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Sample records for prevents p53-mediated apoptosis

  1. Zbtb1 Safeguards Genome Integrity and Prevents p53-Mediated Apoptosis in Proliferating Lymphoid Progenitors.

    PubMed

    Cao, Xin; Lu, Ying; Zhang, Xianyu; Kovalovsky, Damian

    2016-08-15

    Expression of the transcription factor Zbtb1 is required for normal lymphoid development. We report in the present study that Zbtb1 maintains genome integrity in immune progenitors, without which cells undergo increased DNA damage and p53-mediated apoptosis during replication and differentiation. Increased DNA damage in Zbtb1-mutant (ScanT) progenitors was due to increased sensitivity to replication stress, which was a consequence of inefficient activation of the S-phase checkpoint response. Increased p53-mediated apoptosis affected not only lymphoid but also myeloid development in competitive bone marrow chimeras, and prevention of apoptosis by transgenic Bcl2 expression and p53 deficiency rescued lymphoid as well as myeloid development from Zbtb1-mutant progenitors. Interestingly, however, protection from apoptosis rescued only the early stages of T cell development, and thymocytes remained arrested at the double-negative 3 developmental stage, indicating a strict requirement of Zbtb1 at later T cell developmental stages. Collectively, these results indicate that Zbtb1 prevents DNA damage in replicating immune progenitors, allowing the generation of B cells, T cells, and myeloid cells. PMID:27402700

  2. Sodium orthovanadate inhibits p53-mediated apoptosis.

    PubMed

    Morita, Akinori; Yamamoto, Shinichi; Wang, Bing; Tanaka, Kaoru; Suzuki, Norio; Aoki, Shin; Ito, Azusa; Nanao, Tomohisa; Ohya, Soichiro; Yoshino, Minako; Zhu, Jin; Enomoto, Atsushi; Matsumoto, Yoshihisa; Funatsu, Osamu; Hosoi, Yoshio; Ikekita, Masahiko

    2010-01-01

    Sodium orthovanadate (vanadate) inhibits the DNA-binding activity of p53, but its precise effects on p53 function have not been examined. Here, we show that vanadate exerts a potent antiapoptotic activity through both transcription-dependent and transcription-independent mechanisms relative to other p53 inhibitors, including pifithrin (PFT) alpha. We compared the effects of vanadate to PFTalpha and PFTmicro, an inhibitor of transcription-independent apoptosis by p53. Vanadate suppressed p53-associated apoptotic events at the mitochondria, including the loss of mitochondrial membrane potential, the conformational change of Bax and Bak, the mitochondrial translocation of p53, and the interaction of p53 with Bcl-2. Similarly, vanadate suppressed the apoptosis-inducing activity of a mitochondrially targeted temperature-sensitive p53 in stable transfectants of SaOS-2 cells. In radioprotection assays, which rely on p53, vanadate completely protected mice from a sublethal dose of 8 Gy and partially from a lethal dose of 12 Gy. Together, our findings indicated that vanadate effectively suppresses p53-mediated apoptosis by both transcription-dependent and transcription-independent pathways, and suggested that both pathways must be inhibited to completely block p53-mediated apoptosis. PMID:20048077

  3. Modulation of p53-mediated transcriptional repression and apoptosis by the adenovirus E1B 19K protein.

    PubMed Central

    Sabbatini, P; Chiou, S K; Rao, L; White, E

    1995-01-01

    BRK cell lines that stably express adenovirus E1A and a murine temperature-sensitive p53 undergo apoptosis when p53 assumes the wild-type conformation. Expression of the E1B 19,000-molecular-weight (19K) protein rescues cells from this p53-mediated apoptosis and diverts cells to a growth-arrested state. As p53 likely functions as a tumor suppressor by regulating transcription, the ability of the E1B 19K protein to regulate p53-mediated transactivation and transcriptional repression was investigated. In promoter-reporter assays the E1B 19K did not block p53-mediated transactivation but did alleviate p53-mediated transcriptional repression. E1B 19K expression permitted efficient transcriptional activation of the p21/WAF-1/cip-1 mRNA by p53, consistent with maintenance of the growth arrest function of p53. The E1B 19K protein is thereby unique among DNA virus-transforming proteins that target p53 for inactivation in that it selectively modulates the transcriptional properties of p53. The E1B 19K protein also rescued cells from apoptosis induced by inhibitors of transcription and protein synthesis. This suggests that cell death may result from the inhibition of expression of survival factors which function to maintain cell viability. p53 may induce apoptosis through generalized transcriptional repression. In turn, the E1B 19K protein may prevent p53-mediated apoptosis by alleviating p53-mediated transcriptional repression. PMID:7823921

  4. 4-Hydroxynonenal induces p53-mediated apoptosis in retinal pigment epithelial cells

    PubMed Central

    Sharma, Abha; Sharma, Rajendra; Chaudhary, Pankaj; Vatsyayan, Rit; Pearce, Virginia; Jeyabal, Prince V.S.; Zimniak, Piotr; Awasthi, Sanjay; Awasthi, Yogesh C.

    2009-01-01

    4-Hydroxynonenal (4-HNE) has been suggested to be involved in stress-induced signaling for apoptosis. In present studies, we have examined the effects of 4-HNE on the intrinsic apoptotic pathway associated with p53 in human retinal pigment epithelial (RPE and ARPE-19) cells. Our results show that 4-HNE causes induction, phosphorylation, and nuclear accumulation of p53 which is accompanied with down regulation of MDM2, activation of the pro-apoptotic p53 target genes viz. p21 and Bax, JNK, caspase3, and onset of apoptosis in treated RPE cells. Reduced expression of p53 by an efficient silencing of the p53 gene resulted in a significant resistance of these cells to 4-HNE-induced cell death. The effects of 4-HNE on the expression and functions of p53 are blocked in GSTA4-4 over expressing cells indicating that 4-HNE-induced, p53-mediated signaling for apoptosis is regulated by GSTs. Our results also show that the induction of p53 in tissues of mGsta4 (-/-) mice correlate with elevated levels of 4-HNE due to its impaired metabolism. Together, these studies suggest that 4-HNE is involved in p53-mediated signaling in in vitro cell cultures as well as in vivo that can be regulated by GSTs. PMID:18930016

  5. Chk2 regulates transcription-independent p53-mediated apoptosis in response to DNA damage

    SciTech Connect

    Chen Chen; Shimizu, Shigeomi; Tsujimoto, Yoshihide; Motoyama, Noboru . E-mail: motoyama@nils.go.jp

    2005-07-29

    The tumor suppressor protein p53 plays a central role in the induction of apoptosis in response to genotoxic stress. The protein kinase Chk2 is an important regulator of p53 function in mammalian cells exposed to ionizing radiation (IR). Cells derived from Chk2-deficient mice are resistant to the induction of apoptosis by IR, and this resistance has been thought to be a result of the defective transcriptional activation of p53 target genes. It was recently shown, however, that p53 itself and histone H1.2 translocate to mitochondria and thereby induces apoptosis in a transcription-independent manner in response to IR. We have now examined whether Chk2 also regulates the transcription-independent induction of apoptosis by p53 and histone H1.2. The reduced ability of IR to induce p53 stabilization in Chk2-deficient thymocytes was associated with a marked impairment of p53 and histone H1 translocation to mitochondria. These results suggest that Chk2 regulates the transcription-independent mechanism of p53-mediated apoptosis by inducing stabilization of p53 in response to IR.

  6. Marijuana smoke condensate induces p53-mediated apoptosis in human lung epithelial cells.

    PubMed

    Kim, Ha Ryong; Jung, Mi Hyun; Lee, Soo Yeun; Oh, Seung Min; Chung, Kyu Hyuck

    2013-01-01

    Since the largely abused worldwide used of marijuana, there have been many ongoing debates regarding the adverse health effects of marijuana smoking. Marijuana smoking was recently proved to cause pulmonary toxicity by inducing genotoxic effects or generating reactive oxygen species. Because p53, a tumor suppressor gene, has an important pathophysiologic role in the regulation of lung epithelial cell DNA damage responses, we hypothesized that p53 may be involved in the oxidative stress-mediated apoptosis induced by marijuana smoking. First, we confirmed that marijuana smoke condensate (MSC) induces oxidative stress in BEAS-2B cells. We observed that reactive oxygen species (ROS) generation was increased by MSC in the DCFH-DA assay. Also, antioxidant enzyme (superoxide dismutase, catalase) activity and their mRNA expressions were up-regulated by MSC. Second, we investigated p53 involvement in the MSC-induced apoptotic pathway in BEAS-2B cells. The results showed that MSC increased caspase-3 activation and DNA fragmentation as markers of apoptosis. In addition, the mRNA levels of apoptosis-related genes (p53 and Bax) were increased by MSC and phospho-p53, along with the increase of Bax protein expression by MSC. Apoptosis and apoptosis-related gene expression were partially blocked by an inhibitor of p53-dependent transcriptional activation (pifithrin-α). The results indicate that p53 plays a role in MSC-induced apoptosis. Taken together, the findings of the present study suggest that MSC partially induces p53-mediated apoptosis through ROS generation in human lung epithelial cells and this may have broader implications for our understanding of pulmonary diseases. PMID:23665932

  7. HIF-1α antagonizes p53-mediated apoptosis by triggering HIPK2 degradation

    PubMed Central

    Nardinocchi, Lavinia; Puca, Rosa; D'Orazi, Gabriella

    2011-01-01

    Many human diseases are characterized by the development of tissue hypoxia. Hypoxia-inducible factor (HIF) is a transcription factor that regulates fundamental cellular processes in response to changes in oxygen concentration, such as angiogenesis, survival, and alterations in metabolism. The levels of HIF-1α subunit are increased in most solid tumors not only by low oxygen but also by growth factors and oncogenes and correlate with patient prognosis and treatment failure. The link between HIF-1α and apoptosis, a major determinant of cancer progression and treatment outcome, is poorly understood. Here we show that HIF-1α protects against drug-induced apoptosis by antagonizing the function of the tumor suppressor p53. HIF-1α upregulation induced proteasomal degradation of homeodomain-interacting protein kinase-2 (HIPK2), the p53 apoptotic activator. Inhibition of HIF-1α by siRNA, HIF-1α-dominant negative or by zinc re-established the HIPK2 levels and the p53-mediated chemosensitivity in tumor cells. Our findings identify a novel circuitry between HIF-1α and p53, and provide a paradigm for HIPK2 dictating cell response to antitumor therapies. PMID:21248371

  8. Structural proteins of Kaposi's sarcoma-associated herpesvirus antagonize p53-mediated apoptosis.

    PubMed

    Chudasama, P; Konrad, A; Jochmann, R; Lausen, B; Holz, P; Naschberger, E; Neipel, F; Britzen-Laurent, N; Stürzl, M

    2015-01-29

    The tumor suppressor p53 is a central regulatory molecule of apoptosis and is commonly mutated in tumors. Kaposi's sarcoma-associated herpesvirus (KSHV)-related malignancies express wild-type p53. Accordingly, KSHV encodes proteins that counteract the cell death-inducing effects of p53. Here, the effects of all KSHV genes on the p53 signaling pathway were systematically analyzed using the reversely transfected cell microarray technology. With this approach we detected eight KSHV-encoded genes with potent p53 inhibiting activity in addition to the previously described inhibitory effects of KSHV genes ORF50, K10 and K10.5. Interestingly, the three most potent newly identified inhibitors were KSHV structural proteins, namely ORF22 (glycoprotein H), ORF25 (major capsid protein) and ORF64 (tegument protein). Validation of these results with a classical transfection approach showed that these proteins inhibited p53 signaling in a dose-dependent manner and that this effect could be reversed by small interfering RNA-mediated knockdown of the respective viral gene. All three genes inhibited p53-mediated apoptosis in response to Nutlin-3 treatment in non-infected and KSHV-infected cells. Addressing putative mechanisms, we could show that these proteins could also inhibit the transactivation of the promoters of apoptotic mediators of p53 such as BAX and PIG3. Altogether, we demonstrate for the first time that structural proteins of KSHV can counteract p53-induced apoptosis. These proteins are expressed in the late lytic phase of the viral life cycle and are incorporated into the KSHV virion. Accordingly, these genes may inhibit cell death in the productive and in the early entrance phase of KSHV infection. PMID:24469037

  9. p53 Mediates Colistin-Induced Autophagy and Apoptosis in PC-12 Cells.

    PubMed

    Zhang, Ling; Xie, Daoyuan; Chen, Xueping; Hughes, Maria L R; Jiang, Guozheng; Lu, Ziyin; Xia, Chunli; Li, Li; Wang, Jinli; Xu, Wei; Sun, Yuan; Li, Rui; Wang, Rui; Qian, Feng; Li, Jian; Li, Jichang

    2016-09-01

    -induced autophagy and apoptosis are associated with the p53-mediated pathway. PMID:27324771

  10. Induction of p53-mediated transcription and apoptosis by exportin-1 (XPO1) inhibition in mantle cell lymphoma.

    PubMed

    Yoshimura, Mariko; Ishizawa, Jo; Ruvolo, Vivian; Dilip, Archana; Quintás-Cardama, Alfonso; McDonnell, Timothy J; Neelapu, Sattva S; Kwak, Larry W; Shacham, Sharon; Kauffman, Michael; Tabe, Yoko; Yokoo, Masako; Kimura, Shinya; Andreeff, Michael; Kojima, Kensuke

    2014-07-01

    The nuclear transporter exportin-1 (XPO1) is highly expressed in mantle cell lymphoma (MCL) cells, and is believed to be associated with the pathogenesis of this disease. XPO1-selective inhibitors of nuclear export (SINE) compounds have been shown to induce apoptosis in MCL cells. Given that p53 is a cargo protein of XPO1, we sought to determine the significance of p53 activation through XPO1 inhibition in SINE-induced apoptosis of MCL cells. We investigated the prognostic impact of XPO1 expression in MCL cells using Oncomine analysis. The significance of p53 mutational/functional status on sensitivity to XPO1 inhibition in cell models and primary MCL samples, and the functional role of p53-mediated apoptosis signaling, were also examined. Increased XPO1 expression was associated with poor prognosis in MCL patients. The XPO1 inhibitor KPT-185 induced apoptosis in MCL cells through p53-dependent and -independent mechanisms, and p53 status was a critical determinant of its apoptosis induction. The KPT-185-induced, p53-mediated apoptosis in the MCL cells occurred in a transcription-dependent manner. Exportin-1 appears to influence patient survival in MCL, and the SINE XPO1 antagonist KPT-185 effectively activates p53-mediated transcription and apoptosis, which would provide a novel strategy for the therapy of MCL. PMID:24766216

  11. Loss of p53-mediated cell-cycle arrest, senescence and apoptosis promotes genomic instability and premature aging

    PubMed Central

    Li, Tongyuan; Liu, Xiangyu; Jiang, Le; Manfredi, James; Zha, Shan; Gu, Wei

    2016-01-01

    Although p53-mediated cell cycle arrest, senescence and apoptosis are well accepted as major tumor suppression mechanisms, the loss of these functions does not directly lead to tumorigenesis, suggesting that the precise roles of these canonical activities of p53 need to be redefined. Here, we report that the cells derived from the mutant mice expressing p533KR, an acetylation-defective mutant that fails to induce cell-cycle arrest, senescence and apoptosis, exhibit high levels of aneuploidy upon DNA damage. Moreover, the embryonic lethality caused by the deficiency of XRCC4, a key DNA double strand break repair factor, can be fully rescued in the p533KR/3KR background. Notably, despite high levels of genomic instability, p533KR/3KRXRCC4−/− mice, unlike p53−/− XRCC4−/− mice, are not succumbed to pro-B-cell lymphomas. Nevertheless, p533KR/3KR XRCC4−/− mice display aging-like phenotypes including testicular atrophy, kyphosis, and premature death. Further analyses demonstrate that SLC7A11 is downregulated and that p53-mediated ferroptosis is significantly induced in spleens and testis of p533KR/3KRXRCC4−/− mice. These results demonstrate that the direct role of p53-mediated cell cycle arrest, senescence and apoptosis is to control genomic stability in vivo. Our study not only validates the importance of ferroptosis in p53-mediated tumor suppression in vivo but also reveals that the combination of genomic instability and activation of ferroptosis may promote aging-associated phenotypes. PMID:26943586

  12. p53 mediates cigarette smoke-induced apoptosis of pulmonary endothelial cells: inhibitory effects of macrophage migration inhibitor factor.

    PubMed

    Damico, Rachel; Simms, Tiffany; Kim, Bo S; Tekeste, Zenar; Amankwan, Henry; Damarla, Mahendra; Hassoun, Paul M

    2011-03-01

    Exposure to cigarette smoke (CS) is the most common cause of emphysema, a debilitating pulmonary disease histopathologically characterized by the irreversible destruction of lung architecture. Mounting evidence links enhanced endothelial apoptosis causally to the development of emphysema. However, the molecular determinants of human endothelial cell apoptosis and survival in response to CS are not fully defined. Such determinants could represent clinically relevant targets for intervention. We show here that CS extract (CSE) triggers the death of human pulmonary macrovascular endothelial cells (HPAECs) through a caspase 9-dependent apoptotic pathway. Exposure to CSE results in the increased expression of p53 in HPAECs. Using the p53 inhibitor, pifithrin-α (PFT-α), and RNA interference (RNAi) directed at p53, we demonstrate that p53 function and expression are required for CSE-mediated apoptosis. The expression of macrophage migration inhibitory factor (MIF), an antiapoptotic cytokine produced by HPAECs, also increases in response to CSE exposure. The addition of recombinant human MIF prevents cell death from exposure to CSE. Further, the suppression of MIF or its receptor/binding partner, Jun activation domain-binding protein 1 (Jab-1), with RNAi enhances the sensitivity of human pulmonary endothelial cells to CSE via a p53-dependent (PFT-α-inhibitable) pathway. Finally, we demonstrate that MIF is a negative regulator of p53 expression in response to CSE, placing MIF upstream of p53 as an antagonist of CSE-induced apoptosis. We conclude that MIF can protect human vascular endothelium from the toxic effects of CSE via the antagonism of p53-mediated apoptosis. PMID:20448056

  13. Benzo(a)pyrene Induced p53 Mediated Male Germ Cell Apoptosis: Synergistic Protective Effects of Curcumin and Resveratrol

    PubMed Central

    Banerjee, Bhaswati; Chakraborty, Supriya; Ghosh, Debidas; Raha, Sanghamitra; Sen, Parimal C.; Jana, Kuladip

    2016-01-01

    Benzo(a)pyrene (B(a)P) is an environmental toxicant that induces male germ cell apoptosis. Curcumin and resveratrol are phytochemicals with cytoprotective and anti-oxidative properties. At the same time resveratrol is also a natural Aryl hydrocarbon Receptor (AhR) antagonist. Our present study in isolated testicular germ cell population from adult male Wistar rats, highlighted the synergistic protective effect of curcumin and resveratrol against B(a)P induced p53 mediated germ cell apoptosis. Curcumin-resveratrol significantly prevented B(a)P induced decrease in sperm cell count and motility, as well as increased serum testosterone level. Curcumin-resveratrol co-treatment actively protected B(a)P induced testicular germ cell apoptosis. Curcumin-resveratrol co-treatment decreased the expression of pro-apoptotic proteins like cleaved caspase 3, 8 and 9, cleaved PARP, Apaf1, FasL, tBid. Curcumin-resveratrol co-treatment decreased Bax/Bcl2 ratio, mitochondria to cytosolic translocation of cytochrome c and activated the survival protein Akt. Curcumin-resveratrol decreased the expression of p53 dependent apoptotic genes like Fas, FasL, Bax, Bcl2, and Apaf1. B(a)P induced testicular reactive oxygen species (ROS) generation and oxidative stress were significantly ameliorated with curcumin and resveratrol. Curcumin-resveratrol co-treatment prevented B(a)P induced nuclear translocation of AhR and CYP1A1 (Cytochrome P4501A1) expression. The combinatorial treatment significantly inhibited B(a)P induced ERK 1/2, p38 MAPK and JNK 1/2 activation. B(a)P treatment increased the expression of p53 and its phosphorylation (p53 ser 15). Curcumin-resveratrol co-treatment significantly decreased p53 level and its phosphorylation (p53 ser 15). The study concludes that curcumin-resveratrol synergistically modulated MAPKs and p53, prevented oxidative stress, regulated the expression of pro and anti-apoptotic proteins as well as the proteins involved in B(a)P metabolism thus protected germ

  14. Benzo(a)pyrene Induced p53 Mediated Male Germ Cell Apoptosis: Synergistic Protective Effects of Curcumin and Resveratrol.

    PubMed

    Banerjee, Bhaswati; Chakraborty, Supriya; Ghosh, Debidas; Raha, Sanghamitra; Sen, Parimal C; Jana, Kuladip

    2016-01-01

    Benzo(a)pyrene (B(a)P) is an environmental toxicant that induces male germ cell apoptosis. Curcumin and resveratrol are phytochemicals with cytoprotective and anti-oxidative properties. At the same time resveratrol is also a natural Aryl hydrocarbon Receptor (AhR) antagonist. Our present study in isolated testicular germ cell population from adult male Wistar rats, highlighted the synergistic protective effect of curcumin and resveratrol against B(a)P induced p53 mediated germ cell apoptosis. Curcumin-resveratrol significantly prevented B(a)P induced decrease in sperm cell count and motility, as well as increased serum testosterone level. Curcumin-resveratrol co-treatment actively protected B(a)P induced testicular germ cell apoptosis. Curcumin-resveratrol co-treatment decreased the expression of pro-apoptotic proteins like cleaved caspase 3, 8 and 9, cleaved PARP, Apaf1, FasL, tBid. Curcumin-resveratrol co-treatment decreased Bax/Bcl2 ratio, mitochondria to cytosolic translocation of cytochrome c and activated the survival protein Akt. Curcumin-resveratrol decreased the expression of p53 dependent apoptotic genes like Fas, FasL, Bax, Bcl2, and Apaf1. B(a)P induced testicular reactive oxygen species (ROS) generation and oxidative stress were significantly ameliorated with curcumin and resveratrol. Curcumin-resveratrol co-treatment prevented B(a)P induced nuclear translocation of AhR and CYP1A1 (Cytochrome P4501A1) expression. The combinatorial treatment significantly inhibited B(a)P induced ERK 1/2, p38 MAPK and JNK 1/2 activation. B(a)P treatment increased the expression of p53 and its phosphorylation (p53 ser 15). Curcumin-resveratrol co-treatment significantly decreased p53 level and its phosphorylation (p53 ser 15). The study concludes that curcumin-resveratrol synergistically modulated MAPKs and p53, prevented oxidative stress, regulated the expression of pro and anti-apoptotic proteins as well as the proteins involved in B(a)P metabolism thus protected germ

  15. ARF and ATM/ATR cooperate in p53-mediated apoptosis upon oncogenic stress

    SciTech Connect

    Pauklin, Siim . E-mail: spauklin@ut.ee; Kristjuhan, Arnold; Maimets, Toivo; Jaks, Viljar

    2005-08-26

    Induction of apoptosis is pivotal for eliminating cells with damaged DNA or deregulated proliferation. We show that tumor suppressor ARF and ATM/ATR kinase pathways cooperate in the induction of apoptosis in response to elevated expression of c-myc, {beta}-catenin or human papilloma virus E7 oncogenes. Overexpression of oncogenes leads to the formation of phosphorylated H2AX foci, induction of Rad51 protein levels and ATM/ATR-dependent phosphorylation of p53. Inhibition of ATM/ATR kinases abolishes both induction of Rad51 and phosphorylation of p53, and remarkably reduces the level of apoptosis induced by co-expression of oncogenes and ARF. However, the induction of apoptosis is downregulated in p53-/- cells and does not depend on activities of ATM/ATR kinases, indicating that efficient induction of apoptosis by oncogene activation depends on coordinated action of ARF and ATM/ATR pathways in the regulation of p53.

  16. Anticancer Activity of γ-Bisabolene in Human Neuroblastoma Cells via Induction of p53-Mediated Mitochondrial Apoptosis.

    PubMed

    Jou, Yu-Jen; Hua, Chun-Hung; Lin, Chen-Sheng; Wang, Ching-Ying; Wan, Lei; Lin, Ying-Ju; Huang, Su-Hua; Lin, Cheng-Wen

    2016-01-01

    γ-Bisabolene has demonstrated antiproliferative activities against several human cancer cell lines. This study first discloses the antiproliferative and apoptosis induction activities of γ-bisabolene to human neuroblastoma TE671 cells. A CC50 value of γ-bisabolene was 8.2 μM to TE671 cells. Cell cycle analysis with PI staining showed γ-bisabolene elevating the sub-G1 fractions in a time-dependent manner. In addition, annexin V-FITC/PI staining showed γ-bisabolene significantly triggering early (annexin-V positive/PI negative) and late (annexin-V positive/PI positive) apoptosis in dose-dependent manners. γ-Bisabolene induced caspase 3/8/9 activation, intracellular ROS increase, and mitochondrial membrane potential decrease in apoptosis of human neuro-blastoma cells. Moreover, γ-bisabolene increased p53 phosphorylation and up-regulated p53-mediated apoptotic genes Bim and PUMA, as well as decreased the mRNA and protein levels of CK2α. Notably, the results indicated the involvement of CK2α-p53 pathways in mitochondria-mediated apoptosis of human neuroblastoma cells treated with γ-bisabolene. This study elucidated the apoptosis induction pathways of γ-bisabolene-treated neuroblastoma cells, in which could be useful for developing anti-neuroblastoma drugs. PMID:27164076

  17. p53 mediated apoptosis in osteosarcoma MG-63 cells by inhibition of FANCD2 gene expression

    PubMed Central

    Xia, Peng; Sun, Yifu; Zheng, Changjun; Hou, Tingting; Kang, Mingyang; Yang, Xiaoyu

    2015-01-01

    Purpose: The aim of this study was to investigate the association between osteosarcoma (OS) and Fanconi anemia (FA) related pathways and the molecular mechanisms. Methods: siRNA for Fanconi anemia complementation group D2 (FANCD2) was constructed and transfected into the osteosarcoma cell line MG-63 cells. Expression of TP53INP1, p53, p21, caspase-9, and caspase-3 mRNA in MG-63 cells were examined by real-time fluorescence quantitative PCR, and the protein levels were also determined by western blot. Results: After silence of the FANCD2 gene in MG-63 cells, cell proliferation was inhibited, cell cycle was arrested and cell apoptosis was induced. The apoptosis was mediated by the p53 signaling pathway. After FANCD2 expression was inhibited, TP53INP1 gene expression was up-regulated, phosphorylation of p53 was promoted and the p21 protein was activated, leading to cell cycle arrested in G1, finally resulted in caspase-dependent cell apoptosis. Conclusions: Inhibition of FANCD2 gene expression can induce apoptosis of osteosarcoma cells, which indicated that FANCD2 played an important role in the development of osteosarcoma and it might be a potential target for treatment of osteosarcoma. PMID:26379910

  18. A novel TRIM family member, Trim69, regulates zebrafish development through p53-mediated apoptosis.

    PubMed

    Han, Ruiqin; Zhao, Qing; Zong, Shudong; Miao, Shiying; Song, Wei; Wang, Linfang

    2016-05-01

    Trim69 contains the hallmark domains of a tripartite motif (TRIM) protein, including a Ring-finger domain, B-box domain, and coiled-coil domain. Trim69 is structurally and evolutionarily conserved in zebrafish, mouse, rat, human, and chimpanzee. The role of this protein is unclear, however, so we investigated its function in zebrafish development. Trim69 is extensively expressed in zebrafish adults and developing embryos-particularly in the testis, brain, ovary, and heart-and its expression decreases in a time- and stage-dependent manner. Loss of trim69 in zebrafish induces apoptosis and activates apoptosis-related processes; indeed, the tp53 pathway was up-regulated in response to the knockdown. Expression of human trim69 rescued the apoptotic phenotype, while overexpression of trim69 does not increase cellular apoptosis. Taken together, our results suggest that trim69 participates in tp53-mediated apoptosis during zebrafish development. Mol. Reprod. Dev. 83: 442-454, 2016. © 2016 Wiley Periodicals, Inc. PMID:27031046

  19. Integrated high-throughput analysis identifies Sp1 as a crucial determinant of p53-mediated apoptosis

    PubMed Central

    Li, H; Zhang, Y; Ströse, A; Tedesco, D; Gurova, K; Selivanova, G

    2014-01-01

    The restoration of p53 tumor suppressor function is a promising therapeutic strategy to combat cancer. However, the biological outcomes of p53 activation, ranging from the promotion of growth arrest to the induction of cell death, are hard to predict, which limits the clinical application of p53-based therapies. In the present study, we performed an integrated analysis of genome-wide short hairpin RNA screen and gene expression data and uncovered a previously unrecognized role of Sp1 as a central modulator of the transcriptional response induced by p53 that leads to robust induction of apoptosis. Sp1 is indispensable for the pro-apoptotic transcriptional repression by p53, but not for the induction of pro-apoptotic genes. Furthermore, the p53-dependent pro-apoptotic transcriptional repression required the co-binding of Sp1 to p53 target genes. Our results also highlight that Sp1 shares with p53 a common regulator, MDM2, which targets Sp1 for proteasomal degradation. This uncovers a new mechanism of the tight control of apoptosis in cells. Our study advances the understanding of the molecular basis of p53-mediated apoptosis and implicates Sp1 as one of its key modulators. We found that small molecules reactivating p53 can differentially modulate Sp1, thus providing insights into how to manipulate p53 response in a controlled way. PMID:24971482

  20. Targeting GRP75 Improves HSP90 Inhibitor Efficacy by Enhancing p53-Mediated Apoptosis in Hepatocellular Carcinoma

    PubMed Central

    Yang, Ling; Liu, Xiaoyu; E, Qiukai; Gao, Peiye; Ye, Xiaofei; Liu, Wen; Zuo, Ji

    2014-01-01

    Heat shock protein 90 (HSP90) inhibitors are potential drugs for cancer therapy. The inhibition of HSP90 on cancer cell growth largely through degrading client proteins, like Akt and p53, therefore, triggering cancer cell apoptosis. Here, we show that the HSP90 inhibitor 17-AAG can induce the expression of GRP75, a member of heat shock protein 70 (HSP70) family, which, in turn, attenuates the anti-growth effect of HSP90 inhibition on cancer cells. Additionally, 17-AAG enhanced binding of GRP75 and p53, resulting in the retention of p53 in the cytoplasm. Blocking GRP75 with its inhibitor MKT-077 potentiated the anti-tumor effects of 17-AAG by disrupting the formation of GRP75-p53 complexes, thereby facilitating translocation of p53 into the nuclei and leading to the induction of apoptosis-related genes. Finally, dual inhibition of HSP90 and GRP75 was found to significantly inhibit tumor growth in a liver cancer xenograft model. In conclusion, the GRP75 inhibitor MKT-077 enhances 17-AAG-induced apoptosis in HCCs and increases p53-mediated inhibition of tumor growth in vivo. Dual targeting of GRP75 and HSP90 may be a useful strategy for the treatment of HCCs. PMID:24465691

  1. Revisiting DNA damage repair, p53-mediated apoptosis and cisplatin sensitivity in germ cell tumors.

    PubMed

    Cavallo, Francesca; Feldman, Darren R; Barchi, Marco

    2013-01-01

    Testicular germ cell tumors (TGCTs), ie, seminomas and nonseminomas, account for 1% to 3% of all neoplasms in men. They are the most common cancer in young white males and are unique in their responsiveness to cisplatin-based chemotherapy. For this reason, TGCTs are considered a model for curative disease. However, up to now, the molecular mechanisms behind this exceptional responsiveness to DNA-damaging agents have remained unclear. A hypersensitive apoptotic response, as well as a reduction in the proficiency to repair cisplatin-induced DNA damage might account for this behavior. In this review, building on recent findings of p53-induced apoptosis and DNA-repair mechanisms in TGCTs, we will discuss the molecular bases that drive tumor sensitivity to cisplatin, emphasizing the new therapeutic approaches proposed to eventually constrain tumor recurrence, and target TGCTs which are unresponsive to standard therapies. PMID:23784838

  2. NSC-87877 inhibits DUSP26 function in neuroblastoma resulting in p53-mediated apoptosis

    PubMed Central

    Shi, Y; Ma, I T; Patel, R H; Shang, X; Chen, Z; Zhao, Y; Cheng, J; Fan, Y; Rojas, Y; Barbieri, E; Chen, Z; Yu, Y; Jin, J; Kim, E S; Shohet, J M; Vasudevan, S A; Yang, J

    2015-01-01

    Dual specificity protein phosphatase 26 (DUSP26) is overexpressed in high-risk neuroblastoma (NB) and contributes to chemoresistance by inhibiting p53 function. In vitro, DUSP26 has also been shown to effectively inhibit p38 MAP kinase. We hypothesize that inhibiting DUSP26 will result in decreased NB cell growth in a p53 and/or p38-mediated manner. NSC-87877 (8-hydroxy-7-[(6-sulfo-2-naphthyl)azo]-5-quinolinesulfonic acid), a novel DUSP26 small molecule inhibitor, shows effective growth inhibition and induction of apoptosis in NB cell lines. NB cell lines treated with small hairpin RNA (shRNA) targeting DUSP26 also exhibit a proliferation defect both in vitro and in vivo. Treatment of NB cell lines with NSC-87877 results in increased p53 phosphorylation (Ser37 and Ser46) and activation, increased activation of downstream p38 effector proteins (heat shock protein 27 (HSP27) and MAP kinase-activated protein kinase 2 (MAPKAPK2)) and poly ADP ribose polymerase/caspase-3 cleavage. The cytotoxicity resulting from DUSP26 inhibition is partially reversed by knocking down p53 expression with shRNA and also by inhibiting p38 activity with SB203580 (4-[4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-1H-imidazol-5-yl]pyridine). In an intrarenal mouse model of NB, NSC-87877 treatment results in decreased tumor growth and increased p53 and p38 activity. Together, these results suggest that DUSP26 inhibition with NSC-87877 is an effective strategy to induce NB cell cytotoxicity in vitro and in vivo through activation of the p53 and p38 mitogen-activated protein kinase (MAPK) tumor-suppressor pathways. PMID:26247726

  3. Aneuploidy generates proteotoxic stress and DNA damage concurrently with p53-mediated post-mitotic apoptosis in SAC-impaired cells.

    PubMed

    Ohashi, Akihiro; Ohori, Momoko; Iwai, Kenichi; Nakayama, Yusuke; Nambu, Tadahiro; Morishita, Daisuke; Kawamoto, Tomohiro; Miyamoto, Maki; Hirayama, Takaharu; Okaniwa, Masanori; Banno, Hiroshi; Ishikawa, Tomoyasu; Kandori, Hitoshi; Iwata, Kentaro

    2015-01-01

    The molecular mechanism responsible that determines cell fate after mitotic slippage is unclear. Here we investigate the post-mitotic effects of different mitotic aberrations--misaligned chromosomes produced by CENP-E inhibition and monopolar spindles resulting from Eg5 inhibition. Eg5 inhibition in cells with an impaired spindle assembly checkpoint (SAC) induces polyploidy through cytokinesis failure without a strong anti-proliferative effect. In contrast, CENP-E inhibition causes p53-mediated post-mitotic apoptosis triggered by chromosome missegregation. Pharmacological studies reveal that aneuploidy caused by the CENP-E inhibitor, Compound-A, in SAC-attenuated cells causes substantial proteotoxic stress and DNA damage. Polyploidy caused by the Eg5 inhibitor does not produce this effect. Furthermore, p53-mediated post-mitotic apoptosis is accompanied by aneuploidy-associated DNA damage response and unfolded protein response activation. Because Compound-A causes p53 accumulation and antitumour activity in an SAC-impaired xenograft model, CENP-E inhibitors could be potential anticancer drugs effective against SAC-impaired tumours. PMID:26144554

  4. Kaempferol induces ATM/p53-mediated death receptor and mitochondrial apoptosis in human umbilical vein endothelial cells.

    PubMed

    Lee, Chiu-Fang; Yang, Jai-Sing; Tsai, Fuu-Jen; Chiang, Ni-Na; Lu, Chi-Cheng; Huang, Yu-Syuan; Chen, Chun; Chen, Fu-An

    2016-05-01

    Kaempferol is a member of the flavonoid compounds found in vegetables and fruits. It is shown to exhibit biological impact and anticancer activity, but no report exists on the angiogenic effect of kaempferol and induction of cell apoptosis in vitro. In this study, we investigated the role of kaempferol on anti-angiogenic property and the apoptotic mechanism of human umbilical vein endothelial cells (HUVECs). Our results demonstrated that kaempferol decreased HUVEC viability in a time- and concentration-dependent manner. Kaempferol also induced morphological changes and sub-G1 phase cell population (apoptotic cells). Kaempferol triggered apoptosis of HUVECs as detecting by DNA fragmentation, comet assay and immunofluorescent staining for activated caspase-3. The caspase signals, including caspase-8, -9 and -3, were time-dependently activated in HUVECs after kaempferol exposure. Furthermore, pre-treatment with a specific inhibitor of caspase-8 (Z-IETD-FMK) significantly reduced the activity of caspase-8, -9 and -3, indicating that extrinsic pathway is a major signaling pathway in kaempferol-treated HUVECs. Importantly, kaempferol promoted reactive oxygen species (ROS) evaluated using flow cytometric assay in HUVECs. We further investigated the upstream extrinsic pathway and showed that kaempferol stimulated death receptor signals [Fas/CD95, death receptor 4 (DR4) and DR5] through increasing the levels of phosphorylated p53 and phosphorylated ATM pathways in HUVECs, which can be individually confirmed by N-acetylcysteine (NAC), ATM specific inhibitor (caffeine) and p53 siRNA. Based on these results, kaempferol-induced HUVEC apoptosis was involved in an ROS-mediated p53/ATM/death receptor signaling. Kaempferol might possess therapeutic effects on cancer treatment in anti-vascular targeting. PMID:26984266

  5. Berberine induces mitochondrial apoptosis of EBV-transformed B cells through p53-mediated regulation of XAF1 and GADD45α.

    PubMed

    Park, Ga Bin; Park, Sang Hyun; Kim, Daejin; Kim, Yeong Seok; Yoon, Sung Ho; Hur, Dae Young

    2016-07-01

    Berberine exhibits antiproliferative or cytotoxic effects against various cancers. ROS and wild-type p53 play a critical role in berberine-induced cytotoxic effects. In this study, we investigated the correlation between XAF1 and functional p53 in EBV-transformed B cells or cancerous B cells after treatment with berberine. Berberine decreased cell viability and induced apoptosis through a mitochondria-dependent pathway in EBV-transformed B cells and cancerous B cells, but not in normal peripheral blood mononuclear cells. Activated p53 and its downstream targets XAF1 and GADD45α interacted with PUMA, Bax, and Bim in mitochondria after treatment with berberine. Blocking phosphorylation of p38/JNK MAPK and treatment with PFT-α, a selective p53 inhibitor, effectively prevented apoptosis and the upregulation of phosphorylated p53, XAF1, and GADD45α. NAC, a ROS scavenger, also suppressed berberine-induced mitochondria disruption and the whole apoptotic process via restoration of p53-related proteins and proapoptotic Bcl-2 family proteins. Taken together, our results suggest that ROS generation might be a predisposing event in berberine-induced mitochondrial apoptosis in EBV-transformed B cells through the upregulation of XAF1 and GADD45α expression by MAPK and functional p53. PMID:27121748

  6. Activation of p53 mediated glycolytic inhibition-oxidative stress-apoptosis pathway in Dalton's lymphoma by a ruthenium (II)-complex containing 4-carboxy N-ethylbenzamide.

    PubMed

    Koiri, Raj Kumar; Trigun, Surendra Kumar; Mishra, Lallan

    2015-03-01

    There is a general agreement that most of the cancer cells switch over to aerobic glycolysis (Warburg effect) and upregulate antioxidant enzymes to prevent oxidative stress induced apoptosis. Thus, there is an evolving view to target these metabolic alterations by novel anticancer agents to restrict tumor progression in vivo. Previously we have reported that when a non toxic dose (10 mg/kg bw i.p.) of a novel anticancer ruthenium(II)-complex containing 4-carboxy N-ethylbenzamide; Ru(II)-CNEB, was administered to the Dalton's lymphoma (DL) bearing mice, it regressed DL growth by inducing apoptosis in the DL cells. It also inactivated M4-LDH (M4-lactate dehydrogenase), an enzyme that drives anaerobic glycolysis in the tumor cells. In the present study we have investigated whether this compound is able to modulate regulation of glycolytic inhibition-apoptosis pathway in the DL cells in vivo. We observed that Ru(II)-CNEB could decline expression of the inducible form of 6-phosphofructo-2-kinase (iPFK2: PFKFB3), the master regulator of glycolysis in the DL cells. The complex also activated superoxide dismutase (the H2O2 producing enzyme) but declined the levels of catalase and glutathione peroxidase (the two H2O2 degrading enzymes) to impose oxidative stress in the DL cells. This was consistent with the enhanced p53 level, decline in Bcl2/Bax ratio and activation of caspase 9 in those DL cells. The findings suggest that Ru(II)-CNEB is able to activate oxidative stress-apoptosis pathway via p53 (a tumor supressor protein) mediated repression of iPFK2, a key glycolytic regulator, in the DL cells in vivo. PMID:25576833

  7. The MEK/ERK Pathway is the Primary Conduit for Borrelia burgdorferi-Induced Inflammation and P53-Mediated Apoptosis in Oligodendrocytes

    PubMed Central

    Parthasarathy, Geetha; Philipp, Mario T.

    2013-01-01

    Lyme neuroborreliosis (LNB) affects both the central and peripheral nervous systems. In a rhesus macaque model of LNB we had previously shown that brains of rhesus macaques inoculated with Borrelia burgdorferi release inflammatory mediators, and undergo oligodendrocyte and neuronal cell death. In vitro analysis of this phenomenon indicated that while B. burgdorferi can induce inflammation and apoptosis of oligodendrocytes per se, microglia are required for neuronal apoptosis. We hypothesized that the inflammatory milieu elicited by the bacterium in microglia or oligodendrocytes contributes to the apoptosis of neurons and glial cells, respectively, and that downstream signaling events in NFkB and/or MAPK pathways play a role in these phenotypes. To test these hypotheses in oligodendrocytes, several pathway inhibitors were used to determine their effect on inflammation and apoptosis, as induced by B. burgdorferi. In a human oligodendrocyte cell line (MO3.13), inhibition of the ERK pathway in the presence of B. burgdorferi markedly reduced inflammation, followed by the JNK, p38 and NFkB pathway inhibition. In addition to eliciting inflammation, B. burgdorferi also increased total p53 protein levels, and suppression of the ERK pathway mitigated this effect. While inhibition of p53 had a minimal effect in reducing inflammation, suppression of the ERK pathway or p53 reduced apoptosis as measured by active caspase-3 activity and the TUNEL assay. A similar result was seen in primary human oligodendrocytes wherein suppression of ERK or p53 reduced apoptosis. It is possible that inflammation and apoptosis in oligodendrocytes are divergent arms of MAPK pathways, particularly the MEK/ERK pathway. PMID:24114360

  8. Quantitative phosphoproteomic analysis reveals γ-bisabolene inducing p53-mediated apoptosis of human oral squamous cell carcinoma via HDAC2 inhibition and ERK1/2 activation.

    PubMed

    Jou, Yu-Jen; Chen, Chao-Jung; Liu, Yu-Ching; Way, Tzong-Der; Lai, Chih-Ho; Hua, Chun-Hung; Wang, Ching-Ying; Huang, Su-Hua; Kao, Jung-Yie; Lin, Cheng-Wen

    2015-10-01

    γ-Bisabolene, one of main components in cardamom, showed potent in vitro and in vivo anti-proliferative activities against human oral squamous cell carcinoma (OSCC). γ-Bisabolene activated caspases-3/9 and decreased mitochondrial memebrane potential, leading to apoptosis of OSCC cell lines (Ca9-22 and SAS), but not normal oral fibroblast cells. Phosphoproteome profiling of OSCC cells treated with γ-bisabolene was identified using TiO2-PDMS plate and LC-MS/MS, then confirmed using Western blotting and real-time RT-PCR assays. Phosphoproteome profiling revealed that γ-bisabolene increased the phosphorylation of ERK1/2, protein phosphatases 1 (PP1), and p53, as well as decreased the phosphorylation of histone deacetylase 2 (HDAC2) in the process of apoptosis induction. Protein-protein interaction network analysis proposed the involvement of PP1-HDAC2-p53 and ERK1/2-p53 pathways in γ-bisabolene-induced apoptosis. Subsequent assays indicated γ-bisabolene eliciting p53 acetylation that enhanced the expression of p53-regulated apoptotic genes. PP1 inhibitor-2 restored the status of HDAC2 phosphorylation, reducing p53 acetylation and PUMA mRNA expression in γ-bisabolene-treated Ca9-22 and SAS cells. Meanwhile, MEK and ERK inhibitors significantly decreased γ-bisabolene-induced PUMA expression in both cancer cell lines. Notably, the results ascertained the involvement of PP1-HDAC2-p53 and ERK1/2-p53 pathways in mitochondria-mediated apoptosis of γ-bisabolene-treated cells. This study demonstrated γ-bisabolene displaying potent anti-proliferative and apoptosis-inducing activities against OSCC in vitro and in vivo, elucidating molecular mechanisms of γ-bisabolene-induced apoptosis. The novel insight could be useful for developing anti-cancer drugs. PMID:26194454

  9. Induction of p53-mediated apoptosis in splenocytes and thymocytes of C57BL/6 mice exposed to perfluorooctane sulfonate (PFOS)

    SciTech Connect

    Dong, Guang-Hui; Wang, Jing; Zhang, Ying-Hua; Liu, Miao-Miao; Wang, Da; Zheng, Li; Jin, Yi-He

    2012-10-15

    Perfluorooctane sulfonate (PFOS) is a persistent environmental contaminant found in human and wildlife tissues. It has been reported that PFOS can cause atrophy of the immune organs and apoptosis of immunocytes in rodents. However, the mechanism behind such cause is still unclear. To understand the model of cell death and its mechanism on lymphoid cells in vivo, we conducted a dose/response experiment in which 4 groups of male adult C57BL/6 mice (12 mice per group) were dosed daily by oral gavage with PFOS at 0, 0.0167, 0.0833, or 0.8333 mg/kg/day, yielding targeted Total Administered Dose (TAD) of 0, 1, 5, or 50 mg PFOS/kg, respectively, over 60 days. The results showed that spleen and thymus weight were significantly reduced in the highest PFOS-dose-group (TAD 50 mg PFOS/kg) compared to the control group, whereas liver weight was significantly increased. We analyzed the cell death via apoptosis with an annexin-V/propidium iodide assay by flow cytometry, and observed that both the percentage of apoptosis and the expression of the pro-apoptotic proteins p53 in splenocytes and thymocytes increased in a dose-related manner after PFOS treatment. We also observed that PFOS induced p53-dependent apoptosis through the cooperation between the Bcl-xl down regulation without changing the Bcl-2 and Bax expression. The down regulation of Bcl-xl was strongly indicating mitochondrial involvement in apoptosis. It is confirmed by the release of cytochrome c and activation of caspase-3. All of these findings establish an important role of p53 and mitochondrial function in PFOS induced toxic environment in the host. -- Highlights: ► PFOS immunotoxicity is caused by induction of apoptosis via the p53 activation. ► PFOS exposure can induce down regulation of Bcl-xl. ► Mitochondria are involved in PFOS-induced apoptosis. ► PFOS exposure can cause the release of cytochrome c and activation of caspase-3.

  10. Triptolide sensitizes AML cells to TRAIL-induced apoptosis via decrease of XIAP and p53-mediated increase of DR5.

    PubMed

    Carter, Bing Z; Mak, Duncan H; Schober, Wendy D; Dietrich, Martin F; Pinilla, Clemencia; Vassilev, Lyubomir T; Reed, John C; Andreeff, Michael

    2008-04-01

    Acute myeloid leukemia (AML) cells are relatively resistant to tumor necrosis factor alpha-related apoptosis-inducing ligand (TRAIL). We previously reported that triptolide, a potent anticancer agent from a Chinese herb, decreases XIAP in leukemic cells. We evaluated the combination of triptolide and TRAIL and found synergistic promotion of apoptosis in AML cells. XIAP-overexpressing U937 cells (U937XIAP) were more resistant to TRAIL than U937neo cells, and inhibition of XIAP with the small-molecule inhibitor 1396-11 enhanced TRAIL-induced apoptosis, implying XIAP as a resistance factor in AML. Furthermore, triptolide increased DR5 levels in OCI-AML3, while the DR5 increase was blunted in p53-knockdown OCI-AML3 and p53-mutated U937 cells, confirming a role for p53 in the regulation of DR5. In support of this finding, disruption of MDM2-p53 binding with subsequent increase in p53 levels by nutlin3a increased DR5 levels and sensitized OCI-AML3 cells to TRAIL. The combination of 1396-11 plus nutlin3a plus TRAIL was more effective than either the 1396-11 and TRAIL or nutlin3a and TRAIL combinations in OCI-AML3 cells, further supporting the role of triptolide as a sensitizer to TRAIL-induced apoptosis in part by independent modulation of XIAP expression and p53 signaling. Thus, the combination of triptolide and TRAIL may provide a novel strategy for treating AML by overcoming critical mechanisms of apoptosis resistance. PMID:18187663

  11. A human papillomavirus type 18 E6/E7 transgene sensitizes mouse lens cells to human wild-type p53-mediated apoptosis.

    PubMed

    Nakamura, T; Williams-Simons, L; Westphal, H

    1997-06-26

    We have studied the concerted action of factors that influence the balance between cell proliferation and cell death in the developing lens of transgenic mice. We show that a human papillomavirus type 18 (HPV18) E6/E7 transgene that predominantly expresses the viral E7 gene product triggers apoptosis in a dose dependent manner, and causes retardation of lens growth or microphakia. E7 is known to inactivate pRB, the product of the retinoblastoma gene, and to enhance the action of p53. Our earlier work had demonstrated that over-expression of p53 itself can cause apoptosis of lens cells, and that a mutant p53 allele can interfere with this process. In the present study, we examined lenses that simultaneously express different constellations of the HPV18 E6/E7, wild-type and mutant human p53, and wild-type human pRB transgenes. We observed that lens cells expressing the HPV18 transgene are more sensitive to wild-type human p53 action than normal lens cells. As a result, there is severe microphakia in lenses that express both the HPV18 and the wild-type p53 transgenes. By contrast, apoptosis was reduced in lenses that co-expressed the HPV18 and either the pRB or the mutant p53 transgene. We conclude that levels of wild-type p53 are critical, and that any excess of p53 or suppression of pRB can cause cell death. Our results encourage attempts to counteract the deleterious action of human papillomaviruses in cervical cancer by a combination of measures that decrease cell proliferation and enhance apoptosis. PMID:9223662

  12. CPUY201112, a novel synthetic small-molecule compound and inhibitor of heat shock protein Hsp90, induces p53-mediated apoptosis in MCF-7 cells

    PubMed Central

    Xu, Xiao-Li; Bao, Qi-chao; Jia, Jian-Min; Liu, Fang; Guo, Xiao-Ke; Zhang, Ming-ye; Wei, Jin-lian; Lu, Meng-chen; Xu, Li-li; Zhang, Xiao-Jin; You, Qi-Dong; Sun, Hao-Peng

    2016-01-01

    Heat-shock protein 90 (Hsp90) is highly expressed in many tumor cells and is associated with the maintenance of malignant phenotypes. Targeting Hsp90 has had therapeutic success in both solid and hematological malignancies, which has inspired more studies to identify new Hsp90 inhibitors with improved clinical efficacy. Using a fragment-based approach and subsequent structural optimization guided by medicinal chemistry principles, we identified the novel compound CPUY201112 as a potent Hsp90 inhibitor. It binds to the ATP-binding pocket of Hsp90 with a kinetic dissociation (Kd) constant of 27 ± 2.3 nM. It also exhibits potent in vitro antiproliferative effects in a range of solid tumor cells. In MCF-7 cells with high Hsp90 expression, CPUY201112 induces the degradation of Hsp90 client proteins including HER-2, Akt, and c-RAF. We prove that treating MCF-7 cells with CPUY201112 results in cell cycle arrest and apoptosis through the wild-type (wt) p53 pathway. CPUY201112 also synergizes with Nutlin-3a to induce cancer cell apoptosis. CPUY201112 significantly inhibited the growth of MCF-7 xenografts in nude mice without apparent body weight loss. These results demonstrate that CPUY201112 is a novel Hsp90 inhibitor with potential use in treating wild-type p53 related cancers. PMID:26743233

  13. FFA-ROS-P53-mediated mitochondrial apoptosis contributes to reduction of osteoblastogenesis and bone mass in type 2 diabetes mellitus

    PubMed Central

    Li, Jun; He, Wang; Liao, Bo; Yang, Jingyue

    2015-01-01

    This study evaluated the association between free fatty acid (FFA), ROS generation, mitochondrial dysfunction and bone mineral density (BMD) in type 2 diabetic patients and investigated the molecular mechanism. db/db and high fat (HF)-fed mice were treated by Etomoxir, an inhibitor of CPT1, MitoQ, and PFT-α, an inhibitor of P53. Bone metabolic factors were assessed and BMSCs were isolated and induced to osteogenic differentiation. FFA, lipid peroxidation and mtDNA copy number were correlated with BMD in T2DM patients. Etomoxir, MitoQ and PFT-α significantly inhibited the decrease of BMD and bone breaking strength in db/db and HF-fed mice and suppressed the reduction of BMSCs-differentiated osteoblasts. Etomoxir and MitoQ, but not PFT-α, inhibited the increase of mitochondrial ROS generation in db/db and HF-fed mice and osteoblasts. In addition, Etomoxir, MitoQ and PFT-α significantly inhibited mitochondrial dysfunction in osteoblasts. Moreover, mitochondrial apoptosis was activated in osteoblasts derived from db/db and HF-fed mice, which was inhibited by Etomoxir, MitoQ and PFT-α. Furthermore, mitochondrial accumulation of P53 recruited Bax and initiated molecular events of apoptotic events. These results demonstrated that fatty acid oxidation resulted in ROS generation, activating P53/Bax-mediated mitochondrial apoptosis, leading to reduction of osteogenic differentiation and bone loss in T2DM. PMID:26226833

  14. Profiling Dose-Dependent Activation of p53-Mediated Signaling Pathways by Chemicals with Distinct Mechanisms of DNA Damage

    PubMed Central

    Clewell, Rebecca A.; Sun, Bin; Adeleye, Yeyejide; Carmichael, Paul; Efremenko, Alina; McMullen, Patrick D.; Pendse, Salil; Trask, O. J.; White, Andy; Andersen, Melvin E.

