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Sample records for pathway protects cardiomyocytes

  1. MicroRNA-145 protects cardiomyocytes against hydrogen peroxide (H₂O₂-induced apoptosis through targeting the mitochondria apoptotic pathway.

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    Ruotian Li

    Full Text Available MicroRNAs, a class of small and non-encoding RNAs that transcriptionally or post-transcriptionally modulate the expression of their target genes, has been implicated as critical regulatory molecules in many cardiovascular diseases, including ischemia/reperfusion induced cardiac injury. Here, we report microRNA-145, a tumor suppressor miRNA, can protect cardiomyocytes from hydrogen peroxide H₂O₂-induced apoptosis through targeting the mitochondrial pathway. Quantitative real-time PCR (qPCR demonstrated that the expression of miR-145 in either ischemia/reperfused mice myocardial tissues or H₂O₂-treated neonatal rat ventricle myocytes (NRVMs was markedly down-regulated. Over-expression of miR-145 significantly inhibited the H₂O₂-induced cellular apoptosis, ROS production, mitochondrial structure disruption as well as the activation of key signaling proteins in mitochondrial apoptotic pathway. These protective effects of miR-145 were abrogated by over-expression of Bnip3, an initiation factor of the mitochondrial apoptotic pathway in cardiomyocytes. Finally, we utilized both luciferase reporter assay and western blot analysis to identify Bnip3 as a direct target of miR-145. Our results suggest miR-145 plays an important role in regulating mitochondrial apoptotic pathway in heart challenged with oxidative stress. MiR-145 may represent a potential therapeutic target for treatment of oxidative stress-associated cardiovascular diseases, such as myocardial ischemia/reperfusion injury.

  2. miR-138 protects cardiomyocytes from hypoxia-induced apoptosis via MLK3/JNK/c-jun pathway

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    He, Siyi; Liu, Peng; Jian, Zhao; Li, Jingwei; Zhu, Yun; Feng, Zezhou; Xiao, Yingbin, E-mail: xiaoyb@vip.sina.com

    2013-11-29

    Highlights: •First time to find miR-138 is up-regulated in hypoxic cardiomyocytes. •First time to find miR-138 targets MLK3 and regulates JNK/c-jun pathway. •Rare myocardial biopsy of patients with CHD were collected. •Both silence and overexpression of miR-138 were implemented. •Various methods were used to detect cell function. -- Abstract: Cardiomyocytes experience a series of complex endogenous regulatory mechanisms against apoptosis induced by chronic hypoxia. MicroRNAs are a class of endogenous small non-coding RNAs that regulate cellular pathophysiological processes. Recently, microRNA-138 (miR-138) has been found related to hypoxia, and beneficial for cell proliferation. Therefore, we intend to study the role of miR-138 in hypoxic cardiomyocytes and the main mechanism. Myocardial samples of patients with congenital heart disease (CHD) were collected to test miR-138 expression. Agomir or antagomir of miR-138 was transfected into H9C2 cells to investigate its effect on cell apoptosis. Higher miR-138 expression was observed in patients with cyanotic CHD, and its expression gradually increased with prolonged hypoxia time in H9C2 cells. Using MTT and LDH assays, cell growth was significantly greater in the agomir group than in the negative control (NC) group, while antagomir decreased cell survival. Dual luciferase reporter gene and Western-blot results confirmed MLK3 was a direct target of miR-138. It was found that miR-138 attenuated hypoxia-induced apoptosis using TUNEL, Hoechst staining and Annexin V-PE/7-AAD flow cytometry analysis. We further detected expression of apoptosis-related proteins. In the agomir group, the level of pro-apoptotic proteins such as cleaved-caspase-3, cleaved-PARP and Bad significantly reduced, while Bcl-2 and Bcl-2/Bax ratio increased. Opposite changes were observed in the antagomir group. Downstream targets of MLK3, JNK and c-jun, were also suppressed by miR-138. Our study demonstrates that up-regulation of miR-138 plays

  3. Catestatin exerts direct protective effects on rat cardiomyocytes undergoing ischemia/reperfusion by stimulating PI3K-Akt-GSK3β pathway and preserving mitochondrial membrane potential.

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    Eleonora Bassino

    Full Text Available Catestatin (Cst is a 21-amino acid peptide deriving from Chromogranin A. Cst exerts an overall protective effect against an excessive sympathetic stimulation of cardiovascular system, being able to antagonize catecholamine secretion and to reduce their positive inotropic effect, by stimulating the release of nitric oxide (NO from endothelial cells. Moreover, Cst reduces ischemia/reperfusion (I/R injury, improving post-ischemic cardiac function and cardiomyocyte survival. To define the cardioprotective signaling pathways activated by Cst (5 nM we used isolated adult rat cardiomyocytes undergoing simulated I/R. We evaluated cell viability rate with propidium iodide labeling and mitochondrial membrane potential (MMP with the fluorescent probe JC-1. The involvement of Akt, GSK3β, eNOS and phospholamban (PLN cascade was studied by immunofluorescence. The role of PI3K-Akt/NO/cGMP pathway was also investigated by using the pharmacological blockers wortmannin (Wm, L-NMMA and ODQ. Our experiments revealed that Cst increased cell viability rate by 65% and reduced cell contracture in I/R cardiomyocytes. Wm, L-NMMA and ODQ limited the protective effect of Cst. The protective outcome of Cst was related to its ability to maintain MMP and to increase AktSer473, GSK3βSer9, PLNThr17 and eNOSSer1179 phosphorylation, while treatment with Wm abolished these effects. Thus, the present results show that Cst is able to exert a direct action on cardiomyocytes and give new insights into the molecular mechanisms involved in its protective effect, highlighting the PI3K/NO/cGMP pathway as the trigger and the MMP preservation as the end point of its action.

  4. Ginsenoside Rb1 Protects Neonatal Rat Cardiomyocytes from Hypoxia/Ischemia Induced Apoptosis and Inhibits Activation of the Mitochondrial Apoptotic Pathway

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    Xu Yan

    2014-01-01

    Full Text Available Aim. To investigate the effect of Ginsenoside Rb1 (GS-Rb1 on hypoxia/ischemia (H/I injury in cardiomyocytes in vitro and the mitochondrial apoptotic pathway mediated mechanism. Methods. Neonatal rat cardiomyocytes (NRCMs for the H/I groups were kept in DMEM without glucose and serum, and were placed into a hypoxic jar for 24 h. GS-Rb1 at concentrations from 2.5 to 40 µM was given during hypoxic period for 24 h. NRCMs injury was determined by MTT and lactate dehydrogenase (LDH leakage assay. Cell apoptosis, ROS accumulation, and mitochondrial membrane potential (MMP were assessed by flow cytometry. Cytosolic translocation of mitochondrial cytochrome c and Bcl-2 family proteins were determined by Western blot. Caspase-3 and caspase-9 activities were determined by the assay kit. Results. GS-Rb1 significantly reduced cell death and LDH leakage induced by H/I. It also reduced H/I induced NRCMs apoptosis induced by H/I, in accordance with a minimal reactive oxygen species (ROS burst. Moreover, GS-Rb1 markedly decreased the translocation of cytochrome c from the mitochondria to the cytosol, increased the Bcl-2/ Bax ratio, and preserved mitochondrial transmembrane potential (ΔΨm. Its administration also inhibited activities of caspase-9 and caspase-3. Conclusion. Administration of GS-Rb1 during H/I in vitro is involved in cardioprotection by inhibiting apoptosis, which may be due to inhibition of the mitochondrial apoptotic pathway.

  5. Melatonin attenuates angiotensin II-induced cardiomyocyte hypertrophy through the CyPA/CD147 signaling pathway.

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    Su, Hongyan; Li, Jingyuan; Chen, Tongshuai; Li, Na; Xiao, Jie; Wang, Shujian; Guo, Xiaobin; Yang, Yi; Bu, Peili

    2016-11-01

    Melatonin is well known for its cardioprotective effects; however, whether melatonin exerts therapeutic effects on cardiomyocyte hypertrophy remains to be investigated, as do the mechanisms underlying these effects, if they exist. Cyclophilin A (CyPA) and its corresponding receptor, CD147, which exists in a variety of cells, play crucial roles in modulating reactive oxygen species (ROS) production. In this study, we explored the role of the CyPA/CD147 signaling pathway in angiotensin II (Ang II)-induced cardiomyocyte hypertrophy and the protective effects exerted by melatonin against Ang II-induced injury in cultured H9C2 cells. Cyclosporine A, a specific CyPA/CD147 signaling pathway inhibitor, was used to manipulate CyPA/CD147 activity. H9C2 cells were then subjected to Ang II or CyPA treatment in either the absence or presence of melatonin. Our results indicate that Ang II induces cardiomyocyte hypertrophy through the CyPA/CD147 signaling pathway and promotes ROS production, which can be blocked by melatonin pretreatment in a concentration-dependent manner, in cultured H9C2 cells and that CyPA/CD147 signaling pathway inhibition protects against Ang II-induced cardiomyocyte hypertrophy. The protective effects of melatonin against Ang II-induced cardiomyocyte hypertrophy depend at least partially on CyPA/CD147 inhibition.

  6. Insulin protects apoptotic cardiomyocytes from hypoxia/reoxygenation injury through the sphingosine kinase/sphingosine 1-phosphate axis.

