs) using human immunodeficiency virus type 1 (HIV-1)-based lentiviral vectors (LVs) has several promising therapeutic applications. Yet, efficiency, safety, and cost of LV gene therapy could be ameliorated by enhancing target cell transduction levels and reducing the amount of LV used on the cells. Several transduction enhancers already exist such as fibronectin fragments and cationic compounds, but all present limitations. In this study, we describe a new transduction enhancer called Vectofusin-1, which is a short cationic peptide, active on several LV pseudotypes. Vectofusin-1 is used as a soluble additive to safely increase the frequency of transduced HSCs and to augment the level of transduction to one or two copies of vector per cell in a vector dose-dependent manner. Vectofusin-1 acts at the entry step by promoting the adhesion and the fusion between viral and cellular membranes. Vectofusin-1 is therefore a promising additive that could significantly ameliorate hCD34(+) cell-based gene therapy.Molecular Therapy-Nucleic Acids (2013) 2, e90; doi:10.1038/mtna.2013.17; published online 7 May 2013.

  7. ABSCISIC ACID-INSENSITIVE 4 negatively regulates flowering through directly promoting Arabidopsis FLOWERING LOCUS C transcription

    PubMed Central

    Shu, Kai; Chen, Qian; Wu, Yaorong; Liu, Ruijun; Zhang, Huawei; Wang, Shengfu; Tang, Sanyuan; Yang, Wenyu; Xie, Qi

    2016-01-01

    During the life cycle of a plant, one of the major biological processes is the transition from the vegetative to the reproductive stage. In Arabidopsis, flowering time is precisely controlled by extensive environmental and internal cues. Gibberellins (GAs) promote flowering, while abscisic acid (ABA) is considered as a flowering suppressor. However, the detailed mechanism through which ABA inhibits the floral transition is poorly understood. Here, we report that ABSCISIC ACID-INSENSITIVE 4 (ABI4), a key component in the ABA signalling pathway, negatively regulates floral transition by directly promoting FLOWERING LOCUS C (FLC) transcription. The abi4 mutant showed the early flowering phenotype whereas ABI4-overexpressing (OE-ABI4) plants had delayed floral transition. Consistently, quantitative reverse transcription–PCR (qRT–PCR) assay revealed that the FLC transcription level was down-regulated in abi4, but up-regulated in OE-ABI4. The change in FT level was consistent with the pattern of FLC expression. Chromatin immunoprecipitation-qPCR (ChIP-qPCR), electrophoretic mobility shift assay (EMSA), and tobacco transient expression analysis showed that ABI4 promotes FLC expression by directly binding to its promoter. Genetic analysis demonstrated that OE-ABI4::flc-3 could not alter the flc-3 phenotype. OE-FLC::abi4 showed a markedly delayed flowering phenotype, which mimicked OE-FLC::WT, and suggested that ABI4 acts upstream of FLC in the same genetic pathway. Taken together, these findings suggest that ABA inhibits the floral transition by activating FLC transcription through ABI4. PMID:26507894

  8. NLS-RARα promotes proliferation and inhibits differentiation in HL-60 cells.

    PubMed

    Hu, Xiu-Xiu; Zhong, Liang; Zhang, Xi; Gao, Yuan-Mei; Liu, Bei-Zhong

    2014-01-01

    A unique mRNA produced in leukemic cells from a t(15;17) acute promyelocytic leukemia (APL) patient encodes a fusion protein between the retinoic acid receptor α (RARα) and a myeloid gene product called PML. Studies have reported that neutrophil elastase (NE) cleaves bcr-1-derived PML-RARα in early myeloid cells, leaving only the nuclear localization signal (NLS) of PML attached to RARα. The resultant NLS-RARα fusion protein mainly localizes to, and functions within, the cell nucleus. It is speculated that NLS-RARα may act in different ways from the wild-type RARα, but its biological characteristics have not been reported. This study takes two approaches. Firstly, the NLS-RARα was silenced with pNLS-RARα-shRNA. The mRNA and protein expression of NLS-RARα were detected by RT-PCR and Western blot respectively. Cell proliferation in vitro was assessed by MTT assay. Flow cytometry (FCM) was used to detect the differentiation of cells. Secondly, the NLS-RARα was over-expressed by preparation of recombinant adenovirus HL-60/pAd-NLS-RARα. The assays of mRNA and protein expression of NLS-RARα, and cell proliferation, were as above. By contrast, cell differentiation was stimulated by all trans retinoic acid (ATRA) (2.5µmol/L) at 24h after virus infection of pAd-NLS-RARα, and then detected by CD11b labeling two days later. The transcription and translation of C-MYC was detected in HL-60/pAd-NLS-RARα cells which treated by ATRA. Our results showed that compared to the control groups, the expression of NLS-RARα was significantly reduced in the HL-60/pNLS-RARα-shRNA cells, and increased dramatically in the HL-60/pAd-NLS-RARα cells. The proliferation was remarkably inhibited in the HL-60/pNLS-RARα-shRNA cells in a time-dependent manner, but markedly promoted in the HL-60/pAd-NLS-RARα cells. FCM outcome revealed the differentiation increased in HL-60/pNLS-RARα-shRNA cells, and decreased in the HL-60/pAd-NLS-RARα cells treated with 2.5µmol/L ATRA. The

  9. Retinol oxidation to retinoic acid in human thyroid glandular cells.

    PubMed

    Taibi, Gennaro; Gueli, Maria Concetta; Nicotra, Concetta M A; Cocciadiferro, Letizia; Carruba, Giuseppe

    2014-12-01

    Abstract Retinoic acid is regarded as the retinol metabolite that controls proliferation and differentiation of epithelial cells. In the present study, we investigated the potential role of xanthine dehydrogenase (XDH) in retinoic acid biosynthesis in human thyroid glandular cells (HTGC). In particular, we observed that cellular retinoids binding proteins (CRBPs) are also implicated in the biosynthetic pathway leading to retinoic acid formation in primary cultures of HTGC, as we have already reported for human mammary epithelial cells (HMEC). After partial protein purification, the enzyme responsible for retinoic acid biosynthesis was identified and quantified as XDH by immunoassay, by its ability to oxidize xanthine to uric acid and its sensitivity to the inhibitory effect of oxypurinol. The evidence of XDH-driven formation of retinoic acid in HTGC cultures further corroborates the potential role of XDH in retinoic acid biosynthesis in the epithelia. PMID:24506204

  10. Coordinated Regulation of the Neutral Amino Acid Transporter SNAT2 and the Protein Phosphatase Subunit GADD34 Promotes Adaptation to Increased Extracellular Osmolarity*

    PubMed Central

    Krokowski, Dawid; Jobava, Raul; Guan, Bo-Jhih; Farabaugh, Kenneth; Wu, Jing; Majumder, Mithu; Bianchi, Massimiliano G.; Snider, Martin D.; Bussolati, Ovidio; Hatzoglou, Maria

    2015-01-01

    Cells respond to shrinkage induced by increased extracellular osmolarity via programmed changes in gene transcription and mRNA translation. The immediate response to this stress includes the induction of expression of the neutral amino acid transporter SNAT2. Increased SNAT2-mediated uptake of neutral amino acids is an essential adaptive mechanism for restoring cell volume. In contrast, stress-induced phosphorylation of the α subunit of the translation initiation factor eIF2 (eIF2α) can promote apoptosis. Here we show that the response to mild hyperosmotic stress involves regulation of the phosphorylation of eIF2α by increased levels of GADD34, a regulatory subunit of protein phosphatase 1 (PP1). The induction of GADD34 was dependent on transcriptional control by the c-Jun-binding cAMP response element in the GADD34 gene promoter and posttranscriptional stabilization of its mRNA. This mechanism differs from the regulation of GADD34 expression by other stresses that involve activating transcription factor 4 (ATF4). ATF4 was not translated during hyperosmotic stress despite an increase in eIF2α phosphorylation. The SNAT2-mediated increase in amino acid uptake was enhanced by increased GADD34 levels in a manner involving decreased eIF2α phosphorylation. It is proposed that the induction of the SNAT2/GADD34 axis enhances cell survival by promoting the immediate adaptive response to stress. PMID:26041779

  11. Endothelial cells mitigate DNA damage and promote the regeneration of hematopoietic stem cells after radiation injury.

    PubMed

    Zachman, Derek K; Leon, Ronald P; Das, Prerna; Goldman, Devorah C; Hamlin, Kimberly L; Guha, Chandan; Fleming, William H

    2013-11-01

    Endothelial cells (ECs) are an essential component of the hematopoietic microenvironment, which maintains and regulates hematopoietic stem cells (HSCs). Although ECs can support the regeneration of otherwise lethally-irradiated HSCs, the mechanisms are not well understood. To further understand this phenomenon, we studied HSC regeneration from irradiated bone marrow using co-culture with human aortic ECs (HAECs). Co-culture with HAECs induced a 24-fold expansion of long-term HSCs (CD150(+), lineage(lo), Sca-1(+), c-Kit(+); CD150(+)LSK cells) in vitro. These cells gave rise to functional hematopoietic stem and progenitor cells (HSPCs) with colony-forming activity, multilineage reconstitution and serial transplantation potential. Furthermore, HAECs significantly reduced DNA damage in irradiated LSK cells within 24h. Remarkably, we were able to delay the exposure of irradiated bone marrow to the regenerative, HAEC-derived signals for up to 48h and still rescue functional HSCs. G-CSF is the gold standard for promoting hematopoietic regeneration in vivo. However, when compared to HAECs, in vitro G-CSF treatment promoted lineage differentiation and regenerated 5-fold fewer CD150(+)LSK cells. Together, our results show that HAECs are powerful, direct mitigators of HSC injury and DNA damage. Identification of the HAEC-derived factors that rescue HSCs may lead to improved therapies for hematopoietic regeneration after radiation injury. PMID:23939266

  12. Pitx2 expression promotes p21 expression and cell cycle exit in neural stem cells.

    PubMed

    Heldring, Nina; Joseph, Bertrand; Hermanson, Ola; Kioussi, Chrissa

    2012-11-01

    Cortical development is a complex process that involves many events including proliferation, cell cycle exit and differentiation that need to be appropriately synchronized. Neural stem cells (NSCs) isolated from embryonic cortex are characterized by their ability of self-renewal under continued maintenance of multipotency. Cell cycle progression and arrest during development is regulated by numerous factors, including cyclins, cyclin dependent kinases and their inhibitors. In this study, we exogenously expressed the homeodomain transcription factor Pitx2, usually expressed in postmitotic progenitors and neurons of the embryonic cortex, in NSCs with low expression of endogenous Pitx2. We found that Pitx2 expression induced a rapid decrease in proliferation associated with an accumulation of NSCs in G1 phase. A search for potential cell cycle inhibitors responsible for such cell cycle exit of NSCs revealed that Pitx2 expression caused a rapid and dramatic (≉20-fold) increase in expression of the cell cycle inhibitor p21 (WAF1/Cip1). In addition, Pitx2 bound directly to the p21 promoter as assessed by chromatin immunoprecipitation (ChIP) in NSCs. Surprisingly, Pitx2 expression was not associated with an increase in differentiation markers, but instead the expression of nestin, associated with undifferentiated NSCs, was maintained. Our results suggest that Pitx2 promotes p21 expression and induces cell cycle exit in neural progenitors.

  13. [Envelope protein of Jaagsiekte sheep retrovious expressed in NIH3T3 cells promotes cell proliferation].

    PubMed

    DU, Fangyuan; Chen, Dayong; Zhang, Yufei; Sun, Xiaolin; Guo, Wenqing; Liu, Shuying

    2016-09-01

    Objective To explore the influence of the exogenous Jaagsiekte sheep retrovious (exJSRV) envelope protein (Env) on NIH3T3 cell proliferation. Methods A recombinant plasmid pcDNA4/myc-His/exJSRV- env carrying exJSRV- env gene was constructed, and then the correctness of the recombinant plasmid was identified by PCR, restriction enzyme digestion and sequencing. The recombinant plasmid pcDNA4/myc-His/exJSRV- env was transiently transfected into NIH3T3 cells by Lipofectamine(TM) LTX. After the transfection of the recombinant plasmid, the expression of exJSRV- env was detected by reverse transcription PCR and Western blotting. The effect of Env on cell proliferation was investigated by CCK-8 assay and plate colony formation assay. Results The recombinant eukaryotic expression plasmid containing exJSRV- env was successfully constructed as identified by PCR, restriction enzyme identification and sequencing. After the recombinant plasmid was transiently transfected into NIH3T3 cells, reverse transcription PCR and Western blotting showed the expression of exJSRV- env , and Env promoted NIH3T3 cell proliferation significantly. Conclusion JSRV Env was expressed successfully in the NIH3T3 cells and promoted the proliferation of NIH3T3 cells. PMID:27609573

  14. Chronic intermittent psychological stress promotes macrophage reverse cholesterol transport by impairing bile acid absorption in mice

    PubMed Central

    Silvennoinen, Reija; Quesada, Helena; Kareinen, Ilona; Julve, Josep; Kaipiainen, Leena; Gylling, Helena; Blanco-Vaca, Francisco; Escola-Gil, Joan Carles; Kovanen, Petri T; Lee-Rueckert, Miriam

    2015-01-01

    Psychological stress is a risk factor for atherosclerosis, yet the pathophysiological mechanisms involved remain elusive. The transfer of cholesterol from macrophage foam cells to liver and feces (the macrophage-specific reverse cholesterol transport, m-RCT) is an important antiatherogenic pathway. Because exposure of mice to physical restraint, a model of psychological stress, increases serum levels of corticosterone, and as bile acid homeostasis is disrupted in glucocorticoid-treated animals, we investigated if chronic intermittent restraint stress would modify m-RCT by altering the enterohepatic circulation of bile acids. C57Bl/6J mice exposed to intermittent stress for 5 days exhibited increased transit through the large intestine and enhanced fecal bile acid excretion. Of the transcription factors and transporters that regulate bile acid homeostasis, the mRNA expression levels of the hepatic farnesoid X receptor (FXR), the bile salt export pump (BSEP), and the intestinal fibroblast growth factor 15 (FGF15) were reduced, whereas those of the ileal apical sodium-dependent bile acid transporter (ASBT), responsible for active bile acid absorption, remained unchanged. Neither did the hepatic expression of cholesterol 7α-hydroxylase (CYP7A1), the key enzyme regulating bile acid synthesis, change in the stressed mice. Evaluation of the functionality of the m-RCT pathway revealed increased fecal excretion of bile acids that had been synthesized from macrophage-derived cholesterol. Overall, our study reveals that chronic intermittent stress in mice accelerates m-RCT specifically by increasing fecal excretion of bile acids. This novel mechanism of m-RCT induction could have antiatherogenic potential under conditions of chronic stress. PMID:25969465

  15. Chrysophanic Acid Induces Necrosis but not Necroptosis in Human Renal Cell Carcinoma Caki-2 Cells

    PubMed Central

    Choi, Joon-Seok

    2016-01-01

    Background: Chrysophanic acid, also known as chrysophanol, has a number of biological activities. It enhances memory and learning abilities, raises superoxide dismutase activity, and has anti-cancer effects in several model systems. According to previous reports, chrysophanic acid-induced cell death shares features of necrotic cell death. However, the molecular and cellular processes underlying chrysophanic acid-induced cell death remain poorly understood. Methods: Chrysophanic acid-induced cell death was monitored by cell viability assay and Annexin V-propidium iodide (PI) staining of renal cell carcinoma Caki-2 cells. The induction of intracellular reactive oxygen species (ROS) by chrysophanic acid and the suppression of ROS by anti-oxidants were evaluated by 2′,7′-dichlorofluorescin diacetate staining. The expression and phosphorylation of proteins that are involved in apoptosis and necroptosis were detected by immunoblotting. Results: The extent of chrysophanic acid-induced cell death was concentration and time dependent, and dead cells mainly appeared in the PI-positive population, which is a major feature of necrosis, upon fluorescence-activated cell sorting analysis. Chrysophanic acid-induced cell death was associated with the generation of intracellular ROS, and this effect was reversed by pretreatment with N-acetyl cysteine. Chrysophanic acid-induced cell death was not associated with changes in apoptotic or necroptotic marker proteins. Conclusions: The cell death induced by chrysophanic acid resembled neither apoptotic nor necroptotic cell death in human renal cell carcinoma Caki-2 cells. PMID:27390736

  16. Interstitial flows promote an amoeboid cell phenotype and motility of breast cancer cells

    NASA Astrophysics Data System (ADS)

    Tung, Chih-Kuan; Huang, Yu Ling; Zheng, Angela; Wu, Mingming

    2015-03-01

    Lymph nodes, the drainage systems for interstitial flows, are clinically known to be the first metastatic sites of many cancer types including breast and prostate cancers. Here, we demonstrate that breast cancer cell morphology and motility is modulated by interstitial flows in a cell-ECM adhesion dependent manner. The average aspect ratios of the cells are significantly lower (or are more amoeboid like) in the presence of the flow in comparison to the case when the flow is absent. The addition of exogenous adhesion molecules within the extracellular matrix (type I collagen) enhances the overall aspect ratio (or are more mesenchymal like) of the cell population. Using measured cell trajectories, we find that the persistence of the amoeboid cells (aspect ratio less than 2.0) is shorter than that of mesenchymal cells. However, the maximum speed of the amoeboid cells is larger than that of mesenchymal cells. Together these findings provide the novel insight that interstitial flows promote amoeboid cell morphology and motility and highlight the plasticity of tumor cell motility in response to its biophysical environment. Supported by NIH Grant R21CA138366.