    2014-01-01

    As part of a larger effort to provide proof-of-concept in vitro-only risk assessments, we have developed a suite of high-throughput assays for key readouts in the p53 DNA damage response toxicity pathway: double-strand break DNA damage (p-H2AX), permanent chromosomal damage (micronuclei), p53 activation, p53 transcriptional activity, and cell fate (cell cycle arrest, apoptosis, micronuclei). Dose-response studies were performed with these protein and cell fate assays, together with whole genome transcriptomics, for three prototype chemicals: etoposide, quercetin, and methyl methanesulfonate. Data were collected in a human cell line expressing wild-type p53 (HT1080) and results were confirmed in a second p53 competent cell line (HCT 116). At chemical concentrations causing similar increases in p53 protein expression, p53-mediated protein expression and cellular processes showed substantial chemical-specific differences. These chemical-specific differences in the p53 transcriptional response appear to be determined by augmentation of the p53 response by co-regulators. More importantly, dose-response data for each of the chemicals indicate that the p53 transcriptional response does not prevent micronuclei induction at low concentrations. In fact, the no observed effect levels and benchmark doses for micronuclei induction were less than or equal to those for p53-mediated gene transcription regardless of the test chemical, indicating that p53's post-translational responses may be more important than transcriptional activation in the response to low dose DNA damage. This effort demonstrates the process of defining key assays required for a pathway-based, in vitro-only risk assessment, using the p53-mediated DNA damage response pathway as a prototype. PMID:25078064

  15. Anti-lung cancer potential of pure esteric-glycoside condurangogenin A against nonsmall-cell lung cancer cells in vitro via p21/p53 mediated cell cycle modulation and DNA damage-induced apoptosis

    PubMed Central

    Sikdar, Sourav; Mukherjee, Avinaba; Khuda-Bukhsh, Anisur Rahman

    2015-01-01

    Background: Marsdenia condurango (condurango) is a tropical woody vine native to South America. Our earlier study was limited to evaluation of anti-cancer potentials of crude condurango extract and its glycoside-rich components in vitro on lung cancer. Objective: This study aims at evaluating the effect of the single isolated active ingredient condurangogenin A (ConA; C32H42O7) on A549, H522 and H460-nonsmall-cell lung cancer cells. Materials and Methods: ConA was isolated by column chromatography and analyzed by mass spectroscopy, Fourier transform infrared spectroscopy and proton-nuclear magnetic resonance. diphenyltetrazolium bromide assays were conducted on three cell-types using 6%-alcohol as control. Critical studies on cellular morphology, cell-cycle regulation, reactive oxygen species, mitochondrial membrane potential, and DNA-damage were made, and expressions of related signaling markers studied. Results: As IC50 doses of ConA proved to be too high and toxic to both A549 and H522 cells, all experimental studies were carried out on H460 cells with the IC50 dose (32 μg/ml − 24 h). Cellular morphology revealed typical apoptotic features after ConA treatment. At early treatment hours (2 h-12 h), maximum cells were arrested at G0/G1 phase that could be correlated with reduced level of cyclin D1-CDK with p21 up-regulation. At 18 h − 24 h, sub G0/G1 cell population was increased gradually, as revealed from cytochrome-c release and caspase-3 activation, further confirming the apoptosis-inducing ability of ConA at later phases. Gradual increase of TUNEL-positive cells with significant modulation of mitochondria-dependent apoptotic markers at longer time-points would establish apoptosis-induction property of ConA, indicating its potential as a strong candidate for anti-cancer drug formulation. Conclusion: Further studies are warranted against other types of cancer cells and animal models before its possible human use. PMID:26109778

  16. RNF43 interacts with NEDL1 and regulates p53-mediated transcription

    SciTech Connect

    Shinada, Keisuke; Tsukiyama, Tadasuke; Sho, Takuya; Okumura, Fumihiko; Asaka, Masahiro; Hatakeyama, Shigetsugu

    2011-01-07

    Research highlights: {yields} RNF43 binds to NEDD-4-like ubiquitin-protein ligase-1 (NEDL1). {yields} RNF43 interacts with p53 and suppresses transcriptional activity of p53. {yields} RNF43 attenuates apoptosis induced by ultraviolet irradiation. {yields} RNF43 is likely associated with p53-mediated apoptosis in collaboration with NEDL1 in colorectal carcinogenesis. -- Abstract: The ubiquitin-proteasomal system plays a crucial role in oncogenesis in colorectal tissues. Recent studies have shown that stability of {beta}-catenin, which functions as an oncogene for colorectal cancer, is regulated by ubiquitin-mediated degradation. It has been reported that a putative E3 ubiquitin ligase, RNF43, is highly expressed in human colorectal carcinoma and that RNF43 promotes cell growth. However, the involvement of RNF43 in carcinogenesis has not been fully elucidated. In this study, we found by using yeast two-hybrid screening that RNF43 binds to NEDD-4-like ubiquitin-protein ligase-1 (NEDL1), which enhances pro-apoptotic activity by p53. In addition, we found that RNF43 also interacts with p53 and that RNF43 suppresses transcriptional activity of p53 in H1299 cells and attenuates apoptosis induced by ultraviolet irradiation. These findings suggest that RNF43 is associated with p53-mediated apoptosis in collaboration with NEDL1 in colorectal carcinogenesis.

  17. p53-Mediated Cellular Response to DNA Damage in Cells with Replicative Hepatitis B Virus

    NASA Astrophysics Data System (ADS)

    Puisieux, Alain; Ji, Jingwei; Guillot, Celine; Legros, Yann; Soussi, Thierry; Isselbacher, Kurt; Ozturk, Mehmet

    1995-02-01

    Wild-type p53 acts as a tumor suppressor gene by protecting cells from deleterious effects of genotoxic agents through the induction of a G_1/S arrest or apoptosis as a response to DNA damage. Transforming proteins of several oncogenic DNA viruses inactivate tumor suppressor activity of p53 by blocking this cellular response. To test whether hepatitis B virus displays a similar effect, we studied the p53-mediated cellular response to DNA damage in 2215 hepatoma cells with replicative hepatitis B virus. We demonstrate that hepatitis B virus replication does not interfere with known cellular functions of p53 protein.

  18. Development of a mechanistically-based genetically engineered PC12 cell system to detect p53-mediated cytotoxicity.

    PubMed

    van Vliet, Erwin; Eskes, Chantra; Stingele, Silvia; Gartlon, Joanne; Price, Anna; Farina, Massimo; Ponti, Jessica; Hartung, Thomas; Sabbioni, Enrico; Coecke, Sandra

    2007-06-01

    The human wild type p53 gene, key for apoptosis, was introduced into the pheochromocytoma (PC12) cell line, to create a mechanistically-based in vitro test model for the detection of p53-mediated toxicity. Expression of the wt p53 gene was regulated by a system, which allowed or blocked expression p53 by absence or presence of tetracycline in the culture media. Western blot analyses confirmed an inducible and tetracycline-dependent expression of the wt p53 protein. Functionality of the p53 protein was verified by camptothecin treatment, known to induce p53-dependent apoptosis. Results showed that p53-expressing cells were significantly more sensitive to camptothecin induced cytotoxicity compared to non-expressing cells, and presented a significantly higher incidence of apoptosis. A screening study on 31 metal compounds, showed that the classified human carcinogens (NaAsO2, CdSO4 .8H2O, Na2CrO4 .4H2O, MnCl2, (NH4)2PtCl6) significantly increased cytotoxicity in p53-expressing cells compared to non-expressing cells, suggesting that their cytotoxicity was p53-mediated. Finally, acute and subchronic treatment with methyl mercury showed no significant differences in cytotoxicity and the percentage of apoptosis or necrosis between p53-expressing and non-expressing differentiated cells, suggesting that methyl mercury cytotoxicity was p53-independent. PMID:17258428

  19. Simian virus 40 T antigen can regulate p53-mediated transcription independent of binding p53.

    PubMed Central

    Rushton, J J; Jiang, D; Srinivasan, A; Pipas, J M; Robbins, P D

    1997-01-01

    A simian virus 40 (SV40) T-antigen mutant containing only the N-terminal 136 amino acids, able to bind to Rb and p300 but not p53, partially inhibited p53-mediated transcription without affecting the ability of p53 to bind DNA. These results suggest that SV40 T antigen can regulate p53-mediated transcription either directly through protein-protein association or indirectly through interaction with factors which may function to confer p53-mediated transcription. PMID:9188637

  20. ASPP2 involvement in p53-mediated HIV-1 envelope glycoprotein gp120 neurotoxicity in mice cerebrocortical neurons.

    PubMed

    Liu, Zhiying; Zang, Yunjin; Qiao, Luxin; Liu, Kai; Ouyang, Yabo; Zhang, Yulin; Chen, Dexi

    2016-01-01

    The mechanisms behind HIV-1-associated neurocognitive disorders are still unclear. Apoptosis-stimulating protein 2 of p53 (ASPP2) is a damage-inducible p53-binding protein that stimulates p53-mediated apoptosis and transactivates proapoptotic and cell cycle regulatory genes. It has been reported that ASPP2 has a specific regulatory function in the death of retinal ganglion cells and the development of Alzheimer's disease. In this study, we used p53 and ASPP2 knockout mice and primary cerebrocortical neuron culture to analyze the role of the interaction between ASPP2 with p53 in HIV-1 envelope glycoprotein gp120-induced neurotoxicity. The results showed that 10 ng/mL gp120 protein might stimulate p53 overexpression and translocation to the nucleus, and 30 ng/mL gp120 protein could stimulate both p53 and ASPP2 translocation to the nucleus, but only with p53 overexpression. The primary cultured neurons of p53(-/-)ASPP2(+/-) mice had a higher survival rate than p53(-/-) mice under gp120 protein stress. The interaction of ASPP2 with p53 induced by a high dose of gp120 stimulated Bax transcription and contributed to caspase-3 cleavage, and ASPP2-siRNA attenuated gp120 induced neuron death through inhibition of Bax expression. These results suggest that ASPP2 plays an important role in p53-mediated neuronal apoptosis under gp120 stress. PMID:27625111

  1. S-Nitrosylation of parkin as a novel regulator of p53-mediated neuronal cell death in sporadic Parkinson’s disease

    PubMed Central

    2013-01-01

    Background Mutations in the gene encoding parkin, a neuroprotective protein with dual functions as an E3 ubiquitin ligase and transcriptional repressor of p53, are linked to familial forms of Parkinson’s disease (PD). We hypothesized that oxidative posttranslational modification of parkin by environmental toxins may contribute to sporadic PD. Results We first demonstrated that S-nitrosylation of parkin decreased its activity as a repressor of p53 gene expression, leading to upregulation of p53. Chromatin immunoprecipitation as well as gel-shift assays showed that parkin bound to the p53 promoter, and this binding was inhibited by S-nitrosylation of parkin. Additionally, nitrosative stress induced apoptosis in cells expressing parkin, and this death was, at least in part, dependent upon p53. In primary mesencephalic cultures, pesticide-induced apoptosis was prevented by inhibition of nitric oxide synthase (NOS). In a mouse model of pesticide-induced PD, both S-nitrosylated (SNO-)parkin and p53 protein levels were increased, while administration of a NOS inhibitor mitigated neuronal death in these mice. Moreover, the levels of SNO-parkin and p53 were simultaneously elevated in postmortem human PD brain compared to controls. Conclusions Taken together, our data indicate that S-nitrosylation of parkin, leading to p53-mediated neuronal cell death, contributes to the pathophysiology of sporadic PD. PMID:23985028

  2. Titanium dioxide nanoparticles trigger p53-mediated damage response in peripheral blood lymphocytes.

    PubMed

    Kang, Su Jin; Kim, Byeong Mo; Lee, Young Joon; Chung, Hai Won

    2008-06-01

    Titanium dioxide nanoparticles (nano-TiO2) are widely used as a photocatalyst in air and water remediation. These nanoparticles are known to induce toxicity; however, their cytotoxic mechanism is not fully understood. In this study, we investigated the underlying mechanism of nano-TiO2-induced cytotoxicity in peripheral blood lymphocytes. We examined the genotoxic effects of nano-TiO2 in lymphocytes using alkaline single-cell gel electrophoresis (Comet) and cytokinesis-block micronucleus (CBMN) assays. Lymphocytes treated with nano-TiO2 showed significantly increased micronucleus formation and DNA breakage. Western-blot analysis to identify proteins involved in the p53-mediated response to DNA damage revealed the accumulation of p53 and activation of DNA damage checkpoint kinases in nano-TiO2-treated lymphocytes. However, p21 and bax, downstream targets of p53, were not affected, indicating that nano-TiO2 does not stimulate transactivational activity of p53. The generation of reactive oxygen species (ROS) in nano-TiO2-treated cells was also observed, andN-acetylcysteine (NAC) supplementation inhibited the level of nano-TiO2-induced DNA damage. Given that ROS-induced DNA damage leads to p53 activation in the DNA damage response, our results suggest that nano-TiO2 induces ROS generation in lymphocytes, thereby activating p53-mediated DNA damage checkpoint signals. PMID:18418868

  3. Mammary gland-specific ablation of focal adhesion kinase reduces the incidence of p53-mediated mammary tumour formation

    PubMed Central

    van Miltenburg, M H A M; van Nimwegen, M J; Tijdens, I; Lalai, R; Kuiper, R; Klarenbeek, S; Schouten, P C; de Vries, A; Jonkers, J; van de Water, B

    2014-01-01

    kinase deletion reduced proliferative capacity of p53 null and p53R270H mammary epithelial cells but did not lead to increased apoptosis in vivo. Our data identify FAK as an important regulator in mammary epithelial cell proliferation in p53-mediated and p53R270H-induced mammary tumour development. PMID:24809783

  4. P53-mediated rapid induction of apoptosis conveys resistance to viral infection in Drosophila melanogaster

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Arthropod-borne pathogens account for millions of deaths each year. Understanding the genetic mechanisms controlling vector susceptibility to pathogens has profound implications for developing novel strategies for controlling insect transmitted infectious diseases. The fact that many viruses carry...

  5. Dephosphorylation of DBC1 by Protein Phosphatase 4 Is Important for p53-Mediated Cellular Functions

    PubMed Central

    Lee, Jihye; Adelmant, Guillaume; Marto, Jarrod A.; Lee, Dong-Hyun

    2015-01-01

    Deleted in breast cancer-1 (DBC1) contributes to the regulation of cell survival and apoptosis. Recent studies demonstrated that DBC is phosphorylated at Thr454 by ATM/ATR kinases in response to DNA damage, which is a critical event for p53 activation and apoptosis. However, how DBC1 phosphorylation is regulated has not been studied. Here we show that protein phosphatase 4 (PP4) dephosphorylates DBC1, regulating its role in DNA damage response. PP4R2, a regulatory subunit of PP4, mediates the interaction between DBC1 and PP4C, a catalytic subunit. PP4C efficiently dephosphorylates pThr454 on DBC1 in vitro, and the depletion of PP4C/PP4R2 in cells alters the kinetics of DBC1 phosphorylation and p53 activation, and increases apoptosis in response to DNA damage, which are compatible with the expression of the phosphomimetic DBC-1 mutant (T454E). These suggest that the PP4-mediated dephosphorylation of DBC1 is necessary for efficient damage responses in cells. PMID:26194823

  6. Dephosphorylation of DBC1 by Protein Phosphatase 4 Is Important for p53-Mediated Cellular Functions.

    PubMed

    Lee, Jihye; Adelmant, Guillaume; Marto, Jarrod A; Lee, Dong-Hyun

    2015-08-01

    Deleted in breast cancer-1 (DBC1) contributes to the regulation of cell survival and apoptosis. Recent studies demonstrated that DBC is phosphorylated at Thr454 by ATM/ATR kinases in response to DNA damage, which is a critical event for p53 activation and apoptosis. However, how DBC1 phosphorylation is regulated has not been studied. Here we show that protein phosphatase 4 (PP4) dephosphorylates DBC1, regulating its role in DNA damage response. PP4R2, a regulatory subunit of PP4, mediates the interaction between DBC1 and PP4C, a catalytic subunit. PP4C efficiently dephosphorylates pThr454 on DBC1 in vitro, and the depletion of PP4C/PP4R2 in cells alters the kinetics of DBC1 phosphorylation and p53 activation, and increases apoptosis in response to DNA damage, which are compatible with the expression of the phosphomimetic DBC-1 mutant (T454E). These suggest that the PP4-mediated dephosphorylation of DBC1 is necessary for efficient damage responses in cells. PMID:26194823

  7. The p53-mediated cytotoxicity of photodynamic therapy of cancer: Recent advances

    SciTech Connect

    Zawacka-Pankau, Joanna Krachulec, Justyna Grulkowski, Ireneusz Bielawski, Krzysztof P. Selivanova, Galina

    2008-11-01

    Photodynamic therapy (PDT) is a promising modality for the treatment of both pre-malignant and malignant lesions. The mechanism of action converges mainly on the generation of reactive oxygen species which damage cancer cells directly as well as indirectly acting on tumor vasculature. The exact mechanism of PDT action is not fully understood, which is a formidable barrier to its successful clinical application. Elucidation of the mechanisms of cancer cell elimination by PDT might help in establishing highly specific, non-genotoxic anti-cancer treatment of tomorrow. One of the candidate PDT targets is the well-known tumor suppressor p53 protein recognized as the guardian of the genome. Together with its family members, p73 and p63 proteins, p53 is involved in apoptosis induction upon stress stimuli. The wild-type and mutant p53-targeting chemotherapeutics are currently extensively investigated as a promising strategy for highly specific anti-cancer therapy. In photodynamic therapy porphyrinogenic sensitizers are the most widely used compounds due to their potent biophysical and biochemical properties. Recent data suggest that the p53 tumor suppressor protein might play a significant role in porphyrin-PDT-mediated cell death by direct interaction with the drug which leads to its accumulation and induction of p53-dependent cell death both in the dark and upon irradiation. In this review we describe the available evidence on the role of p53 in PDT.

  8. Loss of Atrx sensitizes cells to DNA damaging agents through p53-mediated death pathways.

    PubMed

    Conte, Damiano; Huh, Michael; Goodall, Emma; Delorme, Marilyne; Parks, Robin J; Picketts, David J

    2012-01-01

    Prevalent cell death in forebrain- and Sertoli cell-specific Atrx knockout mice suggest that Atrx is important for cell survival. However, conditional ablation in other tissues is not associated with increased death indicating that diverse cell types respond differently to the loss of this chromatin remodeling protein. Here, primary macrophages isolated from Atrx(f/f) mice were infected with adenovirus expressing Cre recombinase or β-galactosidase, and assayed for cell survival under different experimental conditions. Macrophages survive without Atrx but undergo rapid apoptosis upon lipopolysaccharide (LPS) activation suggesting that chromatin reorganization in response to external stimuli is compromised. Using this system we next tested the effect of different apoptotic stimuli on cell survival. We observed that survival of Atrx-null cells were similar to wild type cells in response to serum withdrawal, anti-Fas antibody, C2 ceramide or dexamethasone treatment but were more sensitive to 5-fluorouracil (5-FU). Cell survival could be rescued by re-introducing Atrx or by removal of p53 demonstrating the cell autonomous nature of the effect and its p53-dependence. Finally, we demonstrate that multiple primary cell types (myoblasts, embryonic fibroblasts and neurospheres) were sensitive to 5-FU, cisplatin, and UV light treatment. Together, our results suggest that cells lacking Atrx are more sensitive to DNA damaging agents and that this may result in enhanced death during development when cells are at their proliferative peak. Moreover, it identifies potential treatment options for cancers associated with ATRX mutations, including glioblastoma and pancreatic neuroendocrine tumors. PMID:23284920

  9. Highly specific in vivo gene delivery for p53-mediated apoptosis and genetic photodynamic therapies of tumour

    PubMed Central

    Tseng, S.-Ja; Liao, Zi-Xian; Kao, Shih-Han; Zeng, Yi-Fang; Huang, Kuo-Yen; Li, Hsin-Jung; Yang, Chung-Lin; Deng, Yu-Fan; Huang, Chi-Feng; Yang, Shuenn-Chen; Yang, Pan-Chyr; Kempson, Ivan M.

    2015-01-01

    Anticancer therapies are often compromised by nonspecific effects and challenged by tumour environments’ inherent physicochemical and biological characteristics. Often, therapeutic effect can be increased by addressing multiple parameters simultaneously. Here we report on exploiting extravasation due to inherent vascular leakiness for the delivery of a pH-sensitive polymer carrier. Tumours’ acidic microenvironment instigates a charge reversal that promotes cellular internalization where endosomes destabilize and gene delivery is achieved. We assess our carrier with an aggressive non-small cell lung carcinoma (NSCLC) in vivo model and achieve >30% transfection efficiency via systemic delivery. Rejuvenation of the p53 apoptotic pathway as well as expression of KillerRed protein for sensitization in photodynamic therapy (PDT) is accomplished. A single administration greatly suppresses tumour growth and extends median animal survival from 28 days in control subjects to 68 days. The carrier has capacity for multiple payloads for greater therapeutic response where inter-individual variability can compromise efficacy. PMID:25739372

  10. Lipopolysaccharide prevents valproic acid-induced apoptosis via activation of nuclear factor-κB and inhibition of p53 activation.

    PubMed

    Tsolmongyn, Bilegtsaikhan; Koide, Naoki; Odkhuu, Erdenezaya; Haque, Abedul; Naiki, Yoshikazu; Komatsu, Takayuki; Yoshida, Tomoaki; Yokochi, Takashi

    2013-04-01

    The effect of lipopolysaccharide (LPS) on valproic acid (VPA)-induced cell death was examined by using mouse RAW 264.7 macrophage cells. LPS inhibited the activation of caspase 3 and poly (ADP-ribose) polymerase and prevented VPA-induced apoptosis. LPS inhibited VPA-induced p53 activation and pifithrin-α as a p53 inhibitor as well as LPS prevented VPA-induced apoptosis. LPS abolished the increase of Bax/Bcl-2 ratio, which is a critical indicator of p53-mediated mitochondrial damage, in response to VPA. The nuclear factor (NF)-κB inhibitors, Bay 11-7082 and parthenolide, abolished the preventive action of LPS on VPA-induced apoptosis. A series of toll-like receptor ligands, Pam3CSK4, poly I:C, and CpG DNA as well as LPS prevented VPA-induced apoptosis. Taken together, LPS was suggested to prevent VPA-induced apoptosis via activation of anti-apoptotic NF-κB and inhibition of pro-apoptotic p53 activation. The detailed inhibitory mechanism of VPA-induced apoptosis by LPS is discussed. PMID:23770718

  11. SETD2 is required for DNA double-strand break repair and activation of the p53-mediated checkpoint

    PubMed Central

    Carvalho, Sílvia; Vítor, Alexandra C; Sridhara, Sreerama C; Martins, Filipa B; Raposo, Ana C; Desterro, Joana MP; Ferreira, João; de Almeida, Sérgio F

    2014-01-01

    Histone modifications establish the chromatin states that coordinate the DNA damage response. In this study, we show that SETD2, the enzyme that trimethylates histone H3 lysine 36 (H3K36me3), is required for ATM activation upon DNA double-strand breaks (DSBs). Moreover, we find that SETD2 is necessary for homologous recombination repair of DSBs by promoting the formation of RAD51 presynaptic filaments. In agreement, SETD2-mutant clear cell renal cell carcinoma (ccRCC) cells displayed impaired DNA damage signaling. However, despite the persistence of DNA lesions, SETD2-deficient cells failed to activate p53, a master guardian of the genome rarely mutated in ccRCC and showed decreased cell survival after DNA damage. We propose that this novel SETD2-dependent role provides a chromatin bookmarking instrument that facilitates signaling and repair of DSBs. In ccRCC, loss of SETD2 may afford an alternative mechanism for the inactivation of the p53-mediated checkpoint without the need for additional genetic mutations in TP53. DOI: http://dx.doi.org/10.7554/eLife.02482.001 PMID:24843002

  12. CHIP stabilizes amyloid precursor protein via proteasomal degradation and p53-mediated trans-repression of β-secretase.

    PubMed

    Singh, Amir Kumar; Pati, Uttam

    2015-08-01

    In patient with Alzheimer's disease (AD), deposition of amyloid-beta Aβ, a proteolytic cleavage of amyloid precursor protein (APP) by β-secretase/BACE1, forms senile plaque in the brain. BACE1 activation is caused due to oxidative stresses and dysfunction of ubiquitin-proteasome system (UPS), which is linked to p53 inactivation. As partial suppression of BACE1 attenuates Aβ generation and AD-related pathology, it might be an ideal target for AD treatment. We have shown that both in neurons and in HEK-APP cells, BACE1 is a new substrate of E3-ligase CHIP and an inverse relation exists between CHIP and BACE1 level. CHIP inhibits ectopic BACE1 level by promoting its ubiquitination and proteasomal degradation, thus reducing APP processing; it stabilizes APP in neurons, thus reducing Aβ. CHIP(U) (box) domain physically interacts with BACE1; however, both U-box and TPR domain are essential for ubiquitination and degradation of BACE1. Further, BACE1 is a downstream target of p53 and overexpression of p53 decreases BACE1 level. In HEK-APP cells, CHIP is shown to negatively regulate BACE1 promoter through stabilization of p53's DNA-binding conformation and its binding upon 5' UTR element (+127 to +150). We have thus discovered that CHIP regulates p53-mediated trans-repression of BACE1 at both transcriptional and post-translational level. We propose that a CHIP-BACE1-p53 feedback loop might control APP stabilization, which could further be utilized for new therapeutic intervention in AD. PMID:25773675

  13. Ets-2 and p53 mediate cAMP-induced MMP-2 expression, activity and trophoblast invasion

    PubMed Central

    2009-01-01

    Background We have previously shown that Matrix metalloproteinase (MMP) -2 is a key-enzyme in early trophoblast invasion and that Protein Kinase A (PKA) increases MMP-2 expression and trophoblast invasion. The aim of this study was to examine MMP -2 regulation by PKA in invasive trophoblasts: JAR choriocarcinoma cell-line and 6-8 w first trimester trophoblasts. Methods The effect of Forskolin (PKA) on MMP-2 expression was assessed by Northern Blot and RT-PCR. Possible transcription factors binding to consensus MMP-2 promoter sequences in response to Forskolin, were detected by EMSA binding assay and their expression assessed by western blot analysis. Antisense transfection of relevant transcription factors was performed and the inhibitory effect assessed on MMP-2 expression (RT-PCR), secretion (zymography) and trophoblast invasiveness (transwell migration assay). Results We found that Forskolin increased MMP-2 mRNA in JAR cells within 24 hours, and induced binding to p53, Ets, C/EBP and AP-2. Transcription factors Ets-2, phospho- p53, C/EBP epsilon, C/EBP lambda and AP-2 alpha bound to their respective binding sequences in response to Forskolin and the expressions of these transcription factors were all elevated in Forskolin- treated cells. Inhibition of Ets-2 and p53 reduced MMP-2 expression, secretion and invasiveness of Forskolin treated cells. Conclusion MMP-2 is regulated by PKA through several binding sites and transcription factors including Ets-2, p53, C/EBP, C/EBP lambda and AP-2 alpha. Ets-2 and p53 mediate cAMP- induced trophoblast invasiveness, through regulation of MMP-2. PMID:19939245

  14. CHIP stabilizes amyloid precursor protein via proteasomal degradation and p53-mediated trans-repression of β-secretase

    PubMed Central

    Singh, Amir Kumar; Pati, Uttam

    2015-01-01

    In patient with Alzheimer’s disease (AD), deposition of amyloid-beta Aβ, a proteolytic cleavage of amyloid precursor protein (APP) by β-secretase/BACE1, forms senile plaque in the brain. BACE1 activation is caused due to oxidative stresses and dysfunction of ubiquitin–proteasome system (UPS), which is linked to p53 inactivation. As partial suppression of BACE1 attenuates Aβ generation and AD-related pathology, it might be an ideal target for AD treatment. We have shown that both in neurons and in HEK-APP cells, BACE1 is a new substrate of E3-ligase CHIP and an inverse relation exists between CHIP and BACE1 level. CHIP inhibits ectopic BACE1 level by promoting its ubiquitination and proteasomal degradation, thus reducing APP processing; it stabilizes APP in neurons, thus reducing Aβ. CHIPUbox domain physically interacts with BACE1; however, both U-box and TPR domain are essential for ubiquitination and degradation of BACE1. Further, BACE1 is a downstream target of p53 and overexpression of p53 decreases BACE1 level. In HEK-APP cells, CHIP is shown to negatively regulate BACE1 promoter through stabilization of p53’s DNA-binding conformation and its binding upon 5′ UTR element (+127 to +150). We have thus discovered that CHIP regulates p53-mediated trans-repression of BACE1 at both transcriptional and post-translational level. We propose that a CHIP–BACE1–p53 feedback loop might control APP stabilization, which could further be utilized for new therapeutic intervention in AD. PMID:25773675

  15. p53-Mediated Biliary Defects Caused by Knockdown of cirh1a, the Zebrafish Homolog of the Gene Responsible for North American Indian Childhood Cirrhosis

    PubMed Central

    Wilkins, Benjamin J.; Lorent, Kristin; Matthews, Randolph P.; Pack, Michael

    2013-01-01

    North American Indian Childhood Cirrhosis (NAIC) is a rare, autosomal recessive, progressive cholestatic disease of infancy affecting the Cree-Ojibway first Nations of Quebec. All NAIC patients are homozygous for a missense mutation (R565W) in CIRH1A, the human homolog of the yeast nucleolar protein Utp4. Utp4 is part of the t-Utp subcomplex of the small subunit (SSU) processome, a ribonucleoprotein complex required for ribosomal RNA processing and small subunit assembly. NAIC has thus been proposed to be a primary ribosomal disorder (ribosomopathy); however, investigation of the pathophysiologic mechanism of this disease has been hindered by lack of an animal model. Here, using a morpholino oligonucleotide (MO)-based loss-of-function strategy, we have generated a model of NAIC in the zebrafish, Danio rerio. Zebrafish Cirhin shows substantial homology to the human homolog, and cirh1a mRNA is expressed in developing hepatocytes and biliary epithelial cells. Injection of two independent MOs directed against cirh1a at the one-cell stage causes defects in canalicular and biliary morphology in 5 dpf larvae. In addition, 5 dpf Cirhin-deficient larvae have dose-dependent defects in hepatobiliary function, as assayed by the metabolism of an ingested fluorescent lipid reporter. Previous yeast and in vitro studies have shown that defects in ribosome biogenesis cause stabilization and nuclear accumulation of p53, which in turn causes p53-mediated cell cycle arrest and/or apoptosis. Thus, the nucleolus appears to function as a cellular stress sensor in some cell types. In accordance with this hypothesis, transcriptional targets of p53 are upregulated in Cirhin-deficient zebrafish embryos, and defects in biliary function seen in Cirhin-deficient larvae are completely abrogated by mutation of tp53. Our data provide the first in vivo evidence of a role for Cirhin in biliary development, and support the hypothesis that congenital defects affecting ribosome biogenesis can activate

  16. The novel tryptamine derivative JNJ-26854165 induces wild-type p53- and E2F1-mediated apoptosis in acute myeloid and lymphoid leukemias

    PubMed Central

    Kojima, Kensuke; Burks, Jared K.; Arts, Janine; Andreeff, Michael

    2010-01-01

    Development of small-molecule activators of p53 is currently focused on malignancies containing a wild-type p53 genotype, which is present in most leukemias. JNJ-26854165 is one of such p53-activating agents, but its mechanism of action remains to be elucidated. Here, we report the effects of JNJ-26854165 in acute leukemias. JNJ-26854165 treatment induced p53-mediated apoptosis in acute leukemia cells with wild-type p53, in which p53 rapidly drives transcription-independent apoptosis followed by activation of a transcription-dependent pathway. JNJ-26854165 accelerated the proteasome-mediated degradation of p21, and antagonized the transcriptional induction of p21 by p53. Interestingly, JNJ-26854165 induced S-phase delay and upregulated E2F1 expression in p53 mutant cells, resulting in apoptosis preferentially of S-phase cells. E2F1 knockdown blocked apoptosis induced by JNJ-26854165 in p53 mutant cells. Apoptotic activity of JNJ-26854165 against primary acute leukemia cells was maintained in leukemia/stroma cocultures, unlike doxorubicin, which has reduced cytrotoxicity in coculture systems. JNJ-26854165 synergizes with AraC or doxorubicin to induce p53-mediated apoptosis. Our data suggest that JNJ-26854165 may provide a novel therapeutic approach for the treatment of acute leukemias. The presence of p53-independent apoptotic activity in addition to p53-mediated apoptosis induction, if operational in vivo, may prevent the selection of p53 mutant subclones during therapy. PMID:20736344

  17. Escape from p53-mediated tumor surveillance in neuroblastoma: switching off the p14(ARF)-MDM2-p53 axis.

    PubMed

    Van Maerken, T; Vandesompele, J; Rihani, A; De Paepe, A; Speleman, F

    2009-12-01

    A primary failsafe program against unrestrained proliferation and oncogenesis is provided by the p53 tumor suppressor protein, inactivation of which is considered as a hallmark of cancer. Intriguingly, mutations of the TP53 gene are rarely encountered in neuroblastoma tumors, suggesting that alternative p53-inactivating lesions account for escape from p53 control in this childhood malignancy. Several recent studies have shed light on the mechanisms by which neuroblastoma cells circumvent the p53-driven antitumor barrier. We review here these mechanisms for evasion of p53-mediated growth control and conclude that deregulation of the p14(ARF)-MDM2-p53 axis seems to be the principal mode of p53 inactivation in neuroblastoma, opening new perspectives for targeted therapeutic intervention. PMID:19779493

  18. Wild-type and mutant p53 mediate cisplatin resistance through interaction and inhibition of active caspase-9.

    PubMed

    Chee, Jacqueline L Y; Saidin, Suzan; Lane, David P; Leong, Sai Mun; Noll, Jacqueline E; Neilsen, Paul M; Phua, Yi Ting; Gabra, Hani; Lim, Tit Meng

    2013-01-15

    The p53 gene has been implicated in many cancers due to its frequent mutations as well as mutations in other genes whose proteins directly affect p53's functions. In addition, high expression of p53 [wild-type (WT) or mutant] has been found in the cytoplasm of many tumor cells, and studies have associated these observations with more aggressive tumors and poor prognosis. Cytoplasmic mis-localization of p53 subsequently reduced its transcriptional activity and this loss-of-function (LOF) was used to explain the lack of response to chemotherapeutic agents. However, this hypothesis seemed inadequate in explaining the apparent selection for tumor cells with high levels of p53 protein, a phenomenon that suggests a gain-of-function (GOF) of these mis-localized p53 proteins. In this study, we explored whether the direct involvement of p53 in the apoptotic response is via regulation of the caspase pathway in the cytoplasm. We demonstrate that p53, when present at high levels in the cytoplasm, has an inhibitory effect on caspase-9. Concurrently, knockdown of endogenous p53 caused an increase in the activity of caspase-9. p53 was found to interact with the p35 fragment of caspase-9, and this interaction inhibits the caspase-9 activity. In a p53-null background, the high-level expression of both exogenous WT and mutant p53 increased the resistance of these cells to cisplatin, and the data showed a correlation between high p53 expression and caspase-9 inhibition. These results suggest the inhibition of caspase-9 as a potential mechanism in evading apoptosis in tumors with high-level p53 expression that is cytoplasmically localized. PMID:23255126

  19. Wild-type and mutant p53 mediate cisplatin resistance through interaction and inhibition of active caspase-9

    PubMed Central

    Chee, Jacqueline L.Y.; Saidin, Suzan; Lane, David P.; Leong, Sai Mun; Noll, Jacqueline E.; Neilsen, Paul M.; Phua, Yi Ting; Gabra, Hani; Lim, Tit Meng

    2013-01-01

    The p53 gene has been implicated in many cancers due to its frequent mutations as well as mutations in other genes whose proteins directly affect p53’s functions. In addition, high expression of p53 [wild-type (WT) or mutant] has been found in the cytoplasm of many tumor cells, and studies have associated these observations with more aggressive tumors and poor prognosis. Cytoplasmic mis-localization of p53 subsequently reduced its transcriptional activity and this loss-of-function (LOF) was used to explain the lack of response to chemotherapeutic agents. However, this hypothesis seemed inadequate in explaining the apparent selection for tumor cells with high levels of p53 protein, a phenomenon that suggests a gain-of-function (GOF) of these mis-localized p53 proteins. In this study, we explored whether the direct involvement of p53 in the apoptotic response is via regulation of the caspase pathway in the cytoplasm. We demonstrate that p53, when present at high levels in the cytoplasm, has an inhibitory effect on caspase-9. Concurrently, knockdown of endogenous p53 caused an increase in the activity of caspase-9. p53 was found to interact with the p35 fragment of caspase-9, and this interaction inhibits the caspase-9 activity. In a p53-null background, the high-level expression of both exogenous WT and mutant p53 increased the resistance of these cells to cisplatin, and the data showed a correlation between high p53 expression and caspase-9 inhibition. These results suggest the inhibition of caspase-9 as a potential mechanism in evading apoptosis in tumors with high-level p53 expression that is cytoplasmically localized. PMID:23255126

  20. A sustained deficiency of mitochondrial respiratory complex III induces an apoptotic cell death through the p53-mediated inhibition of pro-survival activities of the activating transcription factor 4

    PubMed Central

    Evstafieva, A G; Garaeva, A A; Khutornenko, A A; Klepikova, A V; Logacheva, M D; Penin, A A; Novakovsky, G E; Kovaleva, I E; Chumakov, P M

    2014-01-01

    Generation of energy in mitochondria is subjected to physiological regulation at many levels, and its malfunction may result in mitochondrial diseases. Mitochondrial dysfunction is associated with different environmental influences or certain genetic conditions, and can be artificially induced by inhibitors acting at different steps of the mitochondrial electron transport chain (ETC). We found that a short-term (5 h) inhibition of ETC complex III with myxothiazol results in the phosphorylation of translation initiation factor eIF2α and upregulation of mRNA for the activating transcription factor 4 (ATF4) and several ATF4-regulated genes. The changes are characteristic for the adaptive integrated stress response (ISR), which is known to be triggered by unfolded proteins, nutrient and metabolic deficiency, and mitochondrial dysfunctions. However, after a prolonged incubation with myxothiazol (13–17 h), levels of ATF4 mRNA and ATF4-regulated transcripts were found substantially suppressed. The suppression was dependent on the p53 response, which is triggered by the impairment of the complex III-dependent de novo biosynthesis of pyrimidines by mitochondrial dihydroorotate dehydrogenase. The initial adaptive induction of ATF4/ISR acted to promote viability of cells by attenuating apoptosis. In contrast, the induction of p53 upon a sustained inhibition of ETC complex III produced a pro-apoptotic effect, which was additionally stimulated by the p53-mediated abrogation of the pro-survival activities of the ISR. Interestingly, a sustained inhibition of ETC complex I by piericidine did not induce the p53 response and stably maintained the pro-survival activation of ATF4/ISR. We conclude that a downregulation of mitochondrial ETC generally induces adaptive pro-survival responses, which are specifically abrogated by the suicidal p53 response triggered by the genetic risks of the pyrimidine nucleotide deficiency. PMID:25375376

  1. Mimulone-Induced Autophagy through p53-Mediated AMPK/mTOR Pathway Increases Caspase-Mediated Apoptotic Cell Death in A549 Human Lung Cancer Cells

    PubMed Central

    Lee, Ji-Won; Park, Mi-Hyun; Moon, Hyung-In; Park, Shin-Ji; Baik, Ji-Sue; Kim, Cheorl-Ho; Lee, Young-Choon

    2014-01-01

    Anticancer properties and mechanisms of mimulone (MML), C-geranylflavonoid isolated from the Paulownia tomentosa fruits, were firstly elucidated in this study. MML prevented cell proliferation in a dose- and time-dependent way and triggered apoptosis through the extrinsic pathway in A549 human lung adenocarcinoma cells. Furthermore, MML-treated cells displayed autophagic features, such as the formation of autophagic vacuoles, a primary morphological feature of autophagy, and the accumulation of microtubule-associated protein 1 light chain 3 (LC3) puncta, another typical maker of autophagy, as determined by FITC-conjugated immunostaining and monodansylcadaverine (MDC) staining, respectively. The expression levels of LC3-I and LC3-II, specific markers of autophagy, were also augmented by MML treatment. Autophagy inhibition by 3-methyladenine (3-MA), pharmacological autophagy inhibitor, and shRNA knockdown of Beclin-1 reduced apoptotic cell death induced by MML. Autophagic flux was not significantly affected by MML treatment and lysosomal inhibitor, chloroquine (CQ) suppressed MML-induced autophagy and apoptosis. MML-induced autophagy was promoted by decreases in p53 and p-mTOR levels and increase of p-AMPK. Moreover, inhibition of p53 transactivation by pifithrin-α (PFT-α) and knockdown of p53 enhanced induction of autophagy and finally promoted apoptotic cell death. Overall, the results demonstrate that autophagy contributes to the cytotoxicity of MML in cancer cells harboring wild-type p53. This study strongly suggests that MML is a potential candidate for an anticancer agent targeting both autophagy and apoptotic cell death in human lung cancer. Moreover, co-treatment of MML and p53 inhibitor would be more effective in human lung cancer therapy. PMID:25490748

  2. Mimulone-induced autophagy through p53-mediated AMPK/mTOR pathway increases caspase-mediated apoptotic cell death in A549 human lung cancer cells.

    PubMed

    An, Hyun-Kyu; Kim, Kyoung-Sook; Lee, Ji-Won; Park, Mi-Hyun; Moon, Hyung-In; Park, Shin-Ji; Baik, Ji-Sue; Kim, Cheorl-Ho; Lee, Young-Choon

    2014-01-01

    Anticancer properties and mechanisms of mimulone (MML), C-geranylflavonoid isolated from the Paulownia tomentosa fruits, were firstly elucidated in this study. MML prevented cell proliferation in a dose- and time-dependent way and triggered apoptosis through the extrinsic pathway in A549 human lung adenocarcinoma cells. Furthermore, MML-treated cells displayed autophagic features, such as the formation of autophagic vacuoles, a primary morphological feature of autophagy, and the accumulation of microtubule-associated protein 1 light chain 3 (LC3) puncta, another typical maker of autophagy, as determined by FITC-conjugated immunostaining and monodansylcadaverine (MDC) staining, respectively. The expression levels of LC3-I and LC3-II, specific markers of autophagy, were also augmented by MML treatment. Autophagy inhibition by 3-methyladenine (3-MA), pharmacological autophagy inhibitor, and shRNA knockdown of Beclin-1 reduced apoptotic cell death induced by MML. Autophagic flux was not significantly affected by MML treatment and lysosomal inhibitor, chloroquine (CQ) suppressed MML-induced autophagy and apoptosis. MML-induced autophagy was promoted by decreases in p53 and p-mTOR levels and increase of p-AMPK. Moreover, inhibition of p53 transactivation by pifithrin-α (PFT-α) and knockdown of p53 enhanced induction of autophagy and finally promoted apoptotic cell death. Overall, the results demonstrate that autophagy contributes to the cytotoxicity of MML in cancer cells harboring wild-type p53. This study strongly suggests that MML is a potential candidate for an anticancer agent targeting both autophagy and apoptotic cell death in human lung cancer. Moreover, co-treatment of MML and p53 inhibitor would be more effective in human lung cancer therapy. PMID:25490748

  3. AS-2, a novel inhibitor of p53-dependent apoptosis, prevents apoptotic mitochondrial dysfunction in a transcription-independent manner and protects mice from a lethal dose of ionizing radiation.

    PubMed

    Morita, Akinori; Ariyasu, Shinya; Wang, Bing; Asanuma, Tetsuo; Onoda, Takayoshi; Sawa, Akiko; Tanaka, Kaoru; Takahashi, Ippei; Togami, Shotaro; Nenoi, Mitsuru; Inaba, Toshiya; Aoki, Shin

    2014-08-01

    In a previous study, we reported that some tetradentate zinc(II) chelators inhibit p53 through the denaturation of its zinc-requiring structure but a chelator, Bispicen, a potent inhibitor of in vitro apoptosis, failed to show any efficient radioprotective effect against irradiated mice because the toxicity of the chelator to mice. The unsuitability of using tetradentate chelators as radioprotectors prompted us to undertake a more extensive search for p53-inhibiting agents that are weaker zinc(II) chelators and therefore less toxic. Here, we show that an 8-hydroxyquinoline (8HQ) derivative, AS-2, suppresses p53-dependent apoptosis through a transcription-independent mechanism. A mechanistic study using cells with different p53 characteristics revealed that the suppressive effect of AS-2 on apoptosis is specifically mediated through p53. In addition, AS-2 was less effective in preventing p53-mediated transcription-dependent events than pifithrin-μ (PFTμ), an inhibitor of transcription-independent apoptosis by p53. Fluorescence visualization of the extranuclear distribution of AS-2 also supports that it is ineffective on the transcription-dependent pathway. Further investigations revealed that AS-2 suppressed mitochondrial apoptotic events, such as the mitochondrial release of intermembrane proteins and the loss of mitochondrial membrane potential, although AS-2 resulted in an increase in the mitochondrial translocation of p53 as opposed to the decrease of cytosolic p53, and did not affect the apoptotic interaction of p53 with Bcl-2. AS-2 also protected mice that had been exposed to a lethal dose of ionizing radiation. Our findings indicate that some types of bidentate 8HQ chelators could serve as radioprotectors with no substantial toxicity in vivo. PMID:25026551

  4. p53-mediated transcriptional regulation and activation of the actin cytoskeleton regulatory RhoC to LIMK2 signaling pathway promotes cell survival

    PubMed Central

    Croft, Daniel R; Crighton, Diane; Samuel, Michael S; Lourenco, Filipe C; Munro, June; Wood, Jenifer; Bensaad, Karim; Vousden, Karen H; Sansom, Owen J; Ryan, Kevin M; Olson, Michael F

    2011-01-01

    The central arbiter of cell fate in response to DNA damage is p53, which regulates the expression of genes involved in cell cycle arrest, survival and apoptosis. Although many responses initiated by DNA damage have been characterized, the role of actin cytoskeleton regulators is largely unknown. We now show that RhoC and LIM kinase 2 (LIMK2) are direct p53 target genes induced by genotoxic agents. Although RhoC and LIMK2 have well-established roles in actin cytoskeleton regulation, our results indicate that activation of LIMK2 also has a pro-survival function following DNA damage. LIMK inhibition by siRNA-mediated knockdown or selective pharmacological blockade sensitized cells to radio- or chemotherapy, such that treatments that were sub-lethal when administered singly resulted in cell death when combined with LIMK inhibition. Our findings suggest that combining LIMK inhibitors with genotoxic therapies could be more efficacious than single-agent administration, and highlight a novel connection between actin cytoskeleton regulators and DNA damage-induced cell survival mechanisms. PMID:21079653

  5. Quercetin-induced apoptosis prevents EBV infection

    PubMed Central

    Lee, Minjung; Son, Myoungki; Ryu, Eunhyun; Shin, Yu Su; Kim, Jong Gwang; Kang, Byung Woog; Sung, Gi-Ho; Cho, Hyosun; Kang, Hyojeung

    2015-01-01

    Epstein-Barr virus (EBV) is a human gamma-1 herpesvirus that establishes a lifelong latency in over 90% of the world's population. During latency, virus exists predominantly as a chromatin-associated, multicopy episome in the nuclei of a variety of tumor cells derived from B cells, T cells, natural killer (NK) cells, and epithelial cells. Licorice is the root of Glycyrrhiza uralensis or G. glabra that has traditionally cultivated in eastern part of Asia. Licorice was reported to have anti-viral, anti-inflammatory, anti-atopic, hepatoprotective, anti-neurodegenerative, anti-tumor, anti-diabetic effects and so forth. Quercetin and isoliquiritigenin are produced from licorice and highly similar in molecular structure. They have diverse bioactive effects such as antiviral activity, anti-asthmatic activity, anti-cancer activity, anti-inflammation activity, monoamine-oxidase inhibitor, and etc. To determine anti-EBV and anti-EBVaGC (Epstein-Barr virus associated gastric carcinoma) effects of licorice, we investigated antitumor and antiviral effects of quercetin and isoliquiritigenin against EBVaGC. Although both quercetin and isoliquiritigenin are cytotoxic to SNU719 cells, quercetin induced more apoptosis in SNU719 cells than isoliquiritigenin, more completely eliminated DNMT1 and DNMT3A expressions than isoliquiritigenin, and more strongly affects the cell cycle progression of SNU719 than isoliquiritigenin. Both quercetin and isoliquiritigenin induce signal transductions to stimulate apoptosis, and induce EBV gene transcription. Quercetin enhances frequency of F promoter use, whereas isoliquiritigenin enhances frequency of Q promoter use. Quercetin reduces EBV latency, whereas isoliquiritigenin increases the latency. Quercetin increases more the EBV progeny production, and inhibits more EBV infection than isoliquiritigenin. These results indicate that quercetin could be a promising candidate for antiviral and antitumor agents against EBV and human gastric carcinoma

  6. p38 MAPK-induced MDM2 degradation confers paclitaxel resistance through p53-mediated regulation of EGFR in human lung cancer cells

    PubMed Central

    Park, Shin-Hyung; Seong, Myeong-A; Lee, Ho-Young

    2016-01-01

    Paclitaxel (PTX) is a chemotherapeutic agent that is used to treat a variety of cancers, including non-small cell lung cancer (NSCLC). However, the emergence of drug resistance limits the utility of PTX. This study determined the signaling pathway that contributes to PTX resistance. We first established PTX resistant cell lines (H460/R and 226B/R) using a dose-escalating maintenance of PTX. We found that p38 MAPK and epidermal growth factor receptor (EGFR) were constitutively activated in these cell lines. The inhibition of p38 MAPK activity by SB203580 treatment or the transfection of dominant-negative p38 MAPK sensitized both cell lines to PTX treatment. Erlotinib, an EGFR inhibitor, also increased PTX-induced apoptosis in PTX resistant cells, which suggests a role for p38 MAPK and EGFR in the development of PTX resistance. We demonstrated that p38 MAPK enhanced EGFR expression via the induction of the rapid degradation of mouse double-minute 2 homolog (MDM2) and the consequent stabilization of p53, a transcription factor of EGFR. These results suggest for the first time that the p38 MAPK/p53/EGFR axis is crucial for the facilitation of PTX resistance in NSCLCs. We also propose a mechanism for the role of the tumor-suppressor p53 in drug resistance. These results provide a foundation for the future development of potential therapeutic strategies to regulate the p38 MAPK/p53/EGFR pathway for the treatment of lung cancer patients with PTX resistance. PMID:26799187

  7. AS-2, a novel inhibitor of p53-dependent apoptosis, prevents apoptotic mitochondrial dysfunction in a transcription-independent manner and protects mice from a lethal dose of ionizing radiation

    SciTech Connect

    Morita, Akinori; Ariyasu, Shinya; Wang, Bing; Asanuma, Tetsuo; Onoda, Takayoshi; Sawa, Akiko; Tanaka, Kaoru; Takahashi, Ippei; Togami, Shotaro; Nenoi, Mitsuru; Inaba, Toshiya; Aoki, Shin

    2014-08-08

    Highlights: • A bidentate HQ derivative, AS-2, suppresses p53-dependent apoptosis by DNA damage. • AS-2 does not significantly affect nuclear p53 response. • UV-excited blue emission of AS-2 clearly showed its extranuclear localization. • AS-2 prevents mitochondrial dysfunction despite the increase of mitochondrial p53. • AS-2 protects mice from a radiation dose that causes lethal hematopoietic syndrome. - Abstract: In a previous study, we reported that some tetradentate zinc(II) chelators inhibit p53 through the denaturation of its zinc-requiring structure but a chelator, Bispicen, a potent inhibitor of in vitro apoptosis, failed to show any efficient radioprotective effect against irradiated mice because the toxicity of the chelator to mice. The unsuitability of using tetradentate chelators as radioprotectors prompted us to undertake a more extensive search for p53-inhibiting agents that are weaker zinc(II) chelators and therefore less toxic. Here, we show that an 8-hydroxyquinoline (8HQ) derivative, AS-2, suppresses p53-dependent apoptosis through a transcription-independent mechanism. A mechanistic study using cells with different p53 characteristics revealed that the suppressive effect of AS-2 on apoptosis is specifically mediated through p53. In addition, AS-2 was less effective in preventing p53-mediated transcription-dependent events than pifithrin-μ (PFTμ), an inhibitor of transcription-independent apoptosis by p53. Fluorescence visualization of the extranuclear distribution of AS-2 also supports that it is ineffective on the transcription-dependent pathway. Further investigations revealed that AS-2 suppressed mitochondrial apoptotic events, such as the mitochondrial release of intermembrane proteins and the loss of mitochondrial membrane potential, although AS-2 resulted in an increase in the mitochondrial translocation of p53 as opposed to the decrease of cytosolic p53, and did not affect the apoptotic interaction of p53 with Bcl-2. AS-2 also

  8. Metformin prevents methylglyoxal-induced apoptosis of mouse Schwann cells

    SciTech Connect

    Ota, Kimiko; Nakamura, Jiro; Li, Weiguo; Kozakae, Mika; Watarai, Atsuko; Nakamura, Nobuhisa; Yasuda, Yutaka; Nakashima, Eirtaro; Naruse, Keiko; Watabe, Kazuhiko; Kato, Koichi; Oiso, Yutaka; Hamada, Yoji . E-mail: yhama@med.nagoya-u.ac.jp

    2007-05-25

    Methylglyoxal (MG) is involved in the pathogenesis of diabetic complications via the formation of advanced glycation end products (AGEs) and reactive oxygen species (ROS). To clarify whether the antidiabetic drug metformin prevents Schwann cell damage induced by MG, we cultured mouse Schwann cells in the presence of MG and metformin. Cell apoptosis was evaluated using Hoechst 33342 nuclear staining, caspase-3 activity, and c-Jun-N-terminal kinase (JNK) phosphorylation. Intracellular ROS formation was determined by flow cytometry, and AMP-activated kinase (AMPK) phosphorylation was also examined. MG treatment resulted in blunted cell proliferation, an increase in the number of apoptotic cells, and the activation of caspase-3 and JNK along with enhanced intracellular ROS formation. All of these changes were significantly inhibited by metformin. No significant activation of AMPK by MG or metformin was observed. Taken together, metformin likely prevents MG-induced apoptotic signals in mouse Schwann cells by inhibiting the formation of AGEs and ROS.