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    Huan Yu

    Full Text Available OBJECTIVE: Experimental and clinical studies have shown that administration of insulin during reperfusion is cardioprotective, but the mechanisms underlying this effect are still unknown. In this study, the ability of insulin to protect apoptotic cardiomyocytes from hypoxia/reoxygenation injury using the sphingosine kinase/sphingosine 1-phosphate axis was investigated. METHODS AND RESULTS: Rat cardiomyocytes were isolated and subjected to hypoxia and reoxygenation. [γ-32P] ATP was used to assess sphingosine kinase activity. Insulin was found to increase sphingosine kinase activity. Immunocytochemistry and Western blot analysis showed changes in the subcellular location of sphingosine kinase 1 from cytosol to the membrane in cardiomyocytes. Insulin caused cardiomyocytes to accumulate of S1P in a dose-dependent manner. FRET efficiency showed that insulin also transactivates the S1P1 receptor. TUNEL staining showed that administration of insulin during reoxygenation could to reduce the rate of reoxygenation-induced apoptosis, which is a requirement for SphK 1 activity. It also reduced the rate of activation of the S1P receptor and inhibited hypoxia/reoxygenation-induced cell death in cardiomyocytes. CONCLUSION: The sphingosine kinase 1/sphingosine 1-phosphate/S1P receptor axis is one pathway through which insulin protects rat cardiomyocytes from apoptosis induced by hypoxia/reoxygenation injury.

  7. DPP4 deficiency exerts protective effect against H2O2 induced oxidative stress in isolated cardiomyocytes.

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    Hui-Chun Ku

    Full Text Available Apart from the antihyperglycemic effects, DPP4 inhibitors and GLP-1 molecules are involved in the preservation of cardiac functions. We have demonstrated that DPP4-deficient rats possess resistance to endotoxemia and ischemia/reperfusion stress. However, whether the decrease of DPP4 activity simply augmented the GLP-1 signaling or that such decrease resulted in a change of cellular function remain unclear. Accordingly, we investigated the responses of H(2O(2-induced oxidative stress in adult wild-type and DPP4-deficient rats isolated cardiomyocytes. The coadministration of GLP-1 or DPP4 inhibitor was also performed to define the mechanisms. Cell viability, ROS concentration, catalase activity, glucose uptake, prosurvival, proapoptotic signaling, and contractile function were examined after cells exposed to H(2O(2. DPP4-deficient cardiomyocytes were found to be resistant to H(2O(2-induced cell death via activating AKT signaling, enhancing glucose uptake, preserving catalase activity, diminishing ROS level and proapoptotic signaling. GLP-1 concentration-dependently improved cell viability in wild-type cardiomyocyte against ROS stress, and the ceiling response concentration (200 nM was chosen for studies. GLP-1 was shown to decrease H(2O(2-induced cell death by its receptor-dependent AKT pathway in wild-type cardiomyocytes, but failed to cause further activation of AKT in DPP4-deficient cardiomyocytes. Acute treatment of DPP4 inhibitor only augmented the protective effect of low dose GLP-1, but failed to alter fuel utilization or ameliorate cell viability in wild-type cardiomyocytes after H(2O(2 exposure. The improvement of cell viability after H(2O(2 exposure was correlated with the alleviation of cellular contractile dysfunction in both DPP4-deficient and GLP-1 treated wild-type cardiomyocytes. These findings demonstrated that GLP-1 receptor-dependent pathway is important and exert protective effect in wild-type cardiomyocyte. Long term loss of

  8. Cardiomyocytes undergo apoptosis in human immunodeficiency virus cardiomyopathy through mitochondrion- and death receptor-controlled pathways.

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    Twu, Cheryl; Liu, Nancy Q; Popik, Waldemar; Bukrinsky, Michael; Sayre, James; Roberts, Jaclyn; Rania, Shammas; Bramhandam, Vishnu; Roos, Kenneth P; MacLellan, W Robb; Fiala, Milan

    2002-10-29

    We investigated 18 AIDS hearts (5 with and 13 without cardiomyopathy) by using immunocytochemistry and computerized image analysis regarding the roles of HIV-1 proteins and tumor necrosis factor ligands in HIV cardiomyopathy (HIVCM). HIVCM and cardiomyocyte apoptosis were significantly related to each other and to the expression by inflammatory cells of gp120 and tumor necrosis factor-alpha. In HIVCM heart, active caspase 9, a component of the mitochondrion-controlled apoptotic pathway, and the elements of the death receptor-mediated pathway, tumor necrosis factor-alpha and Fas ligand, were expressed strongly on macrophages and weakly on cardiomyocytes. HIVCM showed significantly greater macrophage infiltration and cardiomyocyte apoptosis rate compared with non-HIVCM. HIV-1 entered cultured neonatal rat ventricular myocytes by macropinocytosis but did not replicate. HIV-1- or gp120-induced apoptosis of rat myocytes through a mitochondrion-controlled pathway, which was inhibited by heparin, AOP-RANTES, or pertussis toxin, suggesting that cardiomyocyte apoptosis is induced by signaling through chemokine receptors. In conclusion, in patients with HIVCM, cardiomyocytes die through both mitochondrion- and death receptor-controlled apoptotic pathways.

  9. Exogenous cathepsin V protein protects human cardiomyocytes HCM from angiotensin Ⅱ-Induced hypertrophy.

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    Huang, Kun; Gao, Lu; Yang, Ming; Wang, Jiliang; Wang, Zheng; Wang, Lin; Wang, Guobin; Li, Huili

    2017-08-01

    Angiotensin (Ang) Ⅱ-induced cardiac hypertrophy can deteriorate to heart failure, a leading cause of mortality. Endogenous Cathepsin V (CTSV) has been reported to be cardioprotective against hypertrophy. However, little is known about the effect of exogenous CTSV on cardiac hypertrophy. We used the human cardiomyocytes HCM as a cell model to investigate the effects of exogenous CTSV on Ang Ⅱ-induced cardiac cell hypertrophy. Cell surface area and expression of classical markers of hypertrophy were analyzed. We further explored the mechanism of CTSV cardioprotective by assessing the levels and activities of PI3K/Akt/mTOR and MAPK signaling pathway proteins. We found that pre-treating cardiomyocytes with CTSV could significantly inhibit Ang Ⅱ-induced hypertrophy. The mRNA expression of hypertrophy markers ANP, BNP and β-MHC was obviously elevated in Ang Ⅱ-treated cardiac cells. Whereas, exogenous CTSV effectively halted this elevation. Further study revealed that the protective effects of exogenous CTSV might be mediated by repressing the phosphorylation of proteins in the PI3K/Akt/mTOR and MAPK pathways. Based on our results, we concluded that exogenous CTSV inhibited Ang Ⅱ-induced hypertrophy in HCM cells by inhibiting PI3K/Akt/mTOR. This study provides experimental evidence for the application of CTSV protein for the treatment of cardiac hypertrophy. Copyright © 2017 Elsevier Ltd. All rights reserved.

  10. SurR9C84A protects and recovers human cardiomyocytes from hypoxia induced apoptosis

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    Ashok, Ajay [Nanomedicine-Laboratory of Immunology and Molecular Biomedical Research (NLIMBR), School of Medicine (SoM), Faculty of Health, Centre for Molecular and Medical Research - C-MMR, Deakin University, Waurn Ponds, Victoria 3216 (Australia); Department of Pathology, Case Western Reserve University, 2103 Cornell Rd. WRB 5128, Cleveland, OH 44106-7288 (United States); Kanwar, Jagat Rakesh [Nanomedicine-Laboratory of Immunology and Molecular Biomedical Research (NLIMBR), School of Medicine (SoM), Faculty of Health, Centre for Molecular and Medical Research - C-MMR, Deakin University, Waurn Ponds, Victoria 3216 (Australia); Krishnan, Uma Maheswari [Centre for Nanotechnology & Advanced Biomaterials (CeNTAB), School of Chemical & Biotechnology (SCBT), SASTRA University, Thanjavur 613401 (India); Kanwar, Rupinder Kaur, E-mail: rupinder.kanwar@deakin.edu.au [Nanomedicine-Laboratory of Immunology and Molecular Biomedical Research (NLIMBR), School of Medicine (SoM), Faculty of Health, Centre for Molecular and Medical Research - C-MMR, Deakin University, Waurn Ponds, Victoria 3216 (Australia)

    2017-01-01

    . - Highlights: • Protection/regeneration of dying myocardium post myocardial infarction is important. • Downregulation of survivin induces apoptosis in hypoxic human cardiomyocytes (HCM). • Bio-replenishment with SurR9-C84A reinstates HCM survival, recovery and growth. • SurR9-C84A targets mitochondrial depolarization, fcTnT and ROS generation in HCM. • SurR9-C84A upregulates survivin, PCNA, PI3K/Akt pathway, VEGF and HSP levels. • SurR9-C84A holds promise as a treatment and preventive agent to replenish survivin.

  11. DNA damage-inducible transcript 4 (DDIT4) mediates methamphetamine-induced autophagy and apoptosis through mTOR signaling pathway in cardiomyocytes

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    Chen, Rui; Wang, Bin; Chen, Ling; Cai, Dunpeng; Li, Bing; Chen, Chuanxiang; Huang, Enping; Liu, Chao; Lin, Zhoumeng; Xie, Wei-Bing; Wang, Huijun

    2016-01-01

    Methamphetamine (METH) is an amphetamine-like psychostimulant that is commonly abused. Previous studies have shown that METH can induce damages to the nervous system and recent studies suggest that METH can also cause adverse and potentially lethal effects on the cardiovascular system. Recently, we demonstrated that DNA damage-inducible transcript 4 (DDIT4) regulates METH-induced neurotoxicity. However, the role of DDIT4 in METH-induced cardiotoxicity remains unknown. We hypothesized that DDIT4 may mediate METH-induced autophagy and apoptosis in cardiomyocytes. To test the hypothesis, we examined DDIT4 protein expression in cardiomyocytes and in heart tissues of rats exposed to METH with Western blotting. We also determined the effects on METH-induced autophagy and apoptosis after silencing DDIT4 expression with synthetic siRNA with or without pretreatment of a mTOR inhibitor rapamycin in cardiomyocytes using Western blot analysis, fluorescence microscopy and TUNEL staining. Our results showed that METH exposure increased DDIT4 expression and decreased phosphorylation of mTOR that was accompanied with increased autophagy and apoptosis both in vitro and in vivo. These effects were normalized after silencing DDIT4. On the other hand, rapamycin promoted METH-induced autophagy and apoptosis in DDIT4 knockdown cardiomyocytes. These results suggest that DDIT4 mediates METH-induced autophagy and apoptosis through mTOR signaling pathway in cardiomyocytes. - Highlights: • METH exposure increases DDIT4 expression in cardiomyocytes. • DDIT4 mediates METH-induced autophagy and apoptosis in cardiomyocytes. • DDIT4 silencing protects cardiomyocytes against METH-caused autophagy and apoptosis.