  17. Colorimetric determination of phosphoric acid leakage for phosphoric acid-doped polybenzimidazole membrane fuel cell applications

    NASA Astrophysics Data System (ADS)

    Jeong, Yeon Hun; Jung, Ju Hae; Choi, Euiji; Han, Seungyoon; Begley, Alina Irene; Yoo, Sung Jong; Jang, Jong Hyun; Kim, Hyoung-Juhn; Nam, Suk Woo; Lee, Kwan-Young; Kim, Jin Young

    2015-12-01

    A simple and precise colorimetric method for analyzing phosphoric acid leakage in phosphoric acid-doped polybenzimidazole membrane fuel cells is described. The developed method is based on the colorimetric determination from a rapid formation of molybdenum blue color by the reduction reaction of molybdate ions in the presence of phosphoric acid in the acidic medium. The color is stable up to a few months and can be used for the sensitive and accurate detection of phosphoric acid electrolyte which is discharged from the fuel cell during operation. Tests with a wide concentration range of phosphate compounds showed that it permits determination of phosphoric acid up to nanogram quantities. The developed detection method assists monitoring the phosphoric acid contents and developing stable operation strategies of fuel cells.

  18. Arsenic promotes centrosome abnormalities and cell colony formation in p53 compromised human lung cells

    SciTech Connect

    Liao Weiting; Lin Pinpin; Cheng, T.-S.; Yu, H.-S.; Chang, Louis W.

    2007-12-01

    Epidemiological evidence indicated that residents, especially cigarette smokers, in arseniasis areas had significantly higher lung cancer risk than those living in non-arseniasis areas. Thus, an interaction between arsenic and cigarette smoking in lung carcinogenesis was suspected. p53 dysfunction or mutation in lung epithelial cells was frequently observed in cigarette smokers. Our present study was to explore the differential effects by arsenic on H1355 cells (human lung adenocarcinoma cell line with mutation in p53), BEAS-2B (immortalized lung epithelial cell with functional p53) and pifithrin-{alpha}-treated BEAS-2B cells (p53-inhibited cells). These cells were treated with different doses of sodium arsenite (0, 0.1, 1, 5 and 10 {mu}M) for 48 h. A greater reduction in cell viability was observed in the BEAS-2B cells vs. p53 compromised cells (H1355 or p53-inhibited BEAS-2B). Similar observation was also made on 7-day cell survival (growth) study. TUNEL analysis confirmed that there was indeed a significantly reduced arsenite-induced apoptosis found in p53-compromised cells. Centrosomal abnormality has been attributed to eventual chromosomal missegregation, aneuploidy and tumorigenesis. In our present study, reduced p21 and Gadd45a expressions and increased centrosomal abnormality (atopic and multiple centrosomes) were observed in both arsenite-treated H1355 and p53-inhibited BEAS-2B cells as compared with similarly treated BEAS-2B cells. Increased anchorage-independent growth (colony formation) of BEAS-2B cells co-treated with pifithrin-{alpha} and 5 {mu}M sodium arsenite was also observed in soft agar. Our present investigation demonstrated that arsenic would act specifically on p53 compromised cells (either with p53 dysfunction or inhibited) to induce centrosomal abnormality and colony formation. These findings provided strong evidence on the carcinogenic promotional role of arsenic, especially under the condition of p53 dysfunction.

  19. 2,4-Dichlorophenoxyacetic acid promotes S-nitrosylation and oxidation of actin affecting cytoskeleton and peroxisomal dynamics.

    PubMed

    Rodríguez-Serrano, M; Pazmiño, D M; Sparkes, I; Rochetti, A; Hawes, C; Romero-Puertas, M C; Sandalio, L M

    2014-09-01

    2,4-Dichlorophenoxyacetic acid (2,4-D) is a synthetic auxin used as a herbicide to control weeds in agriculture. A high concentration of 2,4-D promotes leaf epinasty and cell death. In this work, the molecular mechanisms involved in the toxicity of this herbicide are studied by analysing in Arabidopsis plants the accumulation of reactive oxygen species (ROS) and nitric oxide (NO), and their effect on cytoskeleton structure and peroxisome dynamics. 2,4-D (23 mM) promotes leaf epinasty, whereas this process was prevented by EDTA, which can reduce ·OH accumulation. The analysis of ROS accumulation by confocal microscopy showed a 2,4-D-dependent increase in both H2O2 and O2·(-), whereas total NO was not affected by the treatment. The herbicide promotes disturbances on the actin cytoskeleton structure as a result of post-translational modification of actin by oxidation and S-nitrosylation, which could disturb actin polymerization, as suggested by the reduction of the F-actin/G-actin ratio. These effects were reduced by EDTA, and the reduction of ROS production in Arabidopsis mutants deficient in xanthine dehydrogenase (Atxdh) gave rise to a reduction in actin oxidation. Also, 2,4-D alters the dynamics of the peroxisome, slowing the speed and shortening the distances by which these organelles are displaced. It is concluded that 2,4-D promotes oxidative and nitrosative stress, causing disturbances in the actin cytoskeleton, thereby affecting the dynamics of peroxisomes and some other organelles such as the mitochondria, with xanthine dehydrogenase being involved in ROS production under these conditions. These structural changes in turn appear to be responsible for the leaf epinasty.

  20. Modulation of Phagosomal pH by Candida albicans Promotes Hyphal Morphogenesis and Requires Stp2p, a Regulator of Amino Acid Transport

    PubMed Central

    Vylkova, Slavena; Lorenz, Michael C.

    2014-01-01

    Candida albicans, the most important fungal pathogen of humans, has a unique interaction with macrophages in which phagocytosis induces a switch from the yeast to hyphal form, allowing it to escape by rupturing the immune cell. While a variety of factors induce this switch in vitro, including neutral pH, it is not clear what triggers morphogenesis within the macrophage where the acidic environment should inhibit this transition. In vitro, C. albicans grown in similar conditions in which amino acids are the primary carbon source generate large quantities of ammonia to raise the extracellular pH and induce the hyphal switch. We show here that C. albicans cells neutralize the macrophage phagosome and that neutral pH is a key inducer of germination in phagocytosed cells by using a mutant lacking STP2, a transcription factor that regulates the expression of multiple amino acid permeases, that is completely deficient in alkalinization in vitro. Phagocytosed stp2Δ mutant cells showed significant reduction in hypha formation and escaped from macrophages less readily compared to wild type cells; as a result stp2Δ mutant cells were killed at a higher rate and caused less damage to RAW264.7 macrophages. Stp2p-regulated import leads to alkalinization of the phagosome, since the majority of the wild type cells fail to co-localize with acidophilic dyes, whereas the stp2Δ mutant cells were located in acidic phagosomes. Furthermore, stp2Δ mutant cells were able to form hyphae and escape from neutral phagosomes, indicating that the survival defect in these cells was pH dependent. Finally, these defects are reflected in an attenuation of virulence in a mouse model of disseminated candidiasis. Altogether our results suggest that C. albicans utilizes amino acids to promote neutralization of the phagosomal pH, hyphal morphogenesis, and escape from macrophages. PMID:24626429

  1. Industry experience in promoting weekly iron-folic acid supplementation in the Philippines.

    PubMed

    Garcia, Josel; Datol-Barrett, Eva; Dizon, Maynilad

    2005-12-01

    After participating in a pilot project under a government-industry partnership to promote the adoption of weekly iron-folic acid supplementation among women of reproductive age in the Philippines in 1998, United Laboratories (UNILAB), the Philippines' largest private pharmaceutical company, decided in April 2002 to launch a weekly iron-folic acid supplement for pregnant and non-pregnant women under the brand name Femina. The business objective set for the Femina brand was to build the category of preventive iron-folic acid supplements in line with the Philippine Department of Health's advocacy on weekly supplementation as an alternate to daily dosing to reduce the prevalence of anemia in the country. The brand was supported with an integrated mix of traditional advertising media with complementary direct-to-consumer educational programs that aimed to create awareness of iron-deficiency anemia, its causes and effects, and the role of weekly intake of iron-folic acid in preventing the condition. Aggressive marketing support for 1 year was successful in creating awareness among the target women. Significant lessons derived from consumers identified opportunity areas that can be further addressed in developing advocacy programs on weekly iron supplementation implemented on a nationwide scale in the future.

  2. MLK3 promotes metabolic dysfunction induced by saturated fatty acid-enriched diet

    PubMed Central

    Gadang, Vidya; Kohli, Rohit; Myronovych, Andriy; Hui, David Y.; Perez-Tilve, Diego

    2013-01-01

    Saturated fatty acids activate the c-Jun NH2-terminal kinase (JNK) pathway, resulting in chronic low-grade inflammation and the development of insulin resistance. Mixed-lineage kinase 3 (MLK3) is a mitogen-activated protein kinase kinase kinase (MAP3K) that mediates JNK activation in response to saturated fatty acids in vitro; however, the exact mechanism for diet-induced JNK activation in vivo is not known. Here, we have used MLK3-deficient mice to examine the role of MLK3 in a saturated-fat diet model of obesity. MLK3-KO mice fed a high-fat diet enriched in medium-chain saturated fatty acids for 16 wk had decreased body fat compared with wild-type (WT) mice due to increased energy expenditure independently of food consumption and physical activity. Moreover, MLK3 deficiency attenuated palmitate-induced JNK activation and M1 polarization in bone marrow-derived macrophages in vitro, and obesity induced JNK activation, macrophage infiltration into adipose tissue, and expression of proinflammatory cytokines in vivo. In addition, loss of MLK3 improved insulin resistance and decreased hepatic steatosis. Together, these data demonstrate that MLK3 promotes saturated fatty acid-induced JNK activation in vivo and diet-induced metabolic dysfunction. PMID:23860122

  3. Industry experience in promoting weekly iron-folic acid supplementation in the Philippines.

    PubMed

    Garcia, Josel; Datol-Barrett, Eva; Dizon, Maynilad

    2005-12-01

    After participating in a pilot project under a government-industry partnership to promote the adoption of weekly iron-folic acid supplementation among women of reproductive age in the Philippines in 1998, United Laboratories (UNILAB), the Philippines' largest private pharmaceutical company, decided in April 2002 to launch a weekly iron-folic acid supplement for pregnant and non-pregnant women under the brand name Femina. The business objective set for the Femina brand was to build the category of preventive iron-folic acid supplements in line with the Philippine Department of Health's advocacy on weekly supplementation as an alternate to daily dosing to reduce the prevalence of anemia in the country. The brand was supported with an integrated mix of traditional advertising media with complementary direct-to-consumer educational programs that aimed to create awareness of iron-deficiency anemia, its causes and effects, and the role of weekly intake of iron-folic acid in preventing the condition. Aggressive marketing support for 1 year was successful in creating awareness among the target women. Significant lessons derived from consumers identified opportunity areas that can be further addressed in developing advocacy programs on weekly iron supplementation implemented on a nationwide scale in the future. PMID:16466091

  4. HIF-2α dependent lipid storage promotes endoplasmic reticulum homeostasis in clear cell renal cell carcinoma

    PubMed Central

    Qiu, Bo; Ackerman, Daniel; Sanchez, Danielle J.; Li, Bo; Ochocki, Joshua D.; Grazioli, Alison; Bobrovnikova-Marjon, Ekaterina; Diehl, J. Alan; Keith, Brian; Simon, M. Celeste

    2015-01-01

    Two hallmarks of clear cell renal cell carcinoma (ccRCC) are constitutive hypoxia inducible factor (HIF) signaling and abundant intracellular lipid droplets (LDs). However, regulation of lipid storage and its role in ccRCC are incompletely understood. Transcriptional profiling of primary ccRCC samples revealed that expression of the LD coat protein gene PLIN2 was elevated in tumors and correlated with HIF-2α, but not HIF-1α, activation. HIF-2α dependent PLIN2 expression promoted lipid storage, proliferation, and viability in xenograft tumors. Mechanistically, lipid storage maintained integrity of the endoplasmic reticulum (ER), which is functionally and physically associated with LDs. Specifically, PLIN2 dependent lipid storage suppressed cytotoxic ER stress responses that otherwise result from elevated protein synthetic activity characteristic of ccRCC cells. Thus, in addition to promoting ccRCC proliferation and anabolic metabolism, HIF-2α modulates lipid storage to sustain ER homeostasis, particularly under conditions of nutrient and oxygen limitation, thereby promoting tumor cell survival. PMID:25829424

  5. Synthesis of ortho acid ester-type 1,3-dioxolanofullerenes: radical reaction of [60]fullerene with halocarboxylic acids promoted by lead(IV) acetate.

    PubMed

    You, Xun; Li, Fa-Bao; Wang, Guan-Wu

    2014-11-21

    A lead(IV) acetate-promoted radical reaction of [60]fullerene with halocarboxylic acids has been exploited to synthesize rare ortho acid ester-type 1,3-dioxolanofullerenes, the hydroxyl group of which can be further transformed to an ester or ether group. Intriguingly, an ortho acid ester-type 1,3-dioxolanofullerene can also be converted to a 1,4-dioxanonofullerene in the presence of a base or manipulated to another ortho acid ester-type 1,3-dioxolanofullerene by reaction with a stronger halocarboxylic acid. Moreover, two possible reaction pathways leading to the observed products are also proposed.

  6. Epstein-Barr virus EBNA3C represses Cp, the major promoter for EBNA expression, but has no effect on the promoter of the cell gene CD21.

    PubMed Central

    Radkov, S A; Bain, M; Farrell, P J; West, M; Rowe, M; Allday, M J

    1997-01-01

    EBNA3C is a potent repressor of transcription when bound to DNA as a fusion with the DNA binding domain (DBD) of GALA. A survey of promoters has revealed that the wild-type, unfused EBNA3C can specifically repress expression from reporter plasmids containing the Epstein-Barr virus Cp latency-associated promoter. Repression of Cp activity required amino acids 207 to 368, which encompasses a region resembling a basic DBD adjacent to a leucine zipper DNA binding motif and a site which binds to the cellular factor CBF1/RBP-Jkappa. However, amino acids 207 to 368 are dispensable when the protein is bound to DNA as a fusion with the GAL4 DBD, thus implicating this region in DNA binding. Mutation of the CBF1/RBP-Jkappa binding site in EBNA3C abrogated repression, strongly suggesting that CBF1/RBP-Jkappa is necessary for targeting the viral protein to Cp. Consistent with this result, mutation of the EBNA2 response element (a CBF1/RBP-Jkappa binding site) in Cp also prevented significant repression. In addition, amino acids 346 to 543, which were previously defined as important for the repressor activity of the GAL4-EBNA3C fusion proteins, also appear to be necessary for the repression of Cp. Since repression by these fusions was not observed in all cell types, it seems likely that EBNA3C either depends on a corepressor which may interact with amino acids 346 to 543 or is modified in a cell-specific manner in order to repress. These data are consistent with EBNA3C contributing to the regulation of EBNA expression in latently infected B cells through CBF1/RBP-Jkappa and another factor, but this need not directly involve EBNA2. Finally, although it has been reported that EBNA3C can upregulate CD21 in some B cells, we were unable to demonstrate any effect of EBNA3C on reporter plasmids which contain the CD21 promoter. PMID:9343213

  7. Methods for promoting wound healing and muscle regeneration with the cell signaling protein Nell1

    SciTech Connect

    Culiat, Cymbeline T.

    2011-03-22

    The present invention provides methods for promoting wound healing and treating muscle atrophy in a mammal in need. The method comprises administering to the mammal a Nell1 protein or a Nell1 nucleic acid molecule.

  8. Methods for promoting wound healing and muscle regeneration with the cell signaling protein Nell1

    SciTech Connect

    Culiat, Cymbeline T

    2014-11-04

    The present invention provides methods for promoting wound healing and treating muscle atrophy in a mammal in need. The method comprises administering to the mammal a Nell1 protein or a Nell1 nucleic acid molecule.

  9. Methylation of the ATM promoter in glioma cells alters ionizing radiation sensitivity

    SciTech Connect

    Roy, Kanaklata; Wang, Lilin; Makrigiorgos, G. Mike; Price, Brendan D. . E-mail: brendan_price@dfci.harvard.edu

    2006-06-09

    Glioblastomas are among the malignancies most resistant to radiation therapy. In contrast, cells lacking the ATM protein are highly sensitive to ionizing radiation. The relationship between ATM protein expression and radiosensitivity in 3 glioma cell lines was examined. T98G cells exhibited normal levels of ATM protein, whereas U118 and U87 cells had significantly lower levels of ATM and increased (>2-fold) sensitivity to ionizing radiation compared to T98G cells. The ATM promoter was methylated in U87 cells. Demethylation by azacytidine treatment increased ATM protein levels in the U87 cells and decreased their radiosensitivity. In contrast, the ATM promoter in U118 cells was not methylated. Further, expression of exogenous ATM did not significantly alter the radiosensitivity of U118 cells. ATM expression is therefore heterogeneous in the glioma cells examined. In conclusion, methylation of the ATM promoter may account for the variable radiosensitivity and heterogeneous ATM expression in a fraction of glioma cells.

  10. miR-411 regulated ITCH expression and promoted cell proliferation in human hepatocellular carcinoma cells.

    PubMed

    Xia, Kunkun; Zhang, Yi; Cao, Shengli; Wu, Yang; Guo, Wenzhi; Yuan, Weitang; Zhang, Shuijun

    2015-03-01

    Hepatocellular carcinoma (HCC) is one of the most common human malignancies and the third most common cause of cancer mortality in the world. In this study, we report that miR-411 expression is markedly upregulated in HCC cells and HCC tissues compared with normal control tissues and cells. Previous studies have shown that miR-411 plays a crucial role in a variety of biological processes in various human cancer cells. However, the specific function of miR-411 in HCC remains unclear. Ectopic expression of miR-411 promoted the proliferation and anchorage-independent growth of HCC cells, whereas inhibition of miR-411 reduced this effect. Bioinformatics analysis further revealed ITCH, a putative tumor suppressor as a potential target of miR-411. Data from luciferase reporter assays showed that miR-411 directly binds to the 3'-untranslated region (3'-UTR) of ITCH mRNA and repressed expression at both transcriptional and translational levels. In functional assays, miR-411 promoted HCC cell proliferation, which could be suppressed by miR-411-in. Taken together, our data provide convincing evidence that miR-411 functions as an onco-miRNA, which was associated with cell proliferation of HCC, and its oncogenic effect is mediated chiefly through direct suppression of ITCH expression.