  9. Molecular mechanisms of nutlin-induced apoptosis in multiple myeloma

    PubMed Central

    Saha, Manujendra N; Jiang, Hua

    2010-01-01

    Multiple myeloma (MM) is an incurable plasma cell malignancy in which p53 is rarely mutated. Thus, activation of the p53 pathway by a small molecule inhibitor of the p53-MDM2 interaction, nutlin, in MM cells retaining wild type p53 is an attractive therapeutic strategy. Recently we reported that nutlin plus velcade (a proteasome inhibitor) displayed a synergistic response in MM. However, the mechanism of the p53-mediated apoptosis in MM has not been fully understood. Our data show that nutlin-induced apoptosis correlated with reduction in cell viability, upregulation of p53, p21 and MDM2 protein levels with a simultaneous increase in pro-apoptotic targets PUMA, Bax and Bak and downregulation of anti-apoptotic targets Bcl2 and survivin and activation of caspase in MM cells harboring wild type p53. Nutlin-induced apoptosis was inhibited when activation of caspase was blocked by the caspase inhibitor. Nutlin caused mitochondrial translocation of p53 where it binds with Bcl2, leading to cytochrome C release. Moreover, blocking the transcriptional arm of p53 by the p53-specific transcriptional inhibitor, pifithrin-α, not only inhibited nutlin-induced upregulation of p53-transcriptional targets but also augmented apoptosis in MM cells, suggesting an association of transcription-independent pathway of apoptosis. However, inhibitor of mitochondrial translocation of p53, PFT-µ, did not prevent nutlin-induced apoptosis, suggesting that the p53 transcription-dependent pathway was also operational in nutlin-induced apoptosis in MM. Our study provides the evidence that nutlin-induced apoptosis in MM cells is mediated by transcription-dependent and -independent pathways and supports further clinical evaluation of nutlin as a novel therapeutic agent in MM. PMID:20595817

  10. Geniposide prevents rotenone-induced apoptosis in primary cultured neurons

    PubMed Central

    Li, Lin; Zhao, Juan; Liu, Ke; Li, Guang-lai; Han, Yan-qing; Liu, Yue-ze

    2015-01-01

    Geniposide, a monomer extracted from gardenia and widely used in Chinese medicine, is a novel agonist at the glucagon-like peptide-1 receptor. This receptor is involved in neuroprotection. In the present study, we sought to identify an anti-apoptotic mechanism for the treatment of neurodegenerative diseases. Primary cultured neurons were treated with different concentrations of rotenone for 48 hours. Morphological observation, cell counting kit-8 assay, lactate dehydrogenase detection and western blot assay demonstrated that 0.5 nM rotenone increased lactate dehydrogenase release, decreased the expression of procaspase-3 and Bcl-2, and increased cleaved caspase-3 expression in normal neurons. All these effects were prevented by geniposide. Our results indicate that geniposide diminished rotenone-induced injury in primary neurons by suppressing apoptosis. This may be one of the molecular mechanisms underlying the efficacy of geniposide in the treatment of neurodegenerative diseases. PMID:26692859

  11. Avenanthramides Prevent Osteoblast and Osteocyte Apoptosis and Induce Osteoclast Apoptosis in Vitro in an Nrf2-Independent Manner.

    PubMed

    Pellegrini, Gretel G; Morales, Cynthya C; Wallace, Taylor C; Plotkin, Lilian I; Bellido, Teresita

    2016-01-01

    Oats contain unique bioactive compounds known as avenanthramides (AVAs) with antioxidant properties. AVAs might enhance the endogenous antioxidant cellular response by activation of the transcription factor Nrf2. Accumulation of reactive oxygen species plays a critical role in many chronic and degenerative diseases, including osteoporosis. In this disease, there is an imbalance between bone formation by osteoblasts and bone resorption by osteoclasts, which is accompanied by increased osteoblast/osteocyte apoptosis and decreased osteoclast apoptosis. We investigated the ability of the synthethic AVAs 2c, 2f and 2p, to 1-regulate gene expression in bone cells, 2-affect the viability of osteoblasts, osteocytes and osteoclasts, and the generation of osteoclasts from their precursors, and 3-examine the potential involvement of the transcription factor Nrf2 in these actions. All doses of AVA 2c and 1 and 5 µM dose of 2p up-regulated collagen 1A expression. Lower doses of AVAs up-regulated OPG (osteoprotegerin) in OB-6 osteoblastic cells, whereas 100 μM dose of 2f and all concentrations of 2c down-regulated RANKL gene expression in MLO-Y4 osteocytic cells. AVAs did not affect apoptosis of OB-6 osteoblastic cells or MLO-Y4 osteocytic cells; however, they prevented apoptosis induced by the DNA topoisomerase inhibitor etoposide, the glucocorticoid dexamethasone, and hydrogen peroxide. AVAs prevented apoptosis of both wild type (WT) and Nrf2 Knockout (KO) osteoblasts, demonstrating that AVAs-induced survival does not require Nrf2 expression. Further, KO osteoclast precursors produced more mature osteoclasts than WT; and KO cultures exhibited less apoptotic osteoclasts than WT cultures. Although AVAs did not affect WT osteoclasts, AVA 2p reversed the low apoptosis of KO osteoclasts. These in vitro results demonstrate that AVAs regulate, in part, the function of osteoblasts and osteocytes and prevent osteoblast/osteocyte apoptosis and increase osteoclast apoptosis; further

  12. Avenanthramides Prevent Osteoblast and Osteocyte Apoptosis and Induce Osteoclast Apoptosis in Vitro in an Nrf2-Independent Manner

    PubMed Central

    Pellegrini, Gretel G.; Morales, Cynthya C.; Wallace, Taylor C.; Plotkin, Lilian I.; Bellido, Teresita

    2016-01-01

    Oats contain unique bioactive compounds known as avenanthramides (AVAs) with antioxidant properties. AVAs might enhance the endogenous antioxidant cellular response by activation of the transcription factor Nrf2. Accumulation of reactive oxygen species plays a critical role in many chronic and degenerative diseases, including osteoporosis. In this disease, there is an imbalance between bone formation by osteoblasts and bone resorption by osteoclasts, which is accompanied by increased osteoblast/osteocyte apoptosis and decreased osteoclast apoptosis. We investigated the ability of the synthethic AVAs 2c, 2f and 2p, to 1-regulate gene expression in bone cells, 2-affect the viability of osteoblasts, osteocytes and osteoclasts, and the generation of osteoclasts from their precursors, and 3-examine the potential involvement of the transcription factor Nrf2 in these actions. All doses of AVA 2c and 1 and 5 µM dose of 2p up-regulated collagen 1A expression. Lower doses of AVAs up-regulated OPG (osteoprotegerin) in OB-6 osteoblastic cells, whereas 100 μM dose of 2f and all concentrations of 2c down-regulated RANKL gene expression in MLO-Y4 osteocytic cells. AVAs did not affect apoptosis of OB-6 osteoblastic cells or MLO-Y4 osteocytic cells; however, they prevented apoptosis induced by the DNA topoisomerase inhibitor etoposide, the glucocorticoid dexamethasone, and hydrogen peroxide. AVAs prevented apoptosis of both wild type (WT) and Nrf2 Knockout (KO) osteoblasts, demonstrating that AVAs-induced survival does not require Nrf2 expression. Further, KO osteoclast precursors produced more mature osteoclasts than WT; and KO cultures exhibited less apoptotic osteoclasts than WT cultures. Although AVAs did not affect WT osteoclasts, AVA 2p reversed the low apoptosis of KO osteoclasts. These in vitro results demonstrate that AVAs regulate, in part, the function of osteoblasts and osteocytes and prevent osteoblast/osteocyte apoptosis and increase osteoclast apoptosis; further

  13. Acetaminophen prevents oxidative burst and delays apoptosis in human neutrophils.

    PubMed

    Freitas, Marisa; Costa, Vera M; Ribeiro, Daniela; Couto, Diana; Porto, Graça; Carvalho, Félix; Fernandes, Eduarda

    2013-05-23

    Acetaminophen is a frequently prescribed over-the-counter drug to reduce fever and pain in the event of inflammatory process. As neutrophils are relevant cells in inflammatory processes, the putative interaction of acetaminophen with these cells, if present, would be of paramount importance. The present study was undertaken to evaluate the effect of acetaminophen in human neutrophils' oxidative burst and lifespan in vitro. The obtained results demonstrate that acetaminophen efficiently modulates neutrophils' oxidative burst in phorbol myristate acetate-activated neutrophils, in a concentration-dependent manner, at in vivo relevant concentrations. It was clearly demonstrated that acetaminophen is a strong scavenger of HOCl and H2O2, which probably contributed to the effect observed in neutrophils. Acetaminophen also induced the depletion of glutathione in stimulated neutrophils, suggesting its transformation into a reactive intermediate. Obtained results further revealed that acetaminophen affects programmed cell death of human neutrophils, resulting in a delay of previously stimulated neutrophils-mediated apoptosis. Overall, our data suggested that acetaminophen has considerable potential to be included in anti-inflammatory therapeutic strategies, by preventing biological damage induced by an excessive production of reactive species generated in activated neutrophils and by extending the lifespan of neutrophils, favoring the elimination of pathogens, thus contributing to tissue healing and resolution of inflammation. PMID:23518321

  14. Iron dysregulation combined with aging prevents sepsis-induced apoptosis

    PubMed Central

    Javadi, Pardis; Buchman, Timothy G.; Stromberg, Paul E.; Turnbull, Isaiah R.; Vyas, Dinesh; Hotchkiss, Richard S.; Karl, Irene E.; Coopersmith, Craig M.

    2005-01-01

    Background Sepsis, iron loading and aging cause independent increases in gut epithelial and splenic apoptosis. It is unknown how their combination will affect apoptosis and systemic cytokine levels. Methods Hfe−/− mice (a murine homolog of hemochromatosis) abnormally accumulate iron in their tissues. Aged (24–26 months) or mature (16–18 months) Hfe−/− mice and wild type (WT) littermates were subjected to cecal ligation and puncture (CLP) or sham laparotomy. Intestine, spleen, and blood were harvested 24 hours later and assessed for apoptosis and cytokine levels. Results Gut epithelial and splenic apoptosis were low in both aged septic and sham Hfe−/− mice, regardless of the amount of iron in their diet. Mature septic WT mice had increased apoptosis compared to age-matched sham WT mice. Mature septic Hfe−/− mice had similar levels of intestinal cell death to age-matched septic WT mice but higher levels of splenic apoptosis. Apoptosis was significantly lower in septic aged Hfe−/− mice than septic mature Hfe−/− animals. Interleukin-6 was elevated in septic aged Hfe−/− mice compared to sham mice. Conclusions Although sepsis, chronic iron dysregulation, and aging each increase gut and splenic apoptosis, their combination yields cell death levels similar to sham animals despite the fact that aged Hfe−/− mice are able to mount an inflammatory response following CLP and mature Hfe−/− mice have elevated sepsis-induced apoptosis. Combining sepsis with two risk factors that ordinarily increase cell death and increase mortality in CLP yields an apoptotic response that could not have been predicted based upon each element in isolation. PMID:15921699

  15. Preventive effects of bicarbonate on cerivastatin-induced apoptosis.

    PubMed

    Kobayashi, Masaki; Kaido, Fumie; Kagawa, Toshiki; Itagaki, Shirou; Hirano, Takeshi; Iseki, Ken

    2007-08-16

    Although HMG-CoA reductase inhibitors such as statins are the most widely used cholesterol-lowering agents, there is a risk of myopathy or rhabdmyolysis occurring in patients taking these drugs. It has been reported that a number of lipophilic statins cause apoptosis in various cells, but it is still not clear whether intracellular acidification is involved in statin-induced apoptosis. There have been few studies aimed at identifying compounds that suppress statin-induced myotoxicity. In the present study, we examined the relationship between cerivastatin-induced apoptosis and intracellular acidification and the effect of bicarbonate on cerivastatin-induced apoptosis using an RD cell line as a model of in vitro skeletal muscle. Cerivastatin reduced the number of viable cells and caused dramatic morphological changes and DNA fragmentation in a concentration-dependent manner. Moreover, cerivastatin-induced apoptosis was associated with intracellular acidification and caspase-9 and -3/7 activation. On the other hand, bicarbonate suppressed cerivastatin-induced pH alteration, caspase activation, morphological change and reduction of cell viability. Accordingly, bicarbonate suppressed statin-induced apoptosis. The strategy to combine statins with bicarbonate can lead to reduction in the chance of the severe adverse events including myopathy or rhabdmyolysis. PMID:17553641

  16. Naringin prevents ovariectomy-induced osteoporosis and promotes osteoclasts apoptosis through the mitochondria-mediated apoptosis pathway

    SciTech Connect

    Li, Fengbo; Sun, Xiaolei; Ma, Jianxiong; Ma, Xinlong; Zhao, Bin; Zhang, Yang; Tian, Peng; Li, Yanjun; Han, Zhe

    2014-09-26

    Highlights: • Naringin possesses many pharmacological activities, promotes the proliferation of osteoblast. • Undecalcified histological obtain dynamic parameters of callus formation and remodeling. • Naringin regulate osteoclast apoptosis by mitochondrial pathway. - Abstract: Naringin, the primary active compound of the traditional Chinese medicine Rhizoma drynariae, possesses many pharmacological activities. The present study is an effort to explore the anti-osteoporosis potential of naringin in vivo and in vitro. In vivo, we used ovariectomized rats to clarify the mechanisms by which naringin anti-osteoporosis. In vitro, we used osteoclasts to investigate naringin promotes osteoclasts apoptosis. Naringin was effective at enhancing BMD, trabecular thickness, bone mineralization, and mechanical strength in a dose-dependent manner. The result of RT-PCR analysis revealed that naringin down-regulated the mRNA expression levels of BCL-2 and up-regulated BAX, caspase-3 and cytochrome C. In addition, naringin significantly reduced the bone resorption area in vitro. These findings suggest that naringin promotes the apoptosis of osteoclasts by regulating the activity of the mitochondrial apoptosis pathway and prevents OVX-induced osteoporosis in rats.

  17. Preventing retinal apoptosis--is there a common therapeutic theme?

    PubMed

    Doonan, Francesca; Groeger, Gillian; Cotter, Thomas G

    2012-07-01

    There is an urgent need for therapies for retinal diseases; retinitis pigmentosa sufferers have no treatment options available and those targeted at other retinopathies have shown limited effectiveness. The process of programmed cell death or apoptosis although complex, remains a possible target for the treatment of retinal diseases. Having identified apoptosis in the vertebrate retina in populations of immature neurons as an essential part of development it was proposed that re-activation of these developmental cell death pathways might provide insight into the death mechanisms operating in retinal diseases. However, the discovery that numerous factors initiate and mediate the apoptotic cascade in mature photoreceptors has resulted in a relatively untargeted approach to examining and arresting apoptosis in the retina. In the last 5 years, mouse models have been treated with a diverse range of drugs or factors including anti-oxidants, growth factors, steroid hormones, calcium/calpain inhibitors and tetracycline antibiotics. Therefore to draw a unifying theme from these broad research areas is challenging. However, this review focusses on two targets which are currently under investigation, reactive oxygen species and mammalian target of rapamycin, drawing together the common themes of these research areas. PMID:22366479

  18. Molecular mechanisms of nutlin-induced apoptosis in multiple myeloma: evidence for p53-transcription-dependent and -independent pathways.

    PubMed

    Saha, Manujendra N; Jiang, Hua; Chang, Hong

    2010-09-15

    Multiple myeloma (MM) is an incurable plasma cell malignancy in which p53 is rarely mutated. Thus, activation of the p53 pathway by a small molecule inhibitor of the p53-MDM2 interaction, nutlin, in MM cells retaining wild type p53 is an attractive therapeutic strategy. Recently we reported that nutlin plus velcade (a proteasome inhibitor) displayed a synergistic response in MM. However, the mechanism of the p53-mediated apoptosis in MM has not been fully understood. Our data show that nutlin-induced apoptosis correlated with reduction in cell viability, upregulation of p53, p21 and MDM2 protein levels with a simultaneous increase in pro-apoptotic targets PUMA, Bax and Bak and downregulation of anti-apoptotic targets Bcl2 and survivin and activation of caspase in MM cells harboring wild type p53. Nutlin-induced apoptosis was inhibited when activation of caspase was blocked by the caspase inhibitor. Nutlin caused mitochondrial translocation of p53 where it binds with Bcl2, leading to cytochrome C release. Moreover, blocking the transcriptional arm of p53 by the p53-specific transcriptional inhibitor, pifithrin-α, not only inhibited nutlin-induced upregulation of p53-transcriptional targets but also augmented apoptosis in MM cells, suggesting an association of transcription-independent pathway of apoptosis. However, inhibitor of mitochondrial translocation of p53, PFT-μ, did not prevent nutlin-induced apoptosis, suggesting that the p53 transcription-dependent pathway was also operational in nutlin-induced apoptosis in MM. Our study provides the evidence that nutlin-induced apoptosis in MM cells is mediated by transcription-dependent and -independent pathways and supports further clinical evaluation of nutlin as a novel therapeutic agent in MM. PMID:20595817

  19. Baicalin prevents Candida albicans infections via increasing its apoptosis rate

    SciTech Connect

    Yang, Shulong; Fu, Yingyuan Wu, Xiuzhen; Zhou, Zhixing; Xu, Jing; Zeng, Xiaoping; Kuang, Nanzhen; Zeng, Yurong

    2014-08-15

    Highlights: • Baicalin increases the ratio of the G0/G1 stages and C. albicans apoptosis. • Baicalin decreases the proliferation index of C. albicans. • Baicalin inhibits the biosynthesis of DNA, RNA and protein in C. albicans. • Baicalin depresses Succinate Dehydrogenase and Ca{sup 2+}–Mg{sup 2+} ATPase in C. albicans. • Baicalin increases the endocytic free Ca{sup 2+} concentration in C. albicans. - Abstract: Background: These experiments were employed to explore the mechanisms underlying baicalin action on Candida albicans. Methodology and principal findings: We detected the baicalin inhibition effects on three isotope-labeled precursors of {sup 3}H-UdR, {sup 3}H-TdR and {sup 3}H-leucine incorporation into C. albicans using the isotope incorporation technology. The activities of Succinate Dehydrogenase (SDH), cytochrome oxidase (CCO) and Ca{sup 2+}–Mg{sup 2+} ATPase, cytosolic Ca{sup 2+} concentration, the cell cycle and apoptosis, as well as the ultrastructure of C.albicans were also tested. We found that baicalin inhibited {sup 3}H-UdR, {sup 3}H-TdR and {sup 3}H-leucine incorporation into C.albicans (P < 0.005). The activities of the SDH and Ca{sup 2+}–Mg{sup 2+} ATPase of C.albicans in baicalin groups were lower than those in control group (P < 0.05). Ca{sup 2+} concentrations of C. albicans in baicalin groups were much higher than those in control group (P < 0.05). The ratio of C.albicans at the G0/G1 stage increased in baicalin groups in dose dependent manner (P < 0.01). There were a significant differences in the apoptosis rate of C.albicans between baicalin and control groups (P < 0.01). After 12–48 h incubation with baicalin (1 mg/ml), C. albicans shown to be markedly damaged under transmission electron micrographs. Innovation and significance: Baicalin can increase the apoptosis rate of C. albicans. These effects of Baicalin may involved in its inhibiting the activities of the SDH and Ca{sup 2+}–Mg{sup 2+} ATPase, increasing

  20. Neuronal gelsolin prevents apoptosis by enhancing actin depolymerization.

    PubMed

    Harms, Christoph; Bösel, Julian; Lautenschlager, Marion; Harms, Ulrike; Braun, Johann S; Hörtnagl, Heide; Dirnagl, Ulrich; Kwiatkowski, David J; Fink, Klaus; Endres, Matthias

    2004-01-01

    Gelsolin (gsn), an actin-severing protein, protects neurons from excitotoxic cell death via inactivation of membranous Ca(2+) channels. Its role during apoptotic cell death, however, has remained unclear. Using several models of neuronal cell death, we demonstrate that endogenous gelsolin has anti-apoptotic properties that correlate to its dynamic actions on the cytoskeleton. We show that neurons lacking gelsolin (gsn(-/-)) have enhanced apoptosis following exposure to staurosporine, thapsigargin, or the cholinergic toxin ethylcholine aziridinium (AF64A). AF64A-induced loss of mitochondrial membrane potential and activation of caspase-3 was specifically enhanced in gsn(-/-) neurons and could be reversed by pharmacological inhibition of mitochondrial permeability transition. Moreover, increased caspase-3 activation and cell death in AF64A-treated gsn(-/-) neurons were completely reversed by pharmacological depolymerization of actin filaments and further enhanced by their stabilization. In conclusion, actin remodeling by endogenous gelsolin or analogues protects neurons from apoptosis mediated by mitochondria and caspase-3. PMID:14962741

  1. Myc Prevents Apoptosis and Enhances Endoreduplication Induced by Paclitaxel

    PubMed Central

    Gatti, Giuliana; Maresca, Giovanna; Natoli, Manuela; Florenzano, Fulvio; Nicolin, Angelo; Felsani, Armando; D'Agnano, Igea

    2009-01-01

    Background The role of the MYC oncogene in the apoptotic pathways is not fully understood. MYC has been reported to protect cells from apoptosis activation but also to sensitize cells to apoptotic stimuli. We have previously demonstrated that the down-regulation of Myc protein activates apoptosis in melanoma cells and increases the susceptibility of cells to various antitumoral treatments. Beyond the well-known role in the G1→S transition, MYC is also involved in the G2-M cell cycle phases regulation. Methodology/Principal Findings In this study we have investigated how MYC could influence cell survival signalling during G2 and M phases. We used the microtubules damaging agent paclitaxel (PTX), to arrest the cells in the M phase, in a p53 mutated melanoma cell line with modulated Myc level and activity. An overexpression of Myc protein is able to increase endoreduplication favoring the survival of cells exposed to antimitotic poisoning. The PTX-induced endoreduplication is associated in Myc overexpressing cells with a reduced expression of MAD2, essential component of the molecular core of the spindle assembly checkpoint (SAC), indicating an impairment of this checkpoint. In addition, for the first time we have localized Myc protein at the spindle poles (centrosomes) during pro-metaphase in different cell lines. Conclusions The presence of Myc at the poles during the prometaphase could be necessary for the Myc-mediated attenuation of the SAC and the subsequent induction of endoreduplication. In addition, our data strongly suggest that the use of taxane in antitumor therapeutic strategies should be rationally based on the molecular profile of the individual tumor by specifically analyzing Myc expression levels. PMID:19421315

  2. Astragaloside IV prevents high glucose-induced podocyte apoptosis via downregulation of TRPC6.

    PubMed

    Yao, Xing-Mei; Liu, Yu-Jun; Wang, Yun-Man; Wang, Hao; Zhu, Bing-Bing; Liang, Yong-Ping; Yao, Wei-Guo; Yu, Hui; Wang, Nian-Song; Zhang, Xue-Mei; Peng, Wen

    2016-06-01

    Diabetic nephropathy (DN) is one of the most important causes of end‑stage renal disease. Astragaloside IV (AS-IV) is a saponin isolated from Astragalus membranaceus, which possesses various pharmacological activities. AS‑IV prevents podocyte apoptosis and ameliorates renal injury in DN; however, few studies have focused on its effects on ion channels. The transient receptor potential channel 6 (TRPC6) is an important Ca2+‑permeable ion channel in podocytes, which is involved in high glucose (HG)-induced podocyte apoptosis. The aim of the present study was to investigate whether AS‑IV prevented HG‑induced podocyte apoptosis via TRPC6. Cultured podocytes were pre‑treated with 10, 20 or 40 µM AS‑IV for 1 h prior to HG exposure for 24 h. Apoptosis, cell viability, expression of TRPC6, nuclear factor of activated T cells (NFAT2) and B‑cell lymphoma 2‑associated X protein (Bax), as well as the intracellular Ca2+ concentration were subsequently analyzed. The results indicated that HG induced podocyte apoptosis and upregulation of TRPC6, and increased intracellular Ca2+. Furthermore, enhanced NFAT2 and Bax expression was detected. Conversely, AS‑IV protected HG‑induced podocyte apoptosis, downregulated TRPC6 expression and suppressed intracellular Ca2+ in HG-stimulated podocytes. AS‑IV also suppressed NFAT2 and Bax expression. These results suggest that AS‑IV may prevent HG-induced podocyte apoptosis via downregulation of TRPC6, which is possibly mediated via the calcineurin/NFAT signaling pathway. PMID:27109610

  3. Novel Apoptosis Suppressor Apsup from the Baculovirus Lymantria dispar Multiple Nucleopolyhedrovirus Precludes Apoptosis by Preventing Proteolytic Processing of Initiator Caspase Dronc

    PubMed Central

    Yamada, Hayato; Kitaguchi, Koji; Hamajima, Rina; Kobayashi, Michihiro

    2013-01-01

    We previously identified a novel baculovirus-encoded apoptosis suppressor, Apsup, from the baculovirus Lymantria dispar multiple nucleopolyhedrovirus (LdMNPV). Apsup inhibits the apoptosis of L. dispar Ld652Y cells triggered by infection with p35-defective Autographa californica MNPV (vAcΔp35) and exposure to actinomycin D or UV light. Here, we examined the functional role of Apsup in apoptosis regulation in insect cells. Apsup prevented apoptosis and the proteolytic processing of L. dispar initiator caspase Dronc (Ld-Dronc) in Ld652Y cells triggered by overexpression of Ld-Dronc, LdMNPV inhibitor-of-apoptosis 3 (IAP3), or Hyphantria cunea MNPV IAP1. In vAcΔp35-infected apoptotic Ld652Y cells, Apsup restricted apoptosis induction and prevented processing of endogenous Ld-Dronc. Conversely, upon RNA interference (RNAi)-mediated silencing of apsup, LdMNPV-infected Ld652Y cells, which typically support high-titer virus replication, underwent apoptosis, accompanied by the processing of endogenous Ld-Dronc. Furthermore, endogenous Ld-Dronc coimmunoprecipitated with transiently expressed Apsup, indicating that Apsup physically interacts with Ld-Dronc. Apsup prevented the apoptosis of Sf9 cells triggered by vAcΔp35 infection but did not inhibit apoptosis or activation of caspase-3-like protease in vAcΔp35-infected Drosophila melanogaster S2 cells. Apsup also inhibited the proteolytic processing of L. dispar effector caspase Ld-caspase-1 in the transient expression assay but did not physically interact with Ld-caspase-1. These results demonstrate that Apsup inhibits apoptosis in Ld652Y cells by preventing the proteolytic processing of Ld-Dronc. Together with our previous findings showing that Apsup prevents the processing of both overexpressed Ld-Dronc and Bombyx mori Dronc, these results also demonstrate that Apsup functions as an effective apoptotic suppressor in various lepidopteran, but not dipteran, insect cells. PMID:24067961

  4. Akt inhibition improves irinotecan treatment and prevents cell emergence by switching the senescence response to apoptosis

    PubMed Central

    Vétillard, Alexandra; Jonchère, Barbara; Moreau, Marie; Toutain, Bertrand; Henry, Cécile; Fontanel, Simon; Bernard, Anne-Charlotte; Campone, Mario; Guette, Catherine; Coqueret, Olivier

    2015-01-01

    Activated in response to chemotherapy, senescence is a tumor suppressive mechanism that induces a permanent loss of proliferation. However, in response to treatment, it is not really known how cells can escape senescence and how irreversible or incomplete this pathway is. We have recently described that cells that escape senescence are more transformed than non-treated parental cells, they resist anoikis and rely on Mcl-1. In this study, we further characterize this emergence in response to irinotecan, a first line treatment used in colorectal cancer. Our results indicate that Akt was activated as a feedback pathway during the early step of senescence. The inhibition of the kinase prevented cell emergence and improved treatment efficacy, both in vitro and in vivo. This improvement was correlated with senescence inhibition, p21waf1 downregulation and a concomitant activation of apoptosis due to Noxa upregulation and Mcl-1 inactivation. The inactivation of Noxa prevented apoptosis and increased the number of emergent cells. Using either RNA interference or p21waf1-deficient cells, we further confirmed that an intact p53-p21-senescence pathway favored cell emergence and that its downregulation improved treatment efficacy through apoptosis induction. Therefore, although senescence is an efficient suppressive mechanism, it also generates more aggressive cells as a consequence of apoptosis inhibition. We therefore propose that senescence-inducing therapies should be used sequentially with drugs favoring cell death such as Akt inhibitors. This should reduce cell emergence and tumor relapse through a combined induction of senescence and apoptosis. PMID:26485768

  5. Isoniazid prevents Nrf2 translocation by inhibiting ERK1 phosphorylation and induces oxidative stress and apoptosis

    PubMed Central

    Verma, Ajeet Kumar; Yadav, Arti; Dewangan, Jayant; Singh, Sarvendra Vikram; Mishra, Manisha; Singh, Pradhyumna Kumar; Rath, Srikanta Kumar

    2015-01-01

    Isoniazid is used either alone or in combination with other drugs for the treatment of tuberculosis. It is also used for the prevention of tuberculosis. Chronic treatment of Isoniazid may cause severe liver damage leading to acute liver failure. The mechanism through which Isoniazid causes liver damage is investigated. Isoniazid treatment generates reactive oxygen species and induces apoptosis in Hep3B cells. It induces antioxidative and apoptotic genes leading to increase in mRNA expression and protein levels in Hep3B cells. Whole genome expression analysis of Hep3B cells treated with Isoniazid has resulted in differential expression of various genes playing prime role in regulation of apoptotic, antioxidative, DNA damage, cell signaling, cell proliferation and differentiation pathways. Isoniazid increased cytosolic Nrf2 protein level while decreased nuclear Nrf2 protein level. It also decreased ERK1 phosphorylation and treatment of Hep3B cells with ERK inhibitor followed by Isoniazid resulting in increased apoptosis in these cells. Two dimensional gel electrophoresis results have also shown differential expression of various protein species including heat shock proteins, proteins playing important role in oxidative stress, DNA damage, apoptosis, cell proliferation and differentiation. Results suggest that Isoniazid induces apoptosis through oxidative stress and also prevents Nrf2 translocation into the nucleus by reducing ERK1 phosphorylation thus preventing cytoprotective effect. PMID:26202867

  6. Apoptosis by dietary agents for prevention and treatment of prostate cancer

    PubMed Central

    Khan, Naghma; Adhami, Vaqar Mustafa; Mukhtar, Hasan

    2010-01-01

    Accumulating data clearly indicate that induction of apoptosis is an important event for chemoprevention of cancer by naturally occurring dietary agents. In mammalian cells, apoptosis has been divided into two major pathways: the extrinsic pathway, activated by pro-apoptotic receptor signals at the cellular surface; and the intrinsic pathway, which involves the disruption of mitochondrial membrane integrity. This process is strictly controlled in response to integrity of pro-death signaling and plays critical roles in development, maintenance of homeostasis, and host defense in multicellular organisms. For chemoprevention studies, prostate cancer (PCa) represents an ideal disease due to its long latency, its high incidence, tumor marker availability, and identifiable preneoplastic lesions and risk groups. In this article, we highlight the studies of various apoptosis-inducing dietary compounds for prevention of PCa in vitro in cell culture, in preclinical studies in animals, and in human clinical trials. PMID:19926708

  7. Apoptosis by dietary agents for prevention and treatment of prostate cancer.

    PubMed

    Khan, Naghma; Adhami, Vaqar Mustafa; Mukhtar, Hasan

    2010-03-01

    Accumulating data clearly indicate that induction of apoptosis is an important event for chemoprevention of cancer by naturally occurring dietary agents. In mammalian cells, apoptosis has been divided into two major pathways: the extrinsic pathway, activated by pro-apoptotic receptor signals at the cellular surface; and the intrinsic pathway, which involves the disruption of mitochondrial membrane integrity. This process is strictly controlled in response to integrity of pro-death signaling and plays critical roles in development, maintenance of homeostasis, and host defense in multicellular organisms. For chemoprevention studies, prostate cancer (PCa) represents an ideal disease due to its long latency, its high incidence, tumor marker availability, and identifiable preneoplastic lesions and risk groups. In this article, we highlight the studies of various apoptosis-inducing dietary compounds for prevention of PCa in vitro in cell culture, in preclinical studies in animals, and in human clinical trials. PMID:19926708

  8. IL-15 Prevents Apoptosis, Reverses Innate and Adaptive Immune Dysfunction, and Improves Survival in Sepsis

    PubMed Central

    Inoue, Shigeaki; Unsinger, Jacqueline; Davis, Christopher G.; Muenzer, Jared T.; Ferguson, Thomas A.; Chang, Katherine; Osborne, Dale F.; Clark, Andrew T.; Coopersmith, Craig M.; McDunn, Jonathan E.; Hotchkiss, Richard S.

    2010-01-01

    L-15 is a pluripotent antiapoptotic cytokine that signals to cells of both the innate and adaptive immune system and is regarded as a highly promising immunomodulatory agent in cancer therapy. Sepsis is a lethal condition in which apoptosis-induced depletion of immune cells and subsequent immunosuppression are thought to contribute to morbidity and mortality. This study tested the ability of IL-15 to block apoptosis, prevent immunosuppression, and improve survival in sepsis. Mice were made septic using cecal ligation and puncture or Pseudomonas aeruginosa pneumonia. The experiments comprised a 2×2 full factorial design with surgical sepsis versus sham and IL-15 versus vehicle. In addition to survival studies, splenic cellularity, canonical markers of activation and proliferation, intracellular pro- and antiapoptotic Bcl-2 family protein expression, and markers of immune cell apoptosis were evaluated by flow cytometry. Cytokine production was examined both in plasma of treated mice and splenocytes that were stimulated ex vivo. IL-15 blocked sepsis-induced apoptosis of NK cells, dendritic cells, and CD8 T cells. IL-15 also decreased sepsis-induced gut epithelial apoptosis. IL-15 therapy increased the abundance of antiapoptotic Bcl-2 while decreasing proapoptotic Bim and PUMA. IL-15 increased both circulating IFN-γ, as well as the percentage of NK cells that produced IFN-γ. Finally, IL-15 increased survival in both cecal ligation and puncture and P. aeruginosa pneumonia. In conclusion, IL-15 prevents two immunopathologic hallmarks of sepsis, namely, apoptosis and immunosuppression, and improves survival in two different models of sepsis. IL-15 represents a potentially novel therapy of this highly lethal disorder. PMID:20026737

  9. IL-15 prevents apoptosis, reverses innate and adaptive immune dysfunction, and improves survival in sepsis.

    PubMed

    Inoue, Shigeaki; Unsinger, Jacqueline; Davis, Christopher G; Muenzer, Jared T; Ferguson, Thomas A; Chang, Katherine; Osborne, Dale F; Clark, Andrew T; Coopersmith, Craig M; McDunn, Jonathan E; Hotchkiss, Richard S

    2010-02-01

    IL-15 is a pluripotent antiapoptotic cytokine that signals to cells of both the innate and adaptive immune system and is regarded as a highly promising immunomodulatory agent in cancer therapy. Sepsis is a lethal condition in which apoptosis-induced depletion of immune cells and subsequent immunosuppression are thought to contribute to morbidity and mortality. This study tested the ability of IL-15 to block apoptosis, prevent immunosuppression, and improve survival in sepsis. Mice were made septic using cecal ligation and puncture or Pseudomonas aeruginosa pneumonia. The experiments comprised a 2 x 2 full factorial design with surgical sepsis versus sham and IL-15 versus vehicle. In addition to survival studies, splenic cellularity, canonical markers of activation and proliferation, intracellular pro- and antiapoptotic Bcl-2 family protein expression, and markers of immune cell apoptosis were evaluated by flow cytometry. Cytokine production was examined both in plasma of treated mice and splenocytes that were stimulated ex vivo. IL-15 blocked sepsis-induced apoptosis of NK cells, dendritic cells, and CD8 T cells. IL-15 also decreased sepsis-induced gut epithelial apoptosis. IL-15 therapy increased the abundance of antiapoptotic Bcl-2 while decreasing proapoptotic Bim and PUMA. IL-15 increased both circulating IFN-gamma, as well as the percentage of NK cells that produced IFN-gamma. Finally, IL-15 increased survival in both cecal ligation and puncture and P. aeruginosa pneumonia. In conclusion, IL-15 prevents two immunopathologic hallmarks of sepsis, namely, apoptosis and immunosuppression, and improves survival in two different models of sepsis. IL-15 represents a potentially novel therapy of this highly lethal disorder. PMID:20026737

  10. cAMP prevents TNF-induced apoptosis through inhibiting DISC complex formation in rat hepatocytes

    PubMed Central

    Bhattacharjee, Rajesh; Xiang, Wenpei; Wang, Yinna; Zhang, Xiaoying; Billiar, Timothy R.

    2013-01-01

    Tumor necrosis factor α (TNF) is a pleiotropic proinflammatory cytokine that plays a role in immunity and the control of cell proliferation, cell differentiation, and apoptosis. The pleiotropic nature of TNF is due to the formation of different signaling complexes upon the binding of TNF to its receptor, TNF receptor type 1 (TNFR1). TNF induces apoptosis in various mammalian cells when the cells are co-treated with a transcription inhibitor like actinomycin D (ActD). When TNFR1 is activated, it recruits an adaptor protein, TNF receptor-associated protein with death domain (TRADD), through its cytoplasmic death effector domain (DED). TRADD, in turn, recruits other signaling proteins, including TNF receptor-associated protein 2 (TRAF2) and receptor-associated protein kinase (RIPK) 1, to form a complex. Subsequently, this complex combines with FADD and procaspase-8, converts into a death-inducing signaling complex (DISC) to induce apoptosis. Cyclic AMP (cAMP) is a second messenger that regulates various cellular processes such as cell proliferation, gene expression, and apoptosis. cAMP analogues are reported to act as anti-apoptotic agents in various cell types, including hepatocytes. We found that a cAMP analogue, dibutyryl cAMP (db-cAMP), inhibits TNF + ActD-induced apoptosis in rat hepatocytes. The protein kinase A (PKA) inhibitor KT-5720 reverses this inhibitory effect of cAMP on apoptosis. Cytoprotection by cAMP involves down-regulation of various apoptotic signal regulators like TRADD and FADD and inhibition of caspase-8 and caspase-3 cleavage. We also found that cAMP exerts its affect at the proximal level of TNF signaling by inhibiting the formation of the DISC complex upon the binding of TNF to TNFR1. In conclusion, our study shows that cAMP prevents TNF + ActD-induced apoptosis in rat hepatocytes by inhibiting DISC complex formation. PMID:22634003

  11. Podocyte apoptosis is prevented by blocking the Toll-like receptor pathway

    PubMed Central

    Saurus, P; Kuusela, S; Lehtonen, E; Hyvönen, M E; Ristola, M; Fogarty, C L; Tienari, J; Lassenius, M I; Forsblom, C; Lehto, M; Saleem, M A; Groop, P-H; Holthöfer, H; Lehtonen, S

    2015-01-01

    High serum lipopolysaccharide (LPS) activity in normoalbuminuric patients with type 1 diabetes (T1D) predicts the progression of diabetic nephropathy (DN), but the mechanisms behind this remain unclear. We observed that treatment of cultured human podocytes with sera from normoalbuminuric T1D patients with high LPS activity downregulated 3-phosphoinositide-dependent kinase-1 (PDK1), an activator of the Akt cell survival pathway, and induced apoptosis. Knockdown of PDK1 in cultured human podocytes inhibited antiapoptotic Akt pathway, stimulated proapoptotic p38 MAPK pathway, and increased apoptosis demonstrating an antiapoptotic role for PDK1 in podocytes. Interestingly, PDK1 was downregulated in the glomeruli of diabetic rats and patients with type 2 diabetes before the onset of proteinuria, further suggesting that reduced expression of PDK1 associates with podocyte injury and development of DN. Treatment of podocytes in vitro and mice in vivo with LPS reduced PDK1 expression and induced apoptosis, which were prevented by inhibiting the Toll-like receptor (TLR) signaling pathway with the immunomodulatory agent GIT27. Our data show that LPS downregulates the cell survival factor PDK1 and induces podocyte apoptosis, and that blocking the TLR pathway with GIT27 may provide a non-nephrotoxic means to prevent the progression of DN. PMID:25950482

  12. Vanadate prevents glucocorticoid-induced apoptosis of osteoblasts in vitro and osteocytes in vivo

    PubMed Central

    Conradie, M M; de Wet, H; Kotze, D D R; Burrin, J M; Hough, F S; Hulley, P A

    2007-01-01

    Skeletal mass is maintained by a balance between formation and resorption, cell proliferation and apoptosis. In vitro, glucocorticoids (GCs) decrease extracellular signal-regulated kinases (ERK) activation by mitogens, thus inhibiting osteoblast proliferation. Both ERK activity and proliferation are restored by co-treatment with the protein tyrosine phosphatase inhibitor, vanadate. Since ERK signalling may also be anti-apoptotic, we explored the effects of vanadate on GC-induced apoptosis in vitro and in vivo. Apoptosis in MBA-15.4 pre-osteoblasts increased from 6 h and remained up to eightfold higher through 6 days of 10−6 M dexamethasone (Dex) treatment. Co-incubation with 10−7 M vanadate markedly reduced apoptosis at all time points. Vanadate also prevented GC-induced poly-ADP-ribose polymerase cleavage. We assessed the transcriptional profiles of seven anti-apoptotic proteins (Bcl-2, Bcl-XL, inhibitors of apoptosis protein-1 (IAP-1), IAP-2, X-linked IAP (XIAP), Fas-associated death-domain-like IL-1β-converting enzyme-inhibitory protein (FLIPLong) and FLIPShort) in osteoblasts subjected to various stimuli using real-time quantitative PCR. Although these anti-apoptotic genes responded to different mitogenic conditions, Dex failed to repress their expression, and in fact significantly up-regulated Bcl-XL, IAP-2 and XIAP. Dex may therefore induce apoptosis by up-regulating pro-apoptotic gene expression. We have previously demonstrated that rats treated with GC develop low formation osteoporosis (bone histomorphometry and DEXA) and skeletal fragility (breaking strength) that were largely prevented by co-treatment with vanadate. We report here that vertebrae from rats treated with 3·5 mg/kg per day methylprednisolone for 9 weeks showed increased incidence of terminal deoxynucleotidyl transferase-mediated biotin-dUTP nick end-labelling-positive apoptotic osteocytes, which was reduced by vanadate co-treatment. We conclude that vanadate prevents GC

  13. Long story short: p53 mediates innate immunity.

    PubMed

    Miciak, Jessica; Bunz, Fred

    2016-04-01

    The story of p53 and how we came to understand it is punctuated by fundamental insights into the essence of cancer. In the decades since its discovery, p53 has been shown to be centrally involved in most, if not all, of the cellular processes that maintain tissue homeostasis. Extensive functional analyses of p53 and its tumor-associated mutants have illuminated many of the common defects shared by most cancer cells. As the central character in a tale that continues to unfold, p53 has become increasingly familiar and yet remains surprisingly inscrutable. New relationships periodically come to light, and surprising, novel activities continue to emerge, thereby revealing new dimensions and aspects of its function. What lies at the very core of this complex protagonist? What is its prime motivation? As every avid reader knows, the elements of character are profoundly shaped by adversity--originating from within and without. And so it is with p53. This review will briefly recap the coordinated responses of p53 to viral infection, and outline a hypothetical model that would explain how an abundance of seemingly unrelated phenotypic attributes may in the end reflect a singular function. All stories eventually draw to a conclusion. This epic tale may eventually leave us with the realization that p53, most simply described, is a protein that evolved to mediate immune surveillance. PMID:26951863

  14. Skeletal muscle secreted factors prevent glucocorticoid-induced osteocyte apoptosis through activation of β-catenin.

    PubMed

    Jähn, K; Lara-Castillo, N; Brotto, L; Mo, C L; Johnson, M L; Brotto, M; Bonewald, L F

    2012-01-01

    It is a widely held belief that the sole effect of muscle on bone is through mechanical loading. However, as the two tissues are intimately associated, we hypothesized that muscle myokines may have positive effects on bone. We found that factors produced by muscle will protect osteocytes from undergoing cell death induced by dexamethasone (dex), a glucocorticoid known to induce osteocyte apoptosis thereby compromising their capacity to regulate bone remodeling. Both the trypan blue exclusion assay for cell death and nuclear fragmentation assay for apoptosis were used. MLO-Y4 osteocytes, primary osteocytes, and MC3T3 osteoblastic cells were protected against dex-induced apoptosis by C2C12 myotube conditioned media (MT-CM) or by CM from ex vivo electrically stimulated, intact extensor digitorum longus (EDL) or soleus muscle derived from 4 month-old mice. C2C12 MT-CM, but not undifferentiated myoblast CM prevented dex-induced cell apoptosis and was potent down to 0.1 % CM. The CM from EDL muscle electrically stimulated tetanically at 80 Hz was more potent (10 fold) in prevention of dex-induced osteocyte death than CM from soleus muscle stimulated at the same frequency or CM from EDL stimulated at 1 Hz. This suggests that electrical stimulation increases production of factors that preserve osteocyte viability and that type II fibers are greater producers than type I fibers. The muscle factor(s) appears to protect osteocytes from cell death through activation of the Wnt/β-catenin pathway, as MT-CM induces β-catenin nuclear translocation and β-catenin siRNA abrogated the positive effects of MT-CM on dex-induced apoptosis. We conclude that muscle cells naturally secrete factor(s) that preserve osteocyte viability. PMID:22972510

  15. Vascular Endothelial Growth Factor Prevents Apoptosis and Preserves Contractile Function in Hypertrophied Infant Heart

    PubMed Central

    Friehs, Ingeborg; Barillas, Rodrigo; Vasilyev, Nikolay V.; Roy, Nathalie; McGowan, Francis X.; del Nido, Pedro J.

    2012-01-01

    Background Cardiac hypertrophy is an adaptive response to increased workload that, if unrelieved, leads to heart failure. It has been reported that cardiomyocyte apoptosis contributes to failure, and that vascular endothelial growth factor (VEGF) treatment of hypertrophied myocardium increases capillary density and improves myocardial perfusion. In this study we hypothesized that VEGF treatment reduces cardiomyocyte apoptosis and thereby preserves myocardial contractile function. Methods and Results Newborn rabbits underwent aortic banding. At 4 and 6 weeks of age, hypertrophied animals were treated with intrapericardial administration of recombinant VEGF protein. Three groups of animals were investigated: age-matched controls (C), untreated hypertrophied (H), and VEGF-treated hypertrophied hearts (T). Cardiomyocyte apoptosis was determined by TUNEL staining and PARP cleavage (immunoblotting of nuclear extracts) and cardiac function by transthoracic echocardiography. Death attributable to severe heart failure occurred in 14 of 43 untreated and 2 of 29 VEGF-treated animals (P<0.01). TUNEL-positive cardiomyocyte nuclei (n/1000 nuclei) were significantly increased in untreated hearts at 5 weeks (H: 10±1.8 versus T: 3±0.7) and at 7 weeks (H: 13±3.6 versus T: 5±1.5; P<0.05). Increased apoptosis in untreated hypertrophy was also confirmed by the presence of PARP cleavage (H: 74±7 versus T: 41±4 arbitrary densitometry units; P<0.05). VEGF treatment preserved left ventricular mass, prevented dilation (T: 1.01±0.06 versus H: 0.77±0.07; P<0.05), and preserved contractility indices compared with untreated hearts. Conclusions Lack of adaptive capillary growth impairs myocardial perfusion and substrate delivery in hypertrophying myocardium. VEGF treatment reduces myocardial apoptosis and prolongs survival in a model of severe progressive left ventricular hypertrophy. Promoting capillary growth with VEGF reduces apoptosis, preserves myocardial contractile function, and

  16. Prevention of lymphocyte apoptosis in septic mice with cancer increases mortality

    PubMed Central

    Fox, Amy C.; Breed, Elise R; Liang, Zhe; Clark, Andrew T.; Zee-Cheng, Brendan R.; Chang, Katherine C.; Dominguez, Jessica A.; Jung, Enjae; Dunne, W. Michael; Burd, Eileen M.; Farris, Alton B.; Linehan, David C.; Coopersmith, Craig M.