  12. DNA damage-inducible transcript 4 (DDIT4) mediates methamphetamine-induced autophagy and apoptosis through mTOR signaling pathway in cardiomyocytes

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    Chen, Rui [Department of Forensic Medicine, School of Basic Medical Science, Southern Medical University, Guangzhou 510515 (China); Department of Forensic Medicine, Guangdong Medical University, Dongguan 523808 (China); Wang, Bin; Chen, Ling; Cai, Dunpeng; Li, Bing; Chen, Chuanxiang; Huang, Enping [Department of Forensic Medicine, School of Basic Medical Science, Southern Medical University, Guangzhou 510515 (China); Liu, Chao [Guangzhou Forensic Science Institute, Guangzhou 510030 (China); Lin, Zhoumeng [Institute of Computational Comparative Medicine and Department of Anatomy and Physiology, College of Veterinary Medicine, Kansas State University, Manhattan, KS 66506 (United States); Xie, Wei-Bing, E-mail: xieweib@126.com [Department of Forensic Medicine, School of Basic Medical Science, Southern Medical University, Guangzhou 510515 (China); Wang, Huijun, E-mail: hjwang711@yahoo.cn [Department of Forensic Medicine, School of Basic Medical Science, Southern Medical University, Guangzhou 510515 (China)

    2016-03-15

    Methamphetamine (METH) is an amphetamine-like psychostimulant that is commonly abused. Previous studies have shown that METH can induce damages to the nervous system and recent studies suggest that METH can also cause adverse and potentially lethal effects on the cardiovascular system. Recently, we demonstrated that DNA damage-inducible transcript 4 (DDIT4) regulates METH-induced neurotoxicity. However, the role of DDIT4 in METH-induced cardiotoxicity remains unknown. We hypothesized that DDIT4 may mediate METH-induced autophagy and apoptosis in cardiomyocytes. To test the hypothesis, we examined DDIT4 protein expression in cardiomyocytes and in heart tissues of rats exposed to METH with Western blotting. We also determined the effects on METH-induced autophagy and apoptosis after silencing DDIT4 expression with synthetic siRNA with or without pretreatment of a mTOR inhibitor rapamycin in cardiomyocytes using Western blot analysis, fluorescence microscopy and TUNEL staining. Our results showed that METH exposure increased DDIT4 expression and decreased phosphorylation of mTOR that was accompanied with increased autophagy and apoptosis both in vitro and in vivo. These effects were normalized after silencing DDIT4. On the other hand, rapamycin promoted METH-induced autophagy and apoptosis in DDIT4 knockdown cardiomyocytes. These results suggest that DDIT4 mediates METH-induced autophagy and apoptosis through mTOR signaling pathway in cardiomyocytes. - Highlights: • METH exposure increases DDIT4 expression in cardiomyocytes. • DDIT4 mediates METH-induced autophagy and apoptosis in cardiomyocytes. • DDIT4 silencing protects cardiomyocytes against METH-caused autophagy and apoptosis.

  13. TonEBP modulates the protective effect of taurine in ischemia-induced cytotoxicity in cardiomyocytes

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    Yang, Y J; Han, Y Y; Chen, K; Zhang, Y; Liu, X; Li, S; Wang, K Q; Ge, J B; Liu, W; Zuo, J

    2015-01-01

    Taurine, which is found at high concentration in the heart, exerts several protective actions on myocardium. Physically, the high level of taurine in heart is maintained by a taurine transporter (TauT), the expression of which is suppressed under ischemic insult. Although taurine supplementation upregulates TauT expression, elevates the intracellular taurine content and ameliorates the ischemic injury of cardiomyocytes (CMs), little is known about the regulatory mechanisms of taurine governing TauT expression under ischemia. In this study, we describe the TonE (tonicity-responsive element)/TonEBP (TonE-binding protein) pathway involved in the taurine-regulated TauT expression in ischemic CMs. Taurine inhibited the ubiquitin-dependent proteasomal degradation of TonEBP, promoted the translocation of TonEBP into the nucleus, enhanced TauT promoter activity and finally upregulated TauT expression in CMs. In addition, we observed that TonEBP had an anti-apoptotic and anti-oxidative role in CMs under ischemia. Moreover, the protective effects of taurine on myocardial ischemia were TonEBP dependent. Collectively, our findings suggest that TonEBP is a core molecule in the protective mechanism of taurine in CMs under ischemic insult. PMID:26673669

  14. Succinate modulates Ca(2+) transient and cardiomyocyte viability through PKA-dependent pathway.

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    Aguiar, Carla J; Andrade, Vanessa L; Gomes, Enéas R M; Alves, Márcia N M; Ladeira, Marina S; Pinheiro, Ana Cristina N; Gomes, Dawidson A; Almeida, Alvair P; Goes, Alfredo M; Resende, Rodrigo R; Guatimosim, Silvia; Leite, M Fatima

    2010-01-01

    GPR91 is an orphan G-protein-coupled receptor (GPCR) that has been characterized as a receptor for succinate, a citric acid cycle intermediate, in several tissues. In the heart, the role of succinate is unknown. We now report that rat ventricular cardiomyocytes express GPR91. We found that succinate, through GPR91, increases the amplitude and the rate of decline of global Ca(2+) transient, by increasing the phosphorylation levels of ryanodine receptor and phospholamban, two well known Ca(2+) handling proteins. The effects of succinate on Ca(2+) transient were abolished by pre-treatment with adenylyl cyclase and cAMP-dependent protein kinase (PKA) inhibitors. Direct PKA activation by succinate was further confirmed using a FRET-based A-kinase activity reporter. Additionally, succinate decreases cardiomyocyte viability through a caspase-3 activation pathway, effect also prevented by PKA inhibition. Taken together, these observations show that succinate acts as a signaling molecule in cardiomyocytes, modulating global Ca(2+) transient and cell viability through a PKA-dependent pathway. 2009 Elsevier Ltd. All rights reserved.

  15. Early Administration of Glutamine Protects Cardiomyocytes from Post-Cardiac Arrest Acidosis

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    Yan-Ren Lin

    2016-01-01

    Full Text Available Postcardiac arrest acidosis can decrease survival. Effective medications without adverse side effects are still not well characterized. We aimed to analyze whether early administration of glutamine could improve survival and protect cardiomyocytes from postcardiac arrest acidosis using animal and cell models. Forty Wistar rats with postcardiac arrest acidosis (blood pH < 7.2 were included. They were divided into study (500 mg/kg L-alanyl-L-glutamine, n=20 and control (normal saline, n=20 groups. Each of the rats received resuscitation. The outcomes were compared between the two groups. In addition, cardiomyocytes derived from human induced pluripotent stem cells were exposed to HBSS with different pH levels (7.3 or 6.5 or to culture medium (control. Apoptosis-related markers and beating function were analyzed. We found that the duration of survival was significantly longer in the study group (p<0.05. In addition, in pH 6.5 or pH 7.3 HBSS buffer, the expression levels of cell stress (p53 and apoptosis (caspase-3, Bcl-xL markers were significantly lower in cardiomyocytes treated with 50 mM L-glutamine than those without L-glutamine (RT-PCR. L-glutamine also increased the beating function of cardiomyocytes, especially at the lower pH level (6.5. More importantly, glutamine decreased cardiomyocyte apoptosis and increased these cells’ beating function at a low pH level.

  16. Adrenaline in pro-oxidant conditions elicits intracellular survival pathways in isolated rat cardiomyocytes

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    Costa, Vera Marisa; Silva, Renata; Ferreira, Rita; Amado, Francisco; Carvalho, Felix; Bastos, Maria Lourdes de; Albuquerque Carvalho, Rui; Carvalho, Marcia; Remiao, Fernando

    2009-01-01

    In several pathologic conditions, like cardiac ischemia/reperfusion, the sustained elevation of plasma and interstitial catecholamine levels, namely adrenaline (ADR), and the generation of reactive oxygen species (ROS) are hallmarks. The present work aimed to investigate in cardiomyocytes which intracellular signalling pathways are altered by ADR redox ability. To mimic pathologic conditions, freshly isolated calcium tolerant cardiomyocytes from adult rat were incubated with ADR alone or in the presence of a system capable of generating ROS [(xanthine with xanthine oxidase) (X/XO)]. ADR elicited a pro-oxidant signal with generation of reactive species, which was largely magnified by the ROS generating system. However, no change in cardiomyocytes viability was observed. The pro-oxidant signal promoted the translocation to the nucleus of the transcription factors, Heat shock factor-1 (HSF-1) and Nuclear factor-κB (NF-κB). In addition, proteasome activity was compromised in the experimental groups where the generation of reactive species occurred. The decrease in the proteasome activity of the ADR group resulted from its redox sensitivity, since the activity was recovered by adding the ROS scavenger, tiron. Proteasome inhibition seemed to elicit an increase in HSP70 levels. Furthermore, retention of mitochondrial cytochrome c and inhibition of caspase 3 activity were observed by X/XO incubation in presence or absence of ADR. In conclusion, in spite of all the insults inflicted to the cardiomyocytes, they were capable to activate intracellular responses that enabled their survival. These mechanisms, namely the pathways altered by catecholamine proteasome inhibition, should be further characterized, as they could be of relevance in the ischemia preconditioning and the reperfusion injury