  11. HIV-1 Nef promotes the localization of Gag to the cell membrane and facilitates viral cell-to-cell transfer

    PubMed Central

    2013-01-01

    Background Newly synthesized HIV-1 particles assemble at the plasma membrane of infected cells, before being released as free virions or being transferred through direct cell-to-cell contacts to neighboring cells. Localization of HIV-1 Gag precursor at the cell membrane is necessary and sufficient to trigger viral assembly, whereas the GagPol precursor is additionally required to generate a fully matured virion. HIV-1 Nef is an accessory protein that optimizes viral replication through partly defined mechanisms. Whether Nef modulates Gag and/or GagPol localization and assembly at the membrane and facilitates viral cell-to-cell transfer has not been extensively characterized so far. Results We report that Nef increases the total amount of Gag proteins present in infected cells, and promotes Gag localization at the cell membrane. Moreover, the processing of p55 into p24 is improved in the presence of Nef. We also examined the effect of Nef during HIV-1 cell-to-cell transfer. We show that without Nef, viral transfer through direct contacts between infected cells and target cells is impaired. With a nef-deleted virus, the number of HIV-1 positive target cells after a short 2h co-culture is reduced, and viral material transferred to uninfected cells is less matured. At later time points, this defect is associated with a reduction in the productive infection of new target cells. Conclusions Our results highlight a previously unappreciated role of Nef during the viral replication cycle. Nef promotes HIV-1 Gag membrane localization and processing, and facilitates viral cell-to-cell transfer. PMID:23899341

  12. A new type of Schwann cell graft transplantation to promote optic nerve regeneration in adult rats.

    PubMed

    Fang, Yuan; Mo, Xiaofen; Guo, Wenyi; Zhang, Meng; Zhang, Peihua; Wang, Yan; Rong, Xianfang; Tian, Jie; Sun, Xinghuai

    2010-12-01

    Like other parts of the central nervous system, the adult mammalian optic nerve is difficult to regenerate after injury. Transplantation of the peripheral nerve or a Schwann cell (SC) graft can promote injured axonal regrowth. We tried to develop a new type of tissue-engineered SC graft that consisted of SCs seeded onto a poly(lactic-co-glycolic acid)/chitosan conduit. Meanwhile, SCs were transfected along the ciliary neurotrophic factor (CNTF) gene in vitro by electroporation to increase their neurotrophic effect. Four weeks after transplantation, GAP-43 labelled regenerating axons were found in the SC grafts, and axons in the CNTF-SC graft were longer than those in the SC graft. Tissue-engineered SC grafts can provide a feasible environment for optic nerve regeneration and may become an alternative for bridging damaged nerves and repairing nerve defects in the future.

  13. Nanobiocomposite of poly(lactide-co-glycolide)/chitosan electrospun scaffold can promote proliferation and transdifferentiation of Schwann-like cells from human adipose-derived stem cells.

    PubMed

    Razavi, Shahnaz; Zarkesh-Esfahani, Hamid; Morshed, Mohammad; Vaezifar, Sedigheh; Karbasi, Saeed; Golozar, Mohammad Ali

    2015-08-01

    The transdifferentiation of human adipose-derived stem cells (ADSCs) into Schwann-like cells on biocomposite scaffolds may be a critical issue in nerve regeneration medicine. In this study, tissue-engineered scaffold with chitosan (CS) nanopowders and poly(lactide-co-glycolide) (PLGA) was investigated for its potential Schwann cells (SCs) transdifferentiation. The differentiation of human ADSCs into S-like cells was induced with different CS content and direction of nanofibers on PLGA/CS scaffolds. Cell morphology and proliferation of differentiated cells were investigated by scanning electron microscopy and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay respectively. For assessment efficiency of transdifferentiation, the expression of SC markers (glial fibrillary acidic protein and S100), and myelinogenic marker (myelin basic protein) was investigated in different nanochitosan content and direction of nanofibers scaffolds, using immunocytochemistry technique. The nanochitosan can significantly promote cell proliferation of differentiated cells (p < 0.05). The mean percentage of S-like cells on greater CS content nanofibers scaffold was significantly higher than others (p < 0.05). In addition, the align orientation of nanofibers in scaffolds guided the differentiation of ADSCs toward myelinating S-like cells on the constructs. Overall, we found that high CS content and aligned-orientation of nanofibers in biocomposite scaffold (70/30A) can promote differentiation and myelinogenic capacity of S-like cells induced from human ADSCs. PMID:25614290

  14. Abscisic acid promotes proteasome-mediated degradation of the transcription coactivator NPR1 in Arabidopsis thaliana.

    PubMed

    Ding, Yezhang; Dommel, Matthew; Mou, Zhonglin

    2016-04-01

    Proteasome-mediated turnover of the transcription coactivator NPR1 is pivotal for efficient activation of the broad-spectrum plant immune responses known as localized acquired resistance (LAR) and systemic acquired resistance (SAR) in adjacent and systemic tissues, respectively, and requires the CUL3-based E3 ligase and its adaptor proteins, NPR3 and NPR4, which are receptors for the signaling molecule salicylic acid (SA). It has been shown that SA prevents NPR1 turnover under non-inducing and LAR/SAR-inducing conditions, but how cellular NPR1 homeostasis is maintained remains unclear. Here, we show that the phytohormone abscisic acid (ABA) and SA antagonistically influence cellular NPR1 protein levels. ABA promotes NPR1 degradation via the CUL3(NPR) (3/) (NPR) (4) complex-mediated proteasome pathway, whereas SA may protect NPR1 from ABA-promoted degradation through phosphorylation. Furthermore, we demonstrate that the timing and strength of SA and ABA signaling are critical in modulating NPR1 accumulation and target gene expression. Perturbing ABA or SA signaling in adjacent tissues alters the temporal dynamic pattern of NPR1 accumulation and target gene transcription. Finally, we show that sequential SA and ABA treatment leads to dynamic changes in NPR1 protein levels and target gene expression. Our results revealed a tight correlation between sequential SA and ABA signaling and dynamic changes in NPR1 protein levels and NPR1-dependent transcription in plant immune responses. PMID:26865090

  15. Promoter strength of folic acid synthesis genes affects sulfa drug resistance in Saccharomyces cerevisiae.

    PubMed

    Iliades, Peter; Berglez, Janette; Meshnick, Steven; Macreadie, Ian

    2003-01-01

    The enzyme dihydropteroate synthase (DHPS) is an important target for sulfa drugs in both prokaryotic and eukaryotic microbes. However, the understanding of DHPS function and the action of antifolates in eukaryotes has been limited due to technical difficulties and the complexity of DHPS being a part of a bifunctional or trifunctional protein that comprises the upstream enzymes involved in folic acid synthesis (FAS). Here, yeast strains have been constructed to study the effects of FOL1 expression on growth and sulfa drug resistance. A DHPS knockout yeast strain was complemented by yeast vectors expressing the FOL1 gene under the control of promoters of different strengths. An inverse relationship was observed between the growth rate of the strains and FOL1 expression levels. The use of stronger promoters to drive FOL1 expression led to increased sulfamethoxazole resistance when para-aminobenzoic acid (pABA) levels were elevated. However, high FOL1 expression levels resulted in increased susceptibility to sulfamethoxazole in pABA free media. These data suggest that up-regulation of FOL1 expression can lead to sulfa drug resistance in Saccharomyces cerevisiae.

  16. Okadaic Acid Toxin at Sublethal Dose Produced Cell Proliferation in Gastric and Colon Epithelial Cell Lines

    PubMed Central

    del Campo, Miguel; Toledo, Héctor; Lagos, Néstor

    2013-01-01

    The aim of this study was to analyze the effect of Okadaic Acid (OA) on the proliferation of gastric and colon epithelial cells, the main target tissues of the toxin. We hypothesized that OA, at sublethal doses, activates multiple signaling pathways, such as Erk and Akt, through the inhibition of PP2A. To demonstrate this, we carried out curves of doses and time response against OA in AGS, MKN-45 and Caco 2 cell lines, and found an increase in the cell proliferation at sublethal doses, at 24 h or 48 h exposure. Indeed, cells can withstand high concentrations of the toxin at 4 h exposure, the time chosen considering the maximum time before total gastric emptying. We have proved that this increased proliferation is due to an overexpression of Cyclin B, a cyclin that promotes the passage from G2 to mitosis. In addition, we have demonstrated that OA induces activation of Akt and Erk in the three cells lines, showing that OA can activate pathways involved in oncogenesis. In conclusion, this study contributes to the knowledge about the possible effects of chronic OA consumption. PMID:24317467

  17. Downregulation of SENP1 inhibits cell proliferation, migration and promotes apoptosis in human glioma cells

    PubMed Central

    ZHANG, QIU-SHENG; ZHANG, MENG; HUANG, XIAN-JIAN; LIU, XIAO-JIA; LI, WEI-PING

    2016-01-01

    Small ubiquitin-related modifier protein (SUMO) is an evolutionarily conserved protein in a broad range of eukaryotic organisms. De-SUMOylation, the reverse reaction of SUMOylation, is regulated by a family of SUMO-specific proteases (SENPs). SENP1 is a member of the de-SUMOylation protease family involved in the de-SUMOylation of a variety of SUMOylated proteins. The present study demonstrates that small hairpin RNA (shRNA)-mediated downregulation of SENP1 inhibits cell proliferation and migration, and promotes apoptosis in human glioma cells. Firstly, LN-299 cells were transfected with a plasmid expressing SENP1 shRNA (pGenesil-1-SENP1). The messenger RNA and protein expression of SENP1 was detected by reverse transcription-quantitative polymerase chain reaction and western blot analysis, respectively. Cell proliferation in vitro was assessed using a methyl thiazolyl tetrazolium assay. Flow cytometry (FCM) was used to detect the apoptosis of LN-299 cells. The effect of the downregulation of SENP1 on cell migration was detected by a Transwell migration system. The present results showed that, compared with the control shRNA group, the expression of SENP1 was significantly reduced in the SENP1 shRNA group. The proliferation was markedly inhibited in the SENP1 shRNA group. FCM findings revealed that apoptosis increased significantly in the SENP1 shRNA group. In addition, it was found that downregulation of SENP1 evidently suppressed tumor cell migration. Downregulation of SENP1 expression inhibited the proliferation and migration and promoted apoptosis in LN-299 cells. These results indirectly demonstrate that SENP1 is likely to play a critical role in human glioma cells. PMID:27347128

  18. TAZ promotes cell growth and inhibits Celastrol-induced cell apoptosis

    PubMed Central

    Wang, Shuren; Ma, Kai; Chen, Lechuang; Zhu, Hongxia; Liang, Shufang; Liu, Mei; Xu, Ningzhi

    2016-01-01

    Hippo pathway is a highly conservative signalling pathway related to the development of organisms, which has been demonstrated to be strongly linked to the tumorigenesis and tumour progression. As the major downstream effector of Hippo pathway, yes-associated protein (YAP), is a transcriptional activator of target genes that are involved in cell proliferation and survival. As an oncogene, YAP can promote cell growth and inhibit cell apoptosis. Another major downstream effector of Hippo pathway, transcriptional co-activators with PDZ-binding motif (TAZ), is nearly 60% homologous with YAP. In the present study, we assume that TAZ probably has the similar function to YAP. To test this issue, we established an inducible and a stable expression system of TAZ in T-Rex-293 and HEK293 cells respectively. The results of cell growth curves, colony formation assay and tumour xenograft growth showed that overexpression of TAZ could promote cell growth in vitro and in vivo. Meanwhile, we found that up-regulated expression of TAZ could partially restore Celastrol-induced cell apoptosis. Induced overexpression of TAZ could up-regulate its target genes including ankyrin repeat domain-containing protein (ANKRD), cysteine-rich 61 (CYR61) and connective tissue growth factor (CTGF), increase the expression of B-cell lymphoma-2 (Bcl-2), decrease the expression of Bcl-2 associated X protein (Bax) and activate the phosphatidylinositol 3-kinase (PI3K)/protein kinase B (Akt) pathway, which may be the mechanism underlying anti-apoptosis of TAZ. All these findings indicated that TAZ acts as an oncogene that could be a key regulator of cell proliferation and apoptosis. PMID:27515420

  19. Extracorporeal membrane oxygenation promotes long chain fatty acid oxidation in the immature swine heart in vivo

    PubMed Central

    Kajimoto, Masaki; O’Kelly Priddy, Colleen M.; Ledee, Dolena R.; Xu, Chun; Isern, Nancy; Olson, Aaron K.; Portman, Michael A.

    2013-01-01

    Extracorporeal membrane oxygenation (ECMO) supports infants and children with severe cardiopulmonary compromise. Nutritional support for these children includes provision of medium- and long-chain fatty acids (FAs). However, ECMO induces a stress response, which could limit the capacity for FA oxidation. Metabolic impairment could induce new or exacerbate existing myocardial dysfunction. Using a clinically relevant piglet model, we tested the hypothesis that ECMO maintains the myocardial capacity for FA oxidation and preserves myocardial energy state. Provision of 13-Carbon labeled medium-chain FA (octanoate), long-chain free FAs (LCFAs), and lactate into systemic circulation showed that ECMO promoted relative increases in myocardial LCFA oxidation while inhibiting lactate oxidation. Loading of these labeled substrates at high dose into the left coronary artery demonstrated metabolic flexibility as the heart preferentially oxidized octanoate. ECMO preserved this octanoate metabolic response, but also promoted LCFA oxidation and inhibited lactate utilization. Rapid upregulation of pyruvate dehydrogenase kinase-4 (PDK4) protein appeared to participate in this metabolic shift during ECMO. ECMO also increased relative flux from lactate to alanine further supporting the role for pyruvate dehydrogenase inhibition by PDK4. High dose substrate loading during ECMO also elevated the myocardial energy state indexed by phosphocreatine to ATP ratio. ECMO promotes LCFA oxidation in immature hearts, while maintaining myocardial energy state. These data support the appropriateness of FA provision during ECMO support for the immature heart. PMID:23727393

  20. Extracorporeal membrane oxygenation promotes long chain fatty acid oxidation in the immature swine heart in vivo

    SciTech Connect

    Kajimoto, Masaki; O'Kelly-Priddy, Colleen M.; Ledee, Dolena R.; Xu, Chun; Isern, Nancy G.; Olson, Aaron; Portman, Michael A.

    2013-09-01

    Extracorporeal membrane oxygenation (ECMO) supports infants and children with severe cardiopulmonary compromise. Nutritional support for these children includes provision of medium- and long-chain fatty acids (FAs). However, ECMO induces a stress response, which could limit the capacity for FA oxidation. Metabolic impairment could induce new or exacerbate existing myocardial dysfunction. Using a clinically relevant piglet model, we tested the hypothesis that ECMO maintains the myocardial capacity for FA oxidation and preserves myocardial energy state. Provision of 13-Carbon labeled medium-chain FA (octanoate), longchain free FAs (LCFAs), and lactate into systemic circulation showed that ECMO promoted relative increases in myocardial LCFA oxidation while inhibiting lactate oxidation. Loading of these labeled substrates at high dose into the left coronary artery demonstrated metabolic flexibility as the heart preferentially oxidized octanoate. ECMO preserved this octanoate metabolic response, but also promoted LCFA oxidation and inhibited lactate utilization. Rapid upregulation of pyruvate dehydrogenase kinase-4 (PDK4) protein appeared to participate in this metabolic shift during ECMO. ECMO also increased relative flux from lactate to alanine further supporting the role for pyruvate dehydrogenase inhibition by PDK4. High dose substrate loading during ECMO also elevated the myocardial energy state indexed by phosphocreatine to ATP ratio. ECMO promotes LCFA oxidation in immature hearts, while maintaining myocardial energy state. These data support the appropriateness of FA provision during ECMO support for the immature heart.

  1. Designer nucleic acids to probe and program the cell.

    PubMed

    Krishnan, Yamuna; Bathe, Mark

    2012-12-01

    Recent advances in nucleic acid sequencing, structural, and computational technologies have resulted in dramatic progress in our understanding of nucleic acid structure and function in the cell. This knowledge, together with the predictable base-pairing of nucleic acids and powerful synthesis and expression capabilities now offers the unique ability to program nucleic acids to form precise 3D architectures with diverse applications in synthetic and cell biology. The unique modularity of structural motifs that include aptamers, DNAzymes, and ribozymes, together with their well-defined construction rules, enables the synthesis of functional higher-order nucleic acid complexes from these subcomponents. As we illustrate here, these highly programmable, smart complexes are increasingly enabling researchers to probe and program the cell in a sophisticated manner that moves well beyond the use of nucleic acids for conventional genetic manipulation alone.

  2. Resveratrol sensitizes glioblastoma-initiating cells to temozolomide by inducing cell apoptosis and promoting differentiation.