    2011-01-01

    Lymphocyte apoptosis is thought to play a major role in the pathophysiology of sepsis. However, there is a disconnect between animal models of sepsis and patients with the disease, since the former use subjects that were healthy prior to the onset of infection while most patients have underlying comorbidities. The purpose of this study was to determine whether lymphocyte apoptosis prevention is effective in preventing mortality in septic mice with pre-existing cancer. Mice with lymphocyte Bcl-2 overexpression (Bcl-2-Ig) and wild type (WT) mice were injected with a transplantable pancreatic adenocarcinoma cell line. Three weeks later after development of palpable tumors, all animals received an intratracheal injection of Pseudomonas aeruginosa. Despite having decreased sepsis-induced T and B lymphocyte apoptosis, Bcl-2-Ig mice had markedly increased mortality compared to WT mice following Pseudomonas aeruginosa pneumonia (85% vs. 44% seven-day mortality, p=0.004). The worsened survival in Bcl-2-Ig mice was associated with increases in Th1 cytokines TNF-α and IFN-γ in bronchoalveolar lavage fluid and decreased production of the Th2 cytokine IL-10 in stimulated splenocytes. There were no differences in tumor size or pulmonary pathology between Bcl-2-Ig and WT mice. To verify the mortality difference was not specific to Bcl-2 overexpression, similar experiments were performed in Bim-/- mice. Septic Bim-/- mice with cancer also had increased mortality compared to septic WT mice with cancer. These data demonstrate that despite overwhelming evidence that prevention of lymphocyte apoptosis is beneficial in septic hosts without comorbidities, the same strategy worsens survival in mice with cancer that are given pneumonia. PMID:21734077

  17. Prevention of lymphocyte apoptosis in septic mice with cancer increases mortality.

    PubMed

    Fox, Amy C; Breed, Elise R; Liang, Zhe; Clark, Andrew T; Zee-Cheng, Brendan R; Chang, Katherine C; Dominguez, Jessica A; Jung, Enjae; Dunne, W Michael; Burd, Eileen M; Farris, Alton B; Linehan, David C; Coopersmith, Craig M

    2011-08-15

    Lymphocyte apoptosis is thought to have a major role in the pathophysiology of sepsis. However, there is a disconnect between animal models of sepsis and patients with the disease, because the former use subjects that were healthy prior to the onset of infection while most patients have underlying comorbidities. The purpose of this study was to determine whether lymphocyte apoptosis prevention is effective in preventing mortality in septic mice with preexisting cancer. Mice with lymphocyte Bcl-2 overexpression (Bcl-2-Ig) and wild type (WT) mice were injected with a transplantable pancreatic adenocarcinoma cell line. Three weeks later, after development of palpable tumors, all animals received an intratracheal injection of Pseudomonas aeruginosa. Despite having decreased sepsis-induced T and B lymphocyte apoptosis, Bcl-2-Ig mice had markedly increased mortality compared with WT mice following P. aeruginosa pneumonia (85 versus 44% 7-d mortality; p = 0.004). The worsened survival in Bcl-2-Ig mice was associated with increases in Th1 cytokines TNF-α and IFN-γ in bronchoalveolar lavage fluid and decreased production of the Th2 cytokine IL-10 in stimulated splenocytes. There were no differences in tumor size or pulmonary pathology between Bcl-2-Ig and WT mice. To verify that the mortality difference was not specific to Bcl-2 overexpression, similar experiments were performed in Bim(-/-) mice. Septic Bim(-/-) mice with cancer also had increased mortality compared with septic WT mice with cancer. These data demonstrate that, despite overwhelming evidence that prevention of lymphocyte apoptosis is beneficial in septic hosts without comorbidities, the same strategy worsens survival in mice with cancer that are given pneumonia. PMID:21734077

  18. Tamoxifen prevents apoptosis and follicle loss from cyclophosphamide in cultured rat ovaries.

    PubMed

    Piasecka-Srader, Joanna; Blanco, Fernando F; Delman, Devora H; Dixon, Dan A; Geiser, James L; Ciereszko, Renata E; Petroff, Brian K

    2015-05-01

    Recent studies documented that the selective estrogen receptor modulator tamoxifen prevents follicle loss and promotes fertility following in vivo exposure of rodents to irradiation or ovotoxic cancer drugs, cyclophosphamide and doxorubicin. In an effort to characterize the ovarian-sparing mechanisms of tamoxifen in preantral follicle classes, cultured neonatal rat ovaries (Day 4, Sprague Dawley) were treated for 1-7 days with active metabolites of cyclophosphamide (i.e., 4-hydroxycyclophosphamide; CTX) (0, 1, and 10 μM) and tamoxifen (i.e., 4-hydroxytamoxifen; TAM) (0 and 10 μM) in vitro, and both apoptosis and follicle numbers were measured. CTX caused marked follicular apoptosis and follicular loss. TAM treatment decreased follicular loss and apoptosis from CTX in vitro. TAM alone had no effect on these parameters. IGF-1 and IGF-1 receptor were assessed in ovarian tissue showing no impact of TAM or CTX on these endpoints. Targeted mRNA analysis during follicular rescue by TAM revealed decreased expression of multiple genes related to inflammation, including mediators of lipoxygenase and prostaglandin production and signaling (Alox5, Pla2g1b, Ptgfr), cytokine binding (Il1r1, Il2rg ), apoptosis (Tnfrsf1a), second messenger signaling (Mapk1, Mapk14, Plcg1), as well as tissue remodeling and vasodilation (Bdkrb2, Klk15). The results suggest that TAM protects the ovary from CTX-mediated toxicity through direct ovarian actions that oppose follicular loss. PMID:25833159

  19. Nicotine prevents the apoptosis induced by menadione in human lung cancer cells

    SciTech Connect

    Zhang Tao; Lu Heng; Shang Xuan; Tian Yihao; Zheng Congyi; Wang Shiwen; Cheng Hanhua . E-mail: hhcheng@whu.edu.cn; Zhou Rongjia . E-mail: rjzhou@whu.edu.cn

    2006-04-14

    Approximately 50% of long-term cigarette smokers die prematurely from the adverse effects of smoking, including on lung cancer and other illnesses. Nicotine is a main component in tobacco and has been implicated as a potential factor in the pathogenesis of human lung cancer. However, the mechanism of nicotine action in the development of lung cancer remains largely unknown. In the present study, we designed a nicotine-apoptosis system, by pre-treatment of nicotine making lung cancer cell A549 to be in a physiological nicotine environment, and observed that nicotine promoted cell proliferation and prevented the menadione-induced apoptosis, and exerts its role of anti-apoptosis by shift of apoptotic stage induced by menadione from late apoptotic stage to early apoptotic stage, in which NF-{kappa}B was up-regulated. Interference analysis of NF-{kappa}B in A549 cells showed that knock down of NF-{kappa}B resulted in apoptosis promotion and counteracted the protective effect of nicotine. The findings suggest that nicotine has potential effect in lung cancer genesis, especially in patients with undetectable early tumor development and development of specific NF-{kappa}B inhibitors would represent a potentially exciting new pharmacotherapy for tobacco-related lung cancer.

  20. PD150606 protects against ischemia/reperfusion injury by preventing μ-calpain-induced mitochondrial apoptosis.

    PubMed

    Luo, Tao; Yue, Rongchuan; Hu, Houxiang; Zhou, Zhou; Yiu, Kai Hang; Zhang, Shuang; Xu, Lei; Li, Ke; Yu, Zhengping

    2015-11-15

    Calpain plays an important role in myocardial ischemia/reperfusion (I/R) injury. PD150606, a nonpeptide, cell-permeable and noncompetitive calpain inhibitor, has been shown to have protective properties in ischemic disease. The aims of the present study were to investigate whether PD150606 could alleviate myocardial I/R injury and to examine the possible mechanisms involved. The I/R model was established in vivo in C57BL/6 mice and in vitro using neonatal mouse cardiomyocytes, respectively. To evaluate the protective effects of PD150606 on I/R injury, we measured the myocardial infarct area, apoptosis, and expression of cleaved caspase-3. We also investigated the underlying mechanisms by examining mitochondrial function as reflected by the ATP concentration, translocation of cytochrome c, dynamics of mPTP opening, and membrane potential (ΔΨm), coupled with calpain activity. Pretreatment with PD150606 significantly reduced the infarct area and apoptosis caused by I/R. PD150606 pretreatment also reduced mitochondrial dysfunction by inhibiting calpain activation. Moreover, we found that μ-calpain is the main contributor to I/R-induced calpain activation. Knockdown of μ-calpain with siRNA significantly reversed calpain activation, mitochondrial dysfunction, and cardiomyocyte apoptosis caused by I/R in vitro. Our results suggest that PD150606 may protect against I/R injury via preventing μ-calpain-induced mitochondrial apoptosis. PMID:26091952

  1. Phycocyanin prevents methylglyoxal-induced mitochondrial-dependent apoptosis in INS-1 cells by Nrf2.

    PubMed

    Gao, Yingnv; Liu, Chen; Wan, Guoqing; Wang, Xinshuo; Cheng, Xiaodong; Ou, Yu

    2016-02-01

    Methylglyoxal (MG) is a reactive dicarbonyl compound, whose abnormal accumulation in diabetic patients exerts deleterious effects on cells and tissues. The β-cell is the main target cell of Type 2 diabetes, and its insulin secretion injury and cell apoptosis can be due to mitochondrial dysfunction. Previous studies have demonstrated MG induced β-cell apoptosis. However, little is known about the effect of MG on β-cell mitochondrial dysfunction. Phycocyanin (PC) has been demonstrated to possess various biological activities including the effects on diabetic models in vivo. The aim of this study was to determine the protective effect of PC against methylglyoxal (MG)-induced dysfunction in pancreatic β-cell INS-1 and also the mechanism. We demonstrated that MG induced mitochondrial dysfunction by the decline in ATP levels, and the increase of the level of intracellular reactive oxygen species (ROS). Furthermore, MG released cytochrome c and apoptosis-inducing factor (AIF) from the mitochondrion, induced changes in the expression of Bcl-2 family members, activated caspases and increased PARP cleavage. Interestingly, PC activated nuclear erythroid-related factor 2 (Nrf2), and Nrf2 activation as well as antioxidant enzymes HO-1 and glyoxalase 1 (Glo-1) were confirmed to be involved in the mechanisms underlying the protection of PC by RNA interference. Altogether, these results demonstrated that PC prevented mitochondrial-dependent apoptosis in MG-induced INS-1 cells and the effect was associated with Nrf2 activation. PMID:26805012

  2. Tamoxifen Prevents Apoptosis and Follicle Loss from Cyclophosphamide in Cultured Rat Ovaries1

    PubMed Central

    Piasecka-Srader, Joanna; Blanco, Fernando F.; Delman, Devora H.; Dixon, Dan A.; Geiser, James L.; Ciereszko, Renata E.; Petroff, Brian K.

    2015-01-01

    Recent studies documented that the selective estrogen receptor modulator tamoxifen prevents follicle loss and promotes fertility following in vivo exposure of rodents to irradiation or ovotoxic cancer drugs, cyclophosphamide and doxorubicin. In an effort to characterize the ovarian-sparing mechanisms of tamoxifen in preantral follicle classes, cultured neonatal rat ovaries (Day 4, Sprague Dawley) were treated for 1–7 days with active metabolites of cyclophosphamide (i.e., 4-hydroxycyclophosphamide; CTX) (0, 1, and 10 μM) and tamoxifen (i.e., 4-hydroxytamoxifen; TAM) (0 and 10 μM) in vitro, and both apoptosis and follicle numbers were measured. CTX caused marked follicular apoptosis and follicular loss. TAM treatment decreased follicular loss and apoptosis from CTX in vitro. TAM alone had no effect on these parameters. IGF-1 and IGF-1 receptor were assessed in ovarian tissue showing no impact of TAM or CTX on these endpoints. Targeted mRNA analysis during follicular rescue by TAM revealed decreased expression of multiple genes related to inflammation, including mediators of lipoxygenase and prostaglandin production and signaling (Alox5, Pla2g1b, Ptgfr), cytokine binding (Il1r1, Il2rg ), apoptosis (Tnfrsf1a), second messenger signaling (Mapk1, Mapk14, Plcg1), as well as tissue remodeling and vasodilation (Bdkrb2, Klk15). The results suggest that TAM protects the ovary from CTX-mediated toxicity through direct ovarian actions that oppose follicular loss. PMID:25833159

  3. cAMP prevents TNF-induced apoptosis through inhibiting DISC complex formation in rat hepatocytes

    SciTech Connect

    Bhattacharjee, Rajesh; Xiang, Wenpei; Wang, Yinna; Zhang, Xiaoying

    2012-06-22

    that cAMP exerts its affect at the proximal level of TNF signaling by inhibiting the formation of the DISC complex upon the binding of TNF to TNFR1. In conclusion, our study shows that cAMP prevents TNF + ActD-induced apoptosis in rat hepatocytes by inhibiting DISC complex formation.

  4. Cutaneous HPV23 E6 prevents p53 phosphorylation through interaction with HIPK2.

    PubMed

    Muschik, Dorothea; Braspenning-Wesch, Ilona; Stockfleth, Eggert; Rösl, Frank; Hofmann, Thomas G; Nindl, Ingo

    2011-01-01

    Ultraviolet irradiation (UV) is the major risk factor for the development of skin cancer. Moreover, increasing evidence suggests cutaneotropic human papillomaviruses (HPV) from the beta genus to play a causal role as a co-factor in the development of cutaneous squamous cell carcinoma. Homeodomain-interacting protein kinase 2 (HIPK2) operates as a potential suppressor in skin tumorigenesis and is stabilized by UV-damage. HIPK2 is an important regulator of apoptosis, which forms a complex with the tumor suppressor p53, mediating p53 phosphorylation at Ser 46 and thus promoting pro-apoptotic gene expression. In our study, we demonstrate that cutaneous HPV23 E6 protein directly targets HIPK2 function. Accordingly, HPV23 E6 interacts with HIPK2 both in vitro and in vivo. Furthermore, upon massive UVB-damage HPV23 E6 co-localizes with endogenous HIPK2 at nuclear bodies. Functionally, we demonstrate that HPV23 E6 inhibits HIPK2-mediated p53 Ser 46 phosphorylation through enforcing dissociation of the HIPK2/p53 complex. In addition, HPV23 E6 co-accumulates with endogenous HIPK2 upon UV damage suggesting a mechanism by which HPV23 E6 keeps HIPK2 in check after UV damage. Thus, cutaneous HPV23 E6 prevents HIPK2-mediated p53 Ser 46 phosphorylation, which may favour survival of UV-damaged keratinocytes and skin carcinogenesis by apoptosis evasion. PMID:22110707

  5. Subclinical concentrations of sevoflurane reduce oxidative stress but do not prevent hippocampal apoptosis

    PubMed Central

    ZHOU, ZHI-BIN; YANG, XIAO-YU; TANG, YING; ZHOU, XUE; ZHOU, LI-HUA; FENG, XIA

    2016-01-01

    Sevoflurane is generally considered a pro-apoptotic agent in the neonatal brain. However, recent studies have suggested that low levels of sevoflurane anesthesia may be neuroprotective and have a memory enhancing effect. The present study aimed to investigate whether sevoflurane exerts a neuroprotective effect at subclinical concentrations, with regard to oxidative state. In the current study, postnatal day 7 (P7) Sprague-Dawley rats were continuously exposed to 0.3, 1.3, or 2.3% sevoflurane for 6 h. ELISA was used to quantify the levels of superoxide dismutase, glutathione peroxidase (GSH-px) and malondialdehyde (MDA) in the plasma and the hippocampus. Terminal deoxynucleotidyl-transferase dUTP nick-end labeling staining was used to observe hippocampal neuronal apoptosis. Altered object exploration tests for recognition memory were employed to investigate long-term behavioral effects at postnatal day 28. The results demonstrated that a single 6 h exposure to a subclinical concentration (1.3%) of sevoflurane at P7 reduces MDA and GPH-px production in rats. Sevoflurane induced hippocampal apoptosis in a dose-dependent manner and altered recognition memory testing indicated no differences among the groups. Although early exposure to a subclinical concentration of sevoflurane reduced oxidative stress, it did not prevent the process of sevoflurane-induced hippocampal apoptosis. These changes did not affect subsequent recognition memory in juvenile rats. PMID:27222114

  6. Role of lubricin and boundary lubrication in the prevention of chondrocyte apoptosis.

    PubMed

    Waller, Kimberly A; Zhang, Ling X; Elsaid, Khaled A; Fleming, Braden C; Warman, Matthew L; Jay, Gregory D

    2013-04-01

    Osteoarthritis is a complex disease involving the mechanical breakdown of articular cartilage in the presence of altered joint mechanics and chondrocyte death, but the connection between these factors is not well established. Lubricin, a mucinous glycoprotein encoded by the PRG4 gene, provides boundary lubrication in articular joints. Joint friction is elevated and accompanied by accelerated cartilage damage in humans and mice that have genetic deficiency of lubricin. Here, we investigated the relationship between coefficient of friction and chondrocyte death using ex vivo and in vitro measurements of friction and apoptosis. We observed increases in whole-joint friction and cellular apoptosis in lubricin knockout mice compared with wild-type mice. When we used an in vitro bovine explant cartilage-on-cartilage bearing system, we observed a direct correlation between coefficient of friction and chondrocyte apoptosis in the superficial layers of cartilage. In the bovine explant system, the addition of lubricin as a test lubricant significantly lowered the static coefficient of friction and number of apoptotic chondrocytes. These results demonstrate a direct connection between lubricin, boundary lubrication, and cell survival and suggest that supplementation of synovial fluid with lubricin may be an effective treatment to prevent cartilage deterioration in patients with genetic or acquired deficiency of lubricin. PMID:23530215

  7. Subclinical concentrations of sevoflurane reduce oxidative stress but do not prevent hippocampal apoptosis.

    PubMed

    Zhou, Zhi-Bin; Yang, Xiao-Yu; Tang, Ying; Zhou, Xue; Zhou, Li-Hua; Feng, Xia

    2016-07-01

    Sevoflurane is generally considered a pro-apoptotic agent in the neonatal brain. However, recent studies have suggested that low levels of sevoflurane anesthesia may be neuroprotective and have a memory enhancing effect. The present study aimed to investigate whether sevoflurane exerts a neuroprotective effect at subclinical concentrations, with regard to oxidative state. In the current study, postnatal day 7 (P7) Sprague‑Dawley rats were continuously exposed to 0.3, 1.3, or 2.3% sevoflurane for 6 h. ELISA was used to quantify the levels of superoxide dismutase, glutathione peroxidase (GSH‑px) and malondialdehyde (MDA) in the plasma and the hippocampus. Terminal deoxynucleotidyl-transferase dUTP nick-end labeling staining was used to observe hippocampal neuronal apoptosis. Altered object exploration tests for recognition memory were employed to investigate long‑term behavioral effects at postnatal day 28. The results demonstrated that a single 6 h exposure to a subclinical concentration (1.3%) of sevoflurane at P7 reduces MDA and GPH‑px production in rats. Sevoflurane induced hippocampal apoptosis in a dose‑dependent manner and altered recognition memory testing indicated no differences among the groups. Although early exposure to a subclinical concentration of sevoflurane reduced oxidative stress, it did not prevent the process of sevoflurane-induced hippocampal apoptosis. These changes did not affect subsequent recognition memory in juvenile rats. PMID:27222114

  8. Proteasome inhibitors prevent cytochrome c release during apoptosis but not in excitotoxic death of cerebellar granule neurons.

    PubMed

    Bobba, Antonella; Canu, Nadia; Atlante, Anna; Petragallo, Vito; Calissano, Pietro; Marra, Ersilia

    2002-03-27

    In order to find out whether and how proteasomes participate in the processes leading cerebellar granule cells to death either in necrosis, due to glutamate neurotoxicity, or in apoptosis, due to K(+) shift, we measured the three proteasome activities by using specific fluorescent probes and investigated the effect of several proteasome inhibitors, including MG132, on the cytochrome c release taking place in the early phase of both apoptosis and necrosis. We show that differently from apoptosis, the early phase of necrosis does not require proteasome activation. Inhibition of proteasome activity can prevent cytochrome c release in cerebellar granule cells undergoing apoptosis, thus improving cell survival, but not necrosis. These findings show that proteasomes play an important role in the early phase of apoptosis but not that of necrosis, and that these two types of cell death differ from each other in their mechanism of cytochrome c release. PMID:11943185

  9. Modulatory Effects of Polyphenols on Apoptosis Induction: Relevance for Cancer Prevention

    PubMed Central

    D'Archivio, Massimo; Santangelo, Carmela; Scazzocchio, Beatrice; Varì, Rosaria; Filesi, Carmela; Masella, Roberta; Giovannini, Claudio

    2008-01-01

    Polyphenols, occurring in fruit and vegetables, wine, tea, extra virgin olive oil, chocolate and other cocoa products, have been demonstrated to have clear antioxidant properties in vitro, and many of their biological actions have been attributed to their intrinsic reducing capabilities. However, it has become clear that, in complex biological systems, polyphenols exhibit several additional properties which are yet poorly understood. Apoptosis is a genetically controlled and evolutionarily conserved form of cell death of critical importance for the normal embryonic development and for the maintenance of tissue homeostasis in the adult organism. The malfunction of the death machinery may play a primary role in various pathological processes, since too little or too much apoptosis can lead to proliferative or degenerative diseases, respectively. Cancer cells are characterized by a deregulated proliferation, and/or an inability to undergo programmed cell death. A large body of evidence indicates that polyphenols can exert chemopreventive effects towards different organ specific cancers, affecting the overall process of carcinogenesis by several mechanisms: inhibition of DNA synthesis, modulation of ROS production, regulation of cell cycle arrest, modulation of survival/proliferation pathways. In addition, polyphenols can directly influence different points of the apoptotic process, and/or the expression of regulatory proteins. Although the bulk of data has been obtained in in vitro systems, a number of clinical studies suggesting a preventive and therapeutic effectiveness of polyphenols in vivo is available. However, a deeper knowledge of the underlying mechanisms responsible for the modulation of apoptosis by polyphenols, and their real effectiveness, is necessary in order to propose them as potential chemopreventive and chemotherapeutic candidates for cancer treatment. PMID:19325744

  10. Modulatory effects of polyphenols on apoptosis induction: relevance for cancer prevention.

    PubMed

    D'Archivio, Massimo; Santangelo, Carmela; Scazzocchio, Beatrice; Varì, Rosaria; Filesi, Carmela; Masella, Roberta; Giovannini, Claudio

    2008-03-01

    Polyphenols, occurring in fruit and vegetables, wine, tea, extra virgin olive oil, chocolate and other cocoa products, have been demonstrated to have clear antioxidant properties in vitro, and many of their biological actions have been attributed to their intrinsic reducing capabilities. However, it has become clear that, in complex biological systems, polyphenols exhibit several additional properties which are yet poorly understood. Apoptosis is a genetically controlled and evolutionarily conserved form of cell death of critical importance for the normal embryonic development and for the maintenance of tissue homeostasis in the adult organism. The malfunction of the death machinery may play a primary role in various pathological processes, since too little or too much apoptosis can lead to proliferative or degenerative diseases, respectively. Cancer cells are characterized by a deregulated proliferation, and/or an inability to undergo programmed cell death. A large body of evidence indicates that polyphenols can exert chemopreventive effects towards different organ specific cancers, affecting the overall process of carcinogenesis by several mechanisms: inhibition of DNA synthesis, modulation of ROS production, regulation of cell cycle arrest, modulation of survival/proliferation pathways. In addition, polyphenols can directly influence different points of the apoptotic process, and/or the expression of regulatory proteins. Although the bulk of data has been obtained in in vitro systems, a number of clinical studies suggesting a preventive and therapeutic effectiveness of polyphenols in vivo is available. However, a deeper knowledge of the underlying mechanisms responsible for the modulation of apoptosis by polyphenols, and their real effectiveness, is necessary in order to propose them as potential chemopreventive and chemotherapeutic candidates for cancer treatment. PMID:19325744

  11. Heat stress prevents lipopolysaccharide-induced apoptosis in pulmonary microvascular endothelial cells by blocking calpain/p38 MAPK signalling.

    PubMed

    Liu, Zhi-Feng; Zheng, Dong; Fan, Guo-Chang; Peng, Tianqing; Su, Lei

    2016-08-01

    Pulmonary microvascular endothelial cells (PMECs) injury including apoptosis plays an important role in the pathogenesis of acute lung injury during sepsis. Our recent study has demonstrated that calpain activation contributes to apoptosis in PMECs under septic conditions. This study investigated how calpain activation mediated apoptosis and whether heat stress regulated calpain activation in lipopolysaccharides (LPS)-stimulated PMECs. In cultured mouse primary PMECs, incubation with LPS (1 μg/ml, 24 h) increased active caspase-3 fragments and DNA fragmentation, indicative of apoptosis. These effects of LPS were abrogated by pre-treatment with heat stress (43 °C for 2 h). LPS also induced calpain activation and increased phosphorylation of p38 MAPK. Inhibition of calpain and p38 MAPK prevented apoptosis induced by LPS. Furthermore, inhibition of calpain blocked p38 MAPK phosphorylation in LPS-stimulated PMECs. Notably, heat stress decreased the protein levels of calpain-1/2 and calpain activities, and blocked p38 MAPK phosphorylation in response to LPS. Additionally, forced up-regulation of calpain-1 or calpain-2 sufficiently induced p38 MAPK phosphorylation and apoptosis in PMECs, both of which were inhibited by heat stress. In conclusion, heat stress prevents LPS-induced apoptosis in PMECs. This effect of heat stress is associated with down-regulation of calpain expression and activation, and subsequent blockage of p38 MAPK activation in response to LPS. Thus, blocking calpain/p38 MAPK pathway may be a novel mechanism underlying heat stress-mediated inhibition of apoptosis in LPS-stimulated endothelial cells. PMID:27325431

  12. Hydroxycamptothecin induces apoptosis of fibroblasts and prevents intraarticular scar adhesion in rabbits by activating the IRE-1 signal pathway.

    PubMed

    Li, Xiaolei; Sun, Yu; Chen, Hui; Zhu, Gengyao; Liang, Yuan; Wang, Qiang; Wang, Jingcheng; Yan, Lianqi

    2016-06-15

    Hydroxycamptothecin (HCPT) has been proven to prevent intraarticular scar adhesion, but the mechanism is still unclear. ER stress is known to participate in many diseases, and the IRE-1 signal pathway has been reported in fibrotic diseases. The aim of this study was to illustrate the mechanism of HCPT-induced apoptosis in fibroblasts and the prevention of intraarticular scar adhesion. The effects of HCPT on fibroblasts were determined by CCK-8 assay, Hoechst staining and Western blot. The effect of HCPT on intraarticular scar adhesion was detected by macroscopic evaluation, hydroxyproline content, histological evaluation, fibroblast counting and immunohistochemical analysis. HCPT induced apoptosis of fibroblasts, according to CCK-8 assays, Hoechst staining and Western blot analysis. As the concentration of HCPT increased, the expressions of glucose-regulated protein 78 (GRP78), inositol-requiring kinase1 (IRE-1), C/EBP homologous protein (CHOP) and Bax were all increased, but the expression of Bcl-2 was decreased. Knockdown of IRE-1 alleviated the HCPT-induced apoptosis in our fibroblast model. HCPT could prevent intraarticular scar adhesion, according to the results of macroscopic evaluation, hydroxyproline content, histological evaluation and fibroblast counting in a rabbit model. Immunohistochemical analysis showed that IRE-1 expression increased as the concentration increased. The present study showed that the IRE-1 signal pathway might be involved in HCPT-induced apoptosis of fibroblast and might play a role in preventing intraarticular scar adhesion. PMID:27068147

  13. Silencing of the polyamine catabolic key enzyme SSAT prevents CDK inhibitor-induced apoptosis in Caco-2 colon cancer cells.

    PubMed

    Çoker, A; Arısan, E D; Palavan-Ünsal, N

    2012-04-01

    Roscovitine and purvalanol are purine derivative cyclin-dependent kinase (CDK) inhibitors that induce apoptosis in various types of cancer cells. However, their impact on the apoptotic cell death mechanism requires further elucidation. Natural polyamines putrescine, spermidine and spermine play essential roles in the regulation of cell growth and proliferation. Increased levels of polyamines in cells are considered to be involved in cancer progression. Intracellular polyamine levels are under the control of several catabolic enzymes, such as spermidine/spermine-N-acetyl transferase (SSAT), acetylpolyamine oxidase (APAO) and spermine oxidase (SMO), which could be altered by several therapeutic drugs. However, the possible role of polyamines in drug-induced apoptosis has yet to be clarified. In the present study, our aim was to determine the modulation of the polyamine catabolic pathway related to CDK inhibitor-induced apoptosis in Caco-2 cells. We found that roscovitine and purvalanol (each 20 µM) induced apoptosis by activating caspase-9 and -3, and inhibiting the mitochondrial membrane potential in Caco-2 cells. CDK inhibitors decreased the intracellular putrescine and spermine levels without affecting spermidine levels. Although both roscovitine and purvalanol induced SSAT expression, they did not exert a significant effect on the APAO expression profile. SSAT transient silencing prevented roscovitine-induced apoptosis compared to parental cells. Thus, we concluded that roscovitine and purvalanol significantly induce apoptosis in Caco-2 cells by modulating the polyamine catabolism, and that SSAT could be an important target in evaluating the potential role of polyamines in apoptotic cell death. PMID:22294330

  14. Leishmania donovani Exploits Myeloid Cell Leukemia 1 (MCL-1) Protein to Prevent Mitochondria-dependent Host Cell Apoptosis.

    PubMed

    Giri, Jayeeta; Srivastav, Supriya; Basu, Moumita; Palit, Shreyasi; Gupta, Purnima; Ukil, Anindita

    2016-02-12

    Apoptosis is one of the mechanisms used by host cells to remove unwanted intracellular organisms, and often found to be subverted by pathogens through use of host anti-apoptotic proteins. In the present study, with the help of in vitro and in vivo approaches, we documented that the macrophage anti-apoptotic protein myeloid cell leukemia 1 (MCL-1) is exploited by the intra-macrophage parasite Leishmania donovani to protect their "home" from actinomycin D-induced mitochondria-dependent apoptosis. Among all the anti-apoptotic BCL-2 family members, infection preferentially up-regulated expression of MCL-1 at both the mRNA and protein levels and compared with infected control, MCL-1-silenced infected macrophages documented enhanced caspase activity and increased apoptosis when subjected to actinomycin D treatment. Phosphorylation kinetics and ChIP assay demonstrated that infection-induced MCL-1 expression was regulated by transcription factor CREB (cAMP-response element-binding protein) and silencing of CREB resulted in reduced expression of MCL-1 and increased apoptosis. During infection, MCL-1 was found to be localized in mitochondria and this was significantly reduced in Tom70-silenced macrophages, suggesting the active role of TOM70 in MCL-1 transport. In the mitochondria, MCL-1 interacts with the major pro-apoptotic protein BAK and prevents BAK-BAK homo-oligomer formation thereby preventing cytochrome c release-mediated mitochondrial dysfunction. Silencing of MCL-1 in the spleen of infected mice showed decreased parasite burden and increased induction of splenocyte apoptosis. Collectively our results showed that L. donovani exploited the macrophage anti-apoptotic protein MCL-1 to prevent BAK-mediated mitochondria-dependent apoptosis thereby protecting its niche, which is essential for disease progression. PMID:26670606

  15. Folic Acid Protected Neural Cells Against Aluminum-Maltolate-Induced Apoptosis by Preventing miR-19 Downregulation.

    PubMed

    Zhu, Mingming; Li, Bingfei; Ma, Xiao; Huang, Cong; Wu, Rui; Zhu, Weiwei; Li, Xiaoting; Liang, Zhaofeng; Deng, Feifei; Zhu, Jianyun; Xie, Wei; Yang, Xue; Jiang, Ye; Wang, Shijia; Wu, Jieshu; Geng, Shanshan; Xie, Chunfeng; Zhong, Caiyun; Liu, Haiyan

    2016-08-01

    Aluminum (Al)-induced apoptosis is considered as the major cause of its neurotoxicity. Folic acid possesses neuroprotective function by preventing neural cell apoptosis. microRNAs (miRNAs) are important regulators of gene expression participating in cellular processes. As a key component of the miR-17-92 cluster, miR-19 is implicated in regulating apoptotic process, while its role in the neuroprotective effect of folic acid has not been investigated. The present study aimed to investigate the potential involvement and function of miR-19 in the protective action of folic acid against Al-induced neural cell apoptosis. Human SH-SY5Y cells were treated with Al-maltolate (Al-malt) in the presence or absence of folic acid. Results showed that Al-malt-induced apoptosis of SH-SY5Y cells was effectively prevented by folic acid. Al-malt suppressed the expression of miR-19a/19b, along with alterations of miR-19 related apoptotic proteins including PTEN, p-AKT, p53, Bax, Bcl-2, caspase 9 and caspase 3; and these effects were ameliorated by folic acid. miR-19 inhibitor alone induced apoptosis of SH-SY5Y cells. Combination treatment of folic acid and miR-19 inhibitor diminished the neuroprotective effect of folic acid. These findings demonstrated that folic acid protected neuronal cells against Al-malt-induced apoptosis by preventing the downregulation of miR-19 and modulation of miR-19 related downstream PTEN/AKT/p53 pathway. PMID:27113042

  16. cAMP signaling prevents podocyte apoptosis via activation of protein kinase A and mitochondrial fusion.

    PubMed

    Li, Xiaoying; Tao, Hua; Xie, Kewei; Ni, Zhaohui; Yan, Yucheng; Wei, Kai; Chuang, Peter Y; He, John Cijiang; Gu, Leyi

    2014-01-01

    Our previous in vitro studies suggested that cyclic AMP (cAMP) signaling prevents adriamycin (ADR) and puromycin aminonucleoside (PAN)-induced apoptosis in podocytes. As cAMP is an important second messenger and plays a key role in cell proliferation, differentiation and cytoskeleton formation via protein kinase A (PKA) or exchange protein directly activated by cAMP (Epac) pathways, we sought to determine the role of PKA or Epac signaling in cAMP-mediated protection of podocytes. In the ADR nephrosis model, we found that forskolin, a selective activator of adenylate cyclase, attenuated albuminuria and improved the expression of podocyte marker WT-1. When podocytes were treated with pCPT-cAMP (a selective cAMP/PKA activator), PKA activation was increased in a time-dependent manner and prevented PAN-induced podocyte loss and caspase 3 activation, as well as a reduction in mitochondrial membrane potential. We found that PAN and ADR resulted in a decrease in Mfn1 expression and mitochondrial fission in podocytes. pCPT-cAMP restored Mfn1 expression in puromycin or ADR-treated podocytes and induced Drp1 phosphorylation, as well as mitochondrial fusion. Treating podocytes with arachidonic acid resulted in mitochondrial fission, podocyte loss and cleaved caspase 3 production. Arachidonic acid abolished the protective effects of pCPT-cAMP on PAN-treated podocytes. Mdivi, a mitochondrial division inhibitor, prevented PAN-induced cleaved caspase 3 production in podocytes. We conclude that activation of cAMP alleviated murine podocyte caused by ADR. PKA signaling resulted in mitochondrial fusion in podocytes, which at least partially mediated the effects of cAMP. PMID:24642777

  17. Staurosporine-induced apoptosis in astrocytes is prevented by A1 adenosine receptor activation.

    PubMed

    D'Alimonte, Iolanda; Ballerini, Patrizia; Nargi, Eleonora; Buccella, Silvana; Giuliani, Patricia; Di Iorio, Patrizia; Caciagli, Francesco; Ciccarelli, Renata

    2007-05-11

    Astrocyte apoptosis occurs in acute and chronic pathological processes at the central nervous system and the prevention of astrocyte death may represent an efficacious intervention in protecting neurons against degeneration. Our research shows that rat astrocyte exposure to 100 nM staurosporine for 3h caused apoptotic death accompanied by caspase-3, p38 mitogen-ed protein kinase (MAPK) and glycogen synthase kinase-3beta (GSK3beta) activation. N(6)-chlorocyclopentyladenosine (CCPA, 2.5-75 nM), a selective agonist of A(1) adenosine receptors, added to the cultures 1h prior to staurosporine, induced a dose-dependent anti-apoptotic effect, which was inhibited by the A(1) receptor antagonist, 8-cyclopentyl-1,3-dipropylxanthine. CCPA also caused a dose- and time-dependent phosphorylation/activation of Akt, a downstream effector of cell survival promoting phosphatidylinositol 3-kinase (PI3K) pathway, which in turn led to inhibition of staurosporine-induced GSK3beta and p38 MAPK activity. Accordingly, the anti-apoptotic effect of CCPA was abolished by culture pre-treatment with LY294002, a selective PI3K inhibitor, pointing out the prevailing role played by PI3K pathway in the protective effect exerted by A(1) receptor activation. Since an abnormal p38 and GSK3beta activity is implicated in acute (stroke) and chronic (Alzheimer's disease) neurodegenerative diseases, the results of the present study provide a hint to better understand adenosine relevance in these disorders. PMID:17400382

  18. Comprehensive Suppression of All Apoptosis-Induced Proliferation Pathways as a Proposed Approach to Colorectal Cancer Prevention and Therapy

    PubMed Central

    Bordonaro, Michael; Drago, Eric; Atamna, Wafa; Lazarova, Darina L.

    2014-01-01

    Mutations in the WNT/beta-catenin pathway are present in the majority of all sporadic colorectal cancers (CRCs), and histone deacetylase inhibitors induce apoptosis in CRC cells with such mutations. This apoptosis is counteracted by (1) the signaling heterogeneity of CRC cell populations, and (2) the survival pathways induced by mitogens secreted from apoptotic cells. The phenomena of signaling heterogeneity and apoptosis-induced survival constitute the immediate mechanisms of resistance to histone deacetylase inhibitors, and probably other chemotherapeutic agents. We explored the strategy of augmenting CRC cell death by inhibiting all survival pathways induced by the pro-apoptotic agent LBH589, a histone deacetylase inhibitor: AKT, JAK/STAT, and ERK signaling. The apoptosis-enhancing ability of a cocktail of synthetic inhibitors of proliferation was compared to the effects of the natural product propolis. We utilized colorectal adenoma, drug-sensitive and drug-resistant colorectal carcinoma cells to evaluate the apoptotic potential of the combination treatments. The results suggest that an effective approach to CRC combination therapy is to combine apoptosis-inducing drugs (e.g., histone deacetylase inhibitors, such as LBH589) with agents that suppress all compensatory survival pathways induced during apoptosis (such as the cocktail of inhibitors of apoptosis-associated proliferation). The same paradigm can be applied to a CRC prevention approach, as the apoptotic effect of butyrate, a diet-derived histone deacetylase inhibitor, is augmented by other dietary agents that modulate survival pathways (e.g., propolis and coffee extract). Thus, dietary supplements composed by fermentable fiber, propolis, and coffee extract may effectively counteract neoplastic growth in the colon. PMID:25500581

  19. Survivin is expressed in degenerated nucleus pulposus cells and is involved in proliferation and the prevention of apoptosis in vitro

    PubMed Central

    LIN, YAZHOU; YUE, BIN; XIANG, HONGFEI; LIU, YONG; MA, XUEXIAO; CHEN, BOHUA

    2016-01-01

    Survivin is a unique inhibitor of apoptosis, which is frequently present within degenerated human nucleus pulposus (NP) cells. Survivin has been extensively investigated using proliferation and apoptosis assays in tumor cells; however, studies conducted on survivin in degenerative NP cells remain limited to date. The aim of the present study was to investigate survivin expression and its effects on the proliferation and apoptosis of degenerated NP cells in vitro. The expression levels of survivin in the NP cells of patients (>45 years) with lumbar disc degenerative disease and the NP cells of patients (<25 years) with lumbar vertebra fracture were assessed by reverse transcription-quantitative polymerase chain reaction. The effects on in vitro proliferation and apoptosis were investigated through transfection with a specific small interfering (si)RNA. The results of the present study demonstrated that survivin was expressed in the degenerated NP cells, but was undetectable in normal NP cells at the mRNA level. Survivin suppression following transfection with a specific survivin-siRNA reduced the proliferation rate of NP cells and enhanced sensitization to pro-apoptotic stimuli. Therefore, survivin was shown to be expressed and exhibit an important role in the proliferation and prevention of apoptosis of degenerated NP cells. Studies on survivin in NP cells may aid in increasing the understanding of the complex processes underlying NP cell degeneration, and could provide fundamental information for gene therapy to inhibit this degeneration in vitro. PMID:26648308

  20. Simultaneous deletion of Bax and Bak is required to prevent apoptosis and interstitial fibrosis in obstructive nephropathy.

    PubMed

    Jang, Hee-Seong; Padanilam, Babu J

    2015-09-15

    Proximal tubular injury and apoptosis are key mediators of the development of kidney fibrosis, a hallmark of chronic kidney disease. However, the molecular mechanism by which tubular apoptotic cell death leads to kidney fibrosis is poorly understood. In the present study, we tested the roles of Bcl-2-associated X (Bax) and Bcl-2 antagonist/killer (Bak), two crucial proteins involved in intrinsic apoptotic cell death, in the progression of kidney fibrosis. Mice with proximal tubule-specific Bax deletion, systemic deletion of Bak, and dual deletion of Bax and Bak were subjected to unilateral ureteral obstruction (UUO). Dual deficiency of Bax and Bak inhibited tubular apoptosis and atrophy. Consistent with decreased tubular injury, dual ablation of Bax and Bak suppressed UUO-induced inflammation and kidney fibrosis with decreased tubular cell cycle arrest, expression of fibrogenic and inflammatory cytokines, and oxidative stress in the kidney. Bax or Bak deficiency was insufficient to prevent apoptosis and all other aforementioned malevolent effects, suggesting compensatory mediation by each other in the respective signaling pathways. These data suggest that dual ablation of Bax and Bak in the kidney is required to prevent UUO-induced tubular apoptosis and the consequent kidney inflammation and fibrosis. PMID:26180237

  1. Indigofera oblongifolia Prevents Lead Acetate-Induced Hepatotoxicity, Oxidative Stress, Fibrosis and Apoptosis in Rats

    PubMed Central

    Abdel Moneim, Ahmed E.

    2016-01-01

    The current study was aimed to evaluate the preventive effects of Indigofera oblongifolia leaf extract (IOLE) on lead acetate (PbAc)-induced hepatotoxicity in adult male Wistar rats. PbAc was intraperitoneally injected at a dose of 20 mg/kg body weight for 5 days alone or in combination with the IOLE (100 mg/kg). Liver lead concentration and oxidative stress markers such as lipid peroxidation, hydrogen peroxide, nitric oxide, and glutathione content were investigated in addition to the enzymatic antioxidant activities. PbAc injection caused a significant elevation in the liver function parameters, lead level, lipid peroxidation, hydrogen peroxide, and nitric oxide, with a concomitant decline in the glutathione content compared with the control, accompanied by a significant inhibition of antioxidant enzyme activities. The induction of oxidative stress, lead accumulation, and histological alterations in the liver were successfully minimized by pre-administration of IOLE. In addition, the PbAc group showed increase in the levels of Bax, caspase-3, and matrix metalloproteinase-9 proteins, while the expression of Bcl-2 protein was decreased. Prior administration of IOLE significantly mitigated apoptosis and fibrosis in the liver. Finally, the major components in I. oblongifolia extract were identified as polyphenols, flavonoids, and organic acids using liquid chromatography coupled mass spectroscopy. Thus, the findings of the current study revealed that I. oblongifolia had protective, anti-fibrotic, antioxidant, and anti-apoptotic activities on PbAc-induced hepatotoxicity. The beneficial effects of I. oblongifolia were in part mediated by Nrf2/HO-1 pathway. PMID:27391413

  2. Ursodeoxycholic acid and 4-phenylbutyrate prevent endoplasmic reticulum stress-induced podocyte apoptosis in diabetic nephropathy.

    PubMed

    Cao, Ai-Li; Wang, Li; Chen, Xia; Wang, Yun-Man; Guo, Heng-Jiang; Chu, Shuang; Liu, Cheng; Zhang, Xue-Mei; Peng, Wen

    2016-06-01

    Endoplasmic reticulum (ER) stress, resulting from the accumulation of misfolded and/or unfolded proteins in ER membranes, is involved in the pathogenesis of diabetic nephropathy (DN). The aim of this study was to investigate the role of ER stress inhibitors ursodeoxycholic acid (UDCA) and 4-phenylbutyrate (4-PBA) in the treatment of DN in db/db mice. Findings have revealed that diabetic db/db mice were more hyperglycemic than their non-diabetic controls, and exhibited a marked increase in body weight, water intake, urine volume, fasting plasma glucose, systolic blood pressure, glucose and insulin tolerance. UDCA (40 mg/kg/day) or 4-PBA (100 mg/kg/day) treatment for 12 weeks resulted in an improvement in these biochemical and physical parameters. Moreover, UDCA or 4-PBA intervention markedly decreased urinary albuminuria and attenuated mesangial expansion in diabetic db/db mice, compared with db/db mice treated with vehicle. These beneficial effects of UDCA or 4-PBA on DN were associated with the inhibition of ER stress, as evidenced by the decreased expression of BiP, phospho-IRE1α, phospho-eIF2α, CHOP, ATF-6 and spliced X-box binding protein-1 in vitro and in vivo. UDCA or 4-PBA prevented hyperglycemia-induced or high glucose (HG)-induced apoptosis in podocytes in vivo and in vitro via the inhibition of caspase-3 and caspase-12 activation. Autophagy deficiency was also seen in glomeruli in diabetic mice and HG-incubated podocytes, exhibiting decreased expression of LC3B and Beclin-1, which could be restored by UDCA or 4-PBA treatment. Taken together, our results have revealed an important role of ER stress in the development of DN, and UDCA or 4-PBA treatment may be a potential novel therapeutic approach for the treatment of DN. PMID:26999661

  3. Curcumin prevents the non-alcoholic fatty hepatitis via mitochondria protection and apoptosis reduction

    PubMed Central

    Wang, Long; lv, Yisong; Yao, Huixiang; Yin, Li; Shang, Jianhui

    2015-01-01

    Background: Non-alcoholic fatty hepatitis (NASH) is highly prevalent, mitochondria damage is the main pathophysiological characteristic of NASH. However, treatment for mitochondria damage is rarely reported. Methods: NASH model was established in rats, the protective effects of curcumin were evaluated by histological observation; structure and function assessments of mitochondria; and apoptotic genes expression. Results: NASH rats treated with curcumin displayed relatively slight liver damage when compared with NASH livers. The average mitochondrial length and width of NASH (12.0 ± 3.2 and 5.1 ± 1.1 micrometers) were significantly longer than that of normal (6.2 ± 2.1 and 2.1 ± 1.5 micrometers) and NASH treated with curcumin (7.4 ± 1.2 and 3.2 ± 1.5 micrometers) rats. The average malondialdehyde (MDA) and 4-hydroxy nonyl alcohol (HNE) levels in liver homogenates of NASH rats (4.23 ± 0.22 and 19.23 ± 2.3 nmol/Ml) were significantly higher than these in normal (1.32 ± 0.12 and 3.52 ± 0.43 nmol/mL) and NASH treated with curcumin (1.74 ± 0.11 and 4.66 ± 0.99 nmol/mL) rats. The expression levels of CytC, Casp3 and Casp8 of the NASH livers were significantly higher than normal and NASH treated with curcumin rats livers. Conclusion: Our data demonstrated that curcumin prevents the NASH by mitochondria protection and apoptosis reduction and provided a possible novel treatment for NASH. PMID:26617882

  4. Neuronal apoptosis in rats is accompanied by rapid impairment of cellular respiration and is prevented by scavengers of reactive oxygen species.

    PubMed

    Atlante, A; Gagliardi, S; Marra, E; Calissano, P

    1998-04-10

    Apoptosis of cerebellar granule cells induced by potassium withdrawal is accompanied by a very rapid decrease in both cell and mitochondrial respiration supported by glucose and succinate, respectively. The respiratory control ratio, which is an index of oxidative phosphorylation and therefore reflects the ability of mitochondria to produce ATP, is reduced by 50% within the first 2 h after the beginning of apoptosis, insulin-like growth factor I (IGF-I), actinomicin D or cycloheximide, previously reported to inhibit apoptosis, fully prevent the impairment of cellular respiration while scavengers of reactive oxygen species partially inhibit apoptosis and restore cellular respiration. PMID:9605472

  5. Protein Isoaspartate Methyltransferase Prevents Apoptosis Induced by Oxidative Stress in Endothelial Cells: Role of Bcl-Xl Deamidation and Methylation

    PubMed Central

    Cimmino, Amelia; Capasso, Rosanna; Muller, Fabbri; Sambri, Irene; Masella, Lucia; Raimo, Marianna; De Bonis, Maria Luigia; D'Angelo, Stefania; Zappia, Vincenzo; Galletti, Patrizia; Ingrosso, Diego

    2008-01-01

    Background Natural proteins undergo in vivo spontaneous post-biosynthetic deamidation of specific asparagine residues with isoaspartyl formation. Deamidated-isomerized molecules are both structurally and functionally altered. The enzyme isoaspartyl protein carboxyl-O-methyltransferase (PCMT; EC 2.1.1.77) has peculiar substrate specificity towards these deamidated proteins. It catalyzes methyl esterification of the free α-carboxyl group at the isoaspartyl site, thus initiating the repair of these abnormal proteins through the conversion of the isopeptide bond into a normal α-peptide bond. Deamidation occurs slowly during cellular and molecular aging, being accelerated by physical-chemical stresses brought to the living cells. Previous evidence supports a role of protein deamidation in the acquisition of susceptibility to apoptosis. Aim of this work was to shed a light on the role of PCMT in apoptosis clarifying the relevant mechanism(s). Methodology/Principal Findings Endothelial cells transiently transfected with various constructs of PCMT, i.e. overexpressing wild type PCMT or negative dominants, were used to investigate the role of protein methylation during apoptosis induced by oxidative stress (H2O2; 0.1–0.5 mM range). Results show that A) Cells overexpressing “wild type” human PCMT were resistant to apoptosis, whereas overexpression of antisense PCMT induces high sensitivity to apoptosis even at low H2O2 concentrations. B) PCMT protective effect is specifically due to its methyltransferase activity rather than to any other non-enzymatic interactions. In fact negative dominants, overexpressing PCMT mutants devoid of catalytic activity do not prevent apoptosis. C) Cells transfected with antisense PCMT, or overexpressing a PCMT mutant, accumulate isoaspartyl-containing damaged proteins upon H2O2 treatment. Proteomics allowed the identification of proteins, which are both PCMT substrates and apoptosis effectors, whose deamidation occurs under oxidative

  6. Prevention of apoptosis averts glomerular tubular disconnection and podocyte loss in proteinuric kidney disease.

    PubMed

    Burlaka, Ievgeniia; Nilsson, Linnéa M; Scott, Lena; Holtbäck, Ulla; Eklöf, Ann-Christine; Fogo, Agnes B; Brismar, Hjalmar; Aperia, Anita

    2016-07-01

    There is a great need for treatment that arrests progression of chronic kidney disease. Increased albumin in urine leads to apoptosis and fibrosis of podocytes and tubular cells and is a major cause of functional deterioration. There have been many attempts to target fibrosis, but because of the lack of appropriate agents, few have targeted apoptosis. Our group has described an ouabain-activated Na,K-ATPase/IP3R signalosome, which protects from apoptosis. Here we show that albumin uptake in primary rat renal epithelial cells is accompanied by a time- and dose-dependent mitochondrial accumulation of the apoptotic factor Bax, down-regulation of the antiapoptotic factor Bcl-xL and mitochondrial membrane depolarization. Ouabain opposes these effects and protects from apoptosis in albumin-exposed proximal tubule cells and podocytes. The efficacy of ouabain as an antiapoptotic and kidney-protective therapeutic tool was then tested in rats with passive Heymann nephritis, a model of proteinuric chronic kidney disease. Chronic ouabain treatment preserved renal function, protected from renal cortical apoptosis, up-regulated Bax, down-regulated Bcl-xL, and rescued from glomerular tubular disconnection and podocyte loss. Thus we have identified a novel clinically feasible therapeutic tool, which has the potential to protect from apoptosis and rescue from loss of functional tissue in chronic proteinuric kidney disease. PMID:27217195

  7. Chloride channel protein 2 prevents glutamate-induced apoptosis in retinal ganglion cells

    PubMed Central

    Bi, Miao-Miao; Hong, Sen; Ma, Ling-Jun; Zhou, Hong-Yan; Lu, Jia; Zhao, Jing; Zheng, Ya-Juan

    2016-01-01

    Objective(s): The purpose of this study was to investigate the role of chloride channel protein 2 (ClC-2) in glutamate-induced apoptosis in the retinal ganglion cell line (RGC-5). Materials and Methods: RGC-5 cells were treated with 1 mM glutamate for 24 hr. The expression of ClC-2, Bax, and Bcl-2 was detected by western blot analysis. Cell survival and apoptosis were measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and flow cytometry assays, respectively. Caspase-3 and -9 activities were determined by a colorimetric assay. The roles of ClC-2 in glutamate-induced apoptosis were examined by using ClC-2 complementary deoxyribonucleic acid (cDNA) and small inference ribonucleic acid (RNA) transfection technology. Results: Overexpression of ClC-2 in RGC-5 cells significantly decreased glutamate-induced apoptosis and increased cell viability, whereas silencing of ClC-2 with short hairpin (sh) RNA produced opposite effects. ClC-2 overexpression increased the expression of Bcl-2, decreased the expression of Bax, and decreased caspase-3 and -9 activation in RGC-5 cells treated with glutamate, but silencing of ClC-2 produced opposite effects. Conclusion: Our data suggest that ClC-2 chloride channels might play a protective role in glutamate-induced apoptosis in retinal ganglion cells via the mitochondria-dependent apoptosis pathway.