  17. Protective effects of novel single compound, Hirsutine on hypoxic neonatal rat cardiomyocytes.

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    Wu, Li Xin; Gu, Xian Feng; Zhu, Yi Chun; Zhu, Yi Zhun

    2011-01-10

    Uncaria rhynchophylla is a traditional Chinese herb that has been applied in China for treatment of ailments of the cardiovascular system, but little is known about its active constituents and effect in cardiomyocytes. In present study, we investigated the cardioprotective effect of 0.1μΜ, 1μΜ and 10μΜ Hirsutine isolated from the methanolic extracts of Uncaria rhynchophylla by high performance liquid chromatography (HPLC) on neonatal rat cardiomyocytes treated with hypoxia to determine the mechanism underlying the protective effect with regard to cardiac anti-oxidant enzymes and apoptosis genes. Hirsutine significantly increased the viability of cardiomyocytes injured by hypoxia. Gene expression levels of proapoptotic genes (Bax, Fas and caspase-3) were significantly downregulated compared with the hypoxic control group (P<0.05), whereas the expression level of Bcl-2 was upregulated following Hirsutine treatment (P<0.05). Correspondingly, Hirsutine treatment increased Bcl-2 protein level and decreased Bax protein level. Assay investigating cardiac anti-oxidant enzymes provided further evidence for the protective effect of Hirsutine, as indicated by the induction of the anti-oxidant enzymes superoxide dismutase. The results of present study suggest that the mechanism of action of Hirsutine in hypoxic neonatal rat cardiomyocytes may be related to its anti-oxidant and anti-apoptotic properties. This may open an avenue for developing novel candidate compounds with cardioprotectiveeffect from unique Chinese plant. Copyright © 2010 Elsevier B.V. All rights reserved.

  18. A polysaccharide of Dendrobium officinale ameliorates H2O2-induced apoptosis in H9c2 cardiomyocytes via PI3K/AKT and MAPK pathways.

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    Zhang, Jing-Yi; Guo, Ying; Si, Jin-Ping; Sun, Xiao-Bo; Sun, Gui-Bo; Liu, Jing-Jing

    2017-11-01

    Dendrobium officinale is one valuable traditional Chinese medicine, which has skyscraping medicinal value. Polysaccharide is the main active ingredient in D. officinale; its antioxidant activity is a hot research topic nowadays. Oxidative stress plays an important role in the pathological progress of a variety of cardiovascular disease, as one of key factors of cardiomyocyte apoptosis. This research adopts a model of H 2 O 2 induction-H9c2 cardiomyocytes apoptosis, aiming to study the effect of Dendrobium officinale Polysaccharide (DOP-GY) for cardiomyocyte apoptosis caused by oxidative stress and its possible mechanism. Our results showed that pretreatment of DOP-GY (low dose: 6.25μg/mL, medium dose: 12.5μg/mL, high dose: 25μg/mL) followed by a 2h incubation with 200μM H 2 O 2 elevated the survival rate, cutted the LDH leakage, reduced lipid peroxidation damage, improved the activity of the endogenous antioxidant enzymes. In addition, the pretreatment of DOP-GY significantly inhibited the production of ROS, declined of the mitochondrial membrane potential, down-regulated pro-apoptosis protein and up-regulated anti-apoptosis protein. The protective effect was correlated with the PI3K/Akt and MAPK signal pathway. Collectively, these observations suggest that DOY-GY has the potential to exert cardioprotective effects against H 2 O 2 -induced H9c2 cardiomyocyte apoptosis. Copyright © 2017 Elsevier B.V. All rights reserved.

  19. Propofol attenuates H2O2-induced oxidative stress and apoptosis via the mitochondria- and ER-medicated pathways in neonatal rat cardiomyocytes.

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    Liu, Xue-Ru; Cao, Lu; Li, Tao; Chen, Lin-Lin; Yu, Yi-Yan; Huang, Wen-Jun; Liu, Li; Tan, Xiao-Qiu

    2017-05-01

    Previous studies have shown that propofol, an intravenous anesthetic commonly used in clinical practice, protects the myocardium from injury. Mitochondria- and endoplasmic reticulum (ER)-mediated oxidative stress and apoptosis are two important signaling pathways involved in myocardial injury and protection. The present study aimed to test the hypothesis that propofol could exert a cardio-protective effect via the above two pathways. Cultured neonatal rat cardiomyocytes were treated with culture medium (control group), H 2 O 2 at 500 μM (H 2 O 2 group), propofol at 50 μM (propofol group), and H 2 O 2 plus propofol (H 2 O 2  + propofol group), respectively. The oxidative stress, mitochondrial membrane potential (ΔΨm) and apoptosis of the cardiomyocytes were evaluated by a series of assays including ELISA, flow cytometry, immunofluorescence microscopy and Western blotting. Propofol significantly suppressed the H 2 O 2 -induced elevations in the activities of caspases 3, 8, 9 and 12, the ratio of Bax/Bcl-2, and cell apoptosis. Propofol also inhibited the H 2 O 2 -induced reactive oxygen species (ROS) generation, lactic dehydrogenase (LDH) release and mitochondrial transmembrane potential (ΔΨm) depolarization, and restored the H 2 O 2 -induced reductions of glutathione (GSH) and superoxide dismutase (SOD). In addition, propofol decreased the expressions of glucose-regulated protein 78 kDa (Grp78) and inositol-requiring enzyme 1α (IRE1α), two important signaling molecules in the ER-mediated apoptosis pathway. Propofol protects cardiomyocytes from H 2 O 2 -induced injury by inhibiting the mitochondria- and ER-mediated apoptosis signaling pathways.

  20. N-n-butyl haloperidol iodide protects cardiomyocytes against hypoxia/reoxygenation injury by inhibiting autophagy.

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    Wang, Bin; Zhong, Shuping; Zheng, Fuchun; Zhang, Yanmei; Gao, Fenfei; Chen, Yicun; Lu, Binger; Xu, Han; Shi, Ganggang

    2015-09-22

    N-n-butyl haloperidol iodide (F2), a novel compound derived from haloperidol, protects against the damaging effects of ischemia/reperfusion (I/R) injury in vitro and in vivo. In this study, we hypothesized the myocardial protection of F2 on cardiomyocyte hypoxia/reoxygenation (H/R) injury is mediated by inhibiting autophagy in H9c2 cells. The degree of autophagy by treatment with F2 exposed to H/R in H9c2 cell was characterized by monodansylcadaverine, transmission electron microscopy, and expression of autophagy marker protein LC3. Our results indicated that treatment with F2 inhibited autophagy in H9c2 cells exposed to H/R. 3-methyladenine, an inhibitor of autophagy, suppressed H/R-induced autophagy, and decreased apoptosis, whereas rapamycin, a classical autophagy sensitizer, increased autophagy and apoptosis. Mechanistically, macrophage migration inhibitory factor (MIF) was inhibited by F2 treatment after H/R. Accordingly, small interfering RNA (siRNA)-mediated MIF knockdown decreased H/R-induced autophagy. In summary, F2 protects cardiomyocytes during H/R injury through suppressing autophagy activation. Our results provide a new mechanistic insight into a functional role of F2 against H/R-induced cardiomyocyte injury and death.

  1. Protective effect of pomegranate seed oil against H2O2 -induced oxidative stress in cardiomyocytes

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    Mehdi Bihamta

    2017-01-01

    Full Text Available Objective: It has been well documented that oxidative stress is involved in the pathogenesis of cardiac diseases. Previous studies have shown that pomegranate seed oil (PSO has antioxidant properties. This study was designed to investigate probable protective effects of PSO against hydrogen peroxide (H2O2-induced damage in H9c2 cardiomyocytes.Materials and Methods: The cells were pretreated 24 hr with PSO 1 hr before exposure to 200 µM H2O2. Cell viability was evaluated using 3-(4,5-dimethylthiazol-2-yl-2,5-diphenyl tetrazolium (MTT assay. The level of reactive oxygen species (ROS and lipid peroxidation were measured by fluorimetric methods.Results: H2O2 significantly decreased cell viability which was accompanied by an increase in ROS production and lipid peroxidation and a decline in superoxide dismutase activity. Pretreatment with PSO increased viability of cardiomyocytes and decrease the elevated ROS production and lipid peroxidation. Also, PSO was able to restore superoxide dismutase activity.Conclusion: PSO has protective effect against oxidative stress-induced damage in cardiomyocytes and can be considered as a natural cardioprotective agent to prevent cardiovascular diseases.