    PubMed

    Li, Hao; Liu, Yaodong; Jiao, Yumin; Guo, Anchen; Xu, Xiaoxue; Qu, Xianjun; Wang, Shuo; Zhao, Jizong; Li, Ye; Cao, Yong

    2016-01-01

    Glioblastoma-initiating cells play crucial roles in the origin, growth, and recurrence of glioblastoma multiforme. The elimination of glioblastoma-initiating cells is believed to be a key strategy for achieving long-term survival of glioblastoma patients due to the highly resistant property of glioblastoma-initiating cells to temozolomide. Resveratrol, a naturally occurring polyphenol, has been widely studied as a promising candidate for cancer prevention and treatment. Whether resveratrol could enhance the sensitivity of glioblastoma-initiating cells to temozolomide therapy has not yet been reported. Here, using patient-derived glioblastoma-initiating cell lines, we found that resveratrol sensitized glioblastoma-initiating cells to temozolomide both in vitro and in vivo. Furthermore, we showed that resveratrol enhanced glioblastoma-initiating cells to temozolomide-induced apoptosis through DNA double-stranded breaks/pATM/pATR/p53 pathway activation, and promoted glioblastoma-initiating cell differentiation involving p-STAT3 inactivation. Our results propose that temozolomide and resveratrol combination strategy may be effective in the management of glioblastoma patients, particularly for those patients who have been present with a high abundance of glioblastoma-initiating cells in their tumors and show slight responsiveness to temozolomide.

  3. Streptococcal serum opacity factor promotes cholesterol ester metabolism and bile acid secretion in vitro and in vivo.

    PubMed

    Gillard, Baiba K; Rodriguez, Perla J; Fields, David W; Raya, Joe L; Lagor, William R; Rosales, Corina; Courtney, Harry S; Gotto, Antonio M; Pownall, Henry J

    2016-03-01

    Plasma high density lipoprotein-cholesterol (HDL-C) concentrations negatively correlate with atherosclerotic cardiovascular disease. HDL is thought to have several atheroprotective functions, which are likely distinct from the epidemiological inverse relationship between HDL-C levels and risk. Specifically, strategies that reduce HDL-C while promoting reverse cholesterol transport (RCT) may have therapeutic value. The major product of the serum opacity factor (SOF) reaction versus HDL is a cholesteryl ester (CE)-rich microemulsion (CERM), which contains apo E and the CE of ~400,000 HDL particles. Huh7 hepatocytes take up CE faster when delivered as CERM than as HDL, in part via the LDL-receptor (LDLR). Here we compared the final RCT step, hepatic uptake and subsequent intracellular processing to cholesterol and bile salts for radiolabeled HDL-, CERM- and LDL-CE by Huh7 cells and in vivo in C57BL/6J mice. In Huh7 cells, uptake from LDL was greater than from CERM (2-4X) and HDL (5-10X). Halftimes for [(14)C]CE hydrolysis were 3.0±0.2, 4.4±0.6 and 5.4±0.7h respectively for HDL, CERM and LDL-CE. The fraction of sterols secreted as bile acids was ~50% by 8h for all three particles. HDL, CERM and LDL-CE metabolism in mice showed efficient plasma clearance of CERM-CE, liver uptake and metabolism, and secretion as bile acids into the gall bladder. This work supports the therapeutic potential of the SOF reaction, which diverts HDL-CE to the LDLR, thereby increasing hepatic CE uptake, and sterol disposal as bile acids. PMID:26709142

  4. Synergistic transcriptional enhancement does not depend on the number of acidic activation domains bound to the promoter.

    PubMed Central

    Oliviero, S; Struhl, K

    1991-01-01

    Many eukaryotic transcriptional activator proteins contain a DNA-binding domain that interacts with specific promoter sequences and an acidic activation region that is required to stimulate transcription. Transcriptional enhancement by such activator proteins is often synergistic and promiscuous; promoters containing multiple binding sites for an individual protein or even for unrelated proteins can be 10-100 times more active than promoters with single sites. It has been suggested that such synergy reflects a nonlinear response of the basic transcription machinery to the number and/or quality of acidic activation regions. Here, we determine the transcriptional activity of Jun-Fos heterodimers containing one or two GCN4 acidic activation regions on promoters containing one or two Ap-1 target sites. Surprisingly, heterodimers with one or two acidic regions activate transcription with similar efficiency and are equally synergistic (10- to 15-fold) on promoters containing two target sites. Thus, transcriptional synergy does not depend on the number of acidic activation regions but rather on the number of proteins bound to the promoter. This suggests that synergy is mediated either by cooperative DNA binding or by alternative mechanisms in which the DNA-binding domain plays a more direct role in transcription (e.g., changes in DNA structure, nucleosome displacement, or direct interactions with the transcriptional machinery). Images PMID:1898773

  5. Design of Hyaluronic Acid Hydrogels to Promote Neurite Outgrowth in Three Dimensions.

    PubMed

    Tarus, Dominte; Hamard, Lauriane; Caraguel, Flavien; Wion, Didier; Szarpak-Jankowska, Anna; van der Sanden, Boudewijn; Auzély-Velty, Rachel

    2016-09-28

    A hyaluronic acid (HA)-based extracellular matrix (ECM) platform with independently tunable stiffness and density of cell-adhesive peptide (RGD, arginine-glycine-aspartic acid) that mimics key biochemical and mechanical features of brain matrix has been designed. We demonstrated here its utility in elucidating ECM regulation of neural progenitor cell behavior and neurite outgrowth. The analysis of neurite outgrowth in 3-D by two-photon microscopy showed several important results in the development of these hydrogels. First, the ability of neurites to extend deeply into these soft HA-based matrices even in the absence of cell-adhesive ligand further confirms the potential of HA hydrogels for central nervous system (CNS) regeneration. Second, the behavior of hippocampal neural progenitor cells differed markedly between the hydrogels with a storage modulus of 400 Pa and those with a modulus of 800 Pa. We observed an increased outgrowth and density of neurites in the softest hydrogels (G' = 400 Pa). Interestingly, cells seeded on the surface of the hydrogels functionalized with the RGD ligand experienced an optimum in neurite outgrowth as a function of ligand density. Surprinsingly, neurites preferentially progressed inside the gels in a vertical direction, suggesting that outgrowth is directed by the hydrogel structure. This work may provide design principles for the development of hydrogels to facilitate neuronal regeneration in the adult brain.

  6. Design of Hyaluronic Acid Hydrogels to Promote Neurite Outgrowth in Three Dimensions.

    PubMed

    Tarus, Dominte; Hamard, Lauriane; Caraguel, Flavien; Wion, Didier; Szarpak-Jankowska, Anna; van der Sanden, Boudewijn; Auzély-Velty, Rachel

    2016-09-28

    A hyaluronic acid (HA)-based extracellular matrix (ECM) platform with independently tunable stiffness and density of cell-adhesive peptide (RGD, arginine-glycine-aspartic acid) that mimics key biochemical and mechanical features of brain matrix has been designed. We demonstrated here its utility in elucidating ECM regulation of neural progenitor cell behavior and neurite outgrowth. The analysis of neurite outgrowth in 3-D by two-photon microscopy showed several important results in the development of these hydrogels. First, the ability of neurites to extend deeply into these soft HA-based matrices even in the absence of cell-adhesive ligand further confirms the potential of HA hydrogels for central nervous system (CNS) regeneration. Second, the behavior of hippocampal neural progenitor cells differed markedly between the hydrogels with a storage modulus of 400 Pa and those with a modulus of 800 Pa. We observed an increased outgrowth and density of neurites in the softest hydrogels (G' = 400 Pa). Interestingly, cells seeded on the surface of the hydrogels functionalized with the RGD ligand experienced an optimum in neurite outgrowth as a function of ligand density. Surprinsingly, neurites preferentially progressed inside the gels in a vertical direction, suggesting that outgrowth is directed by the hydrogel structure. This work may provide design principles for the development of hydrogels to facilitate neuronal regeneration in the adult brain. PMID:27598554

  7. Human Olfactory Mucosa Multipotent Mesenchymal Stromal Cells Promote Survival, Proliferation, and Differentiation of Human Hematopoietic Cells

    PubMed Central

    Diaz-Solano, Dylana; Wittig, Olga; Ayala-Grosso, Carlos; Pieruzzini, Rosalinda

    2012-01-01

    Multipotent mesenchymal stromal cells (MSCs) from the human olfactory mucosa (OM) are cells that have been proposed as a niche for neural progenitors. OM-MSCs share phenotypic and functional properties with bone marrow (BM) MSCs, which constitute fundamental components of the hematopoietic niche. In this work, we investigated whether human OM-MSCs may promote the survival, proliferation, and differentiation of human hematopoietic stem cells (HSCs). For this purpose, human bone marrow cells (BMCs) were co-cultured with OM-MSCs in the absence of exogenous cytokines. At different intervals, nonadherent cells (NACs) were harvested from BMC/OM-MSC co-cultures, and examined for the expression of blood cell markers by flow cytometry. OM-MSCs supported the survival (cell viability >90%) and proliferation of BMCs, after 54 days of co-culture. At 20 days of co-culture, flow cytometric and microscopic analyses showed a high percentage (73%) of cells expressing the pan-leukocyte marker CD45, and the presence of cells of myeloid origin, including polymorphonuclear leukocytes, monocytes, basophils, eosinophils, erythroid cells, and megakaryocytes. Likewise, T (CD3), B (CD19), and NK (CD56/CD16) cells were detected in the NAC fraction. Colony-forming unit–granulocyte/macrophage (CFU-GM) progenitors and CD34+ cells were found, at 43 days of co-culture. Reverse transcriptase–polymerase chain reaction (RT-PCR) studies showed that OM-MSCs constitutively express early and late-acting hematopoietic cytokines (i.e., stem cell factor [SCF] and granulocyte- macrophage colony-stimulating factor [GM-CSF]). These results constitute the first evidence that OM-MSCs may provide an in vitro microenvironment for HSCs. The capacity of OM-MSCs to support the survival and differentiation of HSCs may be related with the capacity of OM-MSCs to produce hematopoietic cytokines. PMID:22471939

  8. Smooth muscle progenitor cells from peripheral blood promote the neovascularization of endothelial colony-forming cells

    SciTech Connect

    Joo, Hyung Joon; Seo, Ha-Rim; Jeong, Hyo Eun; Choi, Seung-Cheol; Park, Jae Hyung; Yu, Cheol Woong; Hong, Soon Jun; Chung, Seok; Lim, Do-Sun

    2014-07-11

    Highlights: • Two distinct vascular progenitor cells are induced from adult peripheral blood. • ECFCs induce vascular structures in vitro and in vivo. • SMPCs augment the in vitro and in vivo angiogenic potential of ECFCs. • Both cell types have synergistic therapeutic potential in ischemic hindlimb model. - Abstract: Proangiogenic cell therapy using autologous progenitors is a promising strategy for treating ischemic disease. Considering that neovascularization is a harmonized cellular process that involves both endothelial cells and vascular smooth muscle cells, peripheral blood-originating endothelial colony-forming cells (ECFCs) and smooth muscle progenitor cells (SMPCs), which are similar to mature endothelial cells and vascular smooth muscle cells, could be attractive cellular candidates to achieve therapeutic neovascularization. We successfully induced populations of two different vascular progenitor cells (ECFCs and SMPCs) from adult peripheral blood. Both progenitor cell types expressed endothelial-specific or smooth muscle-specific genes and markers, respectively. In a protein array focused on angiogenic cytokines, SMPCs demonstrated significantly higher expression of bFGF, EGF, TIMP2, ENA78, and TIMP1 compared to ECFCs. Conditioned medium from SMPCs and co-culture with SMPCs revealed that SMPCs promoted cell proliferation, migration, and the in vitro angiogenesis of ECFCs. Finally, co-transplantation of ECFCs and SMPCs induced robust in vivo neovascularization, as well as improved blood perfusion and tissue repair, in a mouse ischemic hindlimb model. Taken together, we have provided the first evidence of a cell therapy strategy for therapeutic neovascularization using two different types of autologous progenitors (ECFCs and SMPCs) derived from adult peripheral blood.

  9. Human olfactory mucosa multipotent mesenchymal stromal cells promote survival, proliferation, and differentiation of human hematopoietic cells.

    PubMed

    Diaz-Solano, Dylana; Wittig, Olga; Ayala-Grosso, Carlos; Pieruzzini, Rosalinda; Cardier, Jose E

    2012-11-20

    Multipotent mesenchymal stromal cells (MSCs) from the human olfactory mucosa (OM) are cells that have been proposed as a niche for neural progenitors. OM-MSCs share phenotypic and functional properties with bone marrow (BM) MSCs, which constitute fundamental components of the hematopoietic niche. In this work, we investigated whether human OM-MSCs may promote the survival, proliferation, and differentiation of human hematopoietic stem cells (HSCs). For this purpose, human bone marrow cells (BMCs) were co-cultured with OM-MSCs in the absence of exogenous cytokines. At different intervals, nonadherent cells (NACs) were harvested from BMC/OM-MSC co-cultures, and examined for the expression of blood cell markers by flow cytometry. OM-MSCs supported the survival (cell viability >90%) and proliferation of BMCs, after 54 days of co-culture. At 20 days of co-culture, flow cytometric and microscopic analyses showed a high percentage (73%) of cells expressing the pan-leukocyte marker CD45, and the presence of cells of myeloid origin, including polymorphonuclear leukocytes, monocytes, basophils, eosinophils, erythroid cells, and megakaryocytes. Likewise, T (CD3), B (CD19), and NK (CD56/CD16) cells were detected in the NAC fraction. Colony-forming unit-granulocyte/macrophage (CFU-GM) progenitors and CD34(+) cells were found, at 43 days of co-culture. Reverse transcriptase-polymerase chain reaction (RT-PCR) studies showed that OM-MSCs constitutively express early and late-acting hematopoietic cytokines (i.e., stem cell factor [SCF] and granulocyte- macrophage colony-stimulating factor [GM-CSF]). These results constitute the first evidence that OM-MSCs may provide an in vitro microenvironment for HSCs. The capacity of OM-MSCs to support the survival and differentiation of HSCs may be related with the capacity of OM-MSCs to produce hematopoietic cytokines.

  10. Gallic acid reduces cell viability, proliferation, invasion and angiogenesis in human cervical cancer cells

    PubMed Central

    ZHAO, BING; HU, MENGCAI

    2013-01-01

    Gallic acid is a trihydroxybenzoic acid, also known as 3,4,5-trihydroxybenzoic acid, which is present in plants worldwide, including Chinese medicinal herbs. Gallic acid has been shown to have cytotoxic effects in certain cancer cells, without damaging normal cells. The objective of the present study was to determine whether gallic acid is able to inhibit human cervical cancer cell viability, proliferation and invasion and suppress cervical cancer cell-mediated angiogenesis. Treatment of HeLa and HTB-35 human cancer cells with gallic acid decreased cell viability in a dose-dependent manner. BrdU proliferation and tube formation assays indicated that gallic acid significantly decreased human cervical cancer cell proliferation and tube formation in human umbilical vein endothelial cells, respectively. Additionally, gallic acid decreased HeLa and HTB-35 cell invasion in vitro. Western blot analysis demonstrated that the expression of ADAM17, EGFR, p-Akt and p-Erk was suppressed by gallic acid in the HeLa and HTB-35 cell lines. These data indicate that the suppression of ADAM17 and the downregulation of the EGFR, Akt/p-Akt and Erk/p-Erk signaling pathways may contribute to the suppression of cancer progression by Gallic acid. Gallic acid may be a valuable candidate for the treatment of cervical cancer. PMID:24843386

  11. Ethanol Promotes Chemically Induced Oral Cancer in Mice through Activation of the 5-Lipoxygenase Pathway of Arachidonic Acid Metabolism

    PubMed Central

    Guo, Yizhu; Wang, Xin; Zhang, Xinyan; Sun, Zheng; Chen, Xiaoxin

    2011-01-01

    Alcohol drinking is a known risk factor for oral cancer in humans. However, previous animal studies on the promoting effect of ethanol on oral carcinogenesis were inconclusive. It is necessary to develop an animal model with which the molecular mechanism of ethanol-related oral carcinogenesis may be elucidated in order to develop effective prevention strategies. In this study, mice were first treated with 4-nitroquinoline-1-oxide (4NQO, 100μg/ml in drinking water) for 8 weeks, and then given water or ethanol (8%) as the sole drink for another 16 weeks. During the experiment, 8% ethanol was well tolerated by mice. The incidence of squamous cell carcinoma (SCC) increased from 20% (8/41) to 43% (17/40; p<0.05). Expression of 5-lipoxygenase (5-Lox) and cyclooxygenase 2 (Cox-2) was increased in dysplasia and SCC of 4NQO-treated tongues, and further enhanced by ethanol. Using this mouse model, we further demonstrated that fewer cancers were induced in Alox5−/− mice, as were cell proliferation, inflammation, and angiogenesis in the tongue, as compared with Alox5+/+ mice. Interestingly, Cox-2 expression was induced by ethanol in knockout mice, while 5-Lox and leukotriene A4 hydrolase (LTA4H) expression and leukotriene B4 (LTB4) biosynthesis were dramatically reduced. Moreover, ethanol enhanced expression and nuclear localization of 5-Lox and stimulated LTB4 biosynthesis in human tongue SCC cells (SCC-15 and SCC-4) in vitro. In conclusion, this study clearly demonstrated that ethanol promoted 4NQO-induced oral carcinogenesis, at least in part, through further activation of the 5-Lox pathway of arachidonic acid metabolism. PMID:21881027

  12. L-Rhamnosylation of Listeria monocytogenes Wall Teichoic Acids Promotes Resistance to Antimicrobial Peptides by Delaying Interaction with the Membrane

    PubMed Central

    Carvalho, Filipe; Atilano, Magda L.; Pombinho, Rita; Covas, Gonçalo; Gallo, Richard L.; Filipe, Sérgio R.; Sousa, Sandra; Cabanes, Didier

    2015-01-01

    Listeria monocytogenes is an opportunistic Gram-positive bacterial pathogen responsible for listeriosis, a human foodborne disease. Its cell wall is densely decorated with wall teichoic acids (WTAs), a class of anionic glycopolymers that play key roles in bacterial physiology, including protection against the activity of antimicrobial peptides (AMPs). In other Gram-positive pathogens, WTA modification by amine-containing groups such as D-alanine was largely correlated with resistance to AMPs. However, in L. monocytogenes, where WTA modification is achieved solely via glycosylation, WTA-associated mechanisms of AMP resistance were unknown. Here, we show that the L-rhamnosylation of L. monocytogenes WTAs relies not only on the rmlACBD locus, which encodes the biosynthetic pathway for L-rhamnose, but also on rmlT encoding a putative rhamnosyltransferase. We demonstrate that this WTA tailoring mechanism promotes resistance to AMPs, unveiling a novel link between WTA glycosylation and bacterial resistance to host defense peptides. Using in vitro binding assays, fluorescence-based techniques and electron microscopy, we show that the presence of L-rhamnosylated WTAs at the surface of L. monocytogenes delays the crossing of the cell wall by AMPs and postpones their contact with the listerial membrane. We propose that WTA L-rhamnosylation promotes L. monocytogenes survival by decreasing the cell wall permeability to AMPs, thus hindering their access and detrimental interaction with the plasma membrane. Strikingly, we reveal a key contribution of WTA L-rhamnosylation for L. monocytogenes virulence in a mouse model of infection. PMID:26001194

  13. Lactic acid fermentation in cell-recycle membrane bioreactor.

    PubMed

    Choudhury, B; Swaminathan, T

    2006-02-01

    Traditional lactic acid fermentation suffers from low productivity and low product purity. Cell-recycle fermentation has become one of the methods to obtain high cell density, which results in higher productivity. Lactic acid fermentation was investigated in a cell-recycle membrane bioreactor at higher substrate concentrations of 100 and 120 g/dm3. A maximum cell density of 145 g/dm3 and a maximum productivity of 34 g/(dm3.h) were achieved in cell-recycle fermentation. In spite of complete consumption of substrate, there was a continuous increase in cell density in cell-recycle fermentation. Control of cell density in cell-recycle fermentation was attempted by cell bleeding and reduction in yeast extract concentration.