  8. Apoptosis-promoted tumorigenesis: gamma-irradiation-induced thymic lymphomagenesis requires Puma-driven leukocyte death.

    PubMed

    Michalak, Ewa M; Vandenberg, Cassandra J; Delbridge, Alex R D; Wu, Li; Scott, Clare L; Adams, Jerry M; Strasser, Andreas

    2010-08-01

    Although tumor development requires impaired apoptosis, we describe a novel paradigm of apoptosis-dependent tumorigenesis. Because DNA damage triggers apoptosis through p53-mediated induction of BH3-only proteins Puma and Noxa, we explored their roles in gamma-radiation-induced thymic lymphomagenesis. Surprisingly, whereas Noxa loss accelerated it, Puma loss ablated tumorigenesis. Tumor suppression by Puma deficiency reflected its protection of leukocytes from gamma-irradiation-induced death, because their glucocorticoid-mediated decimation in Puma-deficient mice activated cycling of stem/progenitor cells and restored thymic lymphomagenesis. Our demonstration that cycles of cell attrition and repopulation by stem/progenitor cells can drive tumorigenesis has parallels in human cancers, such as therapy-induced malignancies. PMID:20679396

  9. Bcl6a function is required during optic cup formation to prevent p53-dependent apoptosis and colobomata

    PubMed Central

    Lee, Jiwoon; Lee, Bum-Kyu; Gross, Jeffrey M.

    2013-01-01

    Mutations in BCOR (Bcl6 corepressor) are found in patients with oculo-facio-cardio-dental (OFCD) syndrome, a congenital disorder affecting visual system development, and loss-of-function studies in zebrafish and Xenopus demonstrate a role for Bcor during normal optic cup development in preventing colobomata. The mechanism whereby BCOR functions during eye development to prevent colobomata is not known, but in other contexts it serves as a transcriptional corepressor that potentiates transcriptional repression by B cell leukemia/lymphoma 6 (BCL6). Here, we have explored the function of the zebrafish ortholog of Bcl6, Bcl6a, during eye development, and our results demonstrate that Bcl6a, like Bcor, is required to prevent colobomata during optic cup formation. Our data demonstrate that Bcl6a acts downstream of Vax1 and Vax2, known regulators of ventral optic cup formation and choroid fissure closure, and that bcl6a is a direct target of Vax2. Together, this regulatory network functions to repress p53 expression and thereby suppress apoptosis in the developing optic cup. Furthermore, our data demonstrate that Bcl6a functions cooperatively with Bcor, Rnf2 and Hdac1 in a common gene regulatory network that acts to repress p53 and prevent colobomata. Together, these data support a model in which p53-dependent apoptosis needs to be tightly regulated for normal optic cup formation and that Bcl6a, Bcor, Rnf2 and Hdac1 activities mediate this regulation. PMID:23669349

  10. Lycopene supplementation prevents reactive oxygen species mediated apoptosis in Sertoli cells of adult albino rats exposed to polychlorinated biphenyls

    PubMed Central

    Krishnamoorthy, Gunasekaran; Selvakumar, Kandaswamy; Venkataraman, Prabhu; Elumalai, Perumal

    2013-01-01

    Sertoli cell proliferation is attenuated before attaining puberty and the number is fixed in adult testes. Sertoli cells determine both testis size and daily sperm production by providing physical and metabolic support to spermatogenic cells. Polychlorinated biphenyls (PCBs) exposure disrupts functions of Sertoli cells causing infertility with decreased sperm count. On the other hand, lycopene is improving sperm count and motility by reducing oxidative stress in humans and animals. Hence we hypothesized that PCBs-induced infertility might be due to Sertoli cell apoptosis mediated by oxidative stress and lycopene might prevent PCBs-induced apoptosis by acting against oxidative stress. To test this hypothesis, animals were treated with vehicle control, lycopene, PCBs and PCBs + lycopene for 30 days. After the experimental period, the testes and cauda epididymidis were removed for isolation of Sertoli cells and sperm, respectively. We observed increased levels of oxidative stress markers (H2O2 and LPO) levels, increased expression of apoptotic molecules (caspase-8, Bad, Bid, Bax, cytochrome C and caspase-3), decreased anti-apoptotic (Bcl2) molecule and elevated apoptotic marker activity (caspase-3) in Sertoli cells of PCBs-exposed animals. These results were associated with decreased sperm count and motility in PCBs exposed animals. On the other hand, lycopene prevented the elevation of Sertoli cellular apoptotic parameters and prevented the reduction of sperm parameters (count and motility). The data confirmed that lycopene as an antioxidant scavenged reactive oxygen substances, prevented apoptosis, maintained normal function in Sertoli cells and helped to provide physical and metabolic support for sperm production, thereby treating infertility in men. PMID:24179434

  11. Inhibition of polyamine oxidase prevented cyclin-dependent kinase inhibitor-induced apoptosis in HCT 116 colon carcinoma cells.

    PubMed

    Gürkan, Ajda Coker; Arisan, Elif Damla; Obakan, Pinar; Palavan-Ünsal, Narçin

    2013-12-01

    Roscovitine and purvalanol are novel cyclin-dependent kinase (CDK) inhibitors that prevent cell proliferation and induce apoptotic cell death in various cancer cell lines. Although a number of studies have demonstrated the potential apoptotic role of roscovitine, there is limited data about the therapeutic efficiency of purvalanol on cancer cells. The natural polyamines (PAs) putrescine, spermidine, and spermine have essential roles in the regulation of cell differentiation, growth, and proliferation, and increased levels of these compounds have been associated with cancer progression. Recently, depletion of intracellular PA levels because of modulation of PA catabolic enzymes was shown to be an indicator of the efficacy of chemotherapeutic agents. In this study, our aim was to investigate the potential role of PA catabolic enzymes in CDK inhibitor-induced apoptosis in HCT 116 colon carcinoma cells. Exposure of cells to roscovitine or purvalanol decreased cell viability in a dose- and time-dependent manner. The selected concentrations of roscovitine and purvalanol inhibited cell viability by 50 % compared with control cells and induced apoptosis by activating the mitochondria-mediated pathway in a caspase-dependent manner. However, the apoptotic effect of purvalanol was stronger than that of roscovitine in HCT 116 cells. In addition, we found that CDK inhibitors decreased PA levels and significantly upregulated expression of key PA catabolic enzymes such as polyamine oxidase (PAO) and spermine oxidase (SMO). MDL-72,527, a specific inhibitor of PAO and SMO, decreased apoptotic potential of CDK inhibitors on HCT 116 cells. Moreover, transient silencing of PAO was also reduced prevented CDK inhibitor-induced apoptosis in HCT 116 cells. We conclude that the PA catabolic pathway, especially PAO, is a critical target for understanding the molecular mechanism of CDK inhibitor-induced apoptosis. PMID:23892915

  12. Hydrogen sulfide prevents Abeta-induced neuronal apoptosis by attenuating mitochondrial translocation of PTEN.

    PubMed

    Cui, Weigang; Zhang, Yinghua; Yang, Chenxi; Sun, Yiyuan; Zhang, Min; Wang, Songtao

    2016-06-14

    Neuronal cell apoptosis is an important pathological change in Alzheimer's disease (AD). Hydrogen sulfide (H(2)S) is known to be a novel gaseous signaling molecule and a cytoprotectant in many diseases including AD. However, the molecular mechanism of the antiapoptosis activity of H(2)S in AD is not yet fully understood. The aim of the present study is to evaluate the inhibitory effects of H(2)S on Abeta (Aβ)-induced apoptosis and the molecular mechanisms underlying primary neuron cells. Our results showed that sodium hydrosulfide (NaHS), a donor of H(2)S, significantly ameliorated Aβ-induced cell apoptosis. NaHS also reversed the Aβ-induced translocation of the phosphatase and tensin homologs deleted on chromosome 10 (PTEN) from the cytosol to the mitochondria. Furthermore, H(2)S increased the level of p-AKT/AKT significantly. Interestingly, the antiapoptosis effects of H(2)S were blocked down by specific PI3K/AKT inhibitor wortmannin. In conclusion, these data indicate that H(2)S inhibits Aβ-induced neuronal apoptosis by attenuating mitochondrial translocation of PTEN and that activation of PI3K/AKT signaling pathway plays a critical role in H(2)S-mediated neuronal protection. Our findings provide a novel route into the molecular mechanisms of neuronal apoptosis in AD. PMID:27026591

  13. Prevention of oxidative stress-induced apoptosis of C2C12 myoblasts by a Cichorium intybus root extract.

    PubMed

    Lee, Yong-Hyeon; Kim, Dae-Hyun; Kim, Yoon Suk; Kim, Tack-Joong

    2013-01-01

    Cell injury associated with reactive oxygen species (ROS) has been reported in various muscular disorders. We found that a Cichorium intybus (Cii) extract reduced H(2)O(2)-induced viability loss in C2C12 myoblasts, inhibited oxidative stress-induced apoptosis and increased intracellular heat shock protein 70 (Hsp 70) expression. Cii also inhibited the level of intracellular ceramide. These results indicate that Cii may prevent skeletal muscle atrophy by inducing the expression of Hsp 70 and inhibiting the level of ceramide. PMID:23391909

  14. Nrf2 Protein Up-regulates Antiapoptotic Protein Bcl-2 and Prevents Cellular Apoptosis*

    PubMed Central

    Niture, Suryakant K.; Jaiswal, Anil K.

    2012-01-01

    Nuclear transcription factor Nrf2 regulates the expression and coordinated induction of a battery of genes encoding cytoprotective and drug transporter proteins in response to chemical and radiation stress. This leads to reduced apoptosis, enhanced cell survival, and increased drug resistance. In this study, we investigated the role of Nrf2 in up-regulation of antiapoptotic protein Bcl-2 and its contribution to stress-induced apoptosis and cell survival. Exposure of mouse hepatoma (Hepa-1) and human hepatoblastoma (HepG2) cells to antioxidant tert-butylhydroquinone led to induction of Bcl-2. Mutagenesis and transfection assays identified an antioxidant response element between nucleotides −3148 and −3140 on the reverse strand of the Bcl-2 gene promoter that was essential for activation of Bcl-2 gene expression. Band/supershift and ChIP assays demonstrated binding of Nrf2 to Bcl-2 antioxidant response element. Alterations in Nrf2 led to altered Bcl-2 induction and cellular apoptosis. Moreover, dysfunctional/mutant inhibitor of Nrf2 (INrf2) in human lung cancer cells failed to degrade Nrf2, resulting in an increased Bcl-2 level and decreased etoposide- and UV/γ radiation-mediated DNA fragmentation. In addition, siRNA-mediated down-regulation of Nrf2 also led to decreased apoptosis and increased cell survival. Furthermore, the specific knockdown of Bcl-2 in Nrf2-activated tumor cells led to increased etoposide-induced apoptosis and decreased cell survival and growth/proliferation. These data provide the first evidence of Nrf2 in control of Bcl-2 expression and apoptotic cell death with implications in antioxidant protection, survival of cancer cells, and drug resistance. PMID:22275372

  15. INHIBITION OF CDK9 PREVENTS MECHANICAL INJURY-INDUCED INFLAMMATION, APOPTOSIS AND MATRIX DEGRADATION IN CARTILAGE EXPLANTS

    PubMed Central

    Hu, Z.; Yik, J.H.N.; Cissell, D.D.; Michelier, P.V.; Athanasiou, K.A.; Haudenschild, D.R.

    2016-01-01

    Joint injury often leads to post-traumatic osteoarthritis (PTOA). Acute injury responses to trauma induce production of pro-inflammatory cytokines and catabolic enzymes, which promote chondrocyte apoptosis and degrade cartilage to potentiate PTOA development. Recent studies show that the rate-limiting step for transcriptional activation of injury response genes is controlled by cyclin-dependent kinase 9 (CDK9), and thus it is an attractive target for limiting the injury response. Here, we determined the effects of CDK9 inhibition in suppressing the injury response in mechanically-injured cartilage explants. Bovine cartilage explants were injured by a single compressive load of 30 % strain at 100 %/s, and then treated with the CDK9 inhibitor Flavopiridol. To assess acute injury responses, we measured the mRNA expression of pro-inflammatory cytokines, catabolic enzymes, and apoptotic genes by RT-PCR, and chondrocyte viability and apoptosis by TUNEL staining. For long-term outcome, cartilage matrix degradation was assessed by soluble glycosaminoglycan release, and by determining the mechanical properties with instantaneous and relaxation moduli. Our data showed CDK9 inhibitor markedly reduced injury-induced inflammatory cytokine and catabolic gene expression. CDK9 inhibitor also attenuated chondrocyte apoptosis and reduced cartilage matrix degradation. Lastly, the mechanical properties of the injured explants were preserved by CDK9 inhibitor. Our results provide a temporal profile connecting the chain of events from mechanical impact, acute injury responses, to the subsequent induction of chondrocyte apoptosis and cartilage matrix deterioration. Thus, CDK9 is a potential disease-modifying agent for injury response after knee trauma to prevent or delay PTOA development. PMID:26859911

  16. Selenoprotein S Is Highly Expressed in the Blood Vessels and Prevents Vascular Smooth Muscle Cells From Apoptosis.

    PubMed

    Ye, Yali; Fu, Fen; Li, Xiaoming; Yang, Jie; Liu, Hongmei

    2016-01-01

    Atherosclerosis and related cardiovascular diseases (CVD) represent one of the greatest threats to human health worldwide. The protection of vascular smooth muscle cells (VSMCs) from apoptosis in the plaque has become an important therapeutic target for atherosclerotic plaque stabilization. A significant association of selenoprotein S (SelS) gene polymorphism with atherosclerotic CVD has been reported in epidemiologic studies, but the underlying mechanism remains unknown. In this paper, SelS expression in the thoracic aorta and its role in the protection of VSMCs from apoptosis have been studied. Western blot analysis showed that SelS was highly expressed in rat thoracic aorta. SelS gene silence by small interference RNA (siRNA) rendered VSMCs more sensitive to hydrogen peroxide- or tunicamycin- induced injury and apoptosis, as determined by MTT assay, Hoechst staining, and annexin V/propidium iodide staining. SelS silence aggravated hydrogen peroxide-induced oxidative stress and phosphorylation of p38 MAPK and c-Jun N-terminal kinase (JNK) in VSMCs. Furthermore, SelS silence enhanced endoplasmic reticulum (ER) stress induced by hydrogen peroxide or tunicamycin, as showed by the increased protein levels of ER chaperone 78 kDa glucose-regulated protein (GRP78), ER stress transducer phosphorylated protein kinase RNA like ER kinase (PERK), and the proapoptotic transcription factor C/EBP homologous protein (CHOP). In conclusion, the present study suggested that SelS highly expressed in the blood vessel might protect VSMCs from apoptosis by inhibiting oxidative stress and ER stress. Our finding provided mechanistic insights for the potential preventive role of SelS in atherosclerotic CVD. PMID:26058460

  17. The New Biology of Estrogen-induced Apoptosis Applied to Treat and Prevent Breast Cancer

    PubMed Central

    Jordan, V Craig

    2014-01-01

    The successful use of high dose synthetic estrogens to treat post-menopausal metastatic breast cancer, is the first effective “chemical therapy” proven in clinical trial to treat any cancer. This review documents the clinical use of estrogen for breast cancer treatment or estrogen replacement therapy (ERT) for postmenopausal hysterectomized women which can either result in breast cancer cell growth or breast cancer regression. This has remained a paradox since the 1950s until the discovery of the new biology of estrogen induced apoptosis at the end of the 20th century. The key to triggering apoptosis with estrogen is the selection of breast cancer cell populations that are resistant to long term estrogen deprivation. However, through trial and error estrogen independent growth occurs. At the cellular level, estrogen induced apoptosis is dependent upon the presence of the estrogen receptor (ER) which can be blocked by non-steroidal or steroidal anti-estrogens. The shape of an estrogenic ligand programs the conformation of the ER complex which in turn can modulate estrogen induced apoptosis: class I planar estrogens (eg: estradiol) trigger apoptosis after 24 hours whereas class II angular estrogens (eg: bisphenol triphenylethylene) delay the process until after 72 hours. This contrasts with paclitaxel that causes G2 blockade with immediate apoptosis. The process is complete within 24 hours. Estrogen induced apoptosis is modulated by glucocorticoids and cSrc inhibitors but the target mechanism for estrogen action is genomic and not through a non-genomic pathway. The process is step wise through the creation of endoplasmic reticulum stress and, inflammatory responses that then initiate an unfolded protein response. This in turn initiates apoptosis through the intrinsic pathway (mitochondrial) with subsequent recruitment of the extrinsic pathway (death receptor) to complete the process. The symmetry of the clinical and laboratory studies now permits the creation of

  18. Preventing Friction Induced Chondrocyte Apoptosis: A Comparison of Human Synovial Fluid and Hylan G-F 20

    PubMed Central

    Waller, Kimberly A; Zhang, Ling X; Fleming, Braden C; Jay, Gregory D

    2013-01-01

    Objectives Symptomatic osteoarthritis (OA) is a common painful disease with limited treatment options. A rising number of OA patients have been treated with intraarticular injections of hyaluronic acid, including the high molecular weight hylan G-F 20, which is injected following arthrocentesis. This study investigated the effectiveness of hylan G-F 20 to lower coefficient of friction (COF) and prevent chondrocyte apoptosis in vitro. Methods A disc-on-disc bovine cartilage bearing was used to measure the static and kinetic COF when lubricated with hylan G-F 20, human synovial fluid (HSF) and phosphate buffered saline (PBS). Following friction testing, we stained paraffin embedded sections of these cartilage bearings for activated caspase-3, a marker of apoptosis. Results Bearings lubricated with hylan G-F 20 had kinetic COF values that were similar to bearings lubricated with PBS, but significantly higher than those lubricated with HSF. There were no significant differences in static COF values in bearings lubricated with hylan G-F 20 as compared to PBS or HSF. However, bearings lubricated with HSF had a significantly lower static COF values compared to bearings lubricated with PBS. The mean percentage of caspase-3 positive chondrocytes in the superficial and upper intermediate zones of bearings lubricated with hylan G-F 20 were significantly higher when compared to bearings lubricated with HSF or unloaded controls, but significantly lower than those lubricated with PBS. Conclusion These findings indicate that joint lubrication may prevent chondrocyte apoptosis by lowering the COF. Furthermore, removal of synovial fluid prior to hylan G-F 20 injection may be detrimental to cartilage health. PMID:22660808

  19. Nicorandil Prevents Right Ventricular Remodeling by Inhibiting Apoptosis and Lowering Pressure Overload in Rats with Pulmonary Arterial Hypertension

    PubMed Central

    Yu, Yan-Zhe; Wang, Hui; Bi, Li-Qing; Xie, Wei-Ping; Wang, Hong

    2012-01-01

    Background Most of the deaths among patients with severe pulmonary arterial hypertension (PAH) are caused by progressive right ventricular (RV) pathological remodeling, dysfunction, and failure. Nicorandil can inhibit the development of PAH by reducing pulmonary artery pressure and RV hypertrophy. However, whether nicorandil can inhibit apoptosis in RV cardiomyocytes and prevent RV remodeling has been unclear. Methodology/Principal Findings RV remodeling was induced in rats by intraperitoneal injection of monocrotaline (MCT). RV systolic pressure (RVSP) was measured at the end of each week after MCT injection. Blood samples were drawn for brain natriuretic peptide (BNP) ELISA analysis. The hearts were excised for histopathological, ultrastructural, immunohistochemical, and Western blotting analyses. The MCT-injected rats exhibited greater mortality and less weight gain and showed significantly increased RVSP and RV hypertrophy during the second week. These worsened during the third week. MCT injection for three weeks caused pathological RV remodeling, characterized by hypertrophy, fibrosis, dysfunction, and RV mitochondrial impairment, as indicated by increased levels of apoptosis. Nicorandil improved survival, weight gain, and RV function, ameliorated RV pressure overload, and prevented maladaptive RV remodeling in PAH rats. Nicorandil also reduced the number of apoptotic cardiomyocytes, with a concomitant increase in Bcl-2/Bax ratio. 5-hydroxydecanoate (5-HD) reversed these beneficial effects of nicorandil in MCT-injected rats. Conclusions/Significance Nicorandil inhibits PAH-induced RV remodeling in rats not only by reducing RV pressure overload but also by inhibiting apoptosis in cardiomyocytes through the activation of mitochondrial ATP-sensitive K+ (mitoKATP) channels. The use of a mitoKATP channel opener such as nicorandil for PAH-associated RV remodeling and dysfunction may represent a new therapeutic strategy for the amelioration of RV remodeling during

  20. Azelnidipine prevents cardiac dysfunction in streptozotocin-diabetic rats by reducing intracellular calcium accumulation, oxidative stress and apoptosis

    PubMed Central

    2011-01-01

    Background Numerous evidences suggest that diabetic heart is characterized by compromised ventricular contraction and prolonged relaxation attributable to multiple causative factors including calcium accumulation, oxidative stress and apoptosis. Therapeutic interventions to prevent calcium accumulation and oxidative stress could be therefore helpful in improving the cardiac function under diabetic condition. Methods This study was designed to examine the effect of long-acting calcium channel blocker (CCB), Azelnidipine (AZL) on contractile dysfunction, intracellular calcium (Ca2+) cycling proteins, stress-activated signaling molecules and apoptosis on cardiomyocytes in diabetes. Adult male Wistar rats were made diabetic by a single intraperitoneal (IP) injection of streptozotocin (STZ). Contractile functions were traced from live diabetic rats to isolated individual cardiomyocytes including peak shortening (PS), time-to-PS (TPS), time-to-relengthening (TR90), maximal velocity of shortening/relengthening (± dL/dt) and intracellular Ca2+ fluorescence. Results Diabetic heart showed significantly depressed PS, ± dL/dt, prolonged TPS, TR90 and intracellular Ca2+ clearing and showed an elevated resting intracellular Ca2+. AZL itself exhibited little effect on myocyte mechanics but it significantly alleviated STZ-induced myocyte contractile dysfunction. Diabetes increased the levels of superoxide, enhanced expression of the cardiac damage markers like troponin I, p67phox NADPH oxidase subunit, restored the levels of the mitochondrial superoxide dismutase (Mn-SOD), calcium regulatory proteins RyR2 and SERCA2a, and suppressed the levels of the anti-apoptotic Bcl-2 protein. All of these STZ-induced alterations were reconciled by AZL treatment. Conclusion Collectively, the data suggest beneficial effect of AZL in diabetic cardiomyopathy via altering intracellular Ca2+ handling proteins and preventing apoptosis by its antioxidant property. PMID:22054019

  1. Sodium fluoride induces apoptosis in mouse embryonic stem cells through ROS-dependent and caspase- and JNK-mediated pathways

    SciTech Connect

    Nguyen Ngoc, Tam Dan; Son, Young-Ok; Lim, Shin-Saeng; Shi, Xianglin; Kim, Jong-Ghee; Heo, Jung Sun; Choe, Youngji; Jeon, Young-Mi; Lee, Jeong-Chae

    2012-03-15

    Sodium fluoride (NaF) is used as a source of fluoride ions in diverse applications. Fluoride salt is an effective prophylactic for dental caries and is an essential element required for bone health. However, fluoride is known to cause cytotoxicity in a concentration-dependent manner. Further, no information is available on the effects of NaF on mouse embryonic stem cells (mESCs). We investigated the mode of cell death induced by NaF and the mechanisms involved. NaF treatment greater than 1 mM reduced viability and DNA synthesis in mESCs and induced cell cycle arrest in the G{sub 2}/M phase. The addition of NaF induced cell death mainly by apoptosis rather than necrosis. Catalase (CAT) treatment significantly inhibited the NaF-mediated cell death and also suppressed the NaF-mediated increase in phospho-c-Jun N-terminal kinase (p-JNK) levels. Pre-treatment with SP600125 or z-VAD-fmk significantly attenuated the NaF-mediated reduction in cell viability. In contrast, intracellular free calcium chelator, but not of sodium or calcium ion channel blockers, facilitated NaF-induced toxicity in the cells. A JNK specific inhibitor (SP600125) prevented the NaF-induced increase in growth arrest and the DNA damage-inducible protein 45α. Further, NaF-mediated loss of mitochondrial membrane potential was apparently inhibited by pifithrin-α or CAT inhibitor. These findings suggest that NaF affects viability of mESCs in a concentration-dependent manner, where more than 1 mM NaF causes apoptosis through hydroxyl radical-dependent and caspase- and JNK-mediated pathways. -- Highlights: ► The mode of NaF-induced cell death and the mechanisms involved were examined. ► NaF induced mainly apoptotic death of mouse embryonic stem cells (mESCs). ► NaF induced mitochondrial-mediated and caspase-dependent apoptosis. ► JNK- and p53-mediated pathways are involved in NaF-mediated apoptosis in the cells. ► ROS are the up-stream effector in NaF-mediated activation of JNK and p53 in mESCs.

  2. Long-term caloric restriction in mice may prevent age-related learning impairment via suppression of apoptosis.

    PubMed

    Ma, Lina; Wang, Rong; Dong, Wen; Li, Yun; Xu, Baolei; Zhang, Jingshuang; Zhao, Zhiwei

    2016-12-15

    Caloric restriction (CR) is the most reliable intervention to extend lifespan and prevent age-related disorders in various species from yeast to rodents. However, the underlying mechanisms have not yet been clearly defined. Therefore, we aimed to identify the underlying mechanisms of long-term CR on age-related learning impairment in C57/BL mice. Thirty six-week-old male C57/BL mice were randomly divided into three groups: normal control group (NC group, n=10), high energy group (HE group, n=10), and CR group (n=10). After 10 months, the Morris water maze test was performed to monitor learning abilities. Western blotting, immunohistochemistry and real-time polymerase chain reaction were used to monitor changes in protein and mRNA levels associated with apoptosis-related proteins in the hippocampus. The average escape latency was lower in the CR group compared with the NC group, and the average time taken to first cross the platform in the CR group was significantly shorter than the HE group. Both Bcl-2 protein and mRNA expression levels in the CR group were significantly higher than those of the NC group and HE group. The expression of Bax, Caspase-3 and PARP protein in the CR group was significantly lower than the NC group. Our findings demonstrate that long-term CR may prevent age-related learning impairments via suppressing apoptosis in mice. PMID:27452805

  3. Prevention of neonatal oxygen-induced brain damage by reduction of intrinsic apoptosis

    PubMed Central

    Sifringer, M; Bendix, I; Börner, C; Endesfelder, S; von Haefen, C; Kalb, A; Holifanjaniaina, S; Prager, S; Schlager, G W; Keller, M; Jacotot, E; Felderhoff-Mueser, U

    2012-01-01

    Within the last decade, it became clear that oxygen contributes to the pathogenesis of neonatal brain damage, leading to neurocognitive impairment of prematurely born infants in later life. Recently, we have identified a critical role for receptor-mediated neuronal apoptosis in the immature rodent brain. However, the contribution of the intrinsic apoptotic pathway accompanied by activation of caspase-2 under hyperoxic conditions in the neonatal brain still remains elusive. Inhibition of caspases appears a promising strategy for neuroprotection. In order to assess the influence of specific caspases on the developing brain, we applied a recently developed pentapeptide-based group II caspase inhibitor (5-(2,6-difluoro-phenoxy)-3(R,S)-(2(S)-(2(S)-(3-methoxycarbonyl-2(S)-(3-methyl-2(S)-((quinoline-2-carbonyl)-amino)-butyrylamino)propionylamino)3-methylbutyrylamino)propionylamino)-4-oxo-pentanoic acid methyl ester; TRP601). Here, we report that elevated oxygen (hyperoxia) triggers a marked increase in active caspase-2 expression, resulting in an initiation of the intrinsic apoptotic pathway with upregulation of key proteins, namely, cytochrome c, apoptosis protease-activating factor-1, and the caspase-independent protein apoptosis-inducing factor, whereas BH3-interacting domain death agonist and the anti-apoptotic protein B-cell lymphoma-2 are downregulated. These results coincide with an upregulation of caspase-3 activity and marked neurodegeneration. However, single treatment with TRP601 at the beginning of hyperoxia reversed the detrimental effects in this model. Hyperoxia-mediated neurodegeneration is supported by intrinsic apoptosis, suggesting that the development of highly selective caspase inhibitors will represent a potential useful therapeutic strategy in prematurely born infants. PMID:22237207

  4. Cordyceps sinensis prevents apoptosis in mouse liver with D-galactosamine/lipopolysaccharide-induced fulminant hepatic failure.

    PubMed

    Cheng, Yu-Jung; Cheng, Shiu-Min; Teng, Yi-Hsien; Shyu, Woei-Cherng; Chen, Hsiu-Ling; Lee, Shin-Da

    2014-01-01

    Cordyceps sinensis (C. sinensis) has long been considered to be an herbal medicine and has been used in the treatment of various inflammatory diseases. The present study examined the cytoprotective properties of C. sinensis on D(+)-galactosamine (GalN)/lipopolysaccharide (LPS)-induced fulminant hepatic failure. Mice were randomly assigned into control, GalN/LPS, CS 20 mg and CS 40 mg groups (C. sinensis, oral gavage, five days/week, four weeks). After receiving saline or C. sinensis, mice were intraperitoneally given GalN (800 mg/kg)/LPS (10 μg/kg). The effects of C. sinensis on TNF-α, IL-10, AST, NO, SOD, and apoptoticrelated proteins after the onset of endotoxin intoxication were determined. Data demonstrated that GalN/LPS increased hepatocyte degeneration, circulating AST, TNF-α, IL-10, and hepatic apoptosis and caspase activity. C. sinensis pre-treatment reduced AST, TNF-α, and NO and increased IL-10 and SOD in GalN/LPS induced fulminant hepatic failure. C. sinensis attenuated the apoptosis of hepatocytes, as evidenced by the TUNEL and capase-3, 6 activity analyses. In summary, C. sinensis alleviates GalN/LPS-induced liver injury by modulating the cytokine response and inhibiting apoptosis. PMID:24707872

  5. p14(ARF) Prevents Proliferation of Aneuploid Cells by Inducing p53-Dependent Apoptosis.

    PubMed

    Veneziano, Lorena; Barra, Viviana; Lentini, Laura; Spatafora, Sergio; Di Leonardo, Aldo

    2016-02-01

    Weakening the Spindle Assembly Checkpoint by reduced expression of its components induces chromosome instability and aneuploidy that are hallmarks of cancer cells. The tumor suppressor p14(ARF) is overexpressed in response to oncogenic stimuli to stabilize p53 halting cell progression. Previously, we found that lack or reduced expression of p14(ARF) is involved in the maintenance of aneuploid cells in primary human cells, suggesting that it could be part of a pathway controlling their proliferation. To investigate this aspect further, p14(ARF) was ectopically expressed in HCT116 cells after depletion of the Spindle Assembly Checkpoint MAD2 protein that was used as a trigger for aneuploidy. p14(ARF) Re-expression reduced the number of aneuploid cells in MAD2 post-transcriptionally silenced cells. Also aberrant mitoses, frequently displayed in MAD2-depleted cells, were decreased when p14(ARF) was expressed at the same time. In addition, p14(ARF) ectopic expression in MAD2-depleted cells induced apoptosis associated with increased p53 protein levels. Conversely, p14(ARF) ectopic expression did not induce apoptosis in HCT116 p53KO cells. Collectively, our results suggest that the tumor suppressor p14(ARF) may have an important role in counteracting proliferation of aneuploid cells by activating p53-dependent apoptosis. PMID:25752701

  6. Pharmacologically blocking p53-dependent apoptosis protects intestinal stem cells and mice from radiation.

    PubMed

    Wang, Xinwei; Wei, Liang; Cramer, Julie M; Leibowitz, Brian J; Judge, Colleen; Epperly, Michael; Greenberger, Joel; Wang, Fengchao; Li, Linheng; Stelzner, Matthias G; Dunn, James C Y; Martin, Martin G; Lagasse, Eric; Zhang, Lin; Yu, Jian

    2015-01-01

    Exposure to high levels of ionizing radiation (IR) leads to debilitating and dose-limiting gastrointestinal (GI) toxicity. Using three-dimensional mouse crypt culture, we demonstrated that p53 target PUMA mediates radiation-induced apoptosis via a cell-intrinsic mechanism, and identified the GSK-3 inhibitor CHIR99021 as a potent radioprotector. CHIR99021 treatment improved Lgr5+ cell survival and crypt regeneration after radiation in culture and mice. CHIR99021 treatment specifically blocked apoptosis and PUMA induction and K120 acetylation of p53 mediated by acetyl-transferase Tip60, while it had no effect on p53 stabilization, phosphorylation or p21 induction. CHIR99021 also protected human intestinal cultures from radiation by PUMA but not p21 suppression. These results demonstrate that p53 posttranslational modifications play a key role in the pathological and apoptotic response of the intestinal stem cells to radiation and can be targeted pharmacologically. PMID:25858503

  7. Pharmacologically blocking p53-dependent apoptosis protects intestinal stem cells and mice from radiation

    PubMed Central

    Wang, Xinwei; Wei, Liang; Cramer, Julie M.; Leibowitz, Brian J.; Judge, Colleen; Epperly, Michael; Greenberger, Joel; Wang, Fengchao; Li, Linheng; Stelzner, Matthias G.; Dunn, James C. Y.; Martin, Martin G.; Lagasse, Eric; Zhang, Lin; Yu, Jian

    2015-01-01

    Exposure to high levels of ionizing radiation (IR) leads to debilitating and dose-limiting gastrointestinal (GI) toxicity. Using three-dimensional mouse crypt culture, we demonstrated that p53 target PUMA mediates radiation-induced apoptosis via a cell-intrinsic mechanism, and identified the GSK-3 inhibitor CHIR99021 as a potent radioprotector. CHIR99021 treatment improved Lgr5+ cell survival and crypt regeneration after radiation in culture and mice. CHIR99021 treatment specifically blocked apoptosis and PUMA induction and K120 acetylation of p53 mediated by acetyl-transferase Tip60, while it had no effect on p53 stabilization, phosphorylation or p21 induction. CHIR99021 also protected human intestinal cultures from radiation by PUMA but not p21 suppression. These results demonstrate that p53 posttranslational modifications play a key role in the pathological and apoptotic response of the intestinal stem cells to radiation and can be targeted pharmacologically. PMID:25858503

  8. Salinomycin sensitizes antimitotic drugs-treated cancer cells by increasing apoptosis via the prevention of G2 arrest

    SciTech Connect

    Kim, Ju-Hwa; Yoo, Hye-In; Kang, Han Sung; Ro, Jungsil; Yoon, Sungpil

    2012-02-03

    Highlights: Black-Right-Pointing-Pointer Sal sensitizes antimitotic drugs-treated cancer cells. Black-Right-Pointing-Pointer Sal sensitizes them by prevention of G2 arrest and reduced cyclin D1 levels. Black-Right-Pointing-Pointer Sal also sensitizes them by increasing DNA damage and reducing p21 level. Black-Right-Pointing-Pointer A low concentration of Sal effectively sensitized the cancer cells to antimitotic drugs. -- Abstract: Here, we investigated whether Sal could sensitize cancer cells to antimitotic drugs. We demonstrated that Sal sensitized paclitaxcel (PAC)-, docetaxcel (DOC)-, vinblastin (VIN)-, or colchicine (COL)-treated cancer cell lines, suggesting that Sal has the ability to sensitize the cells to any form of microtubule-targeting drugs. Sensitization to the antimitotic drugs could be achieved with very low concentrations of Sal, suggesting that there is a possibility to minimize Sal toxicity associated with human cancer patient treatments. Sensitization by Sal increased apoptosis, which was observed by C-PARP production. Sal sensitized the cancer cells to antimitotic drugs by preventing G2 arrest, suggesting that Sal contributes to the induction of mitotic catastrophe. Sal generally reduced cyclin D1 levels in PAC-, DOC-, and VIN-treated cells. In addition, Sal treatment increased pH2AX levels and reduced p21 levels in antimitotic drugs-treated cells. These observations suggest that the mechanisms underlying Sal sensitization to DNA-damaging compounds, radiation, and microtubule-targeting drugs are similar. Our data demonstrated that Sal sensitizes cancer cells to antimitotic drugs by increasing apoptosis through the prevention of G2 arrest via conserved Sal-sensitization mechanisms. These results may contribute to the development of Sal-based chemotherapy for cancer patients treated with antimitotic drugs.

  9. Mutation of the Myxoma virus SERP2 P1-site to prevent proteinase inhibition causes apoptosis in cultured RK-13 cells and attenuates disease in rabbits, but mutation to alter specificity causes apoptosis without reducing virulence.

    PubMed

    MacNeill, Amy L; Turner, Peter C; Moyer, Richard W

    Myxoma virus (MYX) prevents apoptosis in RK-13 cells and forms thick dermal lesions with 100% mortality in rabbits. MYX encodes the virulence factor SERP2, a serine proteinase inhibitor (serpin). SERP2 was mutated to evaluate SERP2 function during MYX infection. MYXDeltaSERP2::lacZ (deleted for SERP2) did not inhibit apoptosis in RK-13 cells; infected rabbits had thin dermal lesions and <10% mortality. MYX-SERP2-D294A, a P1-site aspartate to alanine mutant, inactivated the serpin; infection was indistinguishable from MYXDeltaSERP2::lacZ. SERP2-D294E prevented inhibition of caspase-8, caspase-10 and granzyme-B; and MYX-SERP2-D294E failed to block apoptosis in RK-13 cells, but was fully virulent in rabbits. MYXDeltaSERP2::crmA expressed crmA instead of SERP2 and inhibited apoptosis in cell culture, but caused thin lesions and only 70% mortality in rabbits, hence crmA cannot fully substitute for SERP2. Control of apoptosis in culture does not correlate with virulence in rabbits. Virulence may instead depend on inhibition of proinflammatory proteinases by SERP2. PMID:16959285

  10. Rasagiline prevents cyclosporine A-sensitive superoxide flashes induced by PK11195, the initial signal of mitochondrial membrane permeabilization and apoptosis.

    PubMed

    Wu, Yuqiu; Shamoto-Nagai, Masayo; Maruyama, Wakako; Osawa, Toshihiko; Naoi, Makoto

    2016-05-01

    Rasagiline, a neuroprotective inhibitor of type B monoamine oxidase, prevented PK111195-induced apoptosis in SH-SY5Y cells through inhibition of mitochondrial apoptosis signaling (J Neural Transm 120:1539-1551, 2013, J Neural Transm 122:1399-1407, 2015). This paper presents that PK11195 induced superoxide flashes, the transit production burst, mediated by cyclosporine A-sensitive membrane permeability transition. Rasagiline prevented superoxide flashes, calcium efflux, and cell death by PK11195. Regulation of the initial pore formation at the inner mitochondrial membrane was confirmed as the decisive mechanism of neuroprotection by rasagiline. PMID:26931622

  11. Monoamine oxidase inhibition prevents mitochondrial dysfunction and apoptosis in myoblasts from patients with collagen VI myopathies

    PubMed Central

    Sorato, E.; Menazza, S.; Zulian, A.; Sabatelli, P.; Gualandi, F.; Merlini, L.; Bonaldo, P.; Canton, M.; Bernardi, P.; Di Lisa, F.

    2014-01-01

    Although mitochondrial dysfunction and oxidative stress have been proposed to play a crucial role in several types of muscular dystrophy (MD), whether a causal link between these two alterations exists remains an open question. We have documented that mitochondrial dysfunction through opening of the permeability transition pore plays a key role in myoblasts from patients as well as in mouse models of MD, and that oxidative stress caused by monoamine oxidases (MAO) is involved in myofiber damage. In the present study we have tested whether MAO-dependent oxidative stress is a causal determinant of mitochondrial dysfunction and apoptosis in myoblasts from patients affected by collagen VI myopathies. We find that upon incubation with hydrogen peroxide or the MAO substrate tyramine myoblasts from patients upregulate MAO-B expression and display a significant rise in reactive oxygen species (ROS) levels, with concomitant mitochondrial depolarization. MAO inhibition by pargyline significantly reduced both ROS accumulation and mitochondrial dysfunction, and normalized the increased incidence of apoptosis in myoblasts from patients. Thus, MAO-dependent oxidative stress is causally related to mitochondrial dysfunction and cell death in myoblasts from patients affected by collagen VI myopathies, and inhibition of MAO should be explored as a potential treatment for these diseases. PMID:25017965

  12. Fluoxetine enhances cell proliferation and prevents apoptosis in dentate gyrus of maternally separated rats.

    PubMed

    Lee, H J; Kim, J W; Yim, S V; Kim, M J; Kim, S A; Kim, Y J; Kim, C J; Chung, J H

    2001-11-01

    The mother-infant relationship is an instinctive phenomenon, and loss of maternal care in early life influences neonatal development, behavior and physiologic responses.(1,2) Furthermore, the early loss may affect the vulnerability of the infant to neuropsychiatric disorders, such as childhood anxiety disorders, personality disorders and depression, over its lifespan.(3,4) Fluoxetine is prescribed worldwide for depression and is often used in the treatment of childhood mental problems related to maternal separation or loss of maternal care.(5,6) In the present study, fluoxetine was administrated to rats with maternal separation to determine its effects on neuronal development, in particular with respect to cell proliferation and apoptosis in the dentate gyrus of the hippocampus. Rat pups were separated from their mothers and socially isolated on postnatal day 14 and were treated with fluoxetine (5 mg kg(-1)) and 5-bromo-2'-deoxyuridine (BrdU) (50 mg kg(-1)) for 7 days, after which immunohistochemistry and a terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) staining were carried out. In the pups with maternal separation treated with fluoxetine, the number of BrdU-positive cells was significantly increased and that of TUNEL-positive cells was significantly decreased in the dentate gyrus compared to pups with maternal separation that did not receive fluoxetine treatment. These findings indicate that fluoxetine affects new cell proliferation and apoptosis, and we propose that fluoxetine may be useful in the treatment of maternal separation-related diseases. PMID:11673802

  13. Heparin Interaction with the Primed Polymorphonuclear Leukocyte CD11b Induces Apoptosis and Prevents Cell Activation

    PubMed Central

    Cohen-Mazor, Meital; Mazor, Rafi; Kristal, Batya; Kistler, Erik B.; Ziv, Inbal; Chezar, Judith; Sela, Shifra

    2015-01-01

    Heparin is known to have anti-inflammatory effects, yet the mechanisms are not completely understood. In this study, we tested the hypothesis that heparin has a direct effect on activated polymorphonuclear leukocytes (PMNLs), changing their activation state, and can explain its anti-inflammatory effect. To test our hypothesis, we designed both in vitro and ex vivo studies to elucidate the mechanism by which heparin modulates PMNL functions and therefore the inflammatory response. We specifically tested the hypothesis that priming of PMNLs renders them more susceptible to heparin. Amplified levels of CD11b and increased rate of superoxide release manifested PMNL priming. Increase in cell priming resulted in a dose-dependent increase in heparin binding to PMNLs followed by augmented apoptosis. Blocking antibodies to CD11b inhibited heparin binding and abolished the apoptotic response. Moreover, heparin caused a significant dose-dependent decrease in the rate of superoxide release from PMNLs, which was blunted by blocking antibodies to CD11b. Altogether, this study shows that the interaction of heparin with the PMNL CD11b results in cell apoptosis and explains heparin's anti-inflammatory effects. PMID:26819958

  14. Resveratrol prevents rapamycin-induced upregulation of autophagy and selectively induces apoptosis in TSC2-deficient cells

    PubMed Central

    Alayev, Anya; Sun, Yang; Snyder, Rose B; Berger, Sara Malka; Yu, Jane J; Holz, Marina K

    2014-01-01

    The mammalian/mechanistic target of rapamycin complex 1 (mTORC1) signaling pathway is hyperactivated in a variety of cancers and disorders, including lymphangioleiomyomatosis (LAM) and tuberous sclerosis complex (TSC), which are characterized by mutations in tumor suppressors TSC1 or TSC2. The concern with the use of mTORC1 inhibitors, such as rapamycin or its analogs (rapalogs), is that they cause upregulation of autophagy and suppress the negative feedback loop to Akt, which promotes cell survival, causing the therapy to be only partially effective, and relapse occurs upon cessation of treatment. In this study, we investigate the use of rapamycin in combination with resveratrol, a naturally occurring polyphenol, in TSC2-deficient cells. We tested whether such combination would prevent rapamycin-induced upregulation of autophagy and shift the cell fate toward apoptosis. We found that this combination treatment blocked rapamycin-induced upregulation of autophagy and restored inhibition of Akt. Interestingly, the combination of rapamycin and resveratrol selectively promoted apoptosis of TSC2-deficient cells. Thus, the addition of resveratrol to rapamycin treatment may be a promising option for selective and targeted therapy for diseases with TSC loss and mTORC1 hyperactivation. PMID:24304514

  15. Fibronectin prevents endotoxin shock after partial hepatectomy in rats via inhibition of nuclear factor-kappaB and apoptosis.

    PubMed

    Kwon, A-Hon; Qiu, Zeyu; Tsuji, Katsushige; Miyaso, Takeshi; Okumura, Tadayoshi

    2007-07-01

    Fibronectins (Fns) are involved in a number of biologic processes, such as cellular adhesion, motility, differentiation, apoptosis, hemostasis, wound healing, and ischemic injury. We investigated the possible mechanism underlying the protective action of plasma Fn (pFn) on endotoxin shock following partial hepatectomy in rats. Lipopolysaccharide (LPS) was administered intravenously to male Sprague-Dawley rats within 48 hrs of 70% hepatectomy. Prior to LPS administration, pFn or human serum albumin was given intravenously. The survival rate of the pFn-treated group was improved markedly compared with that of the controls. The levels of inflammatory cytokines and nitric oxide (NO) in serum were significantly lower in the pFn-treated group than in the control group. Expression of inducible nitric oxide synthase (iNOS) in hepatocytes also was reduced following pFn treatment. The degree of apoptosis and necrosis in the remnant liver was significantly lower in the pFn-treated rats than the controls. Furthermore, pFn pretreatment greatly inhibited the activation of nuclear factor-kappaB (NF-kappaB), caspase 3 and 8 activities, and cytochrome c release, and caused a decrease in mitochondrial Bcl-x(L). Plasma Fn prevents endotoxin-induced liver injury at least in part through inhibition of NF-kappaB activation, which causes the reduction of iNOS expression and NO production by hepatocytes, and through the downregulation of inflammatory cytokines and promotion of Bcl-x(L) expression. PMID:17609505

  16. Molecular pathways: gene-environment interactions regulating dietary fiber induction of proliferation and apoptosis via butyrate for cancer prevention.

    PubMed

    Bultman, Scott J

    2014-02-15

    Gene-environment interactions are so numerous and biologically complicated that it can be challenging to understand their role in cancer. However, dietary fiber and colorectal cancer prevention may represent a tractable model system. Fiber is fermented by colonic bacteria into short-chain fatty acids such as butyrate. One molecular pathway that has emerged involves butyrate having differential effects depending on its concentration and the metabolic state of the cell. Low-moderate concentrations, which are present near the base of colonic crypts, are readily metabolized in the mitochondria to stimulate cell proliferation via energetics. Higher concentrations, which are present near the lumen, exceed the metabolic capacity of the colonocyte. Unmetabolized butyrate enters the nucleus and functions as a histone deacetylase (HDAC) inhibitor that epigenetically regulates gene expression to inhibit cell proliferation and induce apoptosis as the colonocytes exfoliate into the lumen. Butyrate may therefore play a role in normal homeostasis by promoting turnover of the colonic epithelium. Because cancerous colonocytes undergo the Warburg effect, their preferred energy source is glucose instead of butyrate. Consequently, even moderate concentrations of butyrate accumulate in cancerous colonocytes and function as HDAC inhibitors to inhibit cell proliferation and induce apoptosis. These findings implicate a bacterial metabolite with metaboloepigenetic properties in tumor suppression. PMID:24270685

  17. Fenugreek (Trigonella foenum graecum) seed extract prevents ethanol-induced toxicity and apoptosis in Chang liver cells.