  2. Both cardiomyocyte and endothelial cell Nox4 mediate protection against hemodynamic overload-induced remodelling.

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    Zhang, Min; Mongue-Din, Heloise; Martin, Daniel; Catibog, Norman; Smyrnias, Ioannis; Zhang, Xiaohong; Yu, Bin; Wang, Minshu; Brandes, Ralf P; Schröder, Katrin; Shah, Ajay M

    2018-03-01

    NADPH oxidase-4 (Nox4) is an important reactive oxygen species (ROS) source that is upregulated in the haemodynamically overloaded heart. Our previous studies using global Nox4 knockout (Nox4KO) mice demonstrated a protective role of Nox4 during chronic abdominal aortic banding, involving a paracrine enhancement of myocardial capillary density. However, other authors who studied cardiac-specific Nox4KO mice reported detrimental effects of Nox4 in response to transverse aortic constriction (TAC). It has been speculated that these divergent results are due to cell-specific actions of Nox4 (i.e. cardiomyocyte Nox4 detrimental but endothelial Nox4 beneficial) and/or differences in the model of pressure overload (i.e. abdominal banding vs. TAC). This study aimed to (i) investigate whether the effects of Nox4 on pressure overload-induced cardiac remodelling vary according to the pressure overload model and (ii) compare the roles of cardiomyocyte vs. endothelial cell Nox4. Global Nox4KO mice subjected to TAC developed worse cardiac remodelling and contractile dysfunction than wild-type littermates, consistent with our previous results with abdominal aortic banding. Next, we generated inducible cardiomyocyte-specific Nox4 KO mice (Cardio-Nox4KO) and endothelial-specific Nox4 KO mice (Endo-Nox4KO) and studied their responses to pressure overload. Both Cardio-Nox4KO and Endo-Nox4KO developed worse pressure overload-induced cardiac remodelling and dysfunction than wild-type littermates, associated with significant decrease in protein levels of HIF1α and VEGF and impairment of myocardial capillarization. Cardiomyocyte as well as endothelial cell Nox4 contributes to protection against chronic hemodynamic overload-induced cardiac remodelling, at least in part through common effects on myocardial capillary density. © The Author 2017 Published by Oxford University Press on behalf of the European Society of Cardiology.

  3. The protective effect of lycopene on hypoxia/reoxygenation-induced endoplasmic reticulum stress in H9C2 cardiomyocytes.

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    Gao, Yang; Jia, Pengyu; Shu, WenQi; Jia, Dalin

    2016-03-05

    Nowadays, drugs protecting ischemia/reperfusion (I/R) myocardium become more suitable for clinic. It has been confirmed lycopene has various protections, but lacking the observation of its effect on endoplasmic reticulum stress (ERS)-mediated apoptosis caused by hypoxia/reoxygenation (H/R). This study aims to clarify the protective effect of lycopene on ERS induced by H/R in H9C2 cardiomyocytes. Detect the survival rate, lactic dehydrogenase (LDH) activity, apoptosis ratio, glucose-regulated proteins 78 (GRP78), C/EBP homologous protein (CHOP), c-Jun-N-terminal protein Kinase (JNK) and Caspase-12 mRNA and protein expression and phosphorylation of JNK (p-JNK) protein expression. LDH activity, apoptosis ratio and GRP78 protein expression increase in the H/R group, reduced by lycopene. The survival rate reduces in the H/R and thapsigargin (TG) groups; lycopene and 4-phenyl butyric acid (4-PBA) can improve it caused by H/R, lycopene also can improve it caused by TG. The apoptosis ratio, the expression of GRP78, CHOP and Caspase-12 mRNA and protein and p-JNK protein increase in the H/R and TG groups, weaken in the lycopene+H/R, 4-PBA+H/R and lycopene+TG groups. There is no obvious change in the expression of JNK mRNA or protein. Hence, our results provide the evidence that 10 μM lycopene plays an obviously protective effect on H/R H9C2 cardiomyocytes, realized through reducing ERS and apoptosis. The possible mechanism may be related to CHOP, p-JNK and Caspase-12 pathways. Copyright © 2016. Published by Elsevier B.V.

  4. The ESCRT-III pathway facilitates cardiomyocyte release of cBIN1-containing microparticles.

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    Bing Xu

    2017-08-01

    Full Text Available Microparticles (MPs are cell-cell communication vesicles derived from the cell surface plasma membrane, although they are not known to originate from cardiac ventricular muscle. In ventricular cardiomyocytes, the membrane deformation protein cardiac bridging integrator 1 (cBIN1 or BIN1+13+17 creates transverse-tubule (t-tubule membrane microfolds, which facilitate ion channel trafficking and modulate local ionic concentrations. The microfold-generated microdomains continuously reorganize, adapting in response to stress to modulate the calcium signaling apparatus. We explored the possibility that cBIN1-microfolds are externally released from cardiomyocytes. Using electron microscopy imaging with immunogold labeling, we found in mouse plasma that cBIN1 exists in membrane vesicles about 200 nm in size, which is consistent with the size of MPs. In mice with cardiac-specific heterozygous Bin1 deletion, flow cytometry identified 47% less cBIN1-MPs in plasma, supporting cardiac origin. Cardiac release was also evidenced by the detection of cBIN1-MPs in medium bathing a pure population of isolated adult mouse cardiomyocytes. In human plasma, osmotic shock increased cBIN1 detection by enzyme-linked immunosorbent assay (ELISA, and cBIN1 level decreased in humans with heart failure, a condition with reduced cardiac muscle cBIN1, both of which support cBIN1 release in MPs from human hearts. Exploring putative mechanisms of MP release, we found that the membrane fission complex endosomal sorting complexes required for transport (ESCRT-III subunit charged multivesicular body protein 4B (CHMP4B colocalizes and coimmunoprecipitates with cBIN1, an interaction enhanced by actin stabilization. In HeLa cells with cBIN1 overexpression, knockdown of CHMP4B reduced the release of cBIN1-MPs. Using truncation mutants, we identified that the N-terminal BAR (N-BAR domain in cBIN1 is required for CHMP4B binding and MP release. This study links the BAR protein superfamily

  5. The ESCRT-III pathway facilitates cardiomyocyte release of cBIN1-containing microparticles.

    Science.gov (United States)

    Xu, Bing; Fu, Ying; Liu, Yan; Agvanian, Sosse; Wirka, Robert C; Baum, Rachel; Zhou, Kang; Shaw, Robin M; Hong, TingTing

    2017-08-01

    Microparticles (MPs) are cell-cell communication vesicles derived from the cell surface plasma membrane, although they are not known to originate from cardiac ventricular muscle. In ventricular cardiomyocytes, the membrane deformation protein cardiac bridging integrator 1 (cBIN1 or BIN1+13+17) creates transverse-tubule (t-tubule) membrane microfolds, which facilitate ion channel trafficking and modulate local ionic concentrations. The microfold-generated microdomains continuously reorganize, adapting in response to stress to modulate the calcium signaling apparatus. We explored the possibility that cBIN1-microfolds are externally released from cardiomyocytes. Using electron microscopy imaging with immunogold labeling, we found in mouse plasma that cBIN1 exists in membrane vesicles about 200 nm in size, which is consistent with the size of MPs. In mice with cardiac-specific heterozygous Bin1 deletion, flow cytometry identified 47% less cBIN1-MPs in plasma, supporting cardiac origin. Cardiac release was also evidenced by the detection of cBIN1-MPs in medium bathing a pure population of isolated adult mouse cardiomyocytes. In human plasma, osmotic shock increased cBIN1 detection by enzyme-linked immunosorbent assay (ELISA), and cBIN1 level decreased in humans with heart failure, a condition with reduced cardiac muscle cBIN1, both of which support cBIN1 release in MPs from human hearts. Exploring putative mechanisms of MP release, we found that the membrane fission complex endosomal sorting complexes required for transport (ESCRT)-III subunit charged multivesicular body protein 4B (CHMP4B) colocalizes and coimmunoprecipitates with cBIN1, an interaction enhanced by actin stabilization. In HeLa cells with cBIN1 overexpression, knockdown of CHMP4B reduced the release of cBIN1-MPs. Using truncation mutants, we identified that the N-terminal BAR (N-BAR) domain in cBIN1 is required for CHMP4B binding and MP release. This study links the BAR protein superfamily to the ESCRT

  6. Thymosin beta 4 protects cardiomyocytes from oxidative stress by targeting anti-oxidative enzymes and anti-apoptotic genes.

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    Chuanyu Wei

    Full Text Available Thymosin beta-4 (Tβ4 is a ubiquitous protein with many properties relating to cell proliferation and differentiation that promotes wound healing and modulates inflammatory mediators. The mechanism by which Tβ4 modulates cardiac protection under oxidative stress is not known. The purpose of this study is to dissect the cardioprotective mechanism of Tβ4 on H(2O(2 induced cardiac damage.Rat neonatal cardiomyocytes with or without Tβ4 pretreatment were exposed to H(2O(2 and expression of antioxidant, apoptotic, and anti-inflammatory genes was evaluated by quantitative real-time PCR and western blotting. ROS levels were estimated by DCF-DA using fluorescent microscopy and fluorimetry. Selected antioxidant, anti-inflammatory and antiapoptotic genes were silenced by siRNA transfections in neonatal cardiomyocytes and effect of Tβ4 on H(2O(2-induced cardiac damage was evaluated.Pre-treatment of Tβ4 resulted in reduction of the intracellular ROS levels induced by H(2O(2 in cardiomyocytes. Tβ4 pretreatment also resulted in an increase in the expression of antiapoptotic proteins and reduction of Bax/BCl(2 ratio in the cardiomyocytes. Pretreatment with Tβ4 resulted in stimulating the expression of antioxidant enzymes copper/zinc SOD and catalase in cardiomyocytes at both transcription and translation levels. Tβ4 treatment resulted in the increased expression of anti-apoptotic and anti-inflammatory genes. Silencing of Cu/Zn SOD and catalase gene resulted in apoptotic cell death in the cardiomyocytes which was prevented by treatment with Tβ4.This is the first report that demonstrates the effect of Tβ4 on cardiomyocytes and its capability to selectively upregulate anti-oxidative enzymes, anti-inflammatory genes, and antiapoptotic enzymes in the neonatal cardiomyocytes thus preventing cell death thereby protecting the myocardium. Tβ4 treatment resulted in decreased oxidative stress and inflammation in the myocardium under oxidative stress.