  14. Extravillous trophoblast cells-derived exosomes promote vascular smooth muscle cell migration

    PubMed Central

    Salomon, Carlos; Yee, Sarah; Scholz-Romero, Katherin; Kobayashi, Miharu; Vaswani, Kanchan; Kvaskoff, David; Illanes, Sebastian E.; Mitchell, Murray D.; Rice, Gregory E.

    2014-01-01

    Background: Vascular smooth muscle cells (VSMCs) migration is a critical process during human uterine spiral artery (SpA) remodeling and a successful pregnancy. Extravillous trophoblast cells (EVT) interact with VSMC and enhance their migration, however, the mechanisms by which EVT remodel SpA remain to be fully elucidated. We hypothesize that exosomes released from EVT promote VSMC migration. Methods: JEG-3 and HTR-8/SVneo cell lines were used as models for EVT. Cells were cultured at 37°C and humidified under an atmosphere of 5% CO2-balanced N2 to obtain 8% O2. Cell-conditioned media were collected, and exosomes (exo-JEG-3 and exo- HTR-8/SVneo) isolated by differential and buoyant density centrifugation. The effects of exo-EVT on VSMC migration were established using a real-time, live-cell imaging system (Incucyte™). Exosomal proteins where identified by mass spectrometry and submitted to bioinformatic pathway analysis (Ingenuity software). Results: HTR-8/SVneo cells were significantly more (~30%) invasive than JEG-3 cells. HTR-8/SVneo cells released 2.6-fold more exosomes (6.39 × 108 ± 2.5 × 108 particles/106 cells) compared to JEG-3 (2.86 × 108 ± 0.78 × 108 particles/106 cells). VSMC migration was significantly increased in the presence of exo-JEG-3 and exo-HTR-8/SVneo compared to control (−exosomes) (21.83 ± 0.49 h and 15.57 ± 0.32, respectively, vs. control 25.09 ± 0.58 h, p < 0.05). Sonication completely abolished the effect of exosomes on VSMC migration. Finally, mass spectrometry analysis identified unique exosomal proteins for each EVT cell line-derived exosomes. Conclusion: The data obtained in this study are consistent with the hypothesis that the release, content, and bioactivity of exosomes derived from EVT-like cell lines is cell origin-dependent and differentially regulates VSMC migration. Thus, an EVT exosomal signaling pathway may contribute to SpA remodeling by promoting the migration of VSMC out of the vessel walls. PMID:25157233

  15. The role of growth of normal and preneoplastic cell populations for tumor promotion in rat liver

    PubMed Central

    Schulte-Hermann, R.; Schuppler, J.; Timmermann-Trosiener, I.; Ohde, G.; Bursch, W.; Berger, H.

    1983-01-01

    A number of different compounds, including phenobarbital, hypolipidemic drugs such as clofibrate and nafenopin, the sex steroids progesterone, cyproterone acetate, estradiol and mestranol, chlorinated hydrocarbons such as DDT, hexachlorocyclohexane, and TCDD and the antioxidant butylhydroxytoluene, appears to promote the development of liver tumors from previously induced initiated cells. The mechanisms of tumor promotion by several representative prototypes of these compounds were studied in rat liver in vivo. All liver tumor promoters mentioned above stimulate growth of normal liver. The growth response is due to cellular hypertrophy and/or increased rate of DNA (and cell) replication and/or decreased rate of cell death. Hepatocytes in foci or islands of altered cells (putatively preneoplastic) show higher rates of replication than normal liver cells; various different liver tumor promoters cause a further increase of proliferation of focal cells. The increased proliferative activity is found in different island phenotypes and thus seems to be a useful marker of the putative preneoplastic state. The focal cells respond to several factors limiting proliferation in normal liver, suggesting that they are not autonomous with respect to growth control. Early preneoplastic foci grow slowly without promotion, despite the relatively high rates of cell replication. Thus their cells seem to have a much shorter life-time than normal hepatocytes or to undergo reversion to the normal phenotype. Promoters seem to accelerate island enlargement by increasing cell replication and delaying cell death or remodeling. Thus, tumor promoters enhance the manifestation of the proliferation advantage of the putative initiated cell population. In addition, promoters cause increases in the number of detectable islands. This can partially be explained by enlargement of existing islands, but phenotypic changes that would enhance the probability of detection of remodelling islands and growth

  16. Chlorogenic Acids Biosynthesis in Centella asiatica Cells Is not Stimulated by Salicylic Acid Manipulation.

    PubMed

    Ncube, E N; Steenkamp, P A; Madala, N E; Dubery, I A

    2016-07-01

    Exogenous application of synthetic and natural elicitors of plant defence has been shown to result in mass production of secondary metabolites with nutraceuticals properties in cultured cells. In particular, salicylic acid (SA) treatment has been reported to induce the production of phenylpropanoids, including cinnamic acid derivatives bound to quinic acid (chlorogenic acids). Centella asiatica is an important medicinal plant with several therapeutic properties owing to its wide spectrum of secondary metabolites. We investigated the effect of SA on C. asiatica cells by monitoring perturbation of chlorogenic acids in particular. Different concentrations of SA were used to treat C. asiatica cells, and extracts from both treated and untreated cells were analysed using an optimised UHPLC-QTOF-MS/MS method. Semi-targeted multivariate data analyses with the aid of principal component analysis (PCA) and orthogonal projection to latent structures-discriminant analysis (OPLS-DA) revealed a concentration-dependent metabolic response. Surprisingly, a range of chlorogenic acid derivatives were found to be downregulated as a consequence of SA treatment. Moreover, irbic acid (3,5-O-dicaffeoyl-4-O-malonilquinic acid) was found to be a dominant CGA in C. asiatica cells, although the SA treatment also had a negative effect on its concentration. Overall SA treatment was found to be an ineffective elicitor of CGA production in cultured C. asiatica cells.

  17. Rapid Formation of Cell Aggregates and Spheroids Induced by a "Smart" Boronic Acid Copolymer.

    PubMed

    Amaral, Adérito J R; Pasparakis, George

    2016-09-01

    Cell surface engineering has emerged as a powerful approach to forming cell aggregates/spheroids and cell-biomaterial ensembles with significant uses in tissue engineering and cell therapeutics. Herein, we demonstrate that cell membrane remodeling with a thermoresponsive boronic acid copolymer induces the rapid formation of spheroids using either cancer or cardiac cell lines under conventional cell culture conditions at minute concentrations. It is shown that the formation of well-defined spheroids is accelerated by at least 24 h compared to non-polymer-treated controls, and, more importantly, the polymer allows for fine control of the aggregation kinetics owing to its stimulus response to temperature and glucose content. On the basis of its simplicity and effectiveness to promote cellular aggregation, this platform holds promise in three-dimensional tissue/tumor modeling and tissue engineering applications. PMID:27571512

  18. Effects of omega-3 fatty acids on regulatory T cells in hematologic neoplasms

    PubMed Central

    Betiati, Dayanne da Silva Borges; de Oliveira, Paula Fernanda; Camargo, Carolina de Quadros; Nunes, Everson Araújo; Trindade, Erasmo Benício Santos de Moraes

    2013-01-01

    The development of leukemia and lymphomas is related to the increase in inflammatory process modulators. These, in turn, have divergent actions on the neoplastic process. Populations of T cells have different roles in the neoplastic environment; while interferon-gamma positive T cells have antitumor activity, the FoxP3+interleukin-10 positive population present a pro-tumor activity. Simultaneously, the inflammatory process promotes the mobilization of fatty acids from the cell membrane to produce lipid mediators, which also participate of the inflammatory response. Eicosapentaenoic (EPA) and docosahexaenoic (DHA) omega-3 fatty acids, when incorporated in the plasmatic membrane, decrease the arachidonic acid (AA) metabolism and the production of eicosanoids derived from it. Thus, an alternative family of lipid mediators are produced that are often less inflammatory than those produced from arachidonic acid. Fatty acids can also influence the production of peptide mediators such as cytokines, and the expression of transcription factors, which can determine the production patterns of eicosanoids and cytokines as well as cell differentiation. Due to these properties, the objective of this literature review was to investigate studies published over the last 15 years on the effects of using omega-3 fatty acids on inflammatory markers in leukemia and lymphomas. PMID:23741190

  19. Subcutaneous fatty acid composition of steers finished as weanlings or yearlings with and without growth promotants

    PubMed Central

    2013-01-01

    Background The current study evaluated the subcutaneous fatty acid (FA) composition of calf- and yearling-fed steers with or without growth promoting implants. Crossbred steers (n = 112; 267 ± 5.0 kg) of the same contemporary group were allocated to one of four production system and implant strategy based treatments in a completely randomized design with a 2 × 2 factorial arrangement of treatments. Results There were no interactions (P > 0.05) between production systems and growth promoting implants for the total and individual subcutaneous FA. Yearling as opposed to calf finishing reduced (P < 0.05) subcutaneous proportions of C20:3n-6, trans (t)12-18:1, C14:0, several minor cis-monounsaturated FA (c-MUFA; c9-14:1, c11-16:1, c11-18:1, c12-18:1, c13-18:1, c9-20:1 and c11-20:1), and increased (P < 0 .05) subcutaneous proportions of t11c15-18:2, total and individual branched-chain FA. Subcutaneous fat from steers implanted with growth promotants had higher (P < 0.05) proportions of total polyunsaturated FA (PUFA), total n-6 PUFA, C18:2n-6 and individual t-18:1 isomers (t6 to t10) compared to non-implanted steers. Conclusions Overall, current findings show that production systems and growth promotants led to only minor differences in subcutaneous FA composition of beef steers. PMID:24188642

  20. Identification of the promoter sequences involved in the cell specific expression of the rat somatostatin gene.

    PubMed Central

    Andrisani, O M; Hayes, T E; Roos, B; Dixon, J E

    1987-01-01

    DNA sequences containing the 5' flanking region of the rat somatostatin gene were linked to the coding sequence of the bacterial chloramphenicol acetyl transferase gene. This recombinant plasmid is active in expressing CAT activity in the neuronally derived, somatostatin producing CA-77 cell line. Deletion analyses of the somatostatin promoter show that the sequences proximal to position -60, relative to the cap site are required for expression of this promoter. A 4 base pair deletion of residues -46 through -43 within the somatostatin promoter results in a down mutation in vivo suggesting the existence of an element critical for the expression of the promoter in CA-77 cells. In addition, the somatostatin recombinant and its 5' deletion constructs preferentially express CAT activity in CA-77 cells, whereas only basal level of expression is observed in HeLa, BSC40, and RIN-5F cell lines, pointing to the cell specific nature of this promoter. Images PMID:2886975

  1. Acidic pH promotes oligomerization and membrane insertion of the BclXL apoptotic repressor.

    PubMed

    Bhat, Vikas; Kurouski, Dmitry; Olenick, Max B; McDonald, Caleb B; Mikles, David C; Deegan, Brian J; Seldeen, Kenneth L; Lednev, Igor K; Farooq, Amjad

    2012-12-01

    Solution pH is believed to serve as an intricate regulatory switch in the induction of apoptosis central to embryonic development and cellular homeostasis. Herein, using an array of biophysical techniques, we provide evidence that acidic pH promotes the assembly of BclXL apoptotic repressor into a megadalton oligomer with a plume-like appearance and harboring structural features characteristic of a molten globule. Strikingly, our data reveal that pH tightly modulates not only oligomerization but also ligand binding and membrane insertion of BclXL in a highly subtle manner. Thus, while oligomerization and the accompanying molten globular content of BclXL is least favorable at pH 6, both of these structural features become more pronounced under acidic and alkaline conditions. However, membrane insertion of BclXL appears to be predominantly favored under acidic conditions. In a remarkable contrast, while ligand binding to BclXL optimally occurs at pH 6, it is diminished by an order of magnitude at lower and higher pH. This reciprocal relationship between BclXL oligomerization and ligand binding lends new insights into how pH modulates functional versatility of a key apoptotic regulator and strongly argues that the molten globule may serve as an intermediate primed for membrane insertion in response to apoptotic cues. PMID:22960132

  2. PVM/MA-shelled selol nanocapsules promote cell cycle arrest in A549 lung adenocarcinoma cells

    PubMed Central

    2014-01-01

    Background Selol is an oily mixture of selenitetriacylglycerides that was obtained as a semi-synthetic compound containing selenite. Selol is effective against cancerous cells and less toxic to normal cells compared with inorganic forms of selenite. However, Selol’s hydrophobicity hinders its administration in vivo. Therefore, the present study aimed to produce a formulation of Selol nanocapsules (SPN) and to test its effectiveness against pulmonary adenocarcinoma cells (A549). Results Nanocapsules were produced through an interfacial nanoprecipitation method. The polymer shell was composed of poly(methyl vinyl ether-co-maleic anhydride) (PVM/MA) copolymer. The obtained nanocapsules were monodisperse and stable. Both free Selol (S) and SPN reduced the viability of A549 cells, whereas S induced a greater reduction in non-tumor cell viability than SPN. The suppressor effect of SPN was primarily associated to the G2/M arrest of the cell cycle, as was corroborated by the down-regulations of the CCNB1 and CDC25C genes. Apoptosis and necrosis were induced by Selol in a discrete percentage of A549 cells. SPN also increased the production of reactive oxygen species, leading to oxidative cellular damage and to the overexpression of the GPX1, CYP1A1, BAX and BCL2 genes. Conclusions This study presents a stable formulation of PVM/MA-shelled Selol nanocapsules and provides the first demonstration that Selol promotes G2/M arrest in cancerous cells. PMID:25149827

  3. Synthesis of novel acid electrolytes for phosphoric acid fuel cells

    NASA Astrophysics Data System (ADS)

    Adcock, James L.

    1988-11-01

    A 40 millimole per hour scale aerosol direct fluorination reactor was constructed. F-Methyl F-4-methoxybutanoate and F-4-methoxybutanoyl fluoride were synthesized by aerosol direct fluorination of methyl 4-methoxybutanoate. Basic hydrolysis of the perfluorinated derivatives produce sodium F-4 methoxybutanoate which was pyrolyzed to F-3-methoxy-1-propene. Purification and shipment of 33 grams of F-3-methoxy-1-propene followed. Syntheses by analogous methods allowed production and shipment of 5 grams of F-3-ethoxy 1-propene, 18 grams of F-3-(2-methoxy.ethoxy) 1-propene, and 37 grams of F-3,3-dimethyl 1-butene. Eighteen grams of F-2,2-dimethyl 1-chloropropane was produced directly and shipped. As suggested by other contractors, 5 grams of F-3-methoxy 1-iodopropane, and 5 grams of F-3-(2-methoxy.ethoxy) 1-iodopropane were produced by converting the respective precursor acid sodium salts produced for olefin synthesis to the silver salts and pyrolyzing them with iodine. Each of these compounds was prepared for the first time by the aerosol fluorination process during the course of the contract. These samples were provided to other Gas Research Institute (GRI) contractors for synthesis of perfluorinated sulfur (VI) and phosphorous (V) acids.

  4. Endothelial cell-initiated signaling promotes the survival and self-renewal of cancer stem cells

    PubMed Central

    Krishnamurthy, Sudha; Dong, Zhihong; Vodopyanov, Dmitry; Imai, Atsushi; Helman, Joseph I.; Prince, Mark E.; Wicha, Max S.; Nör, Jacques E.