    PubMed

    Kaviarasan, Subramanian; Ramamurty, Nalini; Gunasekaran, Palani; Varalakshmi, Elango; Anuradha, Carani Venkatraman

    2006-01-01

    The protective effect of a polyphenolic extract of fenugreek seeds (FPEt) against ethanol (EtOH)-induced toxicity was investigated in human Chang liver cells. Cells were incubated with either 30 mM EtOH alone or together in the presence of seed extract for 24 h. Assays were performed in treated cells to evaluate the ability of seeds to prevent the toxic effects of EtOH. EtOH treatment suppressed the growth of Chang liver cells and induced cytotoxicity, oxygen radical formation and mitochondrial dysfunction. Reduced glutathione (GSH) concentration was decreased significantly (P < 0.05) while oxidized glutathione (GSSG) concentration was significantly elevated in EtOH-treated cells as compared with normal cells. Incubation of FPEt along with EtOH significantly increased cell viability in a dose-dependent manner, caused a reduction in lactate dehydrogenase leakage and normalized GSH/GSSG ratio. The extract dose-dependently reduced thiobarbituric acid reactive substances formation. Apoptosis was observed in EtOH-treated cells while FPEt reduced apoptosis by decreasing the accumulation of sub-G1 phase cells. The cytoprotective effects of FPEt were comparable with those of a positive control silymarin, a known hepatoprotective agent. The findings suggest that the polyphenolic compounds of fenugreek seeds can be considered cytoprotective during EtOH-induced liver damage. PMID:16574673

  18. Protocatechuic Acid Prevents oxLDL-Induced Apoptosis by Activating JNK/Nrf2 Survival Signals in Macrophages

    PubMed Central

    Varì, Rosaria; Scazzocchio, Beatrice; Santangelo, Carmela; Filesi, Carmelina; Galvano, Fabio; D'Archivio, Massimo; Masella, Roberta; Giovannini, Claudio

    2015-01-01

    Protocatechuic acid (PCA), one of the main metabolites of complex polyphenols, exerts numerous biological activities including antiapoptotic, anti-inflammatory, and antiatherosclerotic effects. Oxidised LDL have atherogenic properties by damaging arterial wall cells and inducing p53-dependent apoptosis in macrophages. This study was aimed at defining the molecular mechanism responsible for the protective effects of PCA against oxidative and proapoptotic damage exerted by oxLDL in J774 A.1 macrophages. We found that the presence of PCA in cells treated with oxLDL completely inhibited the p53-dependent apoptosis induced by oxLDL. PCA decreased oxLDL-induced ROS overproduction and in particular prevented the early increase of ROS. This decrease seemed to be the main signal responsible for maintaining the intracellular redox homeostasis hindering the activation of p53 induced by ROS, p38MAPK, and PKCδ. Consequently the overexpression of the proapoptotic p53-target genes such as p66Shc protein did not occur. Finally, we demonstrated that PCA induced the activation of JNK, which, in turn, determined the increase of nuclear Nrf2, leading to inhibition of the early ROS overproduction. We concluded that the antiapoptotic mechanism of PCA was most likely related to the activation of the JNK-mediated survival signals that strengthen the cellular antioxidant defences rather than to the PCA antioxidant power. PMID:26180584

  19. Protocatechuic Acid Prevents oxLDL-Induced Apoptosis by Activating JNK/Nrf2 Survival Signals in Macrophages.

    PubMed

    Varì, Rosaria; Scazzocchio, Beatrice; Santangelo, Carmela; Filesi, Carmelina; Galvano, Fabio; D'Archivio, Massimo; Masella, Roberta; Giovannini, Claudio

    2015-01-01

    Protocatechuic acid (PCA), one of the main metabolites of complex polyphenols, exerts numerous biological activities including antiapoptotic, anti-inflammatory, and antiatherosclerotic effects. Oxidised LDL have atherogenic properties by damaging arterial wall cells and inducing p53-dependent apoptosis in macrophages. This study was aimed at defining the molecular mechanism responsible for the protective effects of PCA against oxidative and proapoptotic damage exerted by oxLDL in J774 A.1 macrophages. We found that the presence of PCA in cells treated with oxLDL completely inhibited the p53-dependent apoptosis induced by oxLDL. PCA decreased oxLDL-induced ROS overproduction and in particular prevented the early increase of ROS. This decrease seemed to be the main signal responsible for maintaining the intracellular redox homeostasis hindering the activation of p53 induced by ROS, p38MAPK, and PKCδ. Consequently the overexpression of the proapoptotic p53-target genes such as p66Shc protein did not occur. Finally, we demonstrated that PCA induced the activation of JNK, which, in turn, determined the increase of nuclear Nrf2, leading to inhibition of the early ROS overproduction. We concluded that the antiapoptotic mechanism of PCA was most likely related to the activation of the JNK-mediated survival signals that strengthen the cellular antioxidant defences rather than to the PCA antioxidant power. PMID:26180584

  20. Depletion of cellular poly (A) binding protein prevents protein synthesis and leads to apoptosis in HeLa cells

    SciTech Connect

    Thangima Zannat, Mst.; Bhattacharjee, Rumpa B.; Bag, Jnanankur

    2011-05-13

    Highlights: {yields} Depletion of cellular PABP level arrests mRNA translation in HeLa cells. {yields} PABP knock down leads to apoptotic cell death. {yields} PABP depletion does not affect transcription. {yields} PABP depletion does not lead to nuclear accumulation of mRNA. -- Abstract: The cytoplasmic poly (A) binding protein (PABP) is important in mRNA translation and stability. In yeast, depletion of PABP leads to translation arrest. Similarly, the PABP gene in Drosophila is important for proper development. It is however uncertain, whether mammalian PABP is essential for mRNA translation. Here we showed the effect of PABP depletion on mRNA metabolism in HeLa cells by using a small interfering RNA. Our results suggest that depletion of PABP prevents protein synthesis and consequently leads to cell death through apoptosis. Interestingly, no detectable effect of PABP depletion on transcription, transport and stability of mRNA was observed.

  1. Cellular inhibitor of apoptosis proteins prevent clearance of hepatitis B virus.

    PubMed

    Ebert, Gregor; Preston, Simon; Allison, Cody; Cooney, James; Toe, Jesse G; Stutz, Michael D; Ojaimi, Samar; Scott, Hamish W; Baschuk, Nikola; Nachbur, Ueli; Torresi, Joseph; Chin, Ruth; Colledge, Danielle; Li, Xin; Warner, Nadia; Revill, Peter; Bowden, Scott; Silke, John; Begley, C Glenn; Pellegrini, Marc

    2015-05-01

    Hepatitis B virus (HBV) infection can result in a spectrum of outcomes from immune-mediated control to disease progression, cirrhosis, and liver cancer. The host molecular pathways that influence and contribute to these outcomes need to be defined. Using an immunocompetent mouse model of chronic HBV infection, we identified some of the host cellular and molecular factors that impact on infection outcomes. Here, we show that cellular inhibitor of apoptosis proteins (cIAPs) attenuate TNF signaling during hepatitis B infection, and they restrict the death of infected hepatocytes, thus allowing viral persistence. Animals with a liver-specific cIAP1 and total cIAP2 deficiency efficiently control HBV infection compared with WT mice. This phenotype was partly recapitulated in mice that were deficient in cIAP2 alone. These results indicate that antagonizing the function of cIAPs may promote the clearance of HBV infection. PMID:25902529

  2. Cellular inhibitor of apoptosis proteins prevent clearance of hepatitis B virus

    PubMed Central

    Ebert, Gregor; Preston, Simon; Allison, Cody; Cooney, James; Toe, Jesse G.; Stutz, Michael D.; Ojaimi, Samar; Scott, Hamish W.; Baschuk, Nikola; Nachbur, Ueli; Torresi, Joseph; Chin, Ruth; Colledge, Danielle; Li, Xin; Warner, Nadia; Revill, Peter; Bowden, Scott; Silke, John; Begley, C. Glenn; Pellegrini, Marc

    2015-01-01

    Hepatitis B virus (HBV) infection can result in a spectrum of outcomes from immune-mediated control to disease progression, cirrhosis, and liver cancer. The host molecular pathways that influence and contribute to these outcomes need to be defined. Using an immunocompetent mouse model of chronic HBV infection, we identified some of the host cellular and molecular factors that impact on infection outcomes. Here, we show that cellular inhibitor of apoptosis proteins (cIAPs) attenuate TNF signaling during hepatitis B infection, and they restrict the death of infected hepatocytes, thus allowing viral persistence. Animals with a liver-specific cIAP1 and total cIAP2 deficiency efficiently control HBV infection compared with WT mice. This phenotype was partly recapitulated in mice that were deficient in cIAP2 alone. These results indicate that antagonizing the function of cIAPs may promote the clearance of HBV infection. PMID:25902529

  3. The Mood-Stabilizer Lithium Prevents Hippocampal Apoptosis and Improves Spatial Memory in Experimental Meningitis

    PubMed Central

    Liechti, Fabian D.; Stüdle, Nicolas; Theurillat, Regula; Grandgirard, Denis; Thormann, Wolfgang; Leib, Stephen L.

    2014-01-01

    Pneumococcal meningitis is associated with high morbidity and mortality rates. Brain damage caused by this disease is characterized by apoptosis in the hippocampal dentate gyrus, a morphological correlate of learning deficits in experimental paradigms. The mood stabilizer lithium has previously been found to attenuate brain damage in ischemic and inflammatory diseases of the brain. An infant rat model of pneumococcal meningitis was used to investigate the neuroprotective and neuroregenerative potential of lithium. To assess an effect on the acute disease, LiCl was administered starting five days prior to intracisternal infection with live Streptococcus pneumoniae. Clinical parameters were recorded, cerebrospinal fluid (CSF) was sampled, and the animals were sacrificed 42 hours after infection to harvest the brain and serum. Cryosections of the brains were stained for Nissl substance to quantify brain injury. Hippocampal gene expression of Bcl-2, Bax, p53, and BDNF was analyzed. Lithium concentrations were measured in serum and CSF. The effect of chronic lithium treatment on spatial memory function and cell survival in the dentate gyrus was evaluated in a Morris water maze and by quantification of BrdU incorporation after LiCl treatment during 3 weeks following infection. In the hippocampus, LiCl significantly reduced apoptosis and gene expression of Bax and p53 while it increased expression of Bcl-2. IL-10, MCP-1, and TNF were significantly increased in animals treated with LiCl compared to NaCl. Chronic LiCl treatment improved spatial memory in infected animals. The mood stabilizer lithium may thus be a therapeutic alternative to attenuate neurofunctional deficits as a result of pneumococcal meningitis. PMID:25409333

  4. Sulforaphane prevents microcystin-LR-induced oxidative damage and apoptosis in BALB/c mice

    SciTech Connect

    Sun Xiaoyun; Mi Lixin; Liu Jin; Song Lirong; Chung Funglung; Gan Nanqin

    2011-08-15

    Microcystins (MCs), the products of blooming algae Microcystis, are waterborne environmental toxins that have been implicated in the development of liver cancer, necrosis, and even fatal intrahepatic bleeding. Alternative protective approaches in addition to complete removal of MCs in drinking water are urgently needed. In our previous work, we found that sulforaphane (SFN) protects against microcystin-LR (MC-LR)-induced cytotoxicity by activating the NF-E2-related factor 2 (Nrf2)-mediated defensive response in human hepatoma (HepG2) and NIH 3T3 cells. The purpose of this study was to investigate and confirm efficacy the SFN-induced multi-mechanistic defense system against MC-induced hepatotoxicity in an animal model. We report that SFN protected against MC-LR-induced liver damage and animal death at a nontoxic and physiologically relevant dose in BALB/c mice. The protection by SFN included activities of anti-cytochrome P450 induction, anti-oxidation, anti-inflammation, and anti-apoptosis. Our results suggest that SFN may protect mice against MC-induced hepatotoxicity. This raises the possibility of a similar protective effect in human populations, particularly in developing countries where freshwaters are polluted by blooming algae. - Graphical abstract: Display Omitted Research Highlights: > SFN protected against MC-LR-induced liver damage and animal death in BALB/c mice. > The dose of SFN is at a nontoxic and physiologically relevant dose. > The protection included activities of anti-oxidation, anti-inflammation, and anti-apoptosis. > SFN may protect mice against MC-induced hepatotoxicity.

  5. The transcription-independent mitochondrial p53 program is a major contributor to nutlin-induced apoptosis in tumor cells.

    PubMed

    Vaseva, Angelina V; Marchenko, Natalia D; Moll, Ute M

    2009-06-01

    Strategies to induce p53 activation in tumors that retain wild-type p53 are promising for cancer therapy. Nutlin is a potent and selective pharmacological MDM2 inhibitor that competitively binds to its p53-binding pocket, thereby leading to non-genotoxic p53 stabilization and activation of growth arrest and apoptosis pathways. Nutlin-induced apoptosis is thought to occur via p53's transcriptional program. Here we report that the transcription-independent mitochondrial p53 program plays an important role in Nutlin-induced p53-mediated tumor cell death. Aside from nuclear stabilization, Nutlin causes cytoplasmic p53 accumulation and translocation to mitochondria. Monoubiquitinated p53, originating from a distinct cytoplasmic pool, is the preferred p53 species that translocates to mitochondria in response to stress. Nutlin does not interfere with MDM2's ability to monoubiquitinate p53, due to the fact that MDM2-p53 complexes are only partially disrupted and that Nutlin-stabilized MDM2 retains its E3 ubiquitin ligase activity. Nutlin-induced mitochondrial p53 translocation is rapid and associated with cytochrome C release that precedes induction of p53 target genes. Specific inhibition of mitochondrial p53 translocation by Pifithrin mu reduces the apoptotic Nutlin response by 2.5-fold, underlining the significance of p53's mitochondrial program in Nutlin-induced apoptosis. Surprisingly, blocking the transcriptional arm of p53, either via alpha-Amanitin or the p53-specific transcriptional inhibitor Pifithrin alpha, not only fails to inhibit, but greatly potentiates Nutlin-induced apoptosis. In sum, the direct mitochondrial program is a major mechanism in Nutlin-induced p53-mediated apoptosis. Moreover, at least in some tumors the transcriptional p53 activities in net balance not only are dispensable for the apoptotic Nutlin response, but appear to actively block its therapeutic effect. PMID:19411846

  6. The transcription-independent mitochondrial p53 program is a major contributor to nutlin-induced apoptosis in tumor cells

    PubMed Central

    Vaseva, Angelina V.; Marchenko, Natalia D.; Moll, Ute M.

    2010-01-01

    Strategies to induce p53 activation in tumors that retain wild-type p53 are promising for cancer therapy. Nutlin is a potent and selective pharmacological MDM2 inhibitor that competitively binds to its p53-binding pocket, thereby leading to non-genotoxic p53 stabilization and activation of growth arrest and apoptosis pathways. Nutlin-induced apoptosis is thought to occur via p53’s transcriptional program. Here we report that the transcription-independent mitochondrial p53 program plays an important role in Nutlin-induced p53-mediated tumor cell death. Aside from nuclear stabilization, Nutlin causes cytoplasmic p53 accumulation and translocation to mitochondria. Monoubiquitinated p53, originating from a distinct cytoplasmic pool, is the preferred p53 species that translocates to mitochondria in response to stress. Nutlin does not interfere with MDM2’s ability to monoubiquitinate p53, due to the fact that MDM2-p53 complexes are only partially disrupted and that Nutlin-stabilized MDM2 retains its E3 ubiquitin ligase activity. Nutlin-induced mitochondrial p53 translocation is rapid and associated with cytochrome C release that precedes induction of p53 target genes. Specific inhibition of mitochondrial p53 translocation by Pifithrin μ reduces the apoptotic Nutlin response by 2.5-fold, underlining the significance of p53’s mitochondrial program in Nutlin-induced apoptosis. Surprisingly, blocking the transcriptional arm of p53, either via α-Amanitin or the p53-specific transcriptional inhibitor Pifithrin α, not only fails to inhibit, but greatly potentiates Nutlin-induced apoptosis. In sum, the direct mitochondrial program is a major mechanism in Nutlin-induced p53-mediated apoptosis. Moreover, at least in some tumors the transcriptional p53 activities in net balance not only are dispensable for the apoptotic Nutlin response, but appear to actively block its therapeutic effect. PMID:19411846

  7. Lithium Treatment Prevents Apoptosis in Neonatal Rat Hippocampus Resulting from Sevoflurane Exposure.

    PubMed

    Zhou, Xue; da Li, Wen-; Yuan, Bao-Long; Niu, Li-Jun; Yang, Xiao-Yu; Zhou, Zhi-Bin; Chen, Xiao-Hui; Feng, Xia

    2016-08-01

    We aimed to observe the therapeutic effects of lithium on inhalational anesthetic sevoflurane-induced apoptosis in immature brain hippocampus. From postnatal day 5 (P5) to P28, male Sprague-Dawley pups were intraperitoneally injected with lithium chloride or 0.9 % sodium chloride. On P7 after the injection, pups were exposed to 2.3 % sevoflurane or air for 6 h. Brain tissues were harvested 12 h and 3 weeks after exposure. Cleaved caspase-3, nNOS protein, GSK-3β,p-GSK-3β were assessed by Western blot, and histopathological changes were assessed using Nissl stain and TUNEL stain. From P28, we used the eight-arm radial maze test and step-through test to evaluate the influence of sevoflurane exposure on the learning and memory of juvenile rats. The results showed that neonatal sevoflurane exposure induced caspase-3 activation and histopathological changes in hippocampus can be attenuated by lithium chloride. Sevoflurane increased GSK-3β activity while pretreatment of lithium decreased GSK-3β activity. Moreover, sevoflurane showed possibly slight but temporal influence on the spatial learning and the memory of juvenile rats, and chronic use of lithium chloride might have the therapeutic effect. Our current study suggests that lithium attenuates sevoflurane induced neonatal hippocampual damage by GSK-3β pathway and might improve learning and memory deficits in rats after neonatal exposure. PMID:27068032

  8. Exogenous cardiolipin localizes to mitochondria and prevents TAZ knockdown-induced apoptosis in myeloid progenitor cells.

    PubMed

    Ikon, Nikita; Su, Betty; Hsu, Fong-Fu; Forte, Trudy M; Ryan, Robert O

    2015-08-21

    The concentration and composition of cardiolipin (CL) in mitochondria are altered in age-related heart disease, Barth Syndrome, and other rare genetic disorders, resulting in mitochondrial dysfunction. To explore whether exogenous CL can be delivered to cells, CL was combined with apolipoprotein A-I to generate water-soluble, nanoscale complexes termed nanodisks (ND). Mass spectrometry of HL60 myeloid progenitor cell extracts revealed a 30-fold increase in cellular CL content following incubation with CL-ND. When CL-ND containing a fluorescent CL analogue was employed, confocal microscopy revealed CL localization to mitochondria. The ability of CL-ND to elicit a physiological response was examined in an HL60 cell culture model of Barth Syndrome neutropenia. siRNA knockdown of the phospholipid transacylase, tafazzin (TAZ), induced apoptosis in these cells. When TAZ knockdown cells were incubated with CL-ND, the apoptotic response was attenuated. Thus, CL-ND represent a potential intervention strategy for replenishment of CL in Barth Syndrome, age-related heart disease, and other disorders characterized by depletion of this key mitochondrial phospholipid. PMID:26164234

  9. Chk1 and p21 Cooperate to Prevent Apoptosis during DNA Replication Fork StressD⃞

    PubMed Central

    Rodriguez, Rene; Meuth, Mark

    2006-01-01

    Cells respond to DNA replication stress by triggering cell cycle checkpoints, repair, or death. To understand the role of the DNA damage response pathways in determining whether cells survive replication stress or become committed to death, we examined the effect of loss of these pathways on cellular response to agents that slow or arrest DNA synthesis. We show that replication inhibitors such as excess thymidine, hydroxyurea, and camptothecin are normally poor inducers of apoptosis. However, these agents become potent inducers of death in S-phase cells upon small interfering RNA-mediated depletion of the checkpoint kinase Chk1. This death response is independent of p53 and Chk2. p21-deficient cells, on the other hand, produce a more robust apoptotic response upon Chk1 depletion. p21 is normally induced only late after thymidine treatment. In Chk1-depleted cells p21 induction occurs earlier and does not require p53. Thus, Chk1 plays a primary role in the protection of cells from death induced by replication fork stress, whereas p21 mediates through its role in regulating entry into S phase. These findings are of potential importance to cancer therapy because we demonstrate that the efficacy of clinically relevant agents can be enhanced by manipulation of these signaling pathways. PMID:16280359

  10. Prevention of glucocorticoid induced-apoptosis of osteoblasts and osteocytes by protecting against endoplasmic reticulum (ER) stress in vitro and in vivo in female mice.

    PubMed

    Sato, Amy Y; Tu, Xiaolin; McAndrews, Kevin A; Plotkin, Lilian I; Bellido, Teresita

    2015-04-01

    Endoplasmic reticulum (ER) stress is associated with increased reactive oxygen species (ROS), results from accumulation of misfolded/unfolded proteins, and can trigger apoptosis. ER stress is alleviated by phosphorylation of eukaryotic translation initiation factor 2α (eIF2α), which inhibits protein translation allowing the ER to recover, thus promoting cell viability. We investigated whether osteoblastic cell apoptosis induced by glucocorticoids (GCs) is due to induction of ROS/ER stress and whether inhibition of eIF2α dephosphorylation promotes survival opposing the deleterious effects of GC in vitro and in vivo. Apoptosis of osteocytic MLO-Y4 and osteoblastic OB-6 cells induced by dexamethasone was abolished by ROS inhibitors. Like GC, the ER stress inducing agents brefeldin A and tunicamycin induced osteoblastic cell apoptosis. Salubrinal or guanabenz, specific inhibitors of eIF2α dephosphorylation, blocked apoptosis induced by either GC or ER stress inducers. Moreover, GC markedly decreased mineralization in OB-6 cells or primary osteoblasts; and salubrinal or guanabenz increased mineralization and prevented the inhibitory effect of GC. Furthermore, salubrinal (1 mg/kg/day) abolished osteoblast and osteocyte apoptosis in cancellous and cortical bone and partially prevented the loss of BMD at all sites and the decreased vertebral cancellous bone formation induced by treatment with prednisolone for 28 days (1.4 mg/kg/day). We conclude that part of the pro-apoptotic actions of GC on osteoblastic cells is mediated through ER stress, and that inhibition of eIF2α dephosphorylation protects from GC-induced apoptosis of osteoblasts and osteocytes in vitro and in vivo and from the deleterious effects of GC on the skeleton. PMID:25532480

  11. Prevention of Glucocorticoid Induced-Apoptosis of Osteoblasts and Osteocytes by Protecting against Endoplasmic Reticulum (ER) Stress in vitro and in vivo in Female Mice

    PubMed Central

    Sato, Amy Y.; Tu, Xiaolin; McAndrews, Kevin A.; Plotkin, Lilian I.; Bellido, Teresita

    2014-01-01

    Endoplasmic reticulum (ER) stress is associated with increased reactive oxygen species (ROS), results from accumulation of misfolded/unfolded proteins, and can trigger apoptosis. ER stress is alleviated by phosphorylation of eukaryotic translation initiation factor 2α (eIF2α), which inhibits protein translation allowing the ER to recover, thus promoting cell viability. We investigated whether osteoblastic cell apoptosis induced by glucocorticoids (GC) is due to induction of ROS/ER stress and whether inhibition of eIF2α dephosphorylation promotes survival opposing the deleterious effects of GC in vitro and in vivo. Apoptosis of osteocytic MLO-Y4 and osteoblastic OB-6 cells induced by dexamethasone was abolished by ROS inhibitors. Like GC, the ER stress inducing agents brefeldin A and tunicamycin induced osteoblastic cell apoptosis. Salubrinal or guanabenz, specific inhibitors of eIF2α dephosphorylation, blocked apoptosis induced by either GC or ER stress inducers. Moreover, GC markedly decreased mineralization in OB-6 cells or primary osteoblasts; and salubrinal or guanabenz increased mineralization and prevented the inhibitory effect of GC. Furthermore, salubrinal (1 mg/kg/day) abolished osteoblast and osteocyte apoptosis in cancellous and cortical bone and partially prevented the loss of BMD at all sites and the decreased vertebral cancellous bone formation induced by treatment with prednisolone for 28 days (1.4 mg/kg/day). We conclude that part of the pro-apoptotic actions of GC on osteoblastic cells are mediated through ER stress, and that inhibition of eIF2α dephosphorylation protects from GC-induced apoptosis of osteoblasts and osteocytes in vitro and in vivo and from the deleterious effects of GC on the skeleton. PMID:25532480

  12. GVS-111 prevents oxidative damage and apoptosis in normal and Down's syndrome human cortical neurons.

    PubMed

    Pelsman, Alejandra; Hoyo-Vadillo, Carlos; Gudasheva, Tatiana A; Seredenin, Sergei B; Ostrovskaya, Rita U; Busciglio, Jorge

    2003-05-01

    The neuroprotective activity of a novel N-acylprolyl-containing dipeptide analog of the nootropic 2-oxo-1-pyrrolidine acetamide (Piracetam) designated as GVS-111 (DVD-111/Noopept) was tested in two in vitro models of neuronal degeneration mediated by oxidative stress: normal human cortical neurons treated with H(2)O(2), and Down's syndrome (DS) cortical neurons. Incubation of normal cortical neurons with 50 microM H(2)O(2) for 1h resulted in morphological and structural changes consistent with neuronal apoptosis and in the degeneration of more than 60% of the neurons present in the culture. GVS-111 significantly increased neuronal survival after H(2)O(2)-treatment displaying a dose-dependent neuroprotective activity from 10nM to 100 microM, and an IC(50) value of 1.21+/-0.07 microM. GVS-111 inhibited the accumulation of intracellular free radicals and lipid peroxidation damage in neurons treated with H(2)O(2) or FeSO(4), suggesting an antioxidant mechanism of action. GVS-111 exhibited significantly higher neuroprotection compared to the standard cognition enhancer Piracetam, or to the antioxidants Vitamin E, propyl gallate and N-tert-butyl-2-sulpho-phenylnitrone (s-PBN). In DS cortical cultures, chronic treatment with GVS-111 significantly reduced the appearance of degenerative changes and enhanced neuronal survival. The results suggest that the neuroprotective effect of GVS-111 against oxidative damage and its potential nootropic activity may present a valuable therapeutic combination for the treatment of mental retardation and chronic neurodegenerative disorders. PMID:12711349

  13. The Salmonella effector SopB prevents ROS-induced apoptosis of epithelial cells by retarding TRAF6 recruitment to mitochondria.

    PubMed

    Ruan, Haihua; Zhang, Zhen; Tian, Li; Wang, Suying; Hu, Shuangyan; Qiao, Jian-Jun

    2016-09-16

    Microbial pathogens enter host cells by injecting effector proteins of the Type III secretion system (T3SS), which facilitate pathogen translocation across the host cell membrane. These effector proteins exert their effects by modulating a variety of host innate immune responses, thereby facilitating bacterial replication and systemic infection. Salmonella enterica serovar typhimurium (S.typhimurium) is a clinically important pathogen that causes food poisoning and gastroenteritis. The SopB effector protein of S. typhimurium, encoded by Salmonella pathogenicity islands (SPI)-1 T3SS, protects host epithelial cells from infection-induced apoptosis. However, how SopB influences apoptosis induction remains unclear. Here, we investigated the mechanism of SopB action in host cells. We found that SopB inhibits infection-induced apoptosis by attenuating the production of reactive oxygen species (ROS) in mitochondria, the crucial organelles for apoptosis initiation. Further investigation revealed that SopB binds to cytosolic tumor necrosis factor receptor associated factor 6 (TRAF6) and forms a trap preventing the mitochondrial recruitment of TRAF6, an essential event for ROS generation within mitochondria. By studying the response of Traf6(+/+) and Traf6(-/-)mouse embryonic fibroblasts to S. typhimurium infection, we found that TRAF6 promoted apoptosis by increasing ROS accumulation, which led to increased Bax/Bcl-2 ratio, Bax recruitment to mitochondrial membrane, and release of Cyt c into the cytoplasm. These findings show that SopB suppresses host cell apoptosis by binding to TRAF6 and preventing mitochondrial ROS generation. PMID:27473656

  14. A novel polysaccharide from Sargassum integerrimum induces apoptosis in A549 cells and prevents angiogensis in vitro and in vivo.

    PubMed

    Liu, Ge; Kuang, Shan; Wu, Shimei; Jin, Weihua; Sun, Chaomin

    2016-01-01

    Many polysaccharides isolated from plants have exhibited promising antitumor activities. The aim of this study is to investigate the antitumor activity of the novel polysaccharide named SPS from Sargassum integerrimum, elucidate the underlying anticancer mechanism in a human lung cancer cell line A549, and evaluate its anti-angiogenic activity both in vitro and in vivo. The results show that SPS significantly reduces A549 cells viability in a dose- and time-dependent manner via MTT method. Flow cytometry analysis indicates that SPS could induce cell apoptosis, the loss of mitochondrial membrane potential (MMP), generation of reactive oxygen species (ROS) and G2/M phase cell cycle arrest of A549 cells. Up-regulation of the expressions of P53 and Bax, down-regulation of the expression of Bcl-2, and activation of cleaved caspase-3, caspase-9 and PARP are also detected by western blotting after the treatment of SPS. In addition, SPS inhibits the proliferation, migration and cord formation of human umbilical vein endothelial cells (HUVECs) in vitro, and prevents the vascular development of zebrafish embryos in vivo. Altogether, our data prove the anticancer and anti-angiogenesis properties of SPS, and provide further insights into the potential pharmacological application of SPS as antitumor and anti-angiogenic agent against lung cancer. PMID:27216943

  15. TNF-alpha-induced apoptosis is prevented by erythropoietin treatment on SH-SY5Y cells

    SciTech Connect

    Pregi, Nicolas Wenker, Shirley; Vittori, Daniela; Leiros, Claudia Perez; Nesse, Alcira

    2009-02-01

    The growth factor erythropoietin (Epo) has shown neuronal protective action in addition to its well known proerythroid activity. Furthermore, Epo has dealt with cellular inflammation by inhibiting the expression of several proinflammatory cytokines, such as IL-1 and TNF-{alpha}. The action of TNF can have both apoptotic and antiapoptotic consequences due to altered balance between different cell signalling pathways. This work has focused on the apoptotic effects of this cytokine and the potential protective action of Epo. The model we used was neuroblastoma SH-SY5Y cells cultured in the presence of 25 ng/ml TNF-{alpha} or pretreated with 25 U/ml Epo for 12 h before the addition of TNF-{alpha}. Apoptosis was evaluated by differential cell count after Hoechst staining, analysis of DNA ladder pattern, and measurement of caspase activity. Despite its ability to induce NF-{kappa}B nuclear translocation, TNF-{alpha} induced cell death, which was found to be associated to upregulation of TNF Receptor 1 expression. On the other hand, cells activated by Epo became resistant to cell death. Prevention of death receptor upregulation and caspase activation may explain this antiapoptotic effect of Epo, which may be also favoured by the induction of a higher expression of protective factors, such as Bcl-2 and NF-{kappa}B, through mechanisms involving Jak/STAT and PI3K signalling pathways.

  16. A novel polysaccharide from Sargassum integerrimum induces apoptosis in A549 cells and prevents angiogensis in vitro and in vivo

    PubMed Central

    Liu, Ge; Kuang, Shan; Wu, Shimei; Jin, Weihua; Sun, Chaomin

    2016-01-01

    Many polysaccharides isolated from plants have exhibited promising antitumor activities. The aim of this study is to investigate the antitumor activity of the novel polysaccharide named SPS from Sargassum integerrimum, elucidate the underlying anticancer mechanism in a human lung cancer cell line A549, and evaluate its anti-angiogenic activity both in vitro and in vivo. The results show that SPS significantly reduces A549 cells viability in a dose- and time-dependent manner via MTT method. Flow cytometry analysis indicates that SPS could induce cell apoptosis, the loss of mitochondrial membrane potential (MMP), generation of reactive oxygen species (ROS) and G2/M phase cell cycle arrest of A549 cells. Up-regulation of the expressions of P53 and Bax, down-regulation of the expression of Bcl-2, and activation of cleaved caspase-3, caspase-9 and PARP are also detected by western blotting after the treatment of SPS. In addition, SPS inhibits the proliferation, migration and cord formation of human umbilical vein endothelial cells (HUVECs) in vitro, and prevents the vascular development of zebrafish embryos in vivo. Altogether, our data prove the anticancer and anti-angiogenesis properties of SPS, and provide further insights into the potential pharmacological application of SPS as antitumor and anti-angiogenic agent against lung cancer. PMID:27216943

  17. Ilex latifolia Prevents Amyloid β Protein (25-35)-Induced Memory Impairment by Inhibiting Apoptosis and Tau Phosphorylation in Mice.

    PubMed

    Kim, Joo Youn; Lee, Hong Kyu; Jang, Ji Yeon; Yoo, Jae Kuk; Seong, Yeon Hee

    2015-12-01

    Ilex latifolia Thunb. (Aquifoliaceae), a Chinese bitter tea called "kudingcha," has been widely consumed as a health beverage and found to possess antioxidant, antidiabetic, antihypertensive, anti-inflammatory, and anti-ischemic activities. The aim of the present study was to investigate the neuroprotective effects of an ethanol extract of I. latifolia against amyloid β protein (Aβ)-induced memory impairment in mice and neurotoxicity in cultured rat cortical neurons. Memory impairment in mice was induced by intracerebroventricular injection of 15 nmol Aβ (25-35) and measured by the passive avoidance test and Morris water maze test. Chronic administration of I. latifolia (25-100 mg/kg, p.o.) significantly prevented Aβ (25-35)-induced memory loss. I. latifolia also prevented the decrease of glutathione concentrations, increased lipid peroxidation, expression of phosphorylated tau (p-tau), and changes in apoptosis-associated proteins in the memory-impaired mouse brain. Exposure of cultured cortical neurons to 10 μM Aβ (25-35) for 36 h induced neuronal apoptotic death. The neuronal cell death, elevation of intracellular Ca(2+) concentration, generation of reactive oxygen species, and expression of proapoptotic proteins caused by Aβ (25-35) in the cultured neurons were inhibited by treatment with I. latifolia (1-50 μg/mL). These results suggest that I. latifolia may have a possible therapeutic role in managing cognitive impairment associated with Alzheimer's disease. The underlying mechanism might involve the antiapoptotic effects mediated by antioxidant activity and inhibition of p-tau formation. PMID:26291170

  18. Quercetin Improves Postischemic Recovery of Heart Function in Doxorubicin-Treated Rats and Prevents Doxorubicin-Induced Matrix Metalloproteinase-2 Activation and Apoptosis Induction

    PubMed Central

    Barteková, Monika; Šimončíková, Petra; Fogarassyová, Mária; Ivanová, Monika; Okruhlicová, Ľudmila; Tribulová, Narcisa; Dovinová, Ima; Barančík, Miroslav

    2015-01-01

    Quercetin (QCT) is flavonoid that possesses various biological functions including anti-oxidative and radical-scavenging activities. Moreover, QCT exerts some preventive actions in treatment of cardiovascular diseases. The aim of present study was to explore effects of prolonged administration of QCT on changes induced by repeated application of doxorubicin (DOX) in rat hearts. We focused on the ultrastructure of myocardium, matrix metalloproteinases (MMPs), biometric parameters, and apoptosis induction. Our aim was also to examine effects of QCT on ischemic tolerance in hearts exposed to chronic effects of DOX, and to determine possible mechanisms underlying effects of QCT. Our results showed that QCT prevented several negative chronic effects of DOX: (I) reversed DOX-induced blood pressure increase; (II) mediated improvement of deleterious effects of DOX on ultrastructure of left ventricle; (III) prevented DOX-induced effects on tissue MMP-2 activation; and (iv) reversed effects of DOX on apoptosis induction and superoxide dismutase inhibition. Moreover, we showed that rat hearts exposed to effects of QCT were more resistant to ischemia/reperfusion injury. Effects of QCT on modulation of ischemic tolerance were linked to Akt kinase activation and connexin-43 up-regulation. Taken together, these results demonstrate that prolonged treatment with QCT prevented negative chronic effects of DOX on blood pressure, cellular damage, MMP-2 activation, and apoptosis induction. Moreover, QCT influenced myocardial responses to acute ischemic stress. These facts bring new insights into mechanisms of QCT action on rat hearts exposed to the chronic effects of DOX. PMID:25872140

  19. Brain-derived Neurotrophic Factor Prevents Phencyclidine-induced Apoptosis in Developing Brain by Parallel Activation of both the ERK and PI-3K/Akt Pathways

    PubMed Central

    Xia, Yan; Wang, Cheng Z.; Liu, Jie; Anastasio, Noelle C.; Johnson, Kenneth M.

    2009-01-01

    Summary Phencyclidine is an N-methyl D-aspartate receptor (NMDAR) blocker that has been reported to induce neuronal apoptosis during development and schizophrenia-like behaviors in rats later in life. Brain derived neurotrophic factor (BDNF) has been shown to prevent neuronal death caused by NMDAR blockade, but the precise mechanism is unknown. This study examined the role of the phosphatidylinositol-3 kinase (PI3K)/Akt and extracellular signal-regulated kinase (ERK) pathways in BDNF protection of PCP-induced apoptosis in corticostriatal organotypic cultures. It was observed that BDNF inhibited PCP-induced apoptosis in a concentration dependent fashion. BDNF effectively prevented PCP-induced inhibition of the ERK and PI-3K/Akt pathways and suppressed GSK-3β activation. Blockade of either PI-3K/Akt or ERK activation abolished BDNF protection. Western blot analysis revealed that the PI-3K inhibitor LY294002 prevented the stimulating effect of BDNF on the PI-3K/Akt pathway, but had no effect on the ERK pathway. Similarly, the ERK inhibitor PD98059 prevented the stimulating effect of BDNF on the ERK pathway, but not the PI-3K/Akt pathway. Co-application of LY294002 and PD98059 had no additional effect on BDNF-evoked activation of Akt or ERK. However, concurrent exposure to PD98059 and LY294002 caused much greater inhibition of BDNF-evoked phosphorylation of GSK-3β at serine 9 than did LY294002 alone. Finally, either BDNF or GSK-3β inhibition prevented PCP-induced suppression of cyclic-AMP response element binding protein (CREB) phosphorylation. These data demonstrate that the protective effect of BDNF against PCP-induced apoptosis is mediated by parallel activation of the PI-3K/Akt and ERK pathways, most likely involves inhibition of GSK-3β and activation of CREB. PMID:19887077

  20. The metabolites of glutamine prevent hydroxyl radical-induced apoptosis through inhibiting mitochondria and calcium ion involved pathways in fish erythrocytes.

    PubMed

    Li, Huatao; Jiang, Weidan; Liu, Yang; Jiang, Jun; Zhang, Yongan; Wu, Pei; Zhao, Juan; Duan, Xudong; Zhou, Xiaoqiu; Feng, Lin

    2016-03-01

    The present study explored the apoptosis pathways in hydroxyl radicals ((∙)OH)-induced carp erythrocytes. Carp erythrocytes were treated with the caspase inhibitors in physiological carp saline (PCS) or Ca(2+)-free PCS in the presence of 40μM FeSO4/20μM H2O2. The results showed that the generation of reactive oxygen species (ROS), the release of cytochrome c and DNA fragmentation were caspase-dependent, and Ca(2+) was involved in calpain activation and phosphatidylserine (PS) exposure in (∙)OH-induced carp erythrocytes. Moreover, the results suggested that caspases were involved in PS exposure, and Ca(2+) was involved in DNA fragmentation in (∙)OH-induced fish erythrocytes. These results demonstrated that there might be two apoptosis pathways in fish erythrocytes, one is the caspase and cytochrome c-dependent apoptosis that is similar to that in mammal nucleated cells, the other is the Ca(2+)-involved apoptosis that was similar to that in mammal non-nucleated erythrocytes. So, fish erythrocytes may be used as a model for studying oxidative stress and apoptosis in mammal cells. Furthermore, the present study investigated the effects of glutamine (Gln)'s metabolites [alanine (Ala), citrulline (Cit), proline (Pro) and their combination (Ala10Pro4Cit1)] on the pathways of apoptosis in fish erythrocytes. The results displayed that Ala, Cit, Pro and Ala10Pro4Cit1 effectively suppressed ROS generation, cytochrome c release, activation of caspase-3, caspase-8 and caspase-9 at the physiological concentrations, prevented Ca(2+) influx, calpain activation, PS exposure, DNA fragmentation and the degradation of the cytoskeleton and oxidation of membrane and hemoglobin (Hb) and increased activity of anti-hydroxyl radical (AHR) in (∙)OH-induced carp erythrocytes. Ala10Pro4Cit1 produced a synergistic effect of inhibited oxidative stress and apoptosis in fish erythrocytes. These results demonstrated that Ala, Cit, Pro and their combination can protect mammal erythrocytes

  1. Rapamycin ameliorates cadmium-induced activation of MAPK pathway and neuronal apoptosis by preventing mitochondrial ROS inactivation of PP2A.

    PubMed

    Xu, Chong; Wang, Xiaoxue; Zhu, Yu; Dong, Xiaoqing; Liu, Chunxiao; Zhang, Hai; Liu, Lei; Huang, Shile; Chen, Long

    2016-06-01

    Cadmium (Cd) is a highly toxic metal that affects the central nervous system. Recently we have demonstrated that inhibition of mTOR by rapamycin rescues neuronal cells from Cd-poisoning. Here we show that rapamycin inhibited Cd-induced mitochondrial ROS-dependent neuronal apoptosis. Intriguingly, rapamycin remarkably blocked phosphorylation of JNK, Erk1/2 and p38 in neuronal cells induced by Cd, which was strengthened by co-treatment with Mito-TEMPO. Inhibition of JNK and Erk1/2 by SP600125 and U0126, respectively, potentiated rapamycin's prevention from Cd-induced apoptosis. Consistently, over-expression of dominant negative c-Jun or MKK1 also potently improved the inhibitory effect of rapamycin on Cd neurotoxicity. Furthermore, pretreatment with SP600125 or U0126, or expression of dominant negative c-Jun or MKK1 enhanced the inhibitory effects of rapamycin or Mito-TEMPO on Cd-induced ROS. Further investigation found that co-treatment with Mito-TEMPO/rapamycin more effectively rescued cells by preventing Cd inactivation of PP2A than treatment with rapamycin or Mito-TEMPO alone. Over-expression of wild-type PP2A reinforced rapamycin or Mito-TEMPO suppression of activated JNK and Erk1/2 pathways, as well as ROS production and apoptosis in neuronal cells in response to Cd. The findings indicate that rapamycin ameliorates Cd-evoked neuronal apoptosis by preventing mitochondrial ROS inactivation of PP2A, thereby suppressing activation of JNK and Erk1/2 pathways. Our results underline that rapamycin may have a potential in preventing Cd-induced oxidative stress and neurodegenerative diseases. PMID:26805420

  2. A novel peptide, colivelin, prevents alcohol-induced apoptosis in fetal brain of C57BL/6 mice: signaling pathway investigations

    PubMed Central

    Sari, Youssef; Chiba, Tomohiro; Yamada, Marina; Rebec, George V.; Aiso, Sadakazu

    2009-01-01

    Fetal alcohol exposure is known to induce cell death through apoptosis. We found that colivelin (CLN), a novel peptide with the sequence SALLRSIPAPAGASRLLLLTGEIDLP, prevents this apoptosis. Our initial experiment revealed that CLN enhanced the viability of primary cortical neurons exposed to alcohol. We then used a mouse model of fetal alcohol exposure to identify the intracellular mechanisms underlying these neuroprotective effects. On embryonic day 7 (E7), weight-matched pregnant females were assigned to the following groups: (1) ethanol liquid diet (ALC) 25% (4.49%, v/v) ethanol derived calories; (2) pair-fed control; (3) normal chow; (4) ALC combined with administration (i.p.) of CLN (20 μg/20 g body weight); and (5) pair-fed combined with administration (i.p.) of CLN (20 μg/20 g body weight). On E13, fetal brains were collected and assayed for TUNEL staining, caspase-3 colorimetric assay, ELISA, and MSD electrochemiluminescence. CLN blocked the alcohol-induced decline in brain weight and prevented alcohol-induced: apoptosis, activation of caspase-3 and increases of cytosolic cytochrome c, and decreases of mitochondrial cytochrome c. Analysis of proteins in the upstream signaling pathway revealed that CLN down-regulated the phosphorylation of the c-Jun N-terminal kinase. Moreover, CLN prevented alcohol-induced reduction in phosphorylation of BAD protein. Thus, CLN appears to act directly on upstream signaling proteins to prevent alcohol-induced apoptosis. Further assessment of these proteins and their signaling mechanisms is likely to enhance development of neuroprotective therapies. PMID:19782727

  3. Saponin-rich fraction from Clematis chinensis Osbeck roots protects rabbit chondrocytes against nitric oxide-induced apoptosis via preventing mitochondria impairment and caspase-3 activation.

    PubMed

    Wu, Wenjun; Gao, Xinghua; Xu, Xianxiang; Luo, Yubin; Liu, Mei; Xia, Yufeng; Dai, Yue

    2013-03-01

    Our previous study reported that the saponin-rich fraction from Clematis chinensis Osbeck roots (SFC) could effectively alleviate experimental osteoarthritis induced by monosodium iodoacetate in rats through protecting articular cartilage and inhibiting local inflammation. The present study was performed to investigate the preventive effects of SFC on articular chondrocyte, and explore the underlying mechanisms. Primary rabbit chondrocytes were cultured and exposed to sodium nitroprusside (SNP), a NO donor. After treatment with different concentrations of SFC (30, 100, 300, 1,000 μg/ml) for 24 h, nucleic morphology, apoptotic rate, mitochondrial function and caspase-3 activity of chondrocytes were examined. The results showed that SNP induced remarkable apoptosis of rabbit chondrocytes evidenced by Hoechst 33258 staining and flow cytometry analysis, and SFC prevented the apoptosis in a concentration-dependent manner. Further studies indicated that SFC could prevent the depolarization of mitochondrial membrane potential (∆ψm) in SNP-treated chondrocytes and suppress the activation of caspase-3. It can be concluded that the protection of SFC on articular chondrocytes is associated with the anti-apoptosis effects via inhibiting the mitochondrion impairment and caspase-3 activation. PMID:22821055

  4. Haploinsufficiency for Core Exon Junction Complex Components Disrupts Embryonic Neurogenesis and Causes p53-Mediated Microcephaly.

    PubMed

    Mao, Hanqian; McMahon, John J; Tsai, Yi-Hsuan; Wang, Zefeng; Silver, Debra L

    2016-09-01

    The exon junction complex (EJC) is an RNA binding complex comprised of the core components Magoh, Rbm8a, and Eif4a3. Human mutations in EJC components cause neurodevelopmental pathologies. Further, mice heterozygous for either Magoh or Rbm8a exhibit aberrant neurogenesis and microcephaly. Yet despite the requirement of these genes for neurodevelopment, the pathogenic mechanisms linking EJC dysfunction to microcephaly remain poorly understood. Here we employ mouse genetics, transcriptomic and proteomic analyses to demonstrate that haploinsufficiency for each of the 3 core EJC components causes microcephaly via converging regulation of p53 signaling. Using a new conditional allele, we first show that Eif4a3 haploinsufficiency phenocopies aberrant neurogenesis and microcephaly of Magoh and Rbm8a mutant mice. Transcriptomic and proteomic analyses of embryonic brains at the onset of neurogenesis identifies common pathways altered in each of the 3 EJC mutants, including ribosome, proteasome, and p53 signaling components. We further demonstrate all 3 mutants exhibit defective splicing of RNA regulatory proteins, implying an EJC dependent RNA regulatory network that fine-tunes gene expression. Finally, we show that genetic ablation of one downstream pathway, p53, significantly rescues microcephaly of all 3 EJC mutants. This implicates p53 activation as a major node of neurodevelopmental pathogenesis following EJC impairment. Altogether our study reveals new mechanisms to help explain how EJC mutations influence neurogenesis and underlie neurodevelopmental disease. PMID:27618312

  5. CaMKII inhibition in type II pneumocytes protects from bleomycin-induced pulmonary fibrosis by preventing Ca2+-dependent apoptosis.

    PubMed

    Winters, Christopher J; Koval, Olha; Murthy, Shubha; Allamargot, Chantal; Sebag, Sara C; Paschke, John D; Jaffer, Omar A; Carter, A Brent; Grumbach, Isabella M

    2016-01-01

    The calcium and calmodulin-dependent kinase II (CaMKII) translates increases in intracellular Ca(2+) into downstream signaling events. Its function in pulmonary pathologies remains largely unknown. CaMKII is a well-known mediator of apoptosis and regulator of endoplasmic reticulum (ER) Ca(2+). ER stress and apoptosis of type II pneumocytes lead to aberrant tissue repair and progressive collagen deposition in pulmonary fibrosis. Thus we hypothesized that CaMKII inhibition alleviates fibrosis in response to bleomycin by attenuating apoptosis and ER stress of type II pneumocytes. We first established that CaMKII was strongly expressed in the distal respiratory epithelium, in particular in surfactant protein-C-positive type II pneumocytes, and activated after bleomycin instillation. We generated a novel transgenic model of inducible expression of the CaMKII inhibitor peptide AC3-I limited to type II pneumocytes (Tg SPC-AC3-I). Tg SPC-AC3-I mice were protected from development of pulmonary fibrosis after bleomycin exposure compared with wild-type mice. CaMKII inhibition also provided protection from apoptosis in type II pneumocytes in vitro and in vivo. Moreover, intracellular Ca(2+) levels and ER stress were increased by bleomycin and significantly blunted with CaMKII inhibition in vitro. These data demonstrate that CaMKII inhibition prevents type II pneumocyte apoptosis and development of pulmonary fibrosis in response to bleomycin. CaMKII inhibition may therefore be a promising approach to prevent or ameliorate the progression of pulmonary fibrosis. PMID:26545899

  6. Brca1 is required for embryonic development of the mouse cerebral cortex to normal size by preventing apoptosis of early neural progenitors.

    PubMed

    Pulvers, Jeremy N; Huttner, Wieland B

    2009-06-01

    The extent of apoptosis of neural progenitors is known to influence the size of the cerebral cortex. Mouse embryos lacking Brca1, the ortholog of the human breast cancer susceptibility gene BRCA1, show apoptosis in the neural tube, but the consequences of this for brain development have not been studied. Here we investigated the role of Brca1 during mouse embryonic cortical development by deleting floxed Brca1 using Emx1-Cre, which leads to conditional gene ablation specifically in the dorsal telencephalon after embryonic day (E) 9.5. The postnatal Brca1-ablated cerebral cortex was substantially reduced in size with regard to both cortical thickness and surface area. Remarkably, although the thickness of the cortical layers (except for the upper-most layer) was decreased, cortical layering as such was essentially unperturbed. High levels of apoptosis were found at E11.5 and E13.5, but dropped to near-control levels by E16.5. The apoptosis at the early stage of neurogenesis occurred in both BrdU pulse-labeled neural progenitors and the neurons derived therefrom. No changes were observed in the mitotic index of apical (neuroepithelial, radial glial) progenitors and basal (intermediate) progenitors, indicating that Brca1 ablation did not affect cell cycle progression. Brca1 ablation did, however, result in the nuclear translocation of p53 in neural progenitors, suggesting that their apoptosis involved activation of the p53 pathway. Our results show that Brca1 is required for the cerebral cortex to develop to normal size by preventing the apoptosis of early cortical progenitors and their immediate progeny. PMID:19403657

  7. Targeting Apoptosis Signalling Kinase-1 (ASK-1) Does Not Prevent the Development of Neuropathy in Streptozotocin-Induced Diabetic Mice

    PubMed Central

    Newton, Victoria L.; Ali, Sumia; Duddy, Graham; Whitmarsh, Alan J.; Gardiner, Natalie J.

    2014-01-01

    Apoptosis signal-regulating kinase-1 (ASK1) is a mitogen-activated protein 3 kinase (MAPKKK/MAP3K) which lies upstream of the stress-activated MAPKs, JNK and p38. ASK1 may be activated by a variety of extracellular and intracellular stimuli. MAP kinase activation in the sensory nervous system as a result of diabetes has been shown in numerous preclinical and clinical studies. As a common upstream activator of both p38 and JNK, we hypothesised that activation of ASK1 contributes to nerve dysfunction in diabetic neuropathy. We therefore wanted to characterize the expression of ASK1 in sensory neurons, and determine whether the absence of functional ASK1 would protect against the development of neuropathy in a mouse model of experimental diabetes. ASK1 mRNA and protein is constitutively expressed by multiple populations of sensory neurons of the adult mouse lumbar DRG. Diabetes was induced in male C57BL/6 and transgenic ASK1 kinase-inactive (ASK1n) mice using streptozotocin. Levels of ASK1 do not change in the DRG, spinal cord, or sciatic nerve following induction of diabetes. However, levels of ASK2 mRNA increase in the spinal cord at 4 weeks of diabetes, which could represent a future target for this field. Neither motor nerve conduction velocity deficits, nor thermal or mechanical hypoalgesia were prevented or ameliorated in diabetic ASK1n mice. These results suggest that activation of ASK1 is not responsible for the nerve deficits observed in this mouse model of diabetic neuropathy. PMID:25329046

  8. Inhibitor of apoptosis-stimulating protein of p53 (iASPP) prevents senescence and is required for epithelial stratification

    PubMed Central

    Notari, Mario; Hu, Ying; Koch, Sofia; Lu, Min; Ratnayaka, Indrika; Zhong, Shan; Baer, Caroline; Pagotto, Anna; Goldin, Robert; Salter, Victoria; Candi, Eleonora; Melino, Gerry; Lu, Xin

    2011-01-01

    Inhibitor of apoptosis-stimulating protein of p53 (iASPP) is the most ancient member of the ASPP family of proteins and an evolutionarily conserved inhibitor of p53. iASPP is also a binding partner and negative regulator of p65RelA. Because p65RelA and the p53 family members often have opposite effects in controlling cell fate, it is important to understand the cellular context in which iASPP can regulate their activities. To address this question and to study the biological importance of iASPP in vivo, we generated a transgenic mouse in which iASPP expression is controlled by the Cre/loxP recombination system. We observed that iASPP is able to prevent premature cellular senescence in mouse embryonic fibroblasts. iASPP loss resulted in increased differentiation of primary keratinocytes both in vitro and in vivo. In stratified epithelia, nuclear iASPP often colocalized with p63 in the nuclei of basal keratinocytes. Consistent with this, iASPP bound p63 and inhibited the transcriptional activity of both TAp63α and ΔNp63α in vitro and influenced the expression level of p63-regulated genes such as loricrin and involucrin in vivo. In contrast, under the same conditions, p65RelA was frequently expressed as a cytoplasmic protein in the suprabasal layers of stratified epithelia and rarely colocalized with nuclear iASPP. Thus, iASPP is likely to control epithelial stratification by regulating p63's transcriptional activity, rather than p65RelA's. This study identifies iASPP as an inhibitor of senescence and a key player in controlling epithelial stratification. PMID:21930934

  9. Radiation-Induced Salivary Gland Dysfunction Results From p53-Dependent Apoptosis

    SciTech Connect

    Avila, Jennifer L.; Grundmann, Oliver; Burd, Randy; Limesand, Kirsten H.