  7. Ursodeoxycholic acid protects cardiomyocytes against cobalt chloride induced hypoxia by regulating transcriptional mediator of cells stress hypoxia inducible factor 1α and p53 protein.

    Science.gov (United States)

    Mohamed, Anis Syamimi; Hanafi, Noorul Izzati; Sheikh Abdul Kadir, Siti Hamimah; Md Noor, Julina; Abdul Hamid Hasani, Narimah; Ab Rahim, Sharaniza; Siran, Rosfaiizah

    2017-10-01

    In hepatocytes, ursodeoxycholic acid (UDCA) activates cell signalling pathways such as p53, intracellular calcium ([Ca 2+ ] i ), and sphingosine-1-phosphate (S1P)-receptor via Gα i -coupled-receptor. Recently, UDCA has been shown to protect the heart against hypoxia-reoxygenation injury. However, it is not clear whether UDCA cardioprotection against hypoxia acts through a transcriptional mediator of cells stress, HIF-1α and p53. Therefore, in here, we aimed to investigate whether UDCA could protect cardiomyocytes (CMs) against hypoxia by regulating expression of HIF-1α, p53, [Ca 2+ ] i , and S1P-Gα i -coupled-receptor. Cardiomyocytes were isolated from newborn rats (0-2 days), and hypoxia was induced by using cobalt chloride (CoCl 2 ). Cardiomyocytes were treated with UDCA and cotreated with either FTY720 (S1P-receptor agonist) or pertussis toxin (PTX; Gα i inhibitor). Cells were subjected for proliferation assay, beating frequency, QuantiGene Plex assay, western blot, immunofluorescence, and calcium imaging. Our findings showed that UDCA counteracted the effects of CoCl 2 on cell viability, beating frequency, HIF-1α, and p53 protein expression. We found that these cardioprotection effects of UDCA were similar to FTY720, S1P agonist. Furthermore, we observed that UDCA protects CMs against CoCl 2 -induced [Ca 2+ ] i dynamic alteration. Pharmacological inhibition of the Gα i -sensitive receptor did not abolish the cardioprotection of UDCA against CoCl 2 detrimental effects, except for cell viability and [Ca 2+ ] i . Pertussis toxin is partially effective in inhibiting UDCA protection against CoCl 2 effects on CM cell viability. Interestingly, PTX fully inhibits UDCA cardioprotection on CoCl 2 -induced [Ca 2+ ] i dynamic changes. We conclude that UDCA cardioprotection against CoCl 2 -induced hypoxia is similar to FTY720, and its actions are not fully mediated by the Gα i -coupled protein sensitive pathways. Ursodeoxycholic acid is the most hydrophilic bile

  8. SIRT1 Functions as an Important Regulator of Estrogen-Mediated Cardiomyocyte Protection in Angiotensin II-Induced Heart Hypertrophy

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    Tao Shen

    2014-01-01

    Full Text Available Background. Sirtuin 1 (SIRT1 is a member of the sirtuin family, which could activate cell survival machinery and has been shown to be protective in regulation of heart function. Here, we determined the mechanism by which SIRT1 regulates Angiotensin II- (AngII- induced cardiac hypertrophy and injury in vivo and in vitro. Methods. We analyzed SIRT1 expression in the hearts of control and AngII-induced mouse hypertrophy. Female C57BL/6 mice were ovariectomized and pretreated with 17β-estradiol to measure SIRT1 expression. Protein synthesis, cardiomyocyte surface area analysis, qRT-PCR, TUNEL staining, and Western blot were performed on AngII-induced mouse heart hypertrophy samples and cultured neonatal rat ventricular myocytes (NRVMs to investigate the function of SIRT1. Results. SIRT1 expression was slightly upregulated in AngII-induced mouse heart hypertrophy in vivo and in vitro, accompanied by elevated cardiomyocyte apoptosis. SIRT1 overexpression relieves AngII-induced cardiomyocyte hypertrophy and apoptosis. 17β-Estradiol was able to protect cardiomyocytes from AngII-induced injury with a profound upregulation of SIRT1 and activation of AMPK. Moreover, estrogen receptor inhibitor ICI 182,780 and SIRT1 inhibitor niacinamide could block SIRT1’s protective effect. Conclusions. These results indicate that SIRT1 functions as an important regulator of estrogen-mediated cardiomyocyte protection during AngII-induced heart hypertrophy and injury.

  9. β3-adrenergic receptor activation induces TGFβ1 expression in cardiomyocytes via the PKG/JNK/c-Jun pathway.

    Science.gov (United States)

    Xu, Zhongcheng; Wu, Jimin; Xin, Junzhou; Feng, Yenan; Hu, Guomin; Shen, Jing; Li, Mingzhe; Zhang, Youyi; Xiao, Han; Wang, Li

    2018-06-05

    In heart failure, the expression of cardiac β 3 -adrenergic receptors (β 3 -ARs) increases. However, the precise role of β 3 -AR signaling within cardiomyocytes remains unclear. Transforming growth factor β1 (TGFβ1) is a crucial cytokine mediating the cardiac remodeling that plays a causal role in the progression of heart failure. Here, we set out to determine the effect of β 3 -AR activation on TGFβ1 expression in rat cardiomyocytes and examine the underlying mechanism. The selective β 3 -AR agonist BRL37344 induced an increase in TGFβ1 expression and the phosphorylation of c-Jun N-terminal kinase (JNK) and c-Jun in β 3 -AR-overexpressing cardiomyocytes. Those effects of BRL37344 were suppressed by a β 3 -AR antagonist. Moreover, the inhibition of JNK and c-Jun activity by a JNK inhibitor and c-Jun siRNA blocked the increase in TGFβ1 expression upon β 3 -AR activation. A protein kinase G (PKG) inhibitor also attenuated β 3 -AR-agonist-induced TGFβ1 expression and the phosphorylation of JNK and c-Jun. In conclusion, the β 3 -AR activation in cardiomyocytes increases the expression of TGFβ1 via the PKG/JNK/c-Jun pathway. These results help us further understand the role of β 3 -AR signaling in heart failure. Copyright © 2018 Elsevier Inc. All rights reserved.

  10. [Role and related mechanism of S1P/S1P1 signal pathway during post conditioning of hypertrophic cardiomyocytes].

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    Bao, X H; Li, H X; Tao, J; Li, X M; Yang, Y N; Ma, Y T; Chen, B D

    2016-05-24

    To study the role and mechanism of sphingosine-1-phosphate (S1P)/ sphingosine-1-phosphate receptor 1(S1P1) signal pathway during post conditioning of hypertrophic cardiomyocytes. Neonatal rat cardiomyocytes were isolated and cultured, then stimulated by norepinephrine (NE) to induce cardiomyocytes hypertrophy. Using tri-gas incubator to create hypoxia and reoxygenation enviroment to mimic ischemia-reperfusion and postconditioning. Hypertrophic cardiomyoctyes were divided into five groups according to the presence or absence of various drugs and postconditiong and relevant signal pathways changes were detected: (1) IPost group (hypoxia+ postconditioning); (2) IPost+ S1P group (cells were pretreated with S1P (1 μmol/L) for 2 h before IPost); (3) IPost+ W-146+ S1P group (cells in IPost+ W-146+ S1P group were pretreated with S1P1 inhibitor W-146 (0.4 μmol/L) for 20 min); (4) IPost+ PD98059+ S1P group (cells in IPost+ S1P group were pretreated with MAPK antagonist PD98059 (125 μmol/L) for 20 min); (5) IPost+ LY-294002+ S1P group (cells in IPost+ S1P group were pretreated with PI3K antagonist LY294002 (0.1 μmol/L) for 20 min). Apoptosis was detected by flow cytometry and protein expression of relevant signal pathways were detected by Western blot. (1)Apoptosis rate was significantly increased in hypoxia/reoxygenation (27.90±4.49)% group compared with normal control group (7.97±2.18)%, which could be significantly reduced in IPost group (15.90±1.77)% (all PS1P and IPost+ S1P+ LY-294002 groups than in IPost and IPost+ S1P+ W-146 and IPost+ S1P+ PD98059 group (all PS1P and IPost+ S1P+ LY-294002 group than in IPost and IPost+ S1P+ W-146 group and IPost+ S1P+ PD98059 group (all PS1P+ W-146 and IPost+ S1P+ PD98059 groups. p-ERK1/2 and p-Akt levels in IPost+ S1P+ W-146 group and IPost+ S1P+ PD98059 were similar as in IPost group. S1P can play protective role on NE induced cardiomyocytes hypertrophy during post conditioning through downregulating caspase-3 expression and

  11. Inhibition of MMP-2 Expression with siRNA Increases Baseline Cardiomyocyte Contractility and Protects against Simulated Ischemic Reperfusion Injury

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    Han-Bin Lin

    2014-01-01

    Full Text Available Matrix metalloproteinases (MMPs significantly contribute to ischemia reperfusion (I/R injury, namely, by the degradation of contractile proteins. However, due to the experimental models adopted and lack of isoform specificity of MMP inhibitors, the cellular source and identity of the MMP(s involved in I/R injury remain to be elucidated. Using isolated adult rat cardiomyocytes, subjected to chemically induced I/R-like injury, we show that specific inhibition of MMP-2 expression and activity using MMP-2 siRNA significantly protected cardiomyocyte contractility from I/R-like injury. This was also associated with increased expression of myosin light chains 1 and 2 (MLC1/2 in comparison to scramble siRNA transfection. Moreover, the positive effect of MMP-2 siRNA transfection on cardiomyocyte contractility and MLC1/2 expression levels was also observed under control conditions, suggesting an important additional role for MMP-2 in physiological sarcomeric protein turnover. This study clearly demonstrates that intracellular expression of MMP-2 plays a significant role in sarcomeric protein turnover, such as MLC1 and MLC2, under aerobic (physiological conditions. In addition, this study identifies intracellular/autocrine, cardiomyocyte-produced MMP-2, rather than paracrine/extracellular, as responsible for the degradation of MLC1/2 and consequent contractile dysfunction in cardiomyocytes subjected to I/R injury.