    2010-01-01

    Recent studies have demonstrated that cancer stem cells play an important role in the pathobiology of head and neck squamous cell carcinomas (HNSCC). However, little is known about functional interactions between head and neck cancer stem-like cells (CSC) and surrounding stromal cells. Here, we used Aldehyde Dehydrogenase activity and CD44 expression to sort putative stem cells from primary human HNSCC. Implantation of 1,000 CSC (ALDH+CD44+Lin−) led to tumors in 13 (out of 15) mice, while 10,000 non-cancer stem cells (NCSC; ALDH−CD44−Lin−) resulted in 2 tumors in 15 mice. These data demonstrated that ALDH and CD44 select a sub-population of cells that are highly tumorigenic. The ability to self-renew was confirmed by the observation that ALDH+CD44+Lin− cells sorted from human HNSCC formed more spheroids (orospheres) in 3-D agarose matrices or ultra-low attachment plates than controls and were serially passaged in vivo. We observed that approximately 80% of the CSC were located in close proximity (within 100-µm radius) of blood vessels in human tumors, suggesting the existence of perivascular niches in HNSCC. In vitro studies demonstrated that endothelial cell-secreted factors promoted self-renewal of CSC, as demonstrated by the upregulation of Bmi-1 expression and the increase in the number of orospheres as compared to controls. Notably, selective ablation of tumor-associated endothelial cells stably transduced with a caspase-based artificial death switch (iCaspase-9) caused a marked reduction in the fraction of CSC in xenograft tumors. Collectively, these findings indicate that endothelial cell-initiated signaling can enhance the survival and self-renewal of head and neck cancer stem cells. PMID:21098716

  5. Pseudolaric acid B activates autophagy in MCF-7 human breast cancer cells to prevent cell death

    PubMed Central

    YU, JINGHUA; CHEN, CHUNHAI; XU, TIANYANG; YAN, MINGHUI; XUE, BIANBIAN; WANG, YING; LIU, CHUNYU; ZHONG, TING; WANG, ZENGYAN; MENG, XIANYING; HU, DONGHUA; YU, XIAOFANG

    2016-01-01

    Pseudolaric acid B (PAB) has been demonstrated to exert antitumor effects in MCF-7 human breast cancer cells. The present study aimed to investigate the mechanism of resistance to PAB-induced cell death. Following incubation with 4 µM of PAB for 3 days, the majority of MCF-7 cells became senescent, while some retained the same morphology as control cells, as assessed using a senescence detection kit. Additionally, 36 h of treatment with 4 µM of PAB increased the positive staining of autophagy markers, as shown by monodansylcadaverine and acridine orange staining. Western blot analysis indicated that this treatment also increased expression of the autophagy-related proteins Beclin-1 and microtubule-associated protein 1 light chain 3. Furthermore, treatment with PAB and the autophagy inhibitor 3-methyl adenine significantly decreased the ratio of autophagy, as assessed by flow cytometric analysis of monodansylcadaverine staining density (P<0.001), and increased the ratio of cell death, as assessed by MTT analysis (P<0.001). This indicated that autophagy promotes cell survival as a resistance mechanism to PAB treatment. Additionally, the present study demonstrated that PAB treatment did not affect the mitochondrial membrane potential, which may be related to autophagy. Increased Bcl-2 expression may explain why PAB did not affect the mitochondrial membrane potential. A Bcl-2 binding test demonstrated that PAB treatment inhibits the binding of Bcl-2 and Beclin-1, which may free Beclin-1 to participate in autophagy. Therefore, the present study demonstrated that autophagy may be activated by PAB treatment in human breast cancer MCF-7 cells, contributing to resistance to cell death. PMID:26998069

  6. N-methyl-D-aspartate promotes the survival of cerebellar granule cells in culture.

    PubMed

    Balázs, R; Jørgensen, O S; Hack, N

    1988-11-01

    Our previous studies on the survival-promoting influence of elevated concentrations of extracellular K+ ([K+]e) on cultured cerebellar granule cells led to the proposal that depolarization in vitro mimics the effect of the earliest afferent inputs received by the granule cells in vivo. This, in turn, might be mediated through the stimulation of excitatory amino acid receptors, in particular the N-methyl-D-aspartate-preferring subtype gating ion channels which are also permeable to Ca2+. Here we report that N-methyl-D-aspartate indeed has a dramatic effect on the survival in culture of cells derived from dissociated cerebella of 7-8-day-old rats and cultured in media containing 'low' [K+]e (5-15 mM). In addition to the visual inspection of the cultures, the effect of N-methyl-D-aspartate was quantitatively evaluated, using estimates related to the number of viable cells (determination of DNA and of reduction rate of a tetrazolium salt). Furthermore, proteins which are relatively enriched in either nerve cells (neuronal cell adhesion molecule, D3-protein and synaptin) or in glia (glutamine synthetase) were also measured. The findings showed that the rescue of cells by N-methyl-D-aspartate involved primarily nerve cells and that the survival requirement for N-methyl-D-aspartate, as for high K+, developed between 2 and 4 days in vitro. The effect depended on both the concentration of N-methyl-D-aspartate and the degree of depolarization of the cells: both the potency and the efficacy of N-methyl-D-aspartate were increased as [K+]e was raised from 5 to 15 mM, at which range K+ on its own has little if any influence on granule cell survival. These characteristics are consistent with the voltage-dependence of ion conductance through the N-methyl-D-aspartate receptor-linked channel. The most pronounced effect of N-methyl-D-aspartate was obtained in the presence of 15 mM K+, when cell survival approached that obtained in 'control' cultures (grown in 25 mM K

  7. Abscisic acid-induced gene expression in the liverwort Marchantia polymorpha is mediated by evolutionarily conserved promoter elements.

    PubMed

    Ghosh, Totan K; Kaneko, Midori; Akter, Khaleda; Murai, Shuhei; Komatsu, Kenji; Ishizaki, Kimitsune; Yamato, Katsuyuki T; Kohchi, Takayuki; Takezawa, Daisuke

    2016-04-01

    Abscisic acid (ABA) is a phytohormone widely distributed among members of the land plant lineage (Embryophyta), regulating dormancy, stomata closure and tolerance to environmental stresses. In angiosperms (Magnoliophyta), ABA-induced gene expression is mediated by promoter elements such as the G-box-like ACGT-core motifs recognized by bZIP transcription factors. In contrast, the mode of regulation by ABA of gene expression in liverworts (Marchantiophyta), representing one of the earliest diverging land plant groups, has not been elucidated. In this study, we used promoters of the liverwort Marchantia polymorpha dehydrin and the wheat Em genes fused to the β-glucuronidase (GUS) reporter gene to investigate ABA-induced gene expression in liverworts. Transient assays of cultured cells of Marchantia indicated that ACGT-core motifs proximal to the transcription initiation site play a role in the ABA-induced gene expression. The RY sequence recognized by B3 transcriptional regulators was also shown to be responsible for the ABA-induced gene expression. In transgenic Marchantia plants, ABA treatment elicited an increase in GUS expression in young gemmalings, which was abolished by simultaneous disruption of the ACGT-core and RY elements. ABA-induced GUS expression was less obvious in mature thalli than in young gemmalings, associated with reductions in sensitivity to exogenous ABA during gametophyte growth. In contrast, lunularic acid, which had been suggested to function as an ABA-like substance, had no effect on GUS expression. The results demonstrate the presence of ABA-specific response mechanisms mediated by conserved cis-regulatory elements in liverworts, implying that the mechanisms had been acquired in the common ancestors of embryophytes. PMID:26456006

  8. Retinoic Acid and Histone Deacetylases Regulate Epigenetic Changes in Embryonic Stem Cells*

    PubMed Central

    Urvalek, Alison M.; Gudas, Lorraine J.

    2014-01-01

    All-trans-retinoic acid (RA) is a vitamin A metabolite that plays major roles in regulating stem cell differentiation and development. RA is the ligand of the retinoic acid receptor (RAR) family of transcription factors, which interact with retinoic acid response elements (RAREs) within target gene proximal promoters and enhancers. Although RA-mediated gene activation is well understood, less is known about the mechanisms for repression at RA-regulated genes. Using chromatin immunoprecipitation experiments, we show that in embryonic stem cells in the absence of RA, histone deacetylases (HDACs) differentially bind to various RAREs in proximal promoters or enhancer regions of RA-regulated genes; HDAC1, HDAC2, and HDAC3 bind at RAREs in the Hoxa1 and Cyp26a1 gene regulatory regions, whereas only HDAC1 binds at the RARβ2 RARE. shRNA knockdown of HDAC1, HDAC2, or HDAC3 differentially increases the deposition of the histone 3 lysine 27 acetylation (H3K27ac) epigenetic mark associated with increases in these three transcripts. Importantly, RA treatment differentially mediates the removal of HDACs from the Hoxa1, Cyp26a1, and RARβ2 genes and promotes the deposition of the H3K27ac mark at these genes. Overall, we show that HDACs differentially bind to RA-regulated genes to control key epigenetic marks involved in stem cell differentiation. PMID:24821725

  9. Influence of decenylsuccinic Acid on water permeability of plant cells.

    PubMed

    Lee, O Y; Stadelmann, E J; Weiser, C J

    1972-11-01

    Decenylsuccinic acid altered permeability to water of epidermal cells of bulb scales of Allium cepa and of the leaf midrib of Rhoeo discolor. Water permeability, as determined by deplasmolysis time measurements, was related to the dose of undissociated decenylsuccinic acid (mm undissociated decenylsuccinic acid x minute). No relationship was found between permeability and total dose of decenylsuccinic acid, or dose of dissociated decenylsuccinic acid, suggesting that the undissociated molecule was the active factor in permeability changes and injury.At doses which did not damage cells (0.0008 to 0.6 [mm of the undissociated molecule x minute]) decenylsuccinic acid decreased water permeability. At higher doses (e.g., 4 to 8 [mm x minute]) injury to cells was common and decenylsuccinic acid increased permeability. Doses above the 10 to 20 (mm x minute) range were generally lethal. The plasmolysis form of uninjured cells was altered and protoplasmic swelling occasionally was observed. The dose-dependent reversal of water permeability changes (decreased to increased permeability) may reflect decenylsuccinic acid-induced changes in membrane structure. Reported effects of decenylsuccinic acid on temperature dependence of permeability and frost resistance were not verified. PMID:16658227

  10. The antiretroviral nucleoside analogue Abacavir reduces cell growth and promotes differentiation of human medulloblastoma cells

    PubMed Central

    Rossi, Alessandra; Russo, Giuseppe; Puca, Andrew; La Montagna, Raffaele; Caputo, Mariella; Mattioli, Eliseo; Lopez, Massimo; Giordano, Antonio; Pentimalli, Francesca

    2009-01-01

    Abacavir is one of the most efficacious nucleoside analogues, with a well-characterized inhibitory activity on reverse transcriptase enzymes of retroviral origin, and has been clinically approved for the treatment of AIDS. Recently, Abacavir has been shown to inhibit also the human telomerase activity. Telomerase activity seems to be required in essentially all tumours for the immortalization of a subset of cells, including cancer stem cells. In fact, many cancer cells are dependent on telomerase for their continued replication and therefore telomerase is an attractive target for cancer therapy. Telomerase expression is upregulated in primary primitive neuroectodermal tumours and in the majority of medulloblastomas suggesting that its activation is associated with the development of these diseases. Therefore, we decided to test Abacavir activity on human medulloblastoma cell lines with high telomerase activity. We report that exposure to Abacavir induces a dose-dependent decrease in the proliferation rate of medulloblastoma cells. This is associated with a cell accumulation in the G2/M phase of the cell cycle in the Daoy cell line, and with increased cell death in the D283-MED cell line, and is likely to be dependent on the inhibition of telomerase activity. Interestingly, both cell lines showed features of senescence after Abacavir treatment. Moreover, following Abacavir exposure we detected, by immunofluorescence staining, increased protein expression of the glial marker glial fibrillary acidic protein (GFAP) and the neuronal marker synaptophysin (SYN) in both medulloblastoma cell lines. In conclusion, our results suggest that Abacavir reduces proliferation and induces differentiation of human medulloblastoma cells through the downregulation of telomerase activity. Thus, using Abacavir, alone or in combination with current therapies, might be an effective therapeutic strategy for the treatment of medulloblastoma. PMID:19358275

  11. Wild-type human p53 transactivates the human proliferating cell nuclear antigen promoter

    SciTech Connect

    Shivakumar, C.V.; Brown, D.R.; Deb, S.; Deb, S.P.

    1995-12-01

    The p53 tumor suppressor protein negatively regulates cell growth and somatic mutations in the p53 gene lead to uncontrolled cell growth and oncogenesis. This report describes research which demonstrates, using a number of different cell lines, that at low levels, wild-type p53 transactivates the human proliferating cell nuclear antigen (PCNA) promoter. When expressed at similar levels, tumor-derived p53 mutants did not transactivate the PCNA promoter. It also reports the identification of a wild-type human p53-binding site on the human PCNA promote. 84 refs., 5 figs, 3 tabs.

  12. Retinoic acid stimulate differentiation of hippocampal stem cells into opsin expressing cells in vitro.

    PubMed

    Safari, M; Nobakht, M; Roshandel, N Rahbar; Ghazi, F; Joghataee, M T

    2009-09-01

    The results of several studies have demonstrated that cell differentiation influenced by derivatives of retinoic acid. To determine whether retinoic acid mediate the differentiation of neural stem cells we treated dissociated hippocampal stem cells with different concentrations of all trans or 9-cis retinoic acid and analyzed the effects on cell fate by specific monoclonal antibody for photoreceptors. Addition of exogenous retinoic acid caused a dose dependent specific in the elevation of the cell number that developed as photoreceptors in culture. Also results ofimmunohistochemical studies using monoclonal antibody demonstrated that the primary effect ofretinoic acid was to influence progenitor cells the developed as mature and immature photoreceptors. These results suggest that retinoic acid may play an important effect in the normal development of photoreceptor cells in vitro.

  13. Gallic acid suppresses cell viability, proliferation, invasion and angiogenesis in human glioma cells

    PubMed Central

    Lu, Yong; Jiang, Feng; Jiang, Hao; Wu, Kalina; Zheng, Xuguang; Cai, Yizhong; Katakowski, Mark; Chopp, Michael; To, Shing-Shun Tony

    2010-01-01

    Gallic acid, an organic acid, also known as 3,4,5-trihydroxybenzoic acid, is cytotoxic against certain cancer cells, without harming normal cells. The objective of this study is to evaluate whether gallic acid can inhibit glioma cell viability, proliferation, invasion and reduce glioma cell mediated angiogenesis. Treatment of U87 and U251n glioma cells with gallic acid inhibited cell viability in a dose- and time-dependent manner. BrdU and tube formation assays indicated that gallic acid significantly decreased glioma cell proliferation and tube formation in mouse brain endothelial cells, respectively. In addition, gallic acid decreased U87 cell invasion in vitro. Western blot analysis showed that expression of ADAM17, p-Akt and p-Erk was suppressed by gallic acid in both U87 and U251n cell lines. These data suggest that suppression of ADAM17 and downregulation of PI3K/Akt and Ras/MAPK signaling pathways may contribute to gallic acid-induced decrease of invasiveness. Gallic acid may be a valuable candidate for treatment of brain tumor. PMID:20553913

  14. HCG-Activated Human Peripheral Blood Mononuclear Cells (PBMC) Promote Trophoblast Cell Invasion

    PubMed Central

    Wang, Yaqin; Guo, Yue; Zhou, Danni; Xu, Mei; Ding, Jinli; Yang, Jing

    2015-01-01

    Successful embryo implantation and placentation depend on appropriate trophoblast invasion into the maternal endometrial stroma. Human chorionic gonadotropin (hCG) is one of the earliest embryo-derived secreted signals in the peripheral blood mononuclear cells (PBMC) that abundantly expresses hCG receptors. The aims of this study were to estimate the effect of human embryo–secreted hCG on PBMC function and investigate the role and underlying mechanisms of activated PBMC in trophoblast invasion. Blood samples were collected from women undergoing benign gynecological surgery during the mid-secretory phase. PBMC were isolated and stimulated with or without hCG for 0 or 24 h. Interleukin-1β (IL-1β) and leukemia inhibitory factor (LIF) expressions in PBMC were detected by enzyme-linked immunosorbent assay and real-time polymerase chain reaction (PCR). The JAR cell line served as a model for trophoblast cells and was divided into four groups: control, hCG only, PBMC only, and PBMC with hCG. JAR cell invasive and proliferative abilities were detected by trans-well and CCK8 assays and matrix metalloproteinase (MMP)-2 (MMP-2), MMP-9, vascular endothelial growth factor (VEGF), tissue inhibitor of metalloproteinase (TIMP)-1, and TIMP-2 expressions in JAR cells were detected by western blotting and real-time PCR analysis. We found that hCG can remarkably promote IL-1β and LIF promotion in PBMC after 24-h culture. PBMC activated by hCG significantly increased the number of invasive JAR cells in an invasion assay without affecting proliferation, and hCG-activated PBMC significantly increased MMP-2, MMP-9, and VEGF and decreased TIMP-1 and TIMP-2 expressions in JAR cells in a dose-dependent manner. This study demonstrated that hCG stimulates cytokine secretion in human PBMC and could stimulate trophoblast invasion. PMID:26087261

  15. Methylseleninic acid promotes antitumour effects via nuclear FOXO3a translocation through Akt inhibition

    PubMed Central

    Tarrado-Castellarnau, Míriam; Cortés, Roldán; Zanuy, Miriam; Tarragó-Celada, Josep; Polat, Ibrahim H.; Hill, Richard; Fan, Teresa W.; Link, Wolfgang; Cascante, Marta

    2016-01-01

    Selenium supplement has been shown in clinical trials to reduce the risk of different cancers including lung carcinoma. Previous studies reported that the antiproliferative and pro-apoptotic activities of methylseleninic acid (MSA) in cancer cells could be mediated by inhibition of the PI3K pathway. A better understanding of the downstream cellular targets of MSA will provide information on its mechanism of action and will help to optimise its use in combination therapies with PI3K inhibitors. For this study, the effects of MSA on viability, cell cycle, metabolism, apoptosis, protein and mRNA expression, and Reactive Oxygen Species production were analysed in A549 cells. FOXO3a subcellular localisation was examined in A549 cells and in stably transfected human osteosarcoma U2foxRELOC cells. Our results demonstrate that MSA induces FOXO3a nuclear translocation in A549 cells and in U2OS cells that stably express GFP-FOXO3a. Interestingly, sodium selenite, another selenium compound, did not induce any significant effects on FOXO3a translocation despite inducing apoptosis. Single strand break of DNA, disruption of tumour cell metabolic adaptations, decrease in ROS production, and cell cycle arrest in G1 accompanied by induction of apoptosis are late events occurring after 24 h of MSA treatment in A549 cells. Our findings suggest that FOXO3a is a relevant mediator of the antiproliferative effects of MSA. This new evidence on the mechanistic action of MSA can open new avenues in exploiting its antitumour properties and in the optimal design of novel combination therapies. We present MSA as a promising chemotherapeutic agent with synergistic antiproliferative effects with cisplatin. PMID:26375988