    2009-02-01

    Purpose: Radiotherapy for head-and-neck cancer causes adverse secondary side effects in the salivary glands and results in diminished quality of life for the patient. A previous in vivo study in parotid salivary glands demonstrated that targeted head-and-neck irradiation resulted in marked increases in phosphorylated p53 (serine{sup 18}) and apoptosis, which was suppressed in transgenic mice expressing a constitutively active mutant of Akt1 (myr-Akt1). Methods and Materials: Transgenic and knockout mouse models were exposed to irradiation, and p53-mediated transcription, apoptosis, and salivary gland dysfunction were analyzed. Results: The proapoptotic p53 target genes PUMA and Bax were induced in parotid salivary glands of mice at early time points after therapeutic radiation. This dose-dependent induction requires expression of p53 because no radiation-induced expression of PUMA and Bax was observed in p53-/- mice. Radiation also induced apoptosis in the parotid gland in a dose-dependent manner, which was p53 dependent. Furthermore, expression of p53 was required for the acute and chronic loss of salivary function after irradiation. In contrast, apoptosis was not induced in p53-/- mice, and their salivary function was preserved after radiation exposure. Conclusions: Apoptosis in the salivary glands after therapeutic head-and-neck irradiation is mediated by p53 and corresponds to salivary gland dysfunction in vivo.

  10. EAF2 mediates germinal centre B-cell apoptosis to suppress excessive immune responses and prevent autoimmunity.

    PubMed

    Li, Yingqian; Takahashi, Yoshimasa; Fujii, Shin-ichiro; Zhou, Yang; Hong, Rongjian; Suzuki, Akari; Tsubata, Takeshi; Hase, Koji; Wang, Ji-Yang

    2016-01-01

    Regulated apoptosis of germinal centre (GC) B cells is critical for normal humoral immune responses. ELL-associated factor 2 (EAF2) regulates transcription elongation and has been shown to be an androgen-responsive potential tumour suppressor in prostate by inducing apoptosis. Here we show that EAF2 is selectively upregulated in GC B cells among various immune cell types and promotes apoptosis of GC B cells both in vitro and in vivo. EAF2 deficiency results in enlarged GCs and elevated antibody production during a T-dependent immune response. After immunization with type II collagen, mice lacking EAF2 produce high levels of collagen-specific autoantibodies and rapidly develop severe arthritis. Moreover, the mutant mice spontaneously produce anti-dsDNA, rheumatoid factor and anti-nuclear antibodies as they age. These results demonstrate that EAF2-mediated apoptosis in GC B cells limits excessive humoral immune responses and is important for maintaining self-tolerance. PMID:26935903

  11. Upregulation of lymphocyte apoptosis as a strategy for preventing and treating autoimmune disorders: a role for whole-food vegan diets, fish oil and dopamine agonists.

    PubMed

    McCarty, M F

    2001-08-01

    Induced apoptosis of autoreactive T-lymphocyte precursors in the thymus is crucial for the prevention of autoimmune disorders. IGF-I and prolactin, which are lymphocyte growth factors, may have the potential to suppress apoptosis in thymocytes and thus encourage autoimmunity; conversely, dietary fish oil rich in omega-3 fats appears to upregulate apoptosis in lymphocytes. Since whole-food vegan diets may downregulate systemic IGF-I activity, it is proposed that such a diet, in conjunction with fish oil supplementation and treatment with dopamine agonists capable of suppressing prolactin secretion, may have utility for treating and preventing autoimmune disorders. This prediction is consistent with the extreme rarity of autoimmune disorders among sub-Saharan black Africans as long as they followed their traditional quasi-vegan lifestyles, and with recent ecologic studies correlating risks for IDDM and for multiple sclerosis mortality with animal product and/or saturated fat consumption. Moreover, there is evidence that vegan or quasi-vegan diets are useful in the management of rheumatoid arthritis, multiple sclerosis, and possibly SLE. The dopamine agonist bromocryptine exerts anti-inflammatory effects in rodent models of autoimmunity, and there is preliminary evidence that this drug may be clinically useful in several human autoimmune diseases; better tolerated D2-specific agonists such as cabergoline may prove to be more practical for use in therapy. The moderate clinical utility of supplemental fish oil in rheumatoid arthritis and certain other autoimmune disorders is documented. It is not unlikely that extra-thymic anti-inflammatory effects contribute importantly to the clinical utility of vegan diets, bromocryptine, and fish oil in autoimmunity. The favorable impact of low latitude or high altitude on autoimmune risk may be mediated by superior vitamin D status, which is associated with decreased secretion of parathyroid hormone; there are theoretical grounds

  12. All-trans retinoic acid stimulates gene expression of the cardioprotective natriuretic peptide system and prevents fibrosis and apoptosis in cardiomyocytes of obese ob/ob mice.

    PubMed

    Manolescu, Daniel-Constantin; Jankowski, Marek; Danalache, Bogdan A; Wang, Donghao; Broderick, Tom L; Chiasson, Jean-Louis; Gutkowska, Jolanta

    2014-10-01

    In hypertensive rodents, retinoic acid (RA) prevents adverse cardiac remodelling and improves myocardial infarction outcome, but its role in obesity-related changes of cardiac tissue are unclear. We hypothesized that all-trans RA (ATRA) treatment will improve the cardioprotective oxytocin-natriuretic peptides (OT-NP) system, preventing apoptosis and collagen accumulation in hearts of ob/ob mice, a mouse model of obesity and insulin resistance. Female 9-week-old B6.V-Lep/J ob/ob mice (n = 16) were divided into 2 groups: 1 group (n = 8) treated with 100 μg of ATRA dissolved in 100 μL of corn oil (vehicle) delivered daily (∼2 μg·g body weight(-1)·day(-1)) by stomach intubation for 16 days, and 1 group (n = 8) that received the vehicle alone. A group of nonobese littermate mice (n = 9) served as controls. Ob/ob mice exhibited obesity, hyperglycaemia, and downregulation of the cardiac OT-NP system, including the mRNA for the transcription factor GATA4, OT receptor and brain NP, and the protein expression for endothelial nitric oxide synthase. Hearts from ob/ob mice also demonstrated increased apoptosis and collagen accumulation. ATRA treatment induced weight loss and decreased adipocytes diameter in the visceral fat, thus reducing visceral obesity, which is associated with a high risk for cardiovascular disease. RA treatment was associated with a reduction in hyperglycemia and a normalization of the OT-NP system's expression in the hearts of ob/ob mice. Furthermore, ATRA treatment prevented apoptosis and collagen accumulation in hearts of ob/ob mice. The present study indicates that ATRA treatment was effective in restoring the cardioprotective OT-NP system and in preventing abnormal cardiac remodelling in the ob/ob mice. PMID:25017112

  13. TGF-β1 prevents simulated ischemia/reperfusion-induced cardiac fibroblast apoptosis by activation of both canonical and non-canonical signaling pathways.

    PubMed

    Vivar, Raúl; Humeres, Claudio; Ayala, Pedro; Olmedo, Ivonne; Catalán, Mabel; García, Lorena; Lavandero, Sergio; Díaz-Araya, Guillermo

    2013-06-01

    Ischemia/reperfusion injury is a major cause of myocardial death. In the heart, cardiac fibroblasts play a critical role in healing post myocardial infarction. TGF-β1 has shown cardioprotective effects in cardiac damage; however, if TGF-β1 can prevent cardiac fibroblast death triggered by ischemia/reperfusion is unknown. Therefore, we test this hypothesis, and whether the canonical and/or non-canonical TGF-β1 signaling pathways are involved in this protective effect. Cultured rat cardiac fibroblasts were subjected to simulated ischemia/reperfusion. Cell viability was analyzed by trypan blue exclusion and propidium iodide by flow cytometry. The processing of procaspases 8, 9 and 3 to their active forms was assessed by Western blot, whereas subG1 population was evaluated by flow cytometry. Levels of total and phosphorylated forms of ERK1/2, Akt and Smad2/3 were determined by Western blot. The role of these signaling pathways on the protective effect of TGF-β1 was studied using specific chemical inhibitors. Simulated ischemia over 8h triggers a significant cardiac fibroblast death, which increased by reperfusion, with apoptosis actively involved. These effects were only prevented by the addition of TGF-β1 during reperfusion. TGF-β1 pretreatment increased the levels of phosphorylated forms of ERK1/2, Akt and Smad2/3. The inhibition of ERK1/2, Akt and Smad3 also blocked the preventive effects of TGF-β1 on cardiac fibroblast apoptosis induced by simulated ischemia/reperfusion. Overall, our data suggest that TGF-β1 prevents cardiac fibroblast apoptosis induced by simulated ischemia-reperfusion through the canonical (Smad3) and non canonical (ERK1/2 and Akt) signaling pathways. PMID:23416528

  14. Novel function of CRTH2 in preventing apoptosis of human Th2 cells through activation of the phosphatidylinositol 3-kinase pathway.

    PubMed

    Xue, Luzheng; Barrow, Anna; Pettipher, Roy

    2009-06-15

    It is now well established that interaction of PGD(2) with chemoattractant receptor- homologous molecule expressed on Th2 cells (CRTH2) promotes chemotaxis and proinflammatory cytokine production by Th2 lymphocytes. In this study we show a novel function of CRTH2 in mediating an inhibitory effect of PGD(2) on the apoptosis of human Th2 cells induced by cytokine deprivation. This effect was mimicked by the selective CRTH2 agonist 13,14-dihydro-15-keto-PGD(2), inhibited by the CRTH2 antagonists ramatroban and TM30089, and not observed in CRTH2-negative T cells. D prostanoid receptor 1 (DP(1)) or the thromboxane-like prostanoid (TP) receptor did not play a role in mediating the effects of PGD(2) on the apoptosis of Th2 cells because neither the DP(1) antagonist BW868C nor the TP antagonist SQ29548 had any effect on the antiapoptotic effect of PGD(2). Apoptosis of Th2 cells induced by Fas ligation was not suppressed by treatment with PGD(2), illustrating that activation of CRTH2 only inhibits apoptosis induced by cytokine deprivation. Treatment with PGD(2) induced phosphorylation of Akt and BAD, prevented release of cytochrome c from mitochondria, and suppressed cleavage of caspase-3 and poly(ADP-ribose) polymerase in Th2 cells deprived of IL-2. The PI3K inhibitor LY294002 blocked the effect of PGD(2) both on the signaling events and on the apoptotic death of Th2 cells. These data suggest that in addition to promoting the recruitment and activation of Th2 cells, PGD(2) may also impede the resolution of allergic inflammation through inhibiting apoptosis of Th2 cells. PMID:19494281

  15. Myricitrin attenuates endothelial cell apoptosis to prevent atherosclerosis: An insight into PI3K/Akt activation and STAT3 signaling pathways.

    PubMed

    Qin, Meng; Luo, Yun; Meng, Xiang-bao; Wang, Min; Wang, Hong-wei; Song, Shi-yu; Ye, Jing-xue; Pan, Rui-le; Yao, Fan; Wu, Ping; Sun, Gui-bo; Sun, Xiao-bo

    2015-07-01

    Blood vessel endothelial dysfunction induced by oxidized low-density lipoprotein (ox-LDL) has been implicated in the pathogenesis of atherosclerosis and vasculopathy. The ox-LDL-elicited reactive oxygen species (ROS) release has been assumed to serve a critical function in endothelial damage. Myricitrin (from Myrica cerifera) is a natural antioxidant that has strong anti-oxidative, anti-inflammatory, and anti-nociceptive activities. However, the protective effect of myricitrin on ROS-induced endothelial cell injury and its related molecular mechanisms have never been investigated. This study demonstrates that myricitrin can inhibit ox-LDL-induced endothelial apoptosis and prevent plaque formation at an early stage in an atherosclerotic mouse model. The administration of myricitrin in vivo decreases the thickness of the vascular wall in the aortic arch of ApoE-/- mice. In vitro study shows that ox-LDL-induced human umbilical vein endothelial cell apoptosis can be reduced upon receiving myricitrin pre-treatment. Treatment with myricitrin significantly attenuated ox-LDL-induced endothelial cell apoptosis by inhibiting LOX-1 expression and by increasing the activation of the STAT3 and PI3K/Akt/eNOS signaling pathways. At the same time, our result demonstrates that myricitrin treatment optimizes the balance of pro/anti-apoptosis proteins, including Bax, Bad, XIAP, cIAP-2, and survivin. Our study suggests that myricitrin treatment can effectively protect cells from ox-LDL-induced endothelial cell apoptosis, which results in reduced atherosclerotic plaque formation. This result indicates that myricitrin can be used as a drug candidate for the treatment of cardiovascular diseases. PMID:25849952

  16. Bezafibrate prevents palmitate-induced apoptosis in osteoblastic MC3T3-E1 cells through the NF-κB signaling pathway.

    PubMed

    Zhong, Xing; Xiu, Lingling; Wei, Guohong; Pan, Tianrong; Liu, Yuanyuan; Su, Lei; Li, Yanbing; Xiao, Haipeng

    2011-10-01

    Osteoporosis is a bone condition defined by low bone mass and increase of fracture risk due to imbalance between bone resorption by osteoclasts and bone formation by osteoblasts. Low bone mass is likely to be due to the alteration of the osteoclast and osteoblast lifespan through regulated apoptosis. Saturated fatty acid (SFA) intake is negatively associated with bone mineral density (BMD). Furthermore, SFA induces apoptosis in osteoblastic cell lines. Bezafibrate could increase bone mass in intact male rats principally through increasing periosteal bone formation. At present, it is unknown whether bezafibrate attenuates palmitate-induced apoptosis in MC3T3-E1 cells. In the present study, we found that palmitate stimulated the degradation of IκBα and NF-κB translocation, as well as up-regulation of NF-κB-mediated Fas expression in obsteoblastic MC3T3-E1 cells. Furthermore, the NF-κB inhibitor pyrrolidine dithiocarbamate (PDTC) could restore palmitate-induced caspase-3 decrease and inhibit palmitate-induced cleaved caspase-3 increase. We observed that bezafibrate, a dual ligand for the peroxisome proliferator-activated receptors α (PPARα) and PPARδ, significantly attenuated the palmitate-induced cytotoxicity as determined by the MTT assay and inhibited the palmitate-induced apoptosis as determined by a flow cytometry assay using Annexin V-FITC/PI and assessment of the activity of caspase-3. Pre-treatment of bezafibrate prevented palmitate-induced NF-κB activation. Therefore, these findings indicate that bezafibrate inbibits palmitate-induced apoptosis via the NF-κB signaling pathway. Our results point to bezafibrate as a new strategy to attenuate bone loss associated with high fat diet beyond its lipid-lowering actions. PMID:21687928

  17. Glutamine supplementation prevents exercise-induced neutrophil apoptosis and reduces p38 MAPK and JNK phosphorylation and p53 and caspase 3 expression.

    PubMed

    Lagranha, Claudia J; Hirabara, Sandro M; Curi, Rui; Pithon-Curi, Tania C

    2007-01-01

    We have previously shown that a single session of exercise induces DNA fragmentation, mitochondrial membrane depolarization, increases expression of pro-apoptotic genes (bax and bcl-xS) and decreases expression of anti-apoptotic genes (bcl-xL) in rat neutrophils. Glutamine supplementation had a protective effect in the apoptosis induced by a single session of exercise. The mechanism involved in the effect of single session of exercise to induce apoptosis was investigated by measuring expression of p53 and caspase 3 and phosphorylation of p38 mitogen-activated protein kinases (MAPK) and cJun NH(2)-terminal kinase (JNK) in neutrophils from rats supplemented or not with glutamine. Exercise was carried out on a treadmill for 1 h and the rats were killed by decapitation. Neutrophils were obtained by intraperitoneal (i.p.) lavage with PBS, 4 h after injection of oyster glycogen solution. Glutamine supplementation (1g per Kg b.w.) was given by gavage 1 h before the exercise session. Gene expression and protein phosphorylation were then analyzed by reverse transcriptase chain reaction (RT-PCR) and Western blotting, respectively. A single session of exercise increased p38 MAPK and JNK phosphorylation and p53 and caspase 3 expression. Glutamine supplementation partially prevented the increase in p38 MAPK and JNK phosphorylation and p53 expression, and fully abolished the increase in caspase 3 expression. Thus, neutrophil apoptosis induced by a single session of exercise is accompanied by increased p53 and caspase 3 expression and p38 MAPK and JNK phosphorylation. Glutamine supplementation prevents these effects of exercise and reduces apoptosis. PMID:17542038

  18. Inhibition of NOS-NO System Prevents Autoimmune Orchitis Development in Rats: Relevance of NO Released by Testicular Macrophages in Germ Cell Apoptosis and Testosterone Secretion

    PubMed Central

    Jarazo Dietrich, Sabrina; Fass, Mónica Irina; Jacobo, Patricia Verónica; Sobarzo, Cristian Marcelo Alejandro; Lustig, Livia; Theas, María Susana

    2015-01-01

    Background Although the testis is considered an immunoprivileged organ it can orchestrate immune responses against pathological insults such as infection and trauma. Experimental autoimmune orchitis (EAO) is a model of chronic inflammation whose main histopathological features it shares with human orchitis. In EAO an increased number of macrophages infiltrate the interstitium concomitantly with progressive germ cell degeneration and impaired steroidogenesis. Up-regulation of nitric oxide (NO)-NO synthase (NOS) system occurs, macrophages being the main producers of NO. Objective The aim of our study was to evaluate the role of NO-NOS system in orchitis development and determine the involvement of NO released by testicular macrophages on germ cell apoptosis and testosterone secretion. Method and Results EAO was induced in rats by immunization with testicular homogenate and adjuvants (E group) and a group of untreated normal rats (N) was also studied. Blockage of NOS by i.p. injection of E rats with a competitive inhibitor of NOS, L-NAME (8mg/kg), significantly reduced the incidence and severity of orchitis and lowered testicular nitrite content. L-NAME reduced germ cell apoptosis and restored intratesticular testosterone levels, without variations in serum LH. Co-culture of N testicular fragments with testicular macrophages obtained from EAO rats significantly increased germ cell apoptosis and testosterone secretion, whereas addition of L-NAME lowered both effects and reduced nitrite content. Incubation of testicular fragments from N rats with a NO donor DETA-NOnoate (DETA-NO) induced germ cell apoptosis through external and internal apoptotic pathways, an effect prevented by N-acetyl-L-cysteine (NAC). DETA-NO inhibited testosterone released from Leydig cells, whereas NAC (from 2.5 to 15 mM) did not prevent this effect. Conclusions We demonstrated that NO-NOS system is involved in the impairment of testicular function in orchitis. NO secreted mainly by testicular

  19. MicroRNA-26a prevents endothelial cell apoptosis by directly targeting TRPC6 in the setting of atherosclerosis

    PubMed Central

    Zhang, Yong; Qin, Wei; Zhang, Longyin; Wu, Xianxian; Du, Ning; Hu, Yingying; Li, Xiaoguang; Shen, Nannan; Xiao, Dan; Zhang, Haiying; Li, Zhange; Zhang, Yue; Yang, Huan; Gao, Feng; Du, Zhimin; Xu, Chaoqian; Yang, Baofeng

    2015-01-01

    Atherosclerosis, a chronic inflammatory disease, is the major cause of life-threatening complications such as myocardial infarction and stroke. Endothelial apoptosis plays a vital role in the initiation and progression of atherosclerotic lesions. Although a subset of microRNAs (miRs) have been identified as critical regulators of atherosclerosis, studies on their participation in endothelial apoptosis in atherosclerosis have been limited. In our study, we found that miR-26a expression was substantially reduced in the aortic intima of ApoE−/− mice fed with a high-fat diet (HFD). Treatment of human aortic endothelial cells (HAECs) with oxidized low-density lipoprotein (ox-LDL) suppressed miR-26a expression. Forced expression of miR-26a inhibited endothelial apoptosis as evidenced by MTT assay and TUNEL staining results. Further analysis identified TRPC6 as a target of miR-26a, and TRPC6 overexpression abolished the anti-apoptotic effect of miR-26a. Moreover, the cytosolic calcium and the mitochondrial apoptotic pathway were found to mediate the beneficial effects of miR-26a on endothelial apoptosis. Taken together, our study reveals a novel role of miR-26a in endothelial apoptosis and indicates a therapeutic potential of miR-26a for atherosclerosis associated with apoptotic cell death. PMID:25801675

  20. Preventive effects of imperatorin on perfluorohexanesulfonate-induced neuronal apoptosis via inhibition of intracellular calcium-mediated ERK pathway.

    PubMed

    Lee, Eunkyung; Choi, So-Young; Yang, Jae-Ho; Lee, Youn Ju

    2016-07-01

    Early life neuronal exposure to environmental toxicants has been suggested to be an important etiology of neurodegenerative disease development. Perfluorohexanesulfonate (PFHxS), one of the major perfluoroalkyl compounds, is widely distributed environmental contaminants. We have reported that PFHxS induces neuronal apoptosis via ERK-mediated pathway. Imperatorin is a furanocoumarin found in various edible plants and has a wide range of pharmacological effects including neuroprotection. In this study, the effects of imperatorin on PFHxS-induced neuronal apoptosis and the underlying mechanisms are examined using cerebellar granule cells (CGC). CGC were isolated from seven-day old rats and were grown in culture for seven days. Caspase-3 activity and TUNEL staining were used to determine neuronal apoptosis. PFHxS-induced apoptosis of CGC was significantly reduced by imperatorin and PD98059, an ERK pathway inhibitor. PFHxS induced a persistent increase in intracellular calcium, which was significantly blocked by imperatorin, NMDA receptor antagonist, MK801 and the L-type voltage-dependent calcium channel blockers, diltiazem and nifedipine. The activation of caspase-3 by PFHxS was also inhibited by MK801, diltiazem and nifedipine. PFHxS-increased ERK activation was inhibited by imperatorin, MK801, diltiazem and nifedipine. Taken together, imperatorin protects CGC against PFHxS-induced apoptosis via inhibition of NMDA receptor/intracellular calcium-mediated ERK pathway. PMID:27382356

  1. MicroRNA-26a prevents endothelial cell apoptosis by directly targeting TRPC6 in the setting of atherosclerosis

    NASA Astrophysics Data System (ADS)

    Zhang, Yong; Qin, Wei; Zhang, Longyin; Wu, Xianxian; Du, Ning; Hu, Yingying; Li, Xiaoguang; Shen, Nannan; Xiao, Dan; Zhang, Haiying; Li, Zhange; Zhang, Yue; Yang, Huan; Gao, Feng; Du, Zhimin; Xu, Chaoqian; Yang, Baofeng

    2015-03-01

    Atherosclerosis, a chronic inflammatory disease, is the major cause of life-threatening complications such as myocardial infarction and stroke. Endothelial apoptosis plays a vital role in the initiation and progression of atherosclerotic lesions. Although a subset of microRNAs (miRs) have been identified as critical regulators of atherosclerosis, studies on their participation in endothelial apoptosis in atherosclerosis have been limited. In our study, we found that miR-26a expression was substantially reduced in the aortic intima of ApoE-/- mice fed with a high-fat diet (HFD). Treatment of human aortic endothelial cells (HAECs) with oxidized low-density lipoprotein (ox-LDL) suppressed miR-26a expression. Forced expression of miR-26a inhibited endothelial apoptosis as evidenced by MTT assay and TUNEL staining results. Further analysis identified TRPC6 as a target of miR-26a, and TRPC6 overexpression abolished the anti-apoptotic effect of miR-26a. Moreover, the cytosolic calcium and the mitochondrial apoptotic pathway were found to mediate the beneficial effects of miR-26a on endothelial apoptosis. Taken together, our study reveals a novel role of miR-26a in endothelial apoptosis and indicates a therapeutic potential of miR-26a for atherosclerosis associated with apoptotic cell death.

  2. Preventive effects of imperatorin on perfluorohexanesulfonate-induced neuronal apoptosis via inhibition of intracellular calcium-mediated ERK pathway

    PubMed Central

    Lee, Eunkyung; Choi, So-Young; Yang, Jae-Ho

    2016-01-01

    Early life neuronal exposure to environmental toxicants has been suggested to be an important etiology of neurodegenerative disease development. Perfluorohexanesulfonate (PFHxS), one of the major perfluoroalkyl compounds, is widely distributed environmental contaminants. We have reported that PFHxS induces neuronal apoptosis via ERK-mediated pathway. Imperatorin is a furanocoumarin found in various edible plants and has a wide range of pharmacological effects including neuroprotection. In this study, the effects of imperatorin on PFHxS-induced neuronal apoptosis and the underlying mechanisms are examined using cerebellar granule cells (CGC). CGC were isolated from seven-day old rats and were grown in culture for seven days. Caspase-3 activity and TUNEL staining were used to determine neuronal apoptosis. PFHxS-induced apoptosis of CGC was significantly reduced by imperatorin and PD98059, an ERK pathway inhibitor. PFHxS induced a persistent increase in intracellular calcium, which was significantly blocked by imperatorin, NMDA receptor antagonist, MK801 and the L-type voltage-dependent calcium channel blockers, diltiazem and nifedipine. The activation of caspase-3 by PFHxS was also inhibited by MK801, diltiazem and nifedipine. PFHxS-increased ERK activation was inhibited by imperatorin, MK801, diltiazem and nifedipine. Taken together, imperatorin protects CGC against PFHxS-induced apoptosis via inhibition of NMDA receptor/intracellular calcium-mediated ERK pathway. PMID:27382356

  3. Taxifolin prevents diabetic cardiomyopathy in vivo and in vitro by inhibition of oxidative stress and cell apoptosis.

    PubMed

    Sun, Xiao; Chen, Rong-chang; Yang, Zhi-hong; Sun, Gui-bo; Wang, Min; Ma, Xiao-jun; Yang, Li-juan; Sun, Xiao-bo

    2014-01-01

    Diabetic cardiomyopathy has been increasingly recognized as an important cause of heart failure in diabetic patients. Excessive oxidative stress has been suggested to play a critical role in the development of diabetic cardiomyopathy. The objective of this study was to investigate the potential protective effects and mechanisms of taxifolin on cardiac function of streptozotocin-induced diabetic mice and on hyperglycemia-induced apoptosis of H9c2 cardiac myoblasts. In vivo study revealed that taxifolin improved diastolic dysfunction, ameliorated myocardium structure abnormality, inhibited myocyte apoptosis and enhanced endogenous antioxidant enzymes activities. Interestingly, taxifolin reduced angiotensin II level in myocardium, inhibited NADPH oxidase activity, and increased JAK/STAT3 activation. In vitro investigation demonstrated that taxifolin inhibited 33 mM glucoseinduced H9c2 cells apoptosis by decreasing intracellular ROS level. It also inhibited caspase-3 and caspase-9 activation, restored mitochondrial membrane potential, and regulated the expression of proteins related to the intrinsic pathway of apoptosis, thus inhibiting the release of cytochrome c from mitochondria into the cytoplasm. In conclusion, taxifolin exerted cardioprotective effects against diabetic cardiomyopathy by inhibiting oxidative stress and cardiac myocyte apoptosis and might be a potential agent in the treatment of diabetic cardiomyopathy. PMID:24269735

  4. Molecular Regulation of DNA Damage-Induced Apoptosis in Neurons of Cerebral Cortex

    PubMed Central

    Liu, Zhiping; Pipino, Jacqueline; Chestnut, Barry; Landek, Melissa A.

    2009-01-01

    Cerebral cortical neuron degeneration occurs in brain disorders manifesting throughout life, but the mechanisms are understood poorly. We used cultured embryonic mouse cortical neurons and an in vivo mouse model to study mechanisms of DNA damaged-induced apoptosis in immature and differentiated neurons. p53 drives apoptosis of immature and differentiated cortical neurons through its rapid and prominent activation stimulated by DNA strand breaks induced by topoisomerase-I and -II inhibition. Blocking p53-DNA transactivation with α-pifithrin protects immature neurons; blocking p53-mitochondrial functions with μ-pifithrin protects differentiated neurons. Mitochondrial death proteins are upregulated in apoptotic immature and differentiated neurons and have nonredundant proapoptotic functions; Bak is more dominant than Bax in differentiated neurons. p53 phosphorylation is mediated by ataxia telangiectasia mutated (ATM) kinase. ATM inactivation is antiapoptotic, particularly in differentiated neurons, whereas inhibition of c-Abl protects immature neurons but not differentiated neurons. Cell death protein expression patterns in mouse forebrain are mostly similar to cultured neurons. DNA damage induces prominent p53 activation and apoptosis in cerebral cortex in vivo. Thus, DNA strand breaks in cortical neurons induce rapid p53-mediated apoptosis through actions of upstream ATM and c-Abl kinases and downstream mitochondrial death proteins. This molecular network operates through variations depending on neuron maturity. PMID:18820287

  5. Glycyrrhizic acid pretreatment prevents sepsis-induced acute kidney injury via suppressing inflammation, apoptosis and oxidative stress.

    PubMed

    Zhao, Hongyu; Liu, Zhenning; Shen, Haitao; Jin, Shuai; Zhang, Shun

    2016-06-15

    Glycyrrhizic acid (GA), an active ingredient in licorice, has multiple pharmacological activities. The aim of our study was to investigate the molecular mechanism involved in the protective effects of GA in lipopolysaccharide (LPS) stimulated rat mesangial cells (HBZY-1) and septic rats. Sepsis model was established by injection of 5mg/kg LPS in rats or incubation with 1μg/ml LPS for 24h in HBZY-1 cells. A variety of molecular biological experiments were carried out to assess the effects of GA on inflammation, apoptosis, and oxidative stress. First we found that GA alleviated sepsis-induced kidney injury in vivo. Furthermore, GA suppressed inflammatory response in vivo and in vitro. Additionally, GA inhibited cell apoptosis and the changes in expressions of apoptosis related proteins induced by LPS. Moreover, GA markedly inhibited oxidative stress induced by LPS via activation of ERK signaling pathway. Finally GA could inhibit the activation of NF-κ B induced by LPS. Our present study indicates that GA has a protective effect against sepsis-induced inflammatory response, apoptosis, and oxidative stress damage, which provides a molecular basis for a new medical treatment of septic acute kidney injury. PMID:27063444

  6. Dihydroartemisinin prevents liver fibrosis in bile duct ligated rats by inducing hepatic stellate cell apoptosis through modulating the PI3K/Akt pathway.

    PubMed

    Chen, Qin; Chen, Lianyun; Wu, Xiafei; Zhang, Feng; Jin, Huanhuan; Lu, Chunfeng; Shao, Jiangjuan; Kong, Desong; Wu, Li; Zheng, Shizhong

    2016-03-01

    As a frequent event following chronic insult, liver fibrosis triggers wound healing reactions, with extracellular matrix components accumulated in the liver. During liver fibrogenesis, activation of hepatic stellate cells (HSCs) is the pivotal event. Fibrosis regression can feasibly be treated through pharmacological induction of HSC apoptosis. Herein we showed that dihydroartemisinin (DHA) improved liver histological architecture, decreased hepatic enzyme levels, and inhibited HSCs activation in the fibrotic rat liver. DHA also induced apoptosis of HSCs in such liver, as demonstrated by reduced distribution of α-SMA-positive cells and the presence of high number of cleaved-caspase-3-positive cells in vivo, as well as by down-regulation of Bcl-2 and up-regulation of Bax. In addition, in vitro experiments showed that DHA significantly inhibited HSC proliferation and led to dramatic morphological alterations in HSCs. we found that DHA disrupted mitochondrial functions and led to activation of caspase cascades in HSCs. Mechanistic investigations revealed that DHA induced HSC apoptosis through disrupting the phosphoinositide 3-kinase (PI3K)/Akt pathway and that PI3K specific inhibitor LY294002 mimicked the pro-apoptotic effect of DHA. DHA is a promising candidate for the prevention and treatment of liver fibrosis. PMID:26865509

  7. Reduced ultraviolet-induced DNA damage and apoptosis in human skin with topical application of a photolyase-containing DNA repair enzyme cream: clues to skin cancer prevention.

    PubMed

    Berardesca, Enzo; Bertona, Marco; Altabas, Karmela; Altabas, Velimir; Emanuele, Enzo

    2012-02-01

    The exposure of human skin to ultraviolet radiation (UVR) results in the formation of DNA photolesions that give rise to photoaging, mutations, cell death and the onset of carcinogenic events. Photolyase (EC 4.1.99.3) is a DNA repair enzyme that reverses damage caused by exposure to UVR. We sought to investigate whether addition of photolyase enhances the protection provided by a traditional sunscreen (SS), by reducing the in vivo formation of cyclobutane-type pyrimidine dimers (CPDs) and UVR-induced apoptosis in human skin. Ten volunteers (Fitzpatrick skin type II) were exposed to solar-simulated (ss) UVR at a three times minimal erythema dose for 4 consecutive days. Thirty minutes prior to each exposure, the test materials [vehicle, SS (sun protection factor 50) alone, and SS plus photolyase from Anacystis nidulans] were applied topically to three different sites. One additional site was left untreated and one received ssUVR only. Biopsy specimens were taken 72 h after the last irradiation. The amount of CPDs and the extent of apoptosis were measured by ELISA. Photolyase plus SS was superior to SS alone in reducing both the formation of CPDs and apoptotic cell death (both P<0.001). In conclusion, the addition of photolyase to a traditional SS contributes significantly to the prevention of UVR-induced DNA damage and apoptosis when applied topically to human skin. PMID:22086236

  8. Addition of Exogenous NAD+ Prevents Mefloquine-Induced Neuroaxonal and Hair Cell Degeneration through Reduction of Caspase-3-Mediated Apoptosis in Cochlear Organotypic Cultures

    PubMed Central

    Ding, Dalian; Qi, Weidong; Yu, Dongzhen; Jiang, Haiyan; Han, Chul; Kim, Mi-Jung; Katsuno, Kana; Hsieh, Yun Hua; Miyakawa, Takuya; Salvi, Richard; Tanokura, Masaru; Someya, Shinichi

    2013-01-01

    Background Mefloquine is widely used for the treatment of malaria. However, this drug is known to induce neurological side effects including depression, anxiety, balance disorder, and sensorineural hearing loss. Yet, there is currently no treatment for these side effects. Principal Findings In this study, we show that the coenzyme NAD+, known to play a critical role in maintaining the appropriate cellular redox environment, protects cochlear axons and sensory hair cells from mefloquine-induced degeneration in cultured rat cochleae. Mefloquine alone destroyed hair cells and nerve fiber axons in rat cochlear organotypics cultures in a dose-dependent manner, while treatment with NAD+ protected axons and hair cells from mefloquine-induced degeneration. Furthermore, cochlear organs treated with mefloquine showed increased oxidative stress marker levels, including superoxide and protein carbonyl, and increased apoptosis marker levels, including TUNEL-positive nuclei and caspases-3. Treatment with NAD+ reduced the levels of these oxidative stress and apoptosis markers. Conclusions/Significance Taken together, our findings suggest that that mefloquine disrupts the cellular redox environment and induces oxidative stress in cochlear hair cells and nerve fibers leading to caspases-3-mediated apoptosis of these structures. Exogenous NAD+ suppresses mefloquine-induced oxidative stress and prevents the degeneration of cochlear axons and sensory hair cells caused by mefloquine treatment. PMID:24223197

  9. Protection from palmitate-induced mitochondrial DNA damage prevents from mitochondrial oxidative stress, mitochondrial dysfunction, apoptosis, and impaired insulin signaling in rat L6 skeletal muscle cells.

    PubMed

    Yuzefovych, Larysa V; Solodushko, Viktoriya A; Wilson, Glenn L; Rachek, Lyudmila I

    2012-01-01

    Saturated free fatty acids have been implicated in the increase of oxidative stress, mitochondrial dysfunction, apoptosis, and insulin resistance seen in type 2 diabetes. The purpose of this study was to determine whether palmitate-induced mitochondrial DNA (mtDNA) damage contributed to increased oxidative stress, mitochondrial dysfunction, apoptosis, impaired insulin signaling, and reduced glucose uptake in skeletal muscle cells. Adenoviral vectors were used to deliver the DNA repair enzyme human 8-oxoguanine DNA glycosylase/(apurinic/apyrimidinic) lyase (hOGG1) to mitochondria in L6 myotubes. After palmitate exposure, we evaluated mtDNA damage, mitochondrial function, production of mitochondrial reactive oxygen species, apoptosis, insulin signaling pathways, and glucose uptake. Protection of mtDNA from palmitate-induced damage by overexpression of hOGG1 targeted to mitochondria significantly diminished palmitate-induced mitochondrial superoxide production, restored the decline in ATP levels, reduced activation of c-Jun N-terminal kinase (JNK) kinase, prevented cells from entering apoptosis, increased insulin-stimulated phosphorylation of serine-threonine kinase (Akt) (Ser473) and tyrosine phosphorylation of insulin receptor substrate-1, and thereby enhanced glucose transporter 4 translocation to plasma membrane, and restored insulin signaling. Addition of a specific inhibitor of JNK mimicked the effect of mitochondrial overexpression of hOGG1 and partially restored insulin sensitivity, thus confirming the involvement of mtDNA damage and subsequent increase of oxidative stress and JNK activation in insulin signaling in L6 myotubes. Our results are the first to report that mtDNA damage is the proximal cause in palmitate-induced mitochondrial dysfunction and impaired insulin signaling and provide strong evidence that targeting DNA repair enzymes into mitochondria in skeletal muscles could be a potential therapeutic treatment for insulin resistance. PMID:22128025

  10. Hypoxia-induced p53 modulates both apoptosis and radiosensitivity via AKT

    PubMed Central

    Leszczynska, Katarzyna B.; Foskolou, Iosifina P.; Abraham, Aswin G.; Anbalagan, Selvakumar; Tellier, Céline; Haider, Syed; Span, Paul N.; O’Neill, Eric E.; Buffa, Francesca M.; Hammond, Ester M.

    2015-01-01

    Restoration of hypoxia-induced apoptosis in tumors harboring p53 mutations has been proposed as a potential therapeutic strategy; however, the transcriptional targets that mediate hypoxia-induced p53-dependent apoptosis remain elusive. Here, we demonstrated that hypoxia-induced p53-dependent apoptosis is reliant on the DNA-binding and transactivation domains of p53 but not on the acetylation sites K120 and K164, which, in contrast, are essential for DNA damage–induced, p53-dependent apoptosis. Evaluation of hypoxia-induced transcripts in multiple cell lines identified a group of genes that are hypoxia-inducible proapoptotic targets of p53, including inositol polyphosphate-5-phosphatase (INPP5D), pleckstrin domain–containing A3 (PHLDA3), sulfatase 2 (SULF2), B cell translocation gene 2 (BTG2), cytoplasmic FMR1-interacting protein 2 (CYFIP2), and KN motif and ankyrin repeat domains 3 (KANK3). These targets were also regulated by p53 in human cancers, including breast, brain, colorectal, kidney, bladder, and melanoma cancers. Downregulation of these hypoxia-inducible targets associated with poor prognosis, suggesting that hypoxia-induced apoptosis contributes to p53-mediated tumor suppression and treatment response. Induction of p53 targets, PHLDA3, and a specific INPP5D transcript mediated apoptosis in response to hypoxia through AKT inhibition. Moreover, pharmacological inhibition of AKT led to apoptosis in the hypoxic regions of p53-deficient tumors and consequently increased radiosensitivity. Together, these results identify mediators of hypoxia-induced p53-dependent apoptosis and suggest AKT inhibition may improve radiotherapy response in p53-deficient tumors. PMID:25961455

  11. Maternal antioxidants prevent beta cell apoptosis and promote formation of dual hormone-expressing endocrine cells in male offspring following fetal and neonatal nicotine exposure

    PubMed Central

    BRUIN, Jennifer E; WOYNILLOWICZ, Amanda K; HETTINGA, Bart P; TARNOPOLSKY, Mark A; MORRISON, Katherine M; GERSTEIN, Hertzel C; HOLLOWAY, Alison C

    2013-01-01

    Aim Fetal and neonatal nicotine exposure causes beta cell oxidative stress and apoptosis in neonates, leading to adult-onset dysglycemia. The goal of this study was to determine whether an antioxidant intervention could prevent nicotine-induced beta cell loss. Methods Nulliparous female Wistar rats received daily subcutaneous injections of either saline or nicotine bitartrate (1.0 mg/kg/d) for 2 weeks prior to mating until weaning. Nicotine-exposed dams received either normal chow or diet containing antioxidants (1000 IU/kg vitamin E, 0.25% w/w coenzyme Q10 and 0.1% w/w alpha-lipoic acid) during mating, pregnancy and lactation; saline-exposed dams received normal chow. Pancreas tissue was collected from male offspring at 3 weeks of age to measure beta cell fraction, apoptosis, proliferation and the presence of cells co-expressing insulin and glucagon. Results The birth weight of the offspring born to nicotine-exposed dams receiving dietary antioxidants was significantly reduced. Most interestingly, the antioxidant intervention to nicotine-exposed dams prevented the beta cell loss and apoptosis observed in nicotine exposed male offspring whose mothers did not receive antioxidants. Male pups born to nicotine-treated mothers receiving antioxidants also had a trend towards increased beta cell proliferation and a significant increase in islets containing insulin/glucagon bi-hormonal cells relative to the other two treatment groups. Conclusion This study demonstrates that exposure to maternal antioxidants protects beta cells from the damaging effects of nicotine thus preserving beta cell mass. PMID:22385833

  12. microRNA-29b prevents liver fibrosis by attenuating hepatic stellate cell activation and inducing apoptosis through targeting PI3K/AKT pathway

    PubMed Central

    Wang, Jia; Chu, Eagle S.H.; Chen, Hai-Yong; Man, Kwan; Go, Minnie Y.Y.; Huang, Xiao Ru; Lan, Hui Yao; Sung, Joseph J.Y.; Yu, Jun

    2015-01-01

    microRNA-29b (miR-29b) is known to be associated with TGF-β-mediated fibrosis, but the mechanistic action of miR-29b in liver fibrosis remains unclear and is warranted for investigation. We found that miR-29b was significantly downregulated in human and mice fibrotic liver tissues and in primary activated HSCs. miR-29b downregulation was directly mediated by Smad3 through binding to the promoter of miR-29b in hepatic stellate cell (HSC) line LX1, whilst miR-29b could in turn suppress Smad3 expression. miR-29b transduction in the liver of mice prevented CCl4 induced-fibrogenesis, concomitant with decreased expression of α-SMA, collagen I and TIMP-1. Ectopic expression of miR-29b in activated HSCs (LX-1, HSC-T6) inhibited cell viability and colony formation, and caused cell cycle arrest in G1 phase by downregulating cyclin D1 and p21cip1. Further, miR-29b induced apoptosis in HSCs mediated by caspase-9 and PARP. miR-29b inhibited its downstream effectors of PIK3R1 and AKT3 through direct targeting their 3′UTR regions. Moreover, knockdown of PIK3R1 or AKT3 suppressed α-SMA and collagen I and induced apoptosis in both HSCs and in mice. In conclusion, miR-29b prevents liver fibrogenesis by inhibiting HSC activation and inducing HSC apoptosis through inhibiting PI3K/AKT pathway. These results provide novel mechanistic insights for the anti-fibrotic effect of miR-29b. PMID:25356754

  13. Fermented soybeans, Chungkookjang, prevent hippocampal cell death and β-cell apoptosis by decreasing pro-inflammatory cytokines in gerbils with transient artery occlusion.

    PubMed

    Park, Sunmin; Kim, Da Sol; Kang, Sunna; Moon, Bo Reum

    2016-02-01

    Since Chungkookjang, a short-term fermented soybean, is known to improve glucose metabolism and antioxidant activity, it may prevent the neurological symptoms and glucose disturbance induced by artery occlusion. We investigated the protective effects and mechanisms of traditional (TFC) and standardized Chungkookjang fermented with Bacillus licheniformis (BLFC) against ischemia/reperfusion damage in the hippocampal CA1 region and against hyperglycemia after transient cerebral ischemia in gerbils. Gerbils were subjected to either an occlusion of the bilateral common carotid arteries for 8 min to render them ischemic or a sham operation. Ischemic gerbils were fed either a 40% fat diet containing 10% of either cooked soybean (CSB), TFC, or BLFC for 28 days. Neuronal cell death and cytokine expression in the hippocampus, neurological deficit, serum cytokine levels, and glucose metabolism were measured. TFC and BLFC contained more isoflavonoid aglycones than CSB. Artery occlusion increased the expressions of IL-1β and TNF-α as well as cell death in the hippocampal CA1 region and induced severe neurological symptoms. CSB, TFC, and BLFC prevented the neuronal cell death and the symptoms such as dropped eyelid, bristling hair, reduced muscle tone and flexor reflex, and abnormal posture and walking patterns, and suppressed cytokine expressions. CSB was less effective than TFC and BLFC. Artery occlusion induced glucose intolerance due to decreased insulin secretion and β-cell mass. TFC and BLFC prevented the impairment of glucose metabolism by artery occlusion. Especially TFC and BLFC increased β-cell proliferation and suppressed the β-cell apoptosis by suppressing TNF-α and IL-1β which in turn decreased cleaved caspase-3 that caused apoptosis. In conclusion, TFC and BLFC may prevent and alleviate neuronal cell death in the hippocampal CA1 region and neurological symptoms and poststroke hyperglycemia in gerbils with artery occlusion. This might be associated with

  14. Anti-osteopontin monoclonal antibody prevents ovariectomy-induced osteoporosis in mice by promotion of osteoclast apoptosis

    SciTech Connect

    Zhang, Bo; Dai, Jianxin; Wang, Huaqing; Wei, Huafeng; Zhao, Jian; Guo, Yajun; and others

    2014-09-26

    Highlight: • We first report that anti-osteopontin mAb could protect osteoporosis in mice. • Anti-osteopontin mAb could promote the osteoclast apoptosis. • Targeting osteopontin might have therapeutic potentials for osteoporosis. - Abstract: Osteopontin (OPN) is abundant in mineralized tissues and has long been implicated in bone remodeling. However, the therapeutic effect of targeting OPN in bone loss diseases and the underlying molecular mechanism remain largely unknown. Here, we reported that anti-OPN mAb (23C3) could protect against ovariectomy-induced osteoporosis in mice, demonstrated by microcomputed tomography analysis and histopathology evaluation. In vitro assay showed that 23C3 mAb reduced osteoclasts (OCs)-mediated bone resorption through promotion of mature OC apoptosis. Thus, the study has important implications for understanding the role of OPN in OC bone resorption and survival, and OPN antagonists may have therapeutic potential for osteoporosis and other osteopenic diseases.

  15. Pyruvate kinase M2 prevents apoptosis via modulating Bim stability and associates with poor outcome in hepatocellular carcinoma

    PubMed Central

    Li, Min; Zhang, Chao; Liu, Li-Li; Fu, Jia; Jin, Jie-Tian; Luo, Rong-Zhen; Zhang, Chris Zhiyi; Yun, Jing-Ping

    2015-01-01

    Pyruvate kinase M2 (PKM2) contributes to the Warburg effect, a hallmark of cancer. We showed that PKM2 levels were correlated with overall survival (hazard ration = 1.675, 95% confidence interval: 1.389–2.019, P < 0.001) and disease-free survival (hazard ration = 1.573, 95% confidence interval: 1.214–2.038, P < 0.001) in a cohort of 490 patients with HCC. The correlations were further validated in an independent cohort of 148 HCC patients. Multivariate analyses revealed that PKM2 was an independent indicator of poor outcome in HCC. The knockdown of PKM2 in HCC cells inhibited cell proliferation and induced apoptosis in vitro and in vivo. Bim siRNA markedly abolished the PKM2-depletion-induced apoptosis. PKM2 depletion decreased the degradation of Bim. In clinical samples, PKM2 expression was reversely correlated with Bim expression. Combination of PKM2 and Bim levels had the best prognostic significance. We suggest that PKM2 serves as a promising biomarker for poor prognosis of patients with HCC and its knockdown induces HCC apoptosis by stabilizing Bim. PMID:25788265

  16. MDM2-regulated degradation of HIPK2 prevents p53Ser46 phosphorylation and DNA damage-induced apoptosis.

    PubMed

    Rinaldo, Cinzia; Prodosmo, Andrea; Mancini, Francesca; Iacovelli, Stefano; Sacchi, Ada; Moretti, Fabiola; Soddu, Silvia

    2007-03-01

    In response to DNA damage, p53 induces either cell-cycle arrest or apoptosis by differential transcription of several target genes and through transcription-independent apoptotic functions. p53 phosphorylation at Ser46 by HIPK2 is one determinant of the outcome because it takes place only upon severe, nonrepairable DNA damage that irreversibly drives cells to apoptosis. Here, we show that p53 represses its proapoptotic activator HIPK2 via MDM2-mediated degradation, whereas a degradation-resistant HIPK2 mutant has increased apoptotic activity. Upon cytostatic, nonsevere DNA damage, inhibition of HIPK2 degradation is sufficient to induce p53Ser46 phosphorylation and apoptosis, converting growth-arresting stimuli to apoptotic ones. These findings establish HIPK2 as an MDM2 target and support a model in which, upon nonsevere DNA damage, p53 represses its own phosphorylation at Ser46 due to HIPK2 degradation, supporting the notion that the cell-cycle-arresting functions of p53 include active inhibition of the apoptotic ones. PMID:17349959

  17. Selective cellular uptake and induction of apoptosis of cancer-targeted selenium nanoparticles.

    PubMed

    Huang, Yanyu; He, Lizhen; Liu, Wen; Fan, Cundong; Zheng, Wenjie; Wong, Yum-Shing; Chen, Tianfeng

    2013-09-01

    Selenium nanoparticles (SeNPs) have garnered a great deal of attention as potential cancer therapeutic payloads. However, the in vivo targeting drug delivery has been challenging. Herein, we describe the synthesis of tansferrin (Tf)-conjugated SeNPs and its use as a cancer-targeted drug delivery system to achieve enhanced cellular uptake and anticancer efficacy. Tf as targeting ligand significantly enhances the cellular uptake of doxorubicin (DOX)-loaded SeNPs through clathrin-mediated and caveolae/lipid raft-mediated endocytosis in cancer cells overexpressing transferrin receptor, and increases their selectivity between cancer and normal cells. DOX-loaded and Tf-conjugated SeNPs (Tf-SeNPs) exhibits unprecedented enhanced cytotoxicity toward cancer cells through induction of apoptosis with the involvement of intrinsic and extrinsic pathways. Internalized Tf-SeNPs triggers intracellular ROS overproduction, thus activates p53 and MAPKs pathways to promote cell apoptosis. In the nude mice xenograft experiment, Tf-SeNPs significantly inhibits the tumor growth via induction of p53-mediated apoptosis. This cancer-targeted design of SeNPs opens a new path for synergistic treating of cancer with higher efficacy and decreased side effects. PMID:23800743

  18. Disruption of Smad5 gene induces mitochondria-dependent apoptosis in cardiomyocytes

    SciTech Connect

    Sun Yanxun; Zhou Jiang; Liao Xudong; Lue Yaxin; Deng Chuxia; Huang Peitang; Chen Quan; Yang Xiao . E-mail: yangx@nic.bmi.ac.cn

    2005-05-15

    Our previous studies have shown that SMAD5, an important intracellular mediator of transforming growth factor {beta} (TGF-{beta}) family, is required for normal development of the cardiovascular system in vivo. In the current study, we reported that the lack of the Smad5 gene resulted in apoptosis of cardiac myocytes in vivo. To further investigate the mechanism of the Smad5 gene in cardiomyocyte apoptosis, the embryonic stem (ES) cell differentiation system was employed. We found that the myotubes that differentiated from the homozygous Smad5 {sup ex6/ex6} mutant ES cells underwent collapse and degeneration during the late stages of in vitro differentiation, mimicking the in vivo observation. By electron microscopy, abnormal swollen mitochondria were observed in cardiomyocytes both from Smad5-deficient embryos and from ES-differentiated cells. There was also a significant reduction in mitochondrial membrane potential ({delta}{psi} {sub m}) and a leakage of cytochrome c from mitochondria into the cytosol of myocytes differentiated from Smad5 mutant ES cells. The expression of p53 and p21 was found to be elevated in the differentiated Smad5 mutant myocytes, and this was accompanied by an up-regulation in caspase 3 expression. These results suggest that the Smad5-mediated TGF-{beta} signals may protect cardiomyocytes from apoptosis by maintaining the integrity of the mitochondria, probably through suppression of p53 mediated pathways.