  12. Cdk1, PKCδ and calcineurin-mediated Drp1 pathway contributes to mitochondrial fission-induced cardiomyocyte death

    International Nuclear Information System (INIS)

    Zaja, Ivan; Bai, Xiaowen; Liu, Yanan; Kikuchi, Chika; Dosenovic, Svjetlana; Yan, Yasheng; Canfield, Scott G.; Bosnjak, Zeljko J.

    2014-01-01

    Highlights: • Drp1-mediated increased mitochondrial fission but not fusion is involved the cardiomyocyte death during anoxia-reoxygenation injury. • Reactive oxygen species are upstream initiators of mitochondrial fission. • Increased mitochondrial fission is resulted from Cdk1-, PKCδ-, and calcineurin-mediated Drp1 pathways. - Abstract: Myocardial ischemia–reperfusion (I/R) injury is one of the leading causes of death and disability worldwide. Mitochondrial fission has been shown to be involved in cardiomyocyte death. However, molecular machinery involved in mitochondrial fission during I/R injury has not yet been completely understood. In this study we aimed to investigate molecular mechanisms of controlling activation of dynamin-related protein 1 (Drp1, a key protein in mitochondrial fission) during anoxia-reoxygenation (A/R) injury of HL1 cardiomyocytes. A/R injury induced cardiomyocyte death accompanied by the increases of mitochondrial fission, reactive oxygen species (ROS) production and activated Drp1 (pSer616 Drp1), and decrease of inactivated Drp1 (pSer637 Drp1) while mitochondrial fusion protein levels were not significantly changed. Blocking Drp1 activity with mitochondrial division inhibitor mdivi1 attenuated cell death, mitochondrial fission, and Drp1 activation after A/R. Trolox, a ROS scavenger, decreased pSer616 Drp1 level and mitochondrial fission after A/R. Immunoprecipitation assay further indicates that cyclin dependent kinase 1 (Cdk1) and protein kinase C isoform delta (PKCδ) bind Drp1, thus increasing mitochondrial fission. Inhibiting Cdk1 and PKCδ attenuated the increases in pSer616 Drp1, mitochondrial fission, and cardiomyocyte death. FK506, a calcineurin inhibitor, blocked the decrease in expression of inactivated pSer637 Drp1 and mitochondrial fission. Our findings reveal the following novel molecular mechanisms controlling mitochondrial fission during A/R injury of cardiomyocytes: (1) ROS are upstream initiators of

  13. Cdk1, PKCδ and calcineurin-mediated Drp1 pathway contributes to mitochondrial fission-induced cardiomyocyte death

    Energy Technology Data Exchange (ETDEWEB)

    Zaja, Ivan [Department of Anesthesiology, Medical College of Wisconsin, Milwaukee, WI 53226 (United States); Bai, Xiaowen, E-mail: xibai@mcw.edu [Department of Anesthesiology, Medical College of Wisconsin, Milwaukee, WI 53226 (United States); Liu, Yanan; Kikuchi, Chika; Dosenovic, Svjetlana; Yan, Yasheng; Canfield, Scott G. [Department of Anesthesiology, Medical College of Wisconsin, Milwaukee, WI 53226 (United States); Bosnjak, Zeljko J. [Department of Anesthesiology, Medical College of Wisconsin, Milwaukee, WI 53226 (United States); Department of Physiology, Medical College of Wisconsin, Milwaukee, WI 53226 (United States)

    2014-10-31

    Highlights: • Drp1-mediated increased mitochondrial fission but not fusion is involved the cardiomyocyte death during anoxia-reoxygenation injury. • Reactive oxygen species are upstream initiators of mitochondrial fission. • Increased mitochondrial fission is resulted from Cdk1-, PKCδ-, and calcineurin-mediated Drp1 pathways. - Abstract: Myocardial ischemia–reperfusion (I/R) injury is one of the leading causes of death and disability worldwide. Mitochondrial fission has been shown to be involved in cardiomyocyte death. However, molecular machinery involved in mitochondrial fission during I/R injury has not yet been completely understood. In this study we aimed to investigate molecular mechanisms of controlling activation of dynamin-related protein 1 (Drp1, a key protein in mitochondrial fission) during anoxia-reoxygenation (A/R) injury of HL1 cardiomyocytes. A/R injury induced cardiomyocyte death accompanied by the increases of mitochondrial fission, reactive oxygen species (ROS) production and activated Drp1 (pSer616 Drp1), and decrease of inactivated Drp1 (pSer637 Drp1) while mitochondrial fusion protein levels were not significantly changed. Blocking Drp1 activity with mitochondrial division inhibitor mdivi1 attenuated cell death, mitochondrial fission, and Drp1 activation after A/R. Trolox, a ROS scavenger, decreased pSer616 Drp1 level and mitochondrial fission after A/R. Immunoprecipitation assay further indicates that cyclin dependent kinase 1 (Cdk1) and protein kinase C isoform delta (PKCδ) bind Drp1, thus increasing mitochondrial fission. Inhibiting Cdk1 and PKCδ attenuated the increases in pSer616 Drp1, mitochondrial fission, and cardiomyocyte death. FK506, a calcineurin inhibitor, blocked the decrease in expression of inactivated pSer637 Drp1 and mitochondrial fission. Our findings reveal the following novel molecular mechanisms controlling mitochondrial fission during A/R injury of cardiomyocytes: (1) ROS are upstream initiators of

  14. The Chinese Herb Yi-Qi-Huo-Xue Protects Cardiomyocyte Function in Diabetic Cardiomyopathy

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    Xiangsheng Wang

    2018-01-01

    Full Text Available Aims. To study the effect of the Chinese herb Yi-qi-huo-xue on cardiomyopathy in diabetic rats. Methods. Rats were fed a high fat and high glucose diet and injected with 50 ml/kg streptozotocin (STZ to induce diabetic cardiomyopathy (DCM, followed by treatment with Yi-qi-huo-xue for 4 weeks. We measured the rats’ heart weight index, observed the myocardial morphology using hematoxylin eosin (HE staining, and determined the content of collagen types I and III in the myocardium using enzyme-linked immunosorbent assay (ELISA. We determined Bcl-2, Bax, and P53 protein expression by Western blot analysis and the cardiomyocyte apoptosis rate via a flow cytometry assay. Results. Compared with the rats in the control group, the diabetic rats gained weight and had increased blood sugar levels, an enhanced heart weight index, and increased myocardial pathophysiological damage. There was a decrease in their Bcl-2 expression, and their Bax and P53 expression increased. The Bcl-2/Bax ratio was enhanced, and there was an increase in the content of collagen types I and III in the myocardium. After treatment with Yi-qi-huo-xue, all levels listed above returned to normal. Conclusion. The Chinese herb Yi-qi-huo-xue degraded the myocardial interstitial collagen types I and III to protect the myocardium of the diabetic rats, thus delaying the role of myocardial fibrosis. Yi-qi-huo-xue could play an important role in protecting the myocardium of DCM rats by enhancing the expression of the Bcl-2 protein, inhibiting the expression of the Bax and P53 proteins, increasing the ratio of Bcl-2/Bax, and inhibiting the apoptosis of cardiomyocytes.

  15. Anti-apoptotic effect of heat shock protein 90 on hypoxia-mediated cardiomyocyte damage is mediated via the phosphatidylinositol 3-kinase/AKT pathway.

    Science.gov (United States)

    Wang, Wei; Peng, Yizhi; Wang, Yuanyuan; Zhao, Xiaohui; Yuan, Zhiqiang

    2009-09-01

    1. Hypoxia-induced cardiomyocyte apoptosis contributes significantly to cardiac dysfunction following trauma, shock and burn injury. There is evidence that heat shock protein (HSP) 90 is anti-apoptotic in cardiomyocytes subjected to a variety of apoptotic stimuli. Because HSP90 acts as an upstream regulator of the serine/threonine protein kinase Akt survival pathway during cellular stress, we hypothesized that HSP90 exerts a cardioprotective effect via the phosphatidylinositol 3-kinase (PI3-K)/Akt pathway. 2. Neonatal rat cardiomyocytes were subjected to normoxia or hypoxia in the absence or presence of the HSP90 inhibitor geldanamycin (1 μg/mL). Cardiomyocyte apoptosis was assessed by release of lactate dehydrogenase (LDH), terminal deoxyribonucleotidyl transferase-mediated dUTP-digoxigenin nick end-labelling (TUNEL) staining and caspase 3 activity. Expression of HSP90, Akt, Bad and cytochrome c release was determined by western blot analysis. 3. Following exposure of cells to hypoxia, HSP90 was markedly elevated in a time-dependent manner, reaching a peak at 6 h (eightfold increase). Geldanamycin significantly increased hypoxia-induced release of LDH by 114%, the percentage of apoptotic cardiomyocytes by 102% and caspase 3 activity by 78%. Pretreatment of cells with geldanamycin also suppressed phosphorylation of both Akt and its downstream target Bad, but promoted the mitochondrial release of cytochrome c. 4. In conclusion, HSP90 activity is enhanced in cardiomyocytes following hypoxic insult. The anti-apoptotic effect of HSP90 on cardiomyocytes subjected to hypoxia is mediated, at least in part, by the PI3-K/Akt pathway. Key words: apoptosis, cardiomyocyte, heart failure, heat shock protein 90, hypoxia, phosphatidylinositol 3-kinase/Akt signalling pathway, serine/threonine protein kinase Akt.