  16. G Protein-Coupled Receptor 87 (GPR87) Promotes Cell Proliferation in Human Bladder Cancer Cells.

    PubMed

    Zhang, Xia; Liu, Dage; Hayashida, Yushi; Okazoe, Homare; Hashimoto, Takeshi; Ueda, Nobufumi; Sugimoto, Mikio; Kakehi, Yoshiyuki

    2015-01-01

    G protein-coupled receptor 87 (GPR87) is a newly deorphanized member of the cell surface molecule G protein-coupled receptor family. GPR signaling was shown to play a role in promotion of cell growth and survival, metastasis, and drug resistance. The overexpression of GPR87 has also been reported in many malignant tumors including bladder cancer. The aim of the present study is to examine the effect of silencing GPR87 expression with a replication-deficient recombinant adenoviral vector expressing short hairpin RNA targeting GPR87 (Ad-shGPR87) and to explore the underlying molecular mechanisms in bladder cancer cells. Six GPR87-expressing human bladder cancer cells, HT1197, HT1376, J82, RT112, TCCSUP and UMUC3, were used. Infection with Ad-shGPR87 effectively downregulated the GPR87 expression, and significantly reduced the percentage of viable cells in 4 of 6 cell lines as detected by an MTT assay. Significant inhibition on cell proliferation with Ad-shGPR87 was observed in the wild-type p53 bladder cancer cell lines (HT1197, RT112, TCCSUP and UMUC3), but not in the mutant p53 cells (HT1376 and J82). As represented by a wild-type p53 RT112 cell, Ad-shGPR87 infection significantly enhanced p53 and p21 expression and caused caspase-dependent apoptosis. Furthermore, the treatment with Ad-shGPR87 exerted a significant antitumor effect against the GPR87-expressing RT112 xenografts. GPR87 appeared to be a promising target for gene therapy, and Ad-shGPR87 had strong antitumor effects, specifically anti-proliferative and pro-apoptotic effects, against GPR87-expressing human bladder cancer cells. PMID:26473854

  17. Transformed MDCK cells secrete elevated MMP1 that generates LAMA5 fragments promoting endothelial cell angiogenesis

    PubMed Central

    Gopal, Shashi K.; Greening, David W.; Zhu, Hong-Jian; Simpson, Richard J.; Mathias, Rommel A.

    2016-01-01

    Epithelial-mesenchymal transition (EMT) enhances the migration and invasion of cancer cells, and is regulated by various molecular mechanisms including extracellular matrix metalloproteinase (MMP) activity. Previously, we reported transformation of epithelial Madin-Darby canine kidney (MDCK) cells with oncogenic H-Ras (21D1 cells) induces EMT, and significantly elevates MMP1 expression. To explore the biological significance, in this study we characterized 21D1 cells with knocked-down MMP1 expression (21D1−MMP1). MMP1 silencing diminished 21D1 cell migration, invasion and anchorage-independent growth in vitro. Additionally, 21D1−MMP1 cells displayed reduced tumour volume when grown as in vivo subcutaneous xenografts in mice. Depletion of MMP1 lowered the ability of the cellular secretome (extracellular culture medium) to influence recipient cell behaviour. For example, supplementation with 21D1 secretome elevated cell migration of recipient fibroblasts, and enhanced endothelial cell angiogenesis (vessel length and branching). By contrast, 21D1−MMP1 secretome was less potent in both functional assays. We reveal laminin subunit alpha-5 (LAMA5) as a novel biological substrate of MMP1, that generates internal and C-terminal proteolytic fragments in 21D1 secretome. Furthermore, antibody-based inhibition of integrin αvβ3 on endothelial cells nullified the angiogenic capability of 21D1 secretome. Therefore, we report this as a new VEGF-independent mechanism that oncogenic cells may employ to promote tumour angiogenesis. PMID:27324842

  18. PPARγ Agonists Promote Oligodendrocyte Differentiation of Neural Stem Cells by Modulating Stemness and Differentiation Genes

    PubMed Central

    Kanakasabai, Saravanan; Pestereva, Ecaterina; Chearwae, Wanida; Gupta, Sushil K.; Ansari, Saif; Bright, John J.

    2012-01-01

    Neural stem cells (NSCs) are a small population of resident cells that can grow, migrate and differentiate into neuro-glial cells in the central nervous system (CNS). Peroxisome proliferator-activated receptor gamma (PPARγ) is a nuclear receptor transcription factor that regulates cell growth and differentiation. In this study we analyzed the influence of PPARγ agonists on neural stem cell growth and differentiation in culture. We found that in vitro culture of mouse NSCs in neurobasal medium with B27 in the presence of epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF) induced their growth and expansion as neurospheres. Addition of all-trans retinoic acid (ATRA) and PPARγ agonist ciglitazone or 15-Deoxy-Δ12,14-Prostaglandin J2 (15d-PGJ2) resulted in a dose-dependent inhibition of cell viability and proliferation of NSCs in culture. Interestingly, NSCs cultured with PPARγ agonists, but not ATRA, showed significant increase in oligodendrocyte precursor-specific O4 and NG2 reactivity with a reduction in NSC marker nestin, in 3–7 days. In vitro treatment with PPARγ agonists and ATRA also induced modest increase in the expression of neuronal β-III tubulin and astrocyte-specific GFAP in NSCs in 3–7 days. Further analyses showed that PPARγ agonists and ATRA induced significant alterations in the expression of many stemness and differentiation genes associated with neuro-glial differentiation in NSCs. These findings highlight the influence of PPARγ agonists in promoting neuro-glial differentiation of NSCs and its significance in the treatment of neurodegenerative diseases. PMID:23185633

  19. Clove extract inhibits tumor growth and promotes cell cycle arrest and apoptosis.

    PubMed

    Liu, Haizhou; Schmitz, John C; Wei, Jianteng; Cao, Shousong; Beumer, Jan H; Strychor, Sandra; Cheng, Linyou; Liu, Ming; Wang, Cuicui; Wu, Ning; Zhao, Xiangzhong; Zhang, Yuyan; Liao, Joshua; Chu, Edward; Lin, Xiukun

    2014-01-01

    Cloves (Syzygium aromaticum) have been used as a traditional Chinese medicinal herb for thousands of years. Cloves possess antiseptic, antibacterial, antifungal, and antiviral properties, but their potential anticancer activity remains unknown. In this study, we investigated the in vitro and in vivo antitumor effects and biological mechanisms of ethyl acetate extract of cloves (EAEC) and the potential bioactive components responsible for its antitumor activity. The effects of EAEC on cell growth, cell cycle distribution, and apoptosis were investigated using human cancer cell lines. The molecular changes associated with the effects of EAEC were analyzed by Western blot and (qRT)-PCR analysis. The in vivo effect of EAEC and its bioactive component was investigated using the HT-29 tumor xenograft model. We identified oleanolic acid (OA) as one of the components of EAEC responsible for its antitumor activity. Both EAEC and OA display cytotoxicity against several human cancer cell lines. Interestingly, EAEC was superior to OA and the chemotherapeutic agent 5-fluorouracil at suppressing growth of colon tumor xenografts. EAEC promoted G0/G1 cell cycle arrest and induced apoptosis in a dose-dependent manner. Treatment with EAEC and OA selectively increased protein expression of p21(WAF1/Cip1) and γ-H2AX and downregulated expression of cell cycle-regulated proteins. Moreover, many of these changes were at the mRNA level, suggesting transcriptional regulation by EAEC treatment. Our results demonstrate that clove extract may represent a novel therapeutic herb for the treatment of colorectal cancer, and OA appears to be one of the bioactive components.

  20. Effect on epidermal cell of soybean protein-degraded products and structural determination of the root hair promoting peptide.

    PubMed

    Matsumiya, Yoshiki; Sumiyoshi, Sayoko; Matsukura, Takuma; Kubo, Motoki

    2007-11-01

    Peptide(s) produced from degraded soybean protein by an alkaline protease from Bacillus circulans HA12 (degraded soybean-meal products; DSP) increased the number of both the root hair cells (trichoblasts) and hairless cells (atrichoblasts) of Brassica rapa by about 4.4 times and 1.9 times, respectively. To identify the root hair-promoting peptide(s) in DSP, the origin protein of the root hair-promoting peptide(s) was identified as Kunitz trypsin inhibitor (KTI). The root hair-promoting peptide in the degraded products of KTI was purified and produced a signal of 1,198.2 Da with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) analysis. A search of the amino acid sequence of KTI located the peptide GGIRAAPTGNER, which had a molecular weight identical to 1,198.2 Da. The peptide GGIRAAPTGNER was chemically synthesized, and the synthetic peptide possessed root hair-promoting activity. Thus, it is concluded that this peptide in DSP is the foreign bioactive peptide promoting the differentiation of root hairs.

  1. KDM1A promotes tumor cell invasion by silencing TIMP3 in non-small cell lung cancer cells

    PubMed Central

    Li, Wang; Yang, Yan; Tian, Ye; Wang, Xujun; Chen, Sujun; Yang, Yuxin; Huang, Tianhao; Zhao, Tian; Tang, Liang; Su, Bo; Li, Fei; Liu, X. Shirley; Zhang, Fan

    2016-01-01

    Epigenetic regulation plays an important role in tumor metastasis. KDM1A is a histone demethylase specific for H3K4me2/me1 demethylation, and has been found to be overexpressed in many cancers, including non-small cell lung cancer (NSCLC). However, the role of KDM1A in lung cancer remains unclear. Here, we show that KDM1A promotes cancer metastasis in NSCLC cells by repressing TIMP3 (tissue inhibitor of metalloproteinase 3) expression. Consistently with this, overexpression of TIMP3 inhibited MMP2 expression and JNK phosphorylation, both of which are known to be important for cell invasion and migration. Importantly, knockdown of TIMP3 in KDM1A-deficient cells rescued the metastatic capability of NSCLC cells. These findings were also confirmed by pharmacological inhibition assays. We further demonstrate that KDM1A removes H3K4me2 at the promoter of TIMP3, thus repressing the transcription of TIMP3. Finally, high expression of KDM1A and low expression of TIMP3 significantly correlate with a poor prognosis in NSCLC patients. This study establishes a mechanism by which KDM1A promotes cancer metastasis in NSCLC cells, and we suggest that KDM1A may be a potential therapeutic target for NSCLC treatment. PMID:27058897

  2. A betacellulin mutant promotes differentiation of pancreatic acinar AR42J cells into insulin-producing cells with low affinity of binding to ErbB1.

    PubMed

    Nagaoka, Tadahiro; Fukuda, Takayuki; Hashizume, Toshihiro; Nishiyama, Tomoko; Tada, Hiroko; Yamada, Hidenori; Salomon, David S; Yamada, Satoko; Kojima, Itaru; Seno, Masaharu

    2008-06-27

    Betacellulin (BTC) is one of the members of the epidermal growth factor (EGF) ligand family of ErbB receptor tyrosine kinases. It is a differentiation factor as well as a potent mitogen. BTC promotes the differentiation of pancreatic acinar-derived AR42J cells into insulin-producing cells. It independently and preferentially binds to two type I tyrosine kinase receptors, the EGF receptor (ErbB1) and ErbB4. However, the physiochemical characteristics of BTC that are responsible for its preferential binding to these two receptors have not been fully defined. In this study, to investigate the essential amino acid residues of BTC for binding to the two receptors, we introduced point mutations into the EGF domain of BTC employing error-prone PCR. The receptor binding abilities of 190 mutants expressed in Escherichia coli were assessed by enzyme immunoassay. Replacement of the glutamic acid residue at position 88 with a lysine residue in BTC was found to produce a significant loss of affinity for binding to ErbB1, while the affinity of binding to ErbB4 was unchanged. In addition, the mutant of BTC-E/88/K showed less growth-promoting activity on BALB/c 3T3 cells compared with that of the wild-type BTC protein. Interestingly, the BTC mutant protein promoted differentiation of pancreatic acinar AR42J cells at a high frequency into insulin-producing cells compared with AR42J cells that were treated with wild-type BTC protein. These results indicate the possibility of designing BTC mutants, which have an activity of inducing differentiation only, without facilitating growth promotion. PMID:18508082

  3. Differentiation of rabbit bone mesenchymal stem cells into endothelial cells in vitro and promotion of defective bone regeneration in vivo.

    PubMed

    Liu, Jinzhong; Liu, Chao; Sun, Bin; Shi, Ce; Qiao, Chunyan; Ke, Xiaoliang; Liu, Shutai; Liu, Xia; Sun, Hongchen

    2014-04-01

    Tissue engineering strategies often fail to regenerate bones because of inadequate vascularization, especially in the reconstruction of large segmental bone defects. Large volumes of vascular endothelial cells (ECs) that functionally interact with osteoblasts during osteogenesis are difficult to obtain. In this study, we simulated bone healing by co-culturing differentiated ECs and mesenchymal stem cells (MSCs) either on a culture plate or on a polylactide glycolic acid (PLGA) scaffold in vitro. We also evaluated the effect of osteogenesis in repairing rabbit mandible defects in vivo. In this study, MSCs were separated from rabbit as the seed cells. After passage, the MSCs were cultured in an EC-conditioned medium to differentiate into ECs. Immunohistochemical staining analysis with CD34 showed that the induced cells had the characteristics of ECs and MSC. The induced ECs were co-cultured in vitro, and the induction of MSCs to osteoblast served as the control. Alkaline phosphatase (ALP) and alizarin red (AZR) staining experiments were performed, and the Coomassie brilliant blue total protein and ALP activity were measured. The MSCs proliferated and differentiated into osteoblast-like cells through direct contact between the derived ECs and MSCs. The co-cultured cells were seeded on PLGA scaffold to repair 1 cm mandible defects in the rabbit. The effectiveness of the repairs was assessed through soft X-ray and histological analyses. The main findings indicated that MSCs survived well on the scaffold and that the scaffold is biocompatible and noncytotoxic. The results demonstrated that the co-cultured MSC-derived ECs improved MSC osteogenesis and promoted new bone formation. This study may serve as a basis for the use of in vitro co-culturing techniques as an improvisation to bone tissue engineering for the repair of large bone defects.

  4. Interleukin-8 promotes cell migration through integrin αvβ6 upregulation in colorectal cancer.

    PubMed

    Sun, Qi; Sun, Fengkai; Wang, Ben; Liu, Song; Niu, Weibo; Liu, Enyu; Peng, Cheng; Wang, Jiayong; Gao, Huijie; Liang, Benjia; Niu, Zhengchuan; Zou, Xueqing; Niu, Jun

    2014-11-28

    Colorectal cancer (CRC), which is notorious for high morbidity and mortality around the world, shows a predilection for metastasis to liver. Interleukin-8 (IL-8), a chemokine with a defining CXC amino acid motif, has been reported to promote CRC cell migration and is associated with poor prognosis of CRC. However, the underlying molecular mechanism of IL-8-mediated migration remains obscure. In this study, we first demonstrated the cross talk between IL-8 and integrin αvβ6. We analyzed 139 human CRC samples, and found that the immunohistochemical expression of αvβ6 was significantly correlated with expression of IL-8. Furthermore, IL-8 increased the migration through integrin αvβ6 in human CRC cells, and both CXCR1 and CXCR2 were primarily involved during the process. IL-8 upregulated αvβ6 expression in a dose-dependent manner through activation of ERK and Ets-1 signaling pathway. Taken together, our results indicated that IL-8 enhances the migration of CRC cells by increasing αvβ6 integrin expression through the ERK/Ets-1 pathway. Targeting integrin αvβ6 in IL-8 expressing tumors might be a potential therapeutic strategy for CRC patients.

  5. Nanotopography Promotes Pancreatic Differentiation of Human Embryonic Stem Cells and Induced Pluripotent Stem Cells.

    PubMed

    Kim, Jong Hyun; Kim, Hyung Woo; Cha, Kyoung Je; Han, Jiyou; Jang, Yu Jin; Kim, Dong Sung; Kim, Jong-Hoon

    2016-03-22

    Although previous studies suggest that nanotopographical features influence properties and behaviors of stem cells, only a few studies have attempted to derive clinically useful somatic cells from human pluripotent stem cells using nanopatterned surfaces. In the present study, we report that polystyrene nanopore-patterned surfaces significantly promote the pancreatic differentiation of human embryonic and induced pluripotent stem cells. We compared different diameters of nanopores and showed that 200 nm nanopore-patterned surfaces highly upregulated the expression of PDX1, a critical transcription factor for pancreatic development, leading to an approximately 3-fold increase in the percentage of differentiating PDX1(+) pancreatic progenitors compared with control flat surfaces. Furthermore, in the presence of biochemical factors, 200 nm nanopore-patterned surfaces profoundly enhanced the derivation of pancreatic endocrine cells producing insulin, glucagon, or somatostatin. We also demonstrate that nanopore-patterned surface-induced upregulation of PDX1 is associated with downregulation of TAZ, suggesting the potential role of TAZ in nanopore-patterned surface-mediated mechanotransduction. Our study suggests that appropriate cytokine treatments combined with nanotopographical stimulation could be a powerful tool for deriving a high purity of desired cells from human pluripotent stem cells. PMID:26900863

  6. MicroRNA-196b promotes cell proliferation and suppress cell differentiation in vitro

    SciTech Connect

    Cao, Donglin Hu, Liangshan; Lei, Da; Fang, Xiaolin; Zhang, Zhihong; Wang, Ting; Lin, Maorui; Huang, Jiwei; Yang, Huawen; Zhou, Xuan; Zhong, Limei

    2015-01-30

    Highlights: • miRNA-196b increases proliferation and blocks differentiation of progenitor cell. • miRNA-196b inhibits apoptosis and increases viability of cells lines. • Forced expression of miR-196b blocks the differentiation of THP1 induced by PMA. - Abstract: MicroRNA-196b (miR-196b) is frequently amplified and aberrantly overexpressed in acute leukemias. To investigate the role of miR-196b in acute leukemias, it has been observed that forced expression of this miRNA increases proliferation and inhibits apoptosis in human cell lines. More importantly, we show that this miRNA can significantly increase the colony-forming capacity of mouse normal bone marrow progenitor cells alone, as well as partially blocking the cells from differentiation. Taken together, our studies suggest that miRNA-196b may play an essential role in the development of MLL-associated leukemias through inhibiting cell differentiation and apoptosis, while promoting cell proliferation.