  19. Host Immune Defense Peptide LL-37 Activates Caspase-Independent Apoptosis and Suppresses Colon Cancer

    PubMed Central

    Ren, Shun X.; Cheng, Alfred S.L.; To, Ka F.; Tong, Joanna H.M.; Li, May S.; Shen, Jin; Wong, Clover C.M.; Zhang, Lin; Chan, Ruby L.Y.; Wang, Xiao J.; Ng, Simon S.M.; Chiu, Lawrence C.M.; Marquez, Victor E.; Gallo, Richard L.; Chan, Francis K.L.; Yu, Jun; Sung, Joseph J.Y.; Wu, William K.K.; Cho, Chi H.

    2014-01-01

    Cathelicidins are a family of bacteriocidal polypeptides secreted by macrophages and polymorphonuclear leukocytes (PMN). LL-37, the only human cathelicidin, has been implicated in tumorigenesis, but there has been limited investigation of its expression and function in cancer. Here, we report that LL-37 activates a p53-mediated, caspase-independent apoptotic cascade that contributes to suppression of colon cancer. LL-37 was expressed strongly in normal colon mucosa but downregulated in colon cancer tissues, where in both settings its expression correlated with terminal deoxynucleotidyl transferase–mediated dUTP nick end labeling-positive apoptotic cells. Exposure of colon cancer cells to LL-37 induced phosphatidylserine externalization and DNA fragmentation in a manner independent of caspase activation. Apoptogenic function was mediated by nuclear translocation of the proapoptotic factors, apoptosis-inducing factor (AIF) and endonuclease G (EndoG), through p53-dependent upregulation of Bax and Bak and downregulation of Bcl-2 via a pertussis toxin–sensitive G-protein–coupled receptor (GPCR) pathway. Correspondingly, colonic mucosa of cathelicidin-deficient mice exhibited reduced expression of p53, Bax, and Bak and increased expression of Bcl-2 together with a lower basal level of apoptosis. Cathelicidin-deficient mice exhibited an increased susceptibility to azoxymethane-induced colon tumorigenesis, establishing pathophysiologic relevance in colon cancer. Collectively, our findings show that LL-37 activates a GPCR-p53-Bax/Bak/Bcl-2 signaling cascade that triggers AIF/EndoG–mediated apoptosis in colon cancer cells. PMID:23100468

  20. Soy phytochemicals prevent orthotopic growth and metastasis of bladder cancer in mice by alterations of cancer cell proliferation and apoptosis and tumor angiogenesis.

    PubMed

    Singh, Ajita V; Franke, Adrian A; Blackburn, George L; Zhou, Jin-Rong

    2006-02-01

    A role of dietary bioactive components in bladder cancer prevention is biologically plausible because most substances or metabolites are excreted through the urinary tract and are consequently in direct contact with the mucosa of the bladder. We first determined antigrowth activity of genistein against poorly differentiated 253J B-V human bladder cancer cells in vitro. Genistein inhibited the cell growth in a time- and dose-dependent manner via G(2)-M arrest, down-regulation of nuclear factor kappaB (NF-kappaB), and induction of apoptosis. We also evaluated both genistin, which is a natural form of genistein, and the isoflavone-rich soy phytochemical concentrate (SPC) on the growth and metastasis of 253J B-V tumors in an orthotopic tumor model. Mice treated with genistin and SPC had reduced final tumor weights by 56% (P < 0.05) and 52% (P < 0.05), respectively, associated with induction of tumor cell apoptosis and inhibition of tumor angiogenesis in vivo. In addition, SPC treatment, but not genistin treatment, significantly inhibited lung metastases by 95% (P < 0.01) associated with significant down-regulation of NF-kappaB expression in tumor tissues and reduction of circulating insulin-like growth factor-I levels, suggesting that SPC may contain other bioactive ingredients that have antimetastatic activity. The results from our studies suggest that further clinical investigation should be warranted to apply soy phytochemicals, such as SPC, as a potent prevention regimen for bladder cancer progression. This orthotopic human bladder tumor model also provides a clinically relevant experimental tool for assessing potential preventive activity of other dietary components against bladder tumor growth and metastasis. PMID:16452247

  1. Inhibition of Osteocyte Apoptosis Prevents the Increase in Osteocytic Receptor Activator of Nuclear Factor κB Ligand (RANKL) but Does Not Stop Bone Resorption or the Loss of Bone Induced by Unloading*

    PubMed Central

    Plotkin, Lilian I.; Gortazar, Arancha R.; Davis, Hannah M.; Condon, Keith W.; Gabilondo, Hugo; Maycas, Marta; Allen, Matthew R.; Bellido, Teresita

    2015-01-01

    Apoptosis of osteocytes and osteoblasts precedes bone resorption and bone loss with reduced mechanical stimulation, and receptor activator of NF-κB ligand (RANKL) expression is increased with unloading in mice. Because osteocytes are major RANKL producers, we hypothesized that apoptotic osteocytes signal to neighboring osteocytes to increase RANKL expression, which, in turn, increases osteoclastogenesis and bone resorption. The traditional bisphosphonate (BP) alendronate (Aln) or IG9402, a BP analog that does not inhibit resorption, prevented the increase in osteocyte apoptosis and osteocytic RANKL expression. The BPs also inhibited osteoblast apoptosis but did not prevent the increase in osteoblastic RANKL. Unloaded mice exhibited high serum levels of the bone resorption marker C-telopeptide fragments of type I collagen (CTX), elevated osteoclastogenesis, and increased osteoclasts in bone. Aln, but not IG9402, prevented all of these effects. In addition, Aln prevented the reduction in spinal and femoral bone mineral density, spinal bone volume/tissue volume, trabecular thickness, mechanical strength, and material strength induced by unloading. Although IG9402 did not prevent the loss of bone mass, it partially prevented the loss of strength, suggesting a contribution of osteocyte viability to strength independent of bone mass. These results demonstrate that osteocyte apoptosis leads to increased osteocytic RANKL. However, blockade of these events is not sufficient to restrain osteoclast formation, inhibit resorption, or stop bone loss induced by skeletal unloading. PMID:26085098

  2. Interleukin-10 prevents epithelial cell apoptosis by regulating IFNγ and TNFα expression in rhesus macaque colon explants

    PubMed Central

    Pan, Diganta; Das, Arpita; Lala, Wendy; Kenway-Lynch, Carys S.; Liu, David X.; Veazey, Ronald S.; Pahar, Bapi

    2013-01-01

    Interleukin-10 (IL-10) is an important immunomodulatory cytokine that plays an obligate role in regulating inflammatory responses. Here we demonstrated the role of IL-10 in regulating crypts length and breadth as well as maintaining the survival of epithelial cells using rhesus colon explant cultures. Anti-IL-10 antibody treatment of colon explant cultures induced increased production of inflammatory cytokines/molecules like IFNγ, TNFα, CD107a and perforin as well as increased epithelial cell apoptosis compared to media controls tested. Our results suggest that IL-10 plays a crucial role in maintaining mucosal homeostasis by regulating mucosal IFNγ and TNFα cytokine production. PMID:23867612

  3. Interleukin-10 prevents epithelial cell apoptosis by regulating IFNγ and TNFα expression in rhesus macaque colon explants.

    PubMed

    Pan, Diganta; Das, Arpita; Lala, Wendy; Kenway-Lynch, Carys S; Liu, David X; Veazey, Ronald S; Pahar, Bapi

    2013-10-01

    Interleukin-10 (IL-10) is an important immunomodulatory cytokine that plays an obligate role in regulating inflammatory responses. Here we demonstrated the role of IL-10 in regulating crypts length and breadth as well as maintaining the survival of epithelial cells using rhesus colon explant cultures. Anti-IL-10 antibody treatment of colon explant cultures induced increased production of inflammatory cytokines/molecules like IFNγ, TNFα, CD107a and perforin as well as increased epithelial cell apoptosis compared to media controls tested. Our results suggest that IL-10 plays a crucial role in maintaining mucosal homeostasis by regulating mucosal IFNγ and TNFα cytokine production. PMID:23867612

  4. Cardiac progenitor cell-derived exosomes prevent cardiomyocytes apoptosis through exosomal miR-21 by targeting PDCD4.

    PubMed

    Xiao, J; Pan, Y; Li, X H; Yang, X Y; Feng, Y L; Tan, H H; Jiang, L; Feng, J; Yu, X Y

    2016-01-01

    Cardiac progenitor cells derived from adult heart have emerged as one of the most promising stem cell types for cardiac protection and repair. Exosomes are known to mediate cell-cell communication by transporting cell-derived proteins and nucleic acids, including various microRNAs (miRNAs). Here we investigated the cardiac progenitor cell (CPC)-derived exosomal miRNAs on protecting myocardium under oxidative stress. Sca1(+)CPCs-derived exosomes were purified from conditional medium, and identified by nanoparticle trafficking analysis (NTA), transmission electron microscopy and western blotting using CD63, CD9 and Alix as markers. Exosomes production was measured by NTA, the result showed that oxidative stress-induced CPCs secrete more exosomes compared with normal condition. Although six apoptosis-related miRNAs could be detected in two different treatment-derived exosomes, only miR-21 was significantly upregulated in oxidative stress-induced exosomes compared with normal exosomes. The same oxidative stress could cause low miR-21 and high cleaved caspase-3 expression in H9C2 cardiac cells. But the cleaved caspase-3 was significantly decreased when miR-21 was overexpressed by transfecting miR-21 mimic. Furthermore, miR-21 mimic or inhibitor transfection and luciferase activity assay confirmed that programmed cell death 4 (PDCD4) was a target gene of miR-21, and miR-21/PDCD4 axis has an important role in anti-apoptotic effect of H9C2 cell. Western blotting and Annexin V/PI results demonstrated that exosomes pre-treated H9C2 exhibited increased miR-21 whereas decreased PDCD4, and had more resistant potential to the apoptosis induced by the oxidative stress, compared with non-treated cells. These findings revealed that CPC-derived exosomal miR-21 had an inhibiting role in the apoptosis pathway through downregulating PDCD4. Restored miR-21/PDCD4 pathway using CPC-derived exosomes could protect myocardial cells against oxidative stress-related apoptosis. Therefore

  5. Hyperbaric oxygen treatment prevents nitric oxide-induced apoptosis in articular cartilage injury via enhancement of the expression of heat shock protein 70.

    PubMed

    Ueng, Steve W N; Yuan, Li-Jen; Lin, Song-Shu; Niu, Chi-Chien; Chan, Yi-Sheng; Wang, I-Chun; Yang, Chuen-Yung; Chen, Wen-Jer

    2013-03-01

    Heat shock proteins (HSPs), inflammatory cytokines, nitric oxide (NO), and localized hypoxia-induced apoptosis are thought to be correlated to the degree of cartilage injury. We investigated the effect of hyperbaric oxygen (HBO) on (1) interleukin-1β (IL-1β)-induced NO production and apoptosis of rabbit chondrocytes and (2) healing of articular cartilage defects. For the in vitro study, RT-PCR and Western blotting were performed to detect mRNA and protein expressions of HSP70, inducible NO synthase (iNOS), and caspase 3 in IL-1β-treated chondrocytes. To clarify that the HSP70 was necessary for anti-iNOS and anti-apoptotic activity by HBO, we treated the cells with an HSP70 inhibitor, KNK437. For the in vivo study, cartilage defects were created in rabbits. The HBO group was exposed to 100% oxygen at 2.5 ATA for 1.5 h a day for 10 weeks. The control group was exposed to normal air. After sacrifice, specimen sections were sent for examination using a scoring system. Immunohistochemical analyses were performed to detect the expressions of iNOS, HSP70, and caspase 3. Our results suggested that HBO upregulated the mRNA and protein expressions of HSP70 and suppressed those of iNOS and caspase 3 in chondrocytes. KNK437 inhibited the HBO-induced downregulation of iNOS and casapase 3 activities. The histological scores showed that HBO markedly enhanced cartilage repair. Immunohistostaining showed that HBO enhanced HSP70 expression and suppressed iNOS and caspase 3 expressions in chondrocytes. Accordingly, HBO treatment prevents NO-induced apoptosis in articular cartilage injury via enhancement of the expression of heat shock protein 70. PMID:22991091

  6. Unconventional apoptosis of polymorphonuclear neutrophils (PMN): staurosporine delays exposure of phosphatidylserine and prevents phagocytosis by MΦ-2 macrophages of PMN

    PubMed Central

    Franz, S; Muñoz, L E; Heyder, P; Herrmann, M; Schiller, M

    2015-01-01

    Apoptosis of polymorphonuclear neutrophils (PMN) and subsequent ‘silent’ removal represents an important check-point for the resolution of inflammation. Failure in PMN clearance resulting in secondary necrosis-driven tissue damage has been implicated in conditions of chronic inflammation and autoimmunity. Apoptotic PMN undergo profound biophysical changes that warrant their efficient recognition and uptake by phagocytes before fading to secondary necrosis. In this study, we demonstrate that staurosporine (STS), a non-selective but potent inhibitor of cyclin-dependent kinase and protein kinase C, exerts a drastic impact on PMN apoptosis. PMN treated with STS underwent an unconventional form of cell death characterized by a delayed exposure of aminophospholipids, including phosphatidylserine (PS) and phosphatidylethanolamine and an increased exposure of neo-glycans. STS caused an impaired cellular fragmentation and accelerated DNA fragmentation. Phagocytosis of STS-treated PMN lacking PS on their surfaces was decreased significantly, which highlights the importance of PS for the clearance of apoptotic PMN. Specific opsonization with immune complexes completely restored phagocytosis of STS-treated PMN, demonstrating the efficiency of back-up clearance pathways in the absence of PS exposure. PMID:24995908

  7. Unconventional apoptosis of polymorphonuclear neutrophils (PMN): staurosporine delays exposure of phosphatidylserine and prevents phagocytosis by MΦ-2 macrophages of PMN.

    PubMed

    Franz, S; Muñoz, L E; Heyder, P; Herrmann, M; Schiller, M

    2015-01-01

    Apoptosis of polymorphonuclear neutrophils (PMN) and subsequent 'silent' removal represents an important check-point for the resolution of inflammation. Failure in PMN clearance resulting in secondary necrosis-driven tissue damage has been implicated in conditions of chronic inflammation and autoimmunity. Apoptotic PMN undergo profound biophysical changes that warrant their efficient recognition and uptake by phagocytes before fading to secondary necrosis. In this study, we demonstrate that staurosporine (STS), a non-selective but potent inhibitor of cyclin-dependent kinase and protein kinase C, exerts a drastic impact on PMN apoptosis. PMN treated with STS underwent an unconventional form of cell death characterized by a delayed exposure of aminophospholipids, including phosphatidylserine (PS) and phosphatidylethanolamine and an increased exposure of neo-glycans. STS caused an impaired cellular fragmentation and accelerated DNA fragmentation. Phagocytosis of STS-treated PMN lacking PS on their surfaces was decreased significantly, which highlights the importance of PS for the clearance of apoptotic PMN. Specific opsonization with immune complexes completely restored phagocytosis of STS-treated PMN, demonstrating the efficiency of back-up clearance pathways in the absence of PS exposure. PMID:24995908

  8. Metformin prevents endoplasmic reticulum stress-induced apoptosis through AMPK-PI3K-c-Jun NH2 pathway

    USGS Publications Warehouse

    Jung, T.W.; Lee, M.W.; Lee, Y.-J.; Kim, S.M.

    2012-01-01

    Type 2 diabetes mellitus is thought to be partially associated with endoplasmic reticulum (ER) stress toxicity on pancreatic beta cells and the result of decreased insulin synthesis and secretion. In this study, we showed that a well-known insulin sensitizer, metformin, directly protects against dysfunction and death of ER stress-induced NIT-1 cells (a mouse pancreatic beta cell line) via AMP-activated protein kinase (AMPK) and phosphatidylinositol-3 (PI3) kinase activation. We also showed that exposure of NIT-1 cells to metformin (5mM) increases cellular resistance against ER stress-induced NIT-1 cell dysfunction and death. AMPK and PI3 kinase inhibitors abolished the effect of metformin on cell function and death. Metformin-mediated protective effects on ER stress-induced apoptosis were not a result of an unfolded protein response or the induced inhibitors of apoptotic proteins. In addition, we showed that exposure of ER stressed-induced NIT-1 cells to metformin decreases the phosphorylation of c-Jun NH(2) terminal kinase (JNK). These data suggest that metformin is an important determinant of ER stress-induced apoptosis in NIT-1 cells and may have implications for ER stress-mediated pancreatic beta cell destruction via regulation of the AMPK-PI3 kinase-JNK pathway.

  9. Tetrachloro-p-benzoquinone induces hepatic oxidative damage and inflammatory response, but not apoptosis in mouse: The prevention of curcumin

    SciTech Connect

    Xu, Demei; Hu, Lihua; Su, Chuanyang; Xia, Xiaomin; Zhang, Pu; Fu, Juanli; Wang, Wenchao; Xu, Duo; Du, Hong; Hu, Qiuling; Song, Erqun; Song, Yang

    2014-10-15

    This study investigated the protective effects of curcumin on tetrachloro-p-benzoquinone (TCBQ)-induced hepatotoxicity in mice. TCBQ-treatment causes significant liver injury (the elevation of serum AST and ALT activities, histopathological changes in liver section including centrilobular necrosis and inflammatory cells), oxidative stress (the elevation of TBAR level and the inhibition of SOD and catalase activities) and inflammation (up-regulation of iNOS, COX-2, IL-1β, IL-6, TNF-α and NF-κB). However, these changes were alleviated upon pretreatment with curcumin. Interestingly, TCBQ has no effect on caspase family genes or B-cell lymphoma 2 (Bcl-2)/Bcl-2 associated X (Bax) protein expressions, which implied that TCBQ-induced hepatotoxicity is independent of apoptosis. Moreover, curcumin was shown to induce phase II detoxifying/antioxidant enzymes HO-1 and NQO1 through the activation of nuclear factor erythroid-derived 2-like 2 (Nrf2). In summary, the protective mechanisms of curcumin against TCBQ-induced hepatoxicity may be related to the attenuation of oxidative stress, along with the inhibition of inflammatory response via the activation of Nrf2 signaling. - Highlights: • TCBQ-intoxication significantly increased AST and ALT activities. • TCBQ-intoxication induced oxidative stress in mice liver. • TCBQ-intoxication induced inflammatory response in mice liver. • TCBQ-intoxication induced hepatotoxicity is independent of apoptosis. • Curcumin relieved TCBQ-induced liver damage remarkably.

  10. Glucose-Dependent Insulinotropic Peptide Prevents Serum Deprivation-Induced Apoptosis in Human Bone Marrow-Derived Mesenchymal Stem Cells and Osteoblastic Cells.

    PubMed

    Berlier, J L; Kharroubi, I; Zhang, J; Dalla Valle, A; Rigutto, S; Mathieu, M; Gangji, V; Rasschaert, J

    2015-12-01

    Human bone marrow-derived mesenchymal stem cells (hBMSC) are able to differentiate into cells of connective tissue lineages, including bone and cartilage. They are therefore considered as a promising tool for the treatment of bone degenerative diseases. One of the major issues in regenerative cell therapy is the biosafety of fetal bovine serum used for cell culture. Therefore, the development of a culture medium devoid of serum but preserving hBMSC viability will be of clinical value. The glucose-dependent insulinotropic peptide (GIP) has an anti-apoptotic action in insulin-producing cells. Interestingly, GIP also exerts beneficial effects on bone turnover by acting on osteoblasts and osteoclasts. We therefore evaluated the ability of GIP to prevent cell death in osteoblastic cells cultured in serum-free conditions. In hBMSC and SaOS-2 cells, activation of the GIP receptor increased intracellular cAMP levels. Serum deprivation induced apoptosis in SaOS-2 and hBMSC that was reduced by 30 and 50 %, respectively, in the presence of GIP. The protective effect of GIP involves activation of the adenylate cyclase pathway and inhibition of caspases 3/7 activation. These findings demonstrate that GIP exerts a protective action against apoptosis in hBMSC and suggest a novel approach to preserve viability of hBMSC cultured in the absence of serum. PMID:26254594

  11. Prevention of Anti-microbial Peptide LL-37-Induced Apoptosis and ATP Release in the Urinary Bladder by a Modified Glycosaminoglycan

    PubMed Central

    Lee, Won Yong; Savage, Justin R.; Zhang, Jianxing; Jia, Wanjian; Oottamasathien, Siam; Prestwich, Glenn D.

    2013-01-01

    Interstitial cystitis (IC), often referred to in combination with painful bladder syndrome, is a chronic inflammatory disease of the bladder. Current therapies primarily focus on replenishing urothelial glycosaminoglycan (GAG) layer using GAG analogs and managing pain with supportive therapies. However, the elusive etiology of IC and the lack of animal models to study the disease have been major hurdles developing more effective therapeutics. Previously, we showed an increased urinary concentration of antimicrobial peptide LL-37 in spina bifida patients and used LL-37 to develop a mouse model of cystitis that mimics important clinical findings of IC. Here we investigate (1) the molecular mechanism of LL-37 induced cystitis in cultured human urothelial cells and in mice, (2) the protective effects of GM-0111, a modified GAG, within the context of this mechanism, (3) the physiological and molecular markers that correlate with the severity of the inflammation, and (4) the protective effects of several GAGs using these biomarkers in our LL-37 induced cystitis model. We find that LL-37 quickly induces release of ATP and apoptosis in the urothelium. These changes can be inhibited by a chemically-modified GAG, GM-0111. Furthermore, we also find that GAG analogs provide varying degrees of protection against LL-37 challenge in mice. These findings suggest that GM-0111 and possibly GAG molecules prevent the development of cystitis by blocking the apoptosis and the concurrent release of ATP from the urothelium. PMID:24204996

  12. Cocoa-rich diet prevents azoxymethane-induced colonic preneoplastic lesions in rats by restraining oxidative stress and cell proliferation and inducing apoptosis.

    PubMed

    Rodríguez-Ramiro, Ildefonso; Ramos, Sonia; López-Oliva, Elvira; Agis-Torres, Angel; Gómez-Juaristi, Miren; Mateos, Raquel; Bravo, Laura; Goya, Luis; Martín, María Ángeles

    2011-12-01

    Cocoa is a rich source of bioactive compounds with potential chemopreventive ability but up to date its effectiveness in animal models of colon carcinogenesis has not been addressed. Herein, we investigated the in vivo effect of a cocoa-rich diet in the prevention of azoxymethane (AOM)-induced colon cancer and the mechanisms involved. Our results showed that cocoa feeding significantly reduced AOM-induced colonic aberrant crypt foci formation and crypt multiplicity. Oxidative imbalance in colon tissues seems to be prevented by cocoa as indicated by reduced oxidation markers levels and increased enzymatic and non-enzymatic endogenous defences. Cocoa-rich diet also exhibited antiproliferative effects by decreasing the levels of extracellular regulated kinases, protein kinase B and cyclin D1 together with pro-apoptotic effects evidenced by reduced Bcl-x(L) levels and increased Bax levels and caspase-3 activity. Our findings provide the first in vivo evidence that a cocoa-rich diet may inhibit the early stage of colon carcinogenesis probably by preventing oxidative stress and cell proliferation and by inducing apoptosis. PMID:21953728

  13. Eugenol precludes cutaneous chemical carcinogenesis in mouse by preventing oxidative stress and inflammation and by inducing apoptosis.

    PubMed

    Kaur, Gurpreet; Athar, Mohammad; Alam, M Sarwar

    2010-03-01

    The present study was designed to investigate the protective efficacy of eugenol against skin cancer and probe into the mechanistic aspects. Skin tumors were initiated by applying 160 nmol DMBA and promoted by twice weekly applications of 8.5 nmol TPA for 28 wk. All mice developed tumors by 13 wk of promotion. However, in mice pretreated with 30 microL eugenol, no tumors were detected until 8 wk (following anti-initiation protocol) and until 14 wk (following antipromotion protocol) of tumor promotion. PCNA and TUNEL immunohistochemistry of tumors revealed eugenol to ameliorate cell proliferation and elevate apoptosis respectively. The effect of eugenol was assessed on specific stages of carcinogenesis. Initiation with DMBA led to a significant upregulation of p53 expression with a concomitant increase in p21(WAF1) levels in epidermal cells indicating induction of damage to the DNA. However, pretreatment with eugenol led to overexpression of these genes, which probably helped stimulate apoptosis of the initiated cells. To ascertain the molecular mechanisms implicated in the antitumor promoting activity of eugenol, its effect was investigated on markers of tumor promotion and inflammation: ODC activity and iNOS and COX-2 expression, and on levels of proinflammatory cytokines (IL-6, TNF-alpha, and PGE(2)). Eugenol markedly inhibited all. Eugenol also inhibited the upstream signaling molecule: NF-kappaB, which regulates the expression of these genes. TPA-induced depletion of cutaneous GSH and antioxidant enzymes armory was also precluded by eugenol. From these results, it could be concluded that eugenol markedly protects against chemically induced skin cancer and acts possibly by virtue of its antiproliferative, anti-inflammatory, and antioxidant activities. PMID:20043298

  14. Polyphenols of Cassia tora leaves prevents lenticular apoptosis and modulates cataract pathology in Sprague-Dawley rat pups.

    PubMed

    Sreelakshmi, V; Abraham, Annie

    2016-07-01

    Cataract is a leading cause of visual impairment worldwide with multifactorial etiology and is a significant global health problem with increasing prevalence with age. Currently, no pharmacological measures are discovered to prevent and treat cataract and a significant number of epidemiological studies have suggested the potential role of antioxidants in the prevention of cataract by scavenging free radicals and preventing lens protein derangement and lenticular cell damage. The main goal of the present study is to evaluate Cassia tora leaves; an edible leafy vegetable employed in Ayurvedic and Chinese system of medicine for eye rejuvenation in preventing selenite-induced cataract in rat pups and to identify the active components that produce the effect. ECT pre-treatment effectively restored both enzymatic and metabolic antioxidant levels, membrane integrity and reduced metal accumulation and thus down-regulate epithelial cell death. Gene expression studies also confirmed these findings. ESI-MS analysis of ECT revealed the presence of chrysophanol, emodin, kaemferol, quercetin, stigmasterol and isoquercetin. The study suggests the possible role of C. tora in alleviating cataract pathology and presence of many anthraquinones and flavonoids. As it is an edible plant, the incorporation of these leaves in daily vegetables might prevent or delay the onset and maturation of cataract. PMID:27261615

  15. Resveratrol Induces Apoptosis-Like Death and Prevents In Vitro and In Vivo Virulence of Entamoeba histolytica.

    PubMed

    Pais-Morales, Jonnatan; Betanzos, Abigail; García-Rivera, Guillermina; Chávez-Munguía, Bibiana; Shibayama, Mineko; Orozco, Esther

    2016-01-01

    Entamoeba histolytica causes amoebiasis, an infection that kills 100,000 individuals each year. Metronidazole and its derivatives are currently used against this protozoan, but these drugs present adverse effects on human health. Here, we investigated the effect of resveratrol (a natural compound) on E. histolytica trophozoites viability, as well as its influence on the parasite virulence. Trophozoites growth was arrested by 72 μM resveratrol and the IC50 was determined as 220 μM at 48 h. Cells appeared smaller, rounded and in clusters, with debris-containing vacuoles and with abnormally condensed chromatin. Resveratrol triggered reactive oxygen species production. It caused lipid peroxidation and produced phosphatidylserine externalization and DNA fragmentation this latter evidenced by TUNEL assays. It also provoked an increase of intracellular Ca2+ concentration, activated calpain and decreased superoxide dismutase activity, indicating that an apoptosis-like event occurred; however, autophagy was not detected. Cytopathic activity, phagocytosis, encystment and in vivo virulence were diminished dramatically by pre-incubation of trophozoites with resveratrol, evidencing that resveratrol attenuated the trophozoite virulence in vitro. Interestingly, after the inoculation of virulent trophozoites, animals treated with the drug did not develop or developed very small abscesses. Our findings propose that resveratrol could be an alternative to contend amoebiasis. PMID:26731663

  16. Resveratrol Induces Apoptosis-Like Death and Prevents In Vitro and In Vivo Virulence of Entamoeba histolytica

    PubMed Central

    Pais-Morales, Jonnatan; Betanzos, Abigail; García-Rivera, Guillermina; Chávez-Munguía, Bibiana; Shibayama, Mineko; Orozco, Esther

    2016-01-01

    Entamoeba histolytica causes amoebiasis, an infection that kills 100,000 individuals each year. Metronidazole and its derivatives are currently used against this protozoan, but these drugs present adverse effects on human health. Here, we investigated the effect of resveratrol (a natural compound) on E. histolytica trophozoites viability, as well as its influence on the parasite virulence. Trophozoites growth was arrested by 72 μM resveratrol and the IC50 was determined as 220 μM at 48 h. Cells appeared smaller, rounded and in clusters, with debris-containing vacuoles and with abnormally condensed chromatin. Resveratrol triggered reactive oxygen species production. It caused lipid peroxidation and produced phosphatidylserine externalization and DNA fragmentation this latter evidenced by TUNEL assays. It also provoked an increase of intracellular Ca2+ concentration, activated calpain and decreased superoxide dismutase activity, indicating that an apoptosis-like event occurred; however, autophagy was not detected. Cytopathic activity, phagocytosis, encystment and in vivo virulence were diminished dramatically by pre-incubation of trophozoites with resveratrol, evidencing that resveratrol attenuated the trophozoite virulence in vitro. Interestingly, after the inoculation of virulent trophozoites, animals treated with the drug did not develop or developed very small abscesses. Our findings propose that resveratrol could be an alternative to contend amoebiasis. PMID:26731663

  17. Acetylation of the p53 DNA binding domain regulates apoptosis induction.

    PubMed Central

    Sykes, Stephen M.; Mellert, Hestia S.; Holbert, Marc A.; Li, Keqin; Marmorstein, Ronen; Lane, William S.; McMahon, Steven B.

    2007-01-01

    SUMMARY The ability of p53 to induce apoptosis plays an important role in tumor suppression. Here we describe a previously unknown post-translational modification of the DNA-binding domain of p53. This modification, acetylation of lysine 120, occurs rapidly after DNA damage and is catalyzed by the MYST family acetyltransferases hMOF and TIP60. Mutation of lysine 120 to arginine, as occurs in human cancer, debilitates K120 acetylation and diminishes p53-mediated apoptosis without affecting cell-cycle arrest. The K120R mutation selectively blocks the transcription of pro-apoptotic target genes such as BAX and PUMA while the non-apoptotic targets p21 and hMDM2 remain unaffected. Consistent with this, depletion of hMOF and/or TIP60 inhibits the ability of p53 to activate BAX and PUMA transcription. Furthermore, the acetyl-lysine 120 form of p53 specifically accumulates at pro-apoptotic target genes. These data suggest that K120 acetylation may help distinguish the cell cycle arrest and apoptotic functions of p53. PMID:17189187

  18. Neuroglobin upregulation induced by 17β-estradiol sequesters cytocrome c in the mitochondria preventing H2O2-induced apoptosis of neuroblastoma cells

    PubMed Central

    De Marinis, E; Fiocchetti, M; Acconcia, F; Ascenzi, P; Marino, M

    2013-01-01

    The sex steroid hormone 17β-estradiol (E2) upregulates the levels of neuroglobin (NGB), a new neuroprotectant globin, to elicit its neuroprotective effect against H2O2-induced apoptosis. Several mechanisms could be proposed to justify the NGB involvement in E2 prevention of stress-induced apoptotic cell death. Here, we evaluate the ability of E2 to modulate the intracellular NGB localization and the NGB interaction with mitochondrial cytochrome c following the H2O2-induced toxicity. Present results demonstrate that NGB is expressed in the nuclei, mitochondria, and cytosol of human neuroblastoma SK-N-BE cells. E2, but not H2O2 treatment of SK-N-BE cells, reallocates NGB mainly at the mitochondria and contemporarily reduces the number of apoptotic nuclei and the levels of cleaved caspase-3. Remarkably, the E2 treatment strongly increases NGB–cytochrome c association into mitochondria and reduces the levels of cytochrome c into the cytosol of SK-N-BE cells. Although both estrogen receptors (ERα and ERβ) are expressed in the nucleus, mitochondria, and cytosol of SK-N-BE cells, this E2 effect specifically requires the mitochondrial ERβ activity. As a whole, these data demonstrate that the interception of the intrinsic apoptotic pathway into mitochondria (i.e., the prevention of cytochrome c release) is one of the pivotal mechanisms underlying E2-dependent NGB neuroprotection against H2O2 toxicity. PMID:23429294

  19. Prevention

    MedlinePlus

    ... our e-newsletter! Aging & Health A to Z Prevention Basic Facts & Information Some factors that affect your ... control of the things that you can change. Preventive Recommendations for Adults Aged 65 and Older The ...

  20. GH-Releasing Hormone Promotes Survival and Prevents TNF-α-Induced Apoptosis and Atrophy in C2C12 Myotubes.

    PubMed

    Gallo, Davide; Gesmundo, Iacopo; Trovato, Letizia; Pera, Giulia; Gargantini, Eleonora; Minetto, Marco Alessandro; Ghigo, Ezio; Granata, Riccarda

    2015-09-01

    Skeletal muscle atrophy is a consequence of different chronic diseases, including cancer, heart failure, and diabetes, and also occurs in aging and genetic myopathies. It results from an imbalance between anabolic and catabolic processes, and inflammatory cytokines, such as TNF-α, have been found elevated in muscle atrophy and implicated in its pathogenesis. GHRH, in addition to stimulating GH secretion from the pituitary, exerts survival and antiapoptotic effects in different cell types. Moreover, we and others have recently shown that GHRH displays antiapoptotic effects in isolated cardiac myocytes and protects the isolated heart from ischemia/reperfusion injury and myocardial infarction in vivo. On these bases, we investigated the effects of GHRH on survival and apoptosis of TNF-α-treated C2C12 myotubes along with the underlying mechanisms. GHRH increased myotube survival and prevented TNF-α-induced apoptosis through GHRH receptor-mediated mechanisms. These effects involved activation of phosphoinositide 3-kinase/Akt pathway and inactivation of glycogen synthase kinase-3β, whereas mammalian target of rapamycin was unaffected. GHRH also increased the expression of myosin heavy chain and the myogenic transcription factor myogenin, which were both reduced by the cytokine. Furthermore, GHRH inhibited TNF-α-induced expression of nuclear factor-κB, calpain, and muscle ring finger1, which are all involved in muscle protein degradation. In summary, these results indicate that GHRH exerts survival and antiapoptotic effects in skeletal muscle cells through the activation of anabolic pathways and the inhibition of proteolytic routes. Overall, our findings suggest a novel therapeutic role for GHRH in the treatment of muscle atrophy-associated diseases. PMID:26110916

  1. Phosphorylation of Tip60 by GSK-3 determines the induction of PUMA and apoptosis by p53

    PubMed Central

    Charvet, Céline; Wissler, Manuela; Brauns-Schubert, Prisca; Wang, Shang-Jui; Tang, Yi; Sigloch, Florian C.; Mellert, Hestia; Brandenburg, Martin; Lindner, Silke E.; Breit, Bernhard; Green, Douglas R.; McMahon, Steven B.; Borner, Christoph; Gu, Wei; Maurer, Ulrich

    2011-01-01

    Summary Activation of p53 by DNA damage results in either cell cycle arrest, allowing DNA repair and cell survival, or induction of apoptosis. As these opposite outcomes are both mediated by p53 stabilization, additional mechanisms to determine this decision must exist. Here we show that glycogen synthase kinase-3 (GSK-3) is required for the p53-mediated induction of the pro-apoptotic BH3 only-protein PUMA, an essential mediator of p53-induced apoptosis. Inhibition of GSK-3 protected from cell death induced by DNA damage and promoted increased long-term cell survival. We demonstrate that GSK-3 phosphorylates serine 86 of the p53-acetyltransferase Tip60. A Tip60S86A mutant was less active to induce p53 K120 acetylation, Histone 4 acetylation and expression of PUMA. Our data suggest that GSK-3 mediated Tip60S86-phosphorylation provides a link between PI3K signaling and the choice for or against apoptosis induction by p53. PMID:21658600

  2. ACTIVITY-DEPENDENT NEUROPROTECTIVE PROTEIN–DERIVED PEPTIDE, NAP, PREVENTING ALCOHOL-INDUCED APOPTOSIS IN FETAL BRAIN OF C57BL/6 MOUSE

    PubMed Central

    SARI, Y.

    2012-01-01

    Possible prevention of the effects of prenatal alcohol exposure has been investigated using peptides that were previously shown to be involved in neuroprotection both in vitro and in vivo. I focused in this study on investigating the neuro-protective effects of one of these peptides with regard to the determination of the downstream signaling pathways involved in neuroprotection. This peptide with the sequence NAPVSIPQ, known as NAP, a fragment of activity-dependent neuroprotective protein, demonstrated a potent protective effect against oxidative stress associated with alcohol exposure. On embryonic day 7 (E7), weight-matched C57BL/6 pregnant females were assigned the following groups: (1) Ethanol liquid diet group (ALC) 25% (4.49%, v/v) ethano-derived calories, (2) Pair-fed (PF) control group (3) Chow control group, (4) treatment groups with alcohol alongside i.p. injections of d-NAP (ALC/d-NAP, 20 or 30 μg/20 g body weight), (5) PF/d-NAP control group. On E13, fetal brains were collected and assayed for TdT-mediated dUTP nick end labeling (TUNEL) staining, caspase-3 colorimetric assay and ELISA for cytochrome c detection. My results show that NAP significantly prevented alcohol-induced weight reduction of the fetal brain. Apoptosis was determined by TUNEL staining; NAP administration significantly prevented alcohol-induced increases in TUNEL-positive cells in primordium cingulate cortex and basal ganglia eminence. The investigation of downstream signaling pathways involving NAP neuroprotection revealed that this peptide significantly prevented alcohol-induced increase in the concentrations of caspase-3 in E13 fetal brains. Moreover, ELISA for cytochrome c shows that NAP significantly prevented both alcohol-induced increases in the level of cytosolic cytochrome c and alcohol-induced decreases in the level of mitochondrial cytochrome c. These data provide an understanding of NAP intracellular target, and the downstream mechanisms of action that will pave a path

  3. Retinoic Acid Induced-Autophagic Flux Inhibits ER-Stress Dependent Apoptosis and Prevents Disruption of Blood-Spinal Cord Barrier after Spinal Cord Injury

    PubMed Central

    Zhou, Yulong; Zhang, Hongyu; Zheng, Binbin; Ye, Libing; Zhu, Sipin; Johnson, Noah R; Wang, Zhouguang; Wei, Xiaojie; Chen, Daqing; Cao, Guodong; Fu, Xiaobing; Li, Xiaokun; Xu, Hua-Zi; Xiao, Jian

    2016-01-01

    Spinal cord injury (SCI) induces the disruption of the blood-spinal cord barrier (BSCB) which leads to infiltration of blood cells, an inflammatory response, and neuronal cell death, resulting spinal cord secondary damage. Retinoic acid (RA) has a neuroprotective effect in both ischemic brain injury and SCI, however the relationship between BSCB disruption and RA in SCI is still unclear. In this study, we demonstrated that autophagy and ER stress are involved in the protective effect of RA on the BSCB. RA attenuated BSCB permeability and decreased the loss of tight junction (TJ) molecules such as P120, β-catenin, Occludin and Claudin5 after injury in vivo as well as in Brain Microvascular Endothelial Cells (BMECs). Moreover, RA administration improved functional recovery in the rat model of SCI. RA inhibited the expression of CHOP and caspase-12 by induction of autophagic flux. However, RA had no significant effect on protein expression of GRP78 and PDI. Furthermore, combining RA with the autophagy inhibitor chloroquine (CQ) partially abolished its protective effect on the BSCB via exacerbated ER stress and subsequent loss of tight junctions. Taken together, the neuroprotective role of RA in recovery from SCI is related to prevention of of BSCB disruption via the activation of autophagic flux and the inhibition of ER stress-induced cell apoptosis. These findings lay the groundwork for future translational studies of RA for CNS diseases, especially those related to BSCB disruption. PMID:26722220

  4. Nuclear glutathione S-transferase pi prevents apoptosis by reducing the oxidative stress-induced formation of exocyclic DNA products.

    PubMed

    Kamada, Kensaku; Goto, Shinji; Okunaga, Tomohiro; Ihara, Yoshito; Tsuji, Kentaro; Kawai, Yoshichika; Uchida, Koji; Osawa, Toshihiko; Matsuo, Takayuki; Nagata, Izumi; Kondo, Takahito

    2004-12-01

    We previously found that nuclear glutathione S-transferase pi (GSTpi) accumulates in cancer cells resistant to anticancer drugs, suggesting that it has a role in the acquisition of resistance to anticancer drugs. In the present study, the effect of oxidative stress on the nuclear translocation of GSTpi and its role in the protection of DNA from damage were investigated. In human colonic cancer HCT8 cells, the hydrogen peroxide (H(2)O(2))-induced increase in nuclear condensation, the population of sub-G(1) peak, and the number of TUNEL-positive cells were observed in cells pretreated with edible mushroom lectin, an inhibitor of the nuclear transport of GSTpi. The DNA damage and the formation of lipid peroxide were dependent on the dose of H(2)O(2) and the incubation time. Immunological analysis showed that H(2)O(2) induced the nuclear accumulation of GSTpi but not of glutathione peroxidase. Formation of the 7-(2-oxo-hepyl)-substituted 1,N(2)-etheno-2'-deoxyguanosine adduct by the reaction of 13-hydroperoxyoctadecadienoic acid (13-HPODE) with 2'-deoxyguanosine was inhibited by GSTpi in the presence of glutathione. The conjugation product of 4-oxo-2-nonenal, a lipid aldehyde of 13-HPODE, with GSH in the presence of GSTpi, was identified by LS/MS. These results suggested that nuclear GSTpi prevents H(2)O(2)-induced DNA damage by scavenging the formation of lipid-peroxide-modified DNA. PMID:15528046

  5. p53-dependent apoptosis contributes to di-(2-ethylhexyl) phthalate-induced hepatotoxicity.

    PubMed

    Ha, Mei; Wei, Li; Guan, Xie; Li, Lianbing; Liu, Changjiang

    2016-01-01

    Di-(2-ethylhexyl) phthalate (DEHP) is used extensively in many personal care and consumer products, resulting in widespread non-occupational human exposure through multiple routes and media. DEHP has various deleterious effects including hepatotoxicity. p53 protein is a central sensor in cell apoptosis. In order to clarify the roles of p53 in DEHP-induced hepatotoxicity, Sprague-Dawley (SD) rats were dosed daily with DEHP by gavage for 30 days; BRL cells (rat liver cell line) were treated with DEHP for 24 h after pretreatment with NAC or small interfering RNA (siRNA). Results indicated that after exposure to DEHP, hepatic histological changes such as hepatocyte edema, vacuolation and hepatic sinusoidal dilation, and increased apoptosis index were observed. In the liver, DEHP induced oxidative stress and DNA damage, which activated p53 in vivo and in vitro. Pretreatment with NAC significantly reduced ROS level and p53 expression in BRL cells. The suppressed Mdm2 also contributed to p53 accumulation. Activated p53 mediated hepatocyte apoptosis via the intrinsic mitochondrial pathway, inhibiting anti-apoptotic Bcl-2 and Bcl-xL and inducing pro-apoptotic Bax, cytochrome c and caspases. In p53-silenced BRL cells, hepatocyte apoptosis mediated by p53 was attenuated. PCNA protein level was upregulated after p53 gene silencing. However, the Fas/FasL apoptotic pathway did not exhibit activated signs in DEHP-caused hepatotoxicity. Taken together, DEHP-caused oxidative stress and Mdm2 downregulation contribute to p53 activation. The p53-dependent apoptotic pathway plays critical and indispensable roles in DEHP-induced hepatotoxicity, while the Fas/FasL pathway does not involve in this molecular event. PMID:26549752

  6. Prevention

    MedlinePlus

    ... Prevention Treatment 2003 U.S. Outbreak African Rodent Importation Ban For Clinicians Clinical Recognition Specimen Collection Treatment Smallpox ... Examining Animals with Suspected Monkeypox African Rodent Importation Ban Resources Related Links Poxvirus Molluscum Contagiosum Orf Virus ( ...

  7. Wild-type p53-mediated down-modulation of interleukin 15 and interleukin 15 receptors in human rhabdomyosarcoma cells.

    PubMed Central

    De Giovanni, C.; Nanni, P.; Sacchi, A.; Soddu, S.; Manni, I.; D'Orazi, G.; Bulfone-Paus, S.; Pohl, T.; Landuzzi, L.; Nicoletti, G.; Frabetti, F.; Rossi, I.; Lollini, P. L.

    1998-01-01

    We recently reported that rhabdomyosarcoma cell lines express and secrete interleukin 15 (IL-15), a tightly regulated cytokine with IL-2-like activity. To test whether the p53-impaired function that is frequently found in this tumour type could play a role in the IL-15 production, wild-type p53 gene was transduced in the human rhabdomyosarcoma cell line RD (which harbours a mutated p53 gene), and its effect on proliferation and expression of IL-15 was studied. Arrest of proliferation was induced by wild-type p53; increased proportions of G1-arrested cells and of apoptotic cells were observed. A marked down-modulation of IL-15 expression, at both the mRNA and protein level, was found in p53-transduced cells. Because a direct effect of IL-15 on normal muscle cells has been reported, the presence of IL-15 membrane receptors was studied by cytofluorometric analysis. Rhabdomyosarcoma cells showed IL-15 membrane receptors, which are down-modulated by wild-type p53 transfected gene. In conclusion, wild-type p53 transduction in human rhabdomyosarcoma cells induces the down-modulation of both IL-15 production and IL-15 receptor expression. Images Figure 3 PMID:9862562

  8. Novel small molecule induces p53-dependent apoptosis in human colon cancer cells

    SciTech Connect

    Park, Sang Eun; Min, Yong Ki; Ha, Jae Du; Kim, Bum Tae; Lee, Woo Ghil . E-mail: bigguy@krict.re.kr

    2007-07-06

    Using high-throughput screening with small-molecule libraries, we identified a compound, KCG165 [(2-(3-(2-(pyrrolidin-1-yl)ethoxy)-1,10b-dihydro-[1,2,4]triazolo[1,5-c] quinazolin-5(6H)-one)], which strongly activated p53-mediated transcriptional activity. KCG165-induced phosphorylations of p53 at Ser{sup 6}, Ser{sup 15}, and Ser{sup 20}, which are all key residues involved in the activation and stabilization of p53. Consistent with these findings, KCG165 increased level of p53 protein and led to the accumulation of transcriptionally active p53 in the nucleus with the increased occupancy of p53 in the endogenous promoter region of its downstream target gene, p21{sup WAF1/CIP}. Notably, KCG165-induced p53-dependent apoptosis in cancer cells. Furthermore, we suggested topoisomerase II as the molecular target of KCG165. Together, these results indicate that KCG165 may have potential applications as an antitumor agent.

  9. Damnacanthal is a potent inducer of apoptosis with anticancer activity by stimulating p53 and p21 genes in MCF-7 breast cancer cells.

    PubMed

    Aziz, Muhammad Yusran Abdul; Omar, Abdul Rahman; Subramani, Tamilselvan; Yeap, Swee Keong; Ho, Wan Yong; Ismail, Nor Hadiani; Ahmad, Syahida; Alitheen, Noorjahan Banu

    2014-05-01

    Damnacanthal, an anthraquinone compound, is isolated from the roots of Morinda citrifolia L. (noni), which has been used for traditional therapy in several chronic diseases, including cancer. Although noni has long been consumed in Asian and Polynesian countries, the molecular mechanisms by which it exerts several benefits are starting to emerge. In the present study, the effect of damnacanthal on MCF-7 cell growth regulation was investigated. Treatment of MCF-7 cells with damnacanthal for 72 h indicated an antiproliferative activity. The MTT method confirmed that damnacanthal inhibited the growth of MCF-7 cells at the concentration of 8.2 μg/ml for 72 h. In addition, the drug was found to induce cell cycle arrest at the G1 checkpoint in MCF-7 cells by cell cycle analysis. Damnacanthal induced apoptosis, determined by Annexin V-fluorescein isothiocyanate/propidium iodide (PI) dual-labeling, acridine-orange/PI dyeing and caspase-7 expression. Furthermore, damnacanthal-mediated apoptosis involves the sustained activation of p21, leading to the transcription of p53 and the Bax gene. Overall, the present study provided significant evidence demonstrating that p53-mediated damnacanthal induced apoptosis through the activation of p21 and caspase-7. PMID:24765160

  10. Phosphorylated AKT inhibits the apoptosis induced by DRAM-mediated mitophagy in hepatocellular carcinoma by preventing the translocation of DRAM to mitochondria.

    PubMed

    Liu, K; Shi, Y; Guo, X H; Ouyang, Y B; Wang, S S; Liu, D J; Wang, A N; Li, N; Chen, D X

    2014-01-01

    Increasing autophagy is beneficial for curing hepatocellular carcinoma (HCC). Damage-regulated autophagy modulator (DRAM) was recently reported to induce apoptosis by mediating autophagy. However, the effects of DRAM-mediated autophagy on apoptosis in HCC cells remain unclear. In this study, normal hepatocytes (7702) and HCC cell lines (HepG2, Hep3B and Huh7) were starved for 48 h. Starvation induced apoptosis and autophagy in all cell lines. We determined that starvation also induced DRAM expression and DRAM-mediated autophagy in both normal hepatocytes and HCC cells. However, DRAM-mediated autophagy was involved in apoptosis in normal hepatocytes but not in HCC cells, suggesting that DRAM-mediated autophagy fails to induce apoptosis in hepatoma in response to starvation. Immunoblot and immunofluorescence assays demonstrated that DRAM translocated to mitochondria and induced mitophagy, which led to apoptosis in 7702 cells. In HCC cells, starvation also activated the phosphatidylinositol 3-kinase (PI3K)/AKT pathway, which blocks the translocation of DRAM to mitochondria through the binding of p-AKT to DRAM in the cytoplasm. Inactivation of the PI3K/AKT pathway rescued DRAM translocation to mitochondria; subsequently, mitochondrial DRAM induced apoptosis in HCC cells by mediating mitophagy. Our findings open new avenues for the investigation of the mechanisms of DRAM-mediated autophagy and suggest that promoting DRAM-mediated autophagy together with PI3K/AKT inhibition might be more effective for autophagy-based therapy in hepatoma. PMID:24556693