  16. Role of S100A1 in hypoxia-induced inflammatory response in cardiomyocytes via TLR4/ROS/NF-κB pathway.

    Science.gov (United States)

    Yu, Jiangkun; Lu, Yanyu; Li, Yapeng; Xiao, Lili; Xing, Yu; Li, Yanshen; Wu, Leiming

    2015-09-01

    S100A1 plays a crucial role in hypoxia-induced inflammatory response in cardiomyocytes. However, the role of S100A1 in hypoxia-induced inflammatory response in cardiomyocytes is still unknown. enzyme-linked immunosorbent assay (ELISA) was performed for the determination of inflammatory cytokines. Immunocytochemistry and immunofluorescence, Western blot analysis and Real-time polymerase chain reaction (RT-PCR) were conducted to assess protein or mRNA expressions. Fluorogenic probe dihydroethidium (DHE) was used to evaluate the generation of reactive oxygen species (ROS) while Hoechst 33342 staining for apoptosis. Small interfering RNA (siRNA) for S100A1 was used to evaluate the role of S100A1. The levels of ROS and inflammatory cytokine including tumor necrosis factor (TNF)-α, interleukin (IL)-1β, IL-6 and IL-8 in H9c2 cells were increased remarkably by hypoxia. However, IL-37 protein or mRNA levels were decreased significantly. Both Toll-like receptor 4 (TLR4) inhibitor Ethyl (6R)-6-[N-(2-Chloro-4fluorophenyl)sulfamoyl]cyclohex-1-ene-1-carboxylate (TAK-242) treatment or siRNA S100A1 downregulated TLR4 expression and inflammatory cytokine level and mRNA in H9c2 cells, as well as weakening ROS and phospho-p65 Nuclear factor (NF)-κB levels. Further, S100A1 treatment significantly reduced TNF-α protein or mRNA level whereas enhanced IL-37 protein or mRNA level, and could attenuate ROS and phospho-p65 NF-κB levels. Our results demonstrate that S100A1 can regulate the inflammatory response and oxidative stress in H9C2 cells via TLR4/ROS/NF-κB pathway. These findings provide an interesting strategy for protecting cardiomyocytes from hypoxia-induced inflammatory response. © 2015 Royal Pharmaceutical Society.

  17. Impact of miR-208 and its Target Gene Nemo-Like Kinase on the Protective Effect of Ginsenoside Rb1 in Hypoxia/Ischemia Injuried Cardiomyocytes

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    Xu Yan

    2016-09-01

    Full Text Available Background/Aims: Ginsenoside Rb1 (GS-Rb1 is one of the most important active pharmacological extracts of the Traditional Chinese Medicine ginseng, with extensive evidence of its cardioprotective properties. Mir-208 has been shown to act as a biomarker of acute myocardial infarction in vivo studies including man. However the impact of miR-208 on the protective effect of GS-Rb1 in hypoxia/ischemia injured cardiomyocytes remains unclear. The current study aims to investigate the target gene of miR-208 and the impact on the protective effect of GS-Rb1 in hypoxia/ischemia (H/I injuried cardiomyocytes. Materials and Methods: Primary cultures of neonatal rat cardiomyocytes (NRCMs was subjected to the H/I conditions with or without GS-Rb1. Cell viability was calculated by MTT assay and confirmed by flow cytometry analysis. Mir-208 was then detected by qRT-PCR. Luciferase reporter assay was carried out to detect the target gene of Mir-208. Then the NRCMs were transfected with miR-208 mimics and inhibitors to evaluate the impact on cardioprotective properties of Rb1. Results: The miR-208 expression level was clearly upregulated in the H/I treated NRCMs accompanied by the percentage of the apoptotic cells which could be reversed by GS-Rb1 pretreatment. The nemo-like kinase (NLK mRNA and protein expression levels were decreased in H/I group measured by RT-PCR and western blotting. Luciferase activity assay was then carried out to identify that NLK may be a direct target of mir-208. MTT assay showed that miR-208 inhibitor slightly decreased the protective effect of Rb1 on the H/I impaired NRCMs. However, results showed no statistical difference. Conclusions: These findings proved that NLK was a direct target of mir-208 and miR-208 act indirectly during Rb1 protecting H/I impaired NRCMs and further researches were needed to explore the relationship that microRNAs and other signal pathways in the protective effect of GS-Rb1 on the hypoxia/ischemia injuries in

  18. Lycopene Protects against Hypoxia/Reoxygenation Injury by Alleviating ER Stress Induced Apoptosis in Neonatal Mouse Cardiomyocytes

    Science.gov (United States)

    Xu, Jiqian; Hu, Houxiang; Chen, Bin; Yue, Rongchuan; Zhou, Zhou; Liu, Yin; Zhang, Shuang; Xu, Lei; Wang, Huan; Yu, Zhengping

    2015-01-01

    Endoplasmic reticulum (ER) stress induced apoptosis plays a pivotal role in myocardial ischemia/reperfusion (I/R)-injury. Inhibiting ER stress is a major therapeutic target/strategy in treating cardiovascular diseases. Our previous studies revealed that lycopene exhibits great pharmacological potential in protecting against the I/R-injury in vitro and vivo, but whether attenuation of ER stress (and) or ER stress-induced apoptosis contributes to the effects remains unclear. In the present study, using neonatal mouse cardiomyocytes to establish an in vitro model of hypoxia/reoxygenation (H/R) to mimic myocardium I/R in vivo, we aimed to explore the hypothesis that lycopene could alleviate the ER stress and ER stress-induced apoptosis in H/R-injury. We observed that lycopene alleviated the H/R injury as revealed by improving cell viability and reducing apoptosis, suppressed reactive oxygen species (ROS) generation and improved the phosphorylated AMPK expression, attenuated ER stress as evidenced by decreasing the expression of GRP78, ATF6 mRNA, sXbp-1 mRNA, eIF2α mRNA and eIF2α phosphorylation, alleviated ER stress-induced apoptosis as manifested by reducing CHOP/GADD153 expression, the ratio of Bax/Bcl-2, caspase-12 and caspase-3 activity in H/R-treated cardiomyocytes. Thapsigargin (TG) is a potent ER stress inducer and used to elicit ER stress of cardiomyocytes. Our results showed that lycopene was able to prevent TG-induced ER stress as reflected by attenuating the protein expression of GRP78 and CHOP/GADD153 compared to TG group, significantly improve TG-caused a loss of cell viability and decrease apoptosis in TG-treated cardiomyocytes. These results suggest that the protective effects of lycopene on H/R-injury are, at least in part, through alleviating ER stress and ER stress-induced apoptosis in neonatal mouse cardiomyocytes. PMID:26291709

  19. A novel miR-371a-5p-mediated pathway, leading to BAG3 upregulation in cardiomyocytes in response to epinephrine, is lost in Takotsubo cardiomyopathy.

    Science.gov (United States)

    d'Avenia, M; Citro, R; De Marco, M; Veronese, A; Rosati, A; Visone, R; Leptidis, S; Philippen, L; Vitale, G; Cavallo, A; Silverio, A; Prota, C; Gravina, P; De Cola, A; Carletti, E; Coppola, G; Gallo, S; Provenza, G; Bossone, E; Piscione, F; Hahne, M; De Windt, L J; Turco, M C; De Laurenzi, V

    2015-10-29

    Molecular mechanisms protecting cardiomyocytes from stress-induced death, including tension stress, are essential for cardiac physiology and defects in these protective mechanisms can result in pathological alterations. Bcl2-associated athanogene 3 (BAG3) is expressed in cardiomyocytes and is a component of the chaperone-assisted autophagy pathway, essential for homeostasis of mechanically altered cells. BAG3 ablation in mice results in a lethal cardiomyopathy soon after birth and mutations of this gene have been associated with different cardiomyopathies including stress-induced Takotsubo cardiomyopathy (TTC). The pathogenic mechanism leading to TTC has not been defined, but it has been suggested that the heart can be damaged by excessive epinephrine (epi) spillover in the absence of a protective mechanism. The aim of this study was to provide more evidence for a role of BAG3 in the pathogenesis of TTC. Therefore, we sequenced BAG3 gene in 70 TTC patients and in 81 healthy donors with the absence of evaluable cardiovascular disease. Mutations and polymorphisms detected in the BAG3 gene included a frequent nucleotide change g2252c in the BAG3 3'-untranslated region (3'-UTR) of Takotsubo patients (PBAG3 upregulation on exposure to epi through an ERK-dependent upregulation of miR-371a-5p. In conclusion, the presence of a g2252c polymorphism in the BAG3 3'-UTR determines loss of miR-371a-5p binding and results in an altered response to epi, potentially representing a new molecular mechanism that contributes to TTC pathogenesis.

  20. Doxazosin stimulates galectin-3 expression and collagen synthesis in HL-1 cardiomyocytes independent of protein kinase C pathway

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    Xiaoqian Qian

    2016-12-01

    Full Text Available Doxazosin, a drug commonly prescribed for hypertension and prostate disease, increases heart failure risk. However, the underlying mechanism remains unclear. Galectin-3 is an important mediator that plays a pathogenic role in cardiac hypertrophy and heart failure. In the present study, we investigated whether doxazosin could stimulate galectin-3 expression and collagen synthesis in cultured HL-1 cardiomyocytes. We found that doxazosin dose-dependently induced galectin-3 protein expression, with a statistically significant increase in expression with a dose as low as 0.01 μM. Doxazosin upregulated collagen I and α-smooth muscle actin (α-SMA protein levels and also induced apoptotic protein caspase-3 in HL-1 cardiomyocytes. Although we previously reported that activation of protein kinase C (PKC stimulates galectin-3 expression, blocking the PKC pathway with