  7. Nanotopography Promotes Pancreatic Differentiation of Human Embryonic Stem Cells and Induced Pluripotent Stem Cells.

    PubMed

    Kim, Jong Hyun; Kim, Hyung Woo; Cha, Kyoung Je; Han, Jiyou; Jang, Yu Jin; Kim, Dong Sung; Kim, Jong-Hoon

    2016-03-22

    Although previous studies suggest that nanotopographical features influence properties and behaviors of stem cells, only a few studies have attempted to derive clinically useful somatic cells from human pluripotent stem cells using nanopatterned surfaces. In the present study, we report that polystyrene nanopore-patterned surfaces significantly promote the pancreatic differentiation of human embryonic and induced pluripotent stem cells. We compared different diameters of nanopores and showed that 200 nm nanopore-patterned surfaces highly upregulated the expression of PDX1, a critical transcription factor for pancreatic development, leading to an approximately 3-fold increase in the percentage of differentiating PDX1(+) pancreatic progenitors compared with control flat surfaces. Furthermore, in the presence of biochemical factors, 200 nm nanopore-patterned surfaces profoundly enhanced the derivation of pancreatic endocrine cells producing insulin, glucagon, or somatostatin. We also demonstrate that nanopore-patterned surface-induced upregulation of PDX1 is associated with downregulation of TAZ, suggesting the potential role of TAZ in nanopore-patterned surface-mediated mechanotransduction. Our study suggests that appropriate cytokine treatments combined with nanotopographical stimulation could be a powerful tool for deriving a high purity of desired cells from human pluripotent stem cells.

  8. Retinoic acid inhibits the cytoproliferative response to weak 50-Hz magnetic fields in neuroblastoma cells

    PubMed Central

    TRILLO, MARÍA ÁNGELES; MARTÍNEZ, MARÍA ANTONIA; CID, MARÍA ANTONIA; ÚBEDA, ALEJANDRO

    2012-01-01

    We previously reported that intermittent exposure to a 50-Hz magnetic field (MF) at 100 μT stimulates cell proliferation in the human neuroblastoma cell line NB69. The present study aimed to investigate whether the magnetic field-induced growth promotion also occurs at a lower magnetic flux density of 10 μT. To this purpose, NB69 cells were subjected for 42 h to intermittent exposure, 3 h on/3 h off, to a 50-Hz MF at a 10 or 100 μT magnetic flux density. The field exposure took place either in the presence or in the absence of the antiproliferative agent retinoic acid. At the end of the treatment and/or incubation period, the cell growth was estimated by hemocytometric counting and spectrophotometric analysis of total protein and DNA contents. Potential changes in DNA synthesis were also assessed through proliferating cell nuclear antigen (PCNA) immunolabeling. The results confirmed previously reported data that a 42-h exposure to a 50-Hz sine wave MF at 100 μT promotes cell growth in the NB69 cell line, and showed that 10 μT induces a similar proliferative response. This effect, which was significantly associated and linearly correlated with PCNA expression, was abolished by the presence of retinoic acid in the culture medium. PMID:23292364

  9. MicroRNA-7 Promotes Glycolysis to Protect against 1-Methyl-4-phenylpyridinium-induced Cell Death.

    PubMed

    Chaudhuri, Amrita Datta; Kabaria, Savan; Choi, Doo Chul; Mouradian, M Maral; Junn, Eunsung

    2015-05-01

    Parkinson disease is associated with decreased activity of the mitochondrial electron transport chain. This defect can be recapitulated in vitro by challenging dopaminergic cells with 1-methyl-4-phenylpyridinium (MPP(+)), a neurotoxin that inhibits complex I of electron transport chain. Consequently, oxidative phosphorylation is blocked, and cells become dependent on glycolysis for ATP production. Therefore, increasing the rate of glycolysis might help cells to produce more ATP to meet their energy demands. In the present study, we show that microRNA-7, a non-coding RNA that protects dopaminergic neuronal cells against MPP(+)-induced cell death, promotes glycolysis in dopaminergic SH-SY5Y and differentiated human neural progenitor ReNcell VM cells, as evidenced by increased ATP production, glucose consumption, and lactic acid production. Through a series of experiments, we demonstrate that targeted repression of RelA by microRNA-7, as well as subsequent increase in the neuronal glucose transporter 3 (Glut3), underlies this glycolysis-promoting effect. Consistently, silencing Glut3 expression diminishes the protective effect of microRNA-7 against MPP(+). Further, microRNA-7 fails to prevent MPP(+)-induced cell death when SH-SY5Y cells are cultured in a low glucose medium, as well as when differentiated ReNcell VM cells or primary mouse neurons are treated with the hexokinase inhibitor, 2-deoxy-d-glucose, indicating that a functional glycolytic pathway is required for this protective effect. In conclusion, microRNA-7, by down-regulating RelA, augments Glut3 expression, promotes glycolysis, and subsequently prevents MPP(+)-induced cell death. This protective effect of microRNA-7 could be exploited to correct the defects in oxidative phosphorylation in Parkinson disease. PMID:25814668

  10. GENETIC MODIFICATION OF GIBBERELLIC ACID SIGNALING TO PROMOTE CARBON SEQUESTRATION IN TREE ROOTS AND STEMS

    SciTech Connect

    Busov, Victor

    2013-03-05

    -oxidases predominantly expressed in roots also decreased lateral root formation. GAs negatively affected lateral root formation by inhibiting lateral root primordium initiation. A whole-genome microarray analysis of root development in GA-modified transgenic plants revealed 2069 genes with significantly altered expression. The expression of 1178 genes, including genes that promote cell proliferation, growth, and cell wall loosening, corresponded to the phenotypic severity of the root traits when transgenic events with differential phenotypic expression were compared. The array data and direct hormone measurements suggested crosstalk of GA signaling with other hormone pathways, including auxin and abscisic acid. Transgenic modification of a differentially expressed gene encoding an auxin efflux carrier suggests that GA modulation of lateral root development is at least partly imparted by polar auxin transport modification. These results suggest a mechanism for GA-regulated modulation of lateral root proliferation associated with regulation of plant allometry during the stress response. Here we summarize progress in identification of three classes of genes useful for control of plant architecture: those affecting hormone metabolism and signaling; transcription and other regulatory factors; and the cell cycle. We focus on strong modifiers of stature and form that may be useful for directed modification of plant architecture, rather than the detailed mechanisms of gene action. Gibberellin (GA) metabolic and response genes are particularly attractive targets for manipulation because many act in a dose-dependent manner; similar phenotypic effects can be readily achieved in heterologous species; and induced pleiotropic effects--such as on nitrogen assimilation, photosynthesis, and lateral root production--are usually positive with respect to crop performance. Genes encoding transcription factors represent strong candidates for manipulation of plant architecture. For example

  11. Retinoic Acid Stimulates Regeneration of Mammalian Auditory Hair Cells

    NASA Astrophysics Data System (ADS)

    Lefebvre, Philippe P.; Malgrange, Brigitte; Staecker, Hinrich; Moonen, Gustave; van de Water, Thomas R.

    1993-04-01

    Sensorineural hearing loss resulting from the loss of auditory hair cells is thought to be irreversible in mammals. This study provides evidence that retinoic acid can stimulate the regeneration in vitro of mammalian auditory hair cells in ototoxic-poisoned organ of Corti explants in the rat. In contrast, treatment with retinoic acid does not stimulate the formation of extra hair cells in control cultures of Corti's organ. Retinoic acid-stimulated hair cell regeneration can be blocked by cytosine arabinoside, which suggests that a period of mitosis is required for the regeneration of auditory hair cells in this system. These results provide hope for a recovery of hearing function in mammals after auditory hair cell damage.

  12. Nucleic Acid Binding by Mason-Pfizer Monkey Virus CA Promotes Virus Assembly and Genome Packaging

    PubMed Central

    Füzik, Tibor; Píchalová, Růžena; Schur, Florian K. M.; Strohalmová, Karolína; Křížová, Ivana; Hadravová, Romana; Rumlová, Michaela; Briggs, John A. G.

    2016-01-01

    ABSTRACT The Gag polyprotein of retroviruses drives immature virus assembly by forming hexameric protein lattices. The assembly is primarily mediated by protein-protein interactions between capsid (CA) domains and by interactions between nucleocapsid (NC) domains and RNA. Specific interactions between NC and the viral RNA are required for genome packaging. Previously reported cryoelectron microscopy analysis of immature Mason-Pfizer monkey virus (M-PMV) particles suggested that a basic region (residues RKK) in CA may serve as an additional binding site for nucleic acids. Here, we have introduced mutations into the RKK region in both bacterial and proviral M-PMV vectors and have assessed their impact on M-PMV assembly, structure, RNA binding, budding/release, nuclear trafficking, and infectivity using in vitro and in vivo systems. Our data indicate that the RKK region binds and structures nucleic acid that serves to promote virus particle assembly in the cytoplasm. Moreover, the RKK region appears to be important for recruitment of viral genomic RNA into Gag particles, and this function could be linked to changes in nuclear trafficking. Together these observations suggest that in M-PMV, direct interactions between CA and nucleic acid play important functions in the late stages of the viral life cycle. IMPORTANCE Assembly of retrovirus particles is driven by the Gag polyprotein, which can self-assemble to form virus particles and interact with RNA to recruit the viral genome into the particles. Generally, the capsid domains of Gag contribute to essential protein-protein interactions during assembly, while the nucleocapsid domain interacts with RNA. The interactions between the nucleocapsid domain and RNA are important both for identifying the genome and for self-assembly of Gag molecules. Here, we show that a region of basic residues in the capsid protein of the betaretrovirus Mason-Pfizer monkey virus (M-PMV) contributes to interaction of Gag with nucleic acid. This

  13. Retinoic acid activates human inducible nitric oxide synthase gene through binding of RAR{alpha}/RXR{alpha} heterodimer to a novel retinoic acid response element in the promoter

    SciTech Connect

    Zou Fang; Liu Yan; Liu Li; Wu Kailang; Wei Wei; Zhu Ying . E-mail: yingzhu@whu.edu.cn; Wu Jianguo . E-mail: wu9988@vip.sina.com

    2007-04-06

    Human inducible nitric oxide synthase (hiNOS) catalyzes nitric oxide (NO) which has a significant effect on tumor suppression and cancer therapy. Here we revealed the detailed molecular mechanism involved in the regulation of hiNOS expression induced by retinoic acid (RA). We showed that RAR{alpha}/RXR{alpha} heterodimer was important in hiNOS promoter activation, hiNOS protein expression, and NO production. Serial deletion and site-directed mutation analysis revealed two half-sites of retinoic acid response element (RARE) spaced by 5 bp located at -172 to -156 in the hiNOS promoter. EMSA and ChIP assays demonstrated that RAR{alpha}/RXR{alpha} directly bound to this RARE of hiNOS promoter. Our results suggested the identification of a novel RARE in the hiNOS promoter and the roles of the nuclear receptors (RAR{alpha}/RXR{alpha}) in the induction of hiNOS by RA.

  14. Sucrose Loading in Isolated Veins of Pisum sativum: Regulation by Abscisic Acid, Gibberellic Acid, and Cell Turgor.

    PubMed

    Estruch, J J; Peretó, J G; Vercher, Y; Beltrán, J P

    1989-09-01

    Enzymatically isolated vein networks from mature pea (Pisum sativum L. cv Alaska) leaves were employed to investigate the properties of sucrose loading and the effect of phytohormones and cell turgor on this process. The sucrose uptake showed two components: a saturable and a first-order kinetics system. The high affinity system (K(m), 3.3 millimolar) was located at the plasmalemma (p-chloromercuriphenylsulfonic acid and orthovanadate sensitivity). Further characterization of this system, including pH dependence and effects of energy metabolism inhibitors, supported the H(+)-sugar symport concept for sucrose loading. Within a physiological range (0.1-100 micromolar) and after 90 min, abscisic acid (ABA) inhibited and gibberellic acid (GA(3)) promoted 1 millimolar sucrose uptake. These responses were partially (ABA) or totally (GA(3)) turgor-dependent. In experiments of combined hormonal treatments, ABA counteracted the GA(3) positive effects on sucrose uptake. The abolishment of these responses by p-chloromercuriphenylsulfonic acid and experiments on proton flux suggest that both factors (cell turgor and hormones) are modulating the H(+) ATPase plasmalemma activity. The results are discussed in terms of their physiological relevance. PMID:16667007

  15. Lipophilic caffeic and ferulic acid derivatives presenting cytotoxicity against human breast cancer cells.

    PubMed

    Serafim, Teresa L; Carvalho, Filipa S; Marques, Maria P M; Calheiros, Rita; Silva, Tiago; Garrido, Jorge; Milhazes, Nuno; Borges, Fernanda; Roleira, Fernanda; Silva, Elisiaario T; Holy, Jon; Oliveira, Paulo J

    2011-05-16

    In the present work, lipophilic caffeic and ferulic acid derivatives were synthesized, and their cytotoxicity on cultured breast cancer cells was compared. A total of six compounds were initially evaluated: caffeic acid (CA), hexyl caffeate (HC), caffeoylhexylamide (HCA), ferulic acid (FA), hexyl ferulate (HF), and feruloylhexylamide (HFA). Cell proliferation, cell cycle progression, and apoptotic signaling were investigated in three human breast cancer cell lines, including estrogen-sensitive (MCF-7) and insensitive (MDA-MB-231 and HS578T). Furthermore, direct mitochondrial effects of parent and modified compounds were investigated by using isolated liver mitochondria. The results indicated that although the parent compounds presented no cytotoxicity, the new compounds inhibited cell proliferation and induced cell cycle alterations and cell death, with a predominant effect on MCF-7 cells. Interestingly, cell cycle data indicates that effects on nontumor BJ fibroblasts were predominantly cytostatic and not cytotoxic. The parent compounds and derivatives also promoted direct alterations on hepatic mitochondrial bioenergetics, although the most unexpected and never before reported one was that FA induces the mitochondrial permeability transition. The results show that the new caffeic and ferulic acid lipophilic derivatives show increased cytotoxicity toward human breast cancer cell lines, although the magnitude and type of effects appear to be dependent on the cell type. Mitochondrial data had no direct correspondence with effects on intact cells suggesting that this organelle may not be a critical component of the cellular effects observed. The data provide a rational approach to the design of effective cytotoxic lipophilic hydroxycinnamic derivatives that in the future could be profitably applied for chemopreventive and/or chemotherapeutic purposes.

  16. RPS27a promotes proliferation, regulates cell cycle progression and inhibits apoptosis of leukemia cells

    SciTech Connect

    Wang, Houcai; Yu, Jing; Zhang, Lixia; Xiong, Yuanyuan; Chen, Shuying; Xing, Haiyan; Tian, Zheng; Tang, Kejing; Wei, Hui; Rao, Qing; Wang, Min; Wang, Jianxiang

    2014-04-18

    Highlights: • RPS27a expression was up-regulated in advanced-phase CML and AL patients. • RPS27a knockdown changed biological property of K562 and K562/G01 cells. • RPS27a knockdown affected Raf/MEK/ERK, P21 and BCL-2 signaling pathways. • RPS27a knockdown may be applicable for new combination therapy in CML patients. - Abstract: Ribosomal protein S27a (RPS27a) could perform extra-ribosomal functions besides imparting a role in ribosome biogenesis and post-translational modifications of proteins. The high expression level of RPS27a was reported in solid tumors, and we found that the expression level of RPS27a was up-regulated in advanced-phase chronic myeloid leukemia (CML) and acute leukemia (AL) patients. In this study, we explored the function of RPS27a in leukemia cells by using CML cell line K562 cells and its imatinib resistant cell line K562/G01 cells. It was observed that the expression level of RPS27a was high in K562 cells and even higher in K562/G01 cells. Further analysis revealed that RPS27a knockdown by shRNA in both K562 and K562G01 cells inhibited the cell viability, induced cell cycle arrest at S and G2/M phases and increased cell apoptosis induced by imatinib. Combination of shRNA with imatinib treatment could lead to more cleaved PARP and cleaved caspase-3 expression in RPS27a knockdown cells. Further, it was found that phospho-ERK(p-ERK) and BCL-2 were down-regulated and P21 up-regulated in RPS27a knockdown cells. In conclusion, RPS27a promotes proliferation, regulates cell cycle progression and inhibits apoptosis of leukemia cells. It appears that drugs targeting RPS27a combining with tyrosine kinase inhibitor (TKI) might represent a novel therapy strategy in TKI resistant CML patients.

  17. Cdk1 Phosphorylates SPAT-1/Bora to Promote Plk1 Activation in C. elegans and Human Cells.

    PubMed

    Thomas, Yann; Cirillo, Luca; Panbianco, Costanza; Martino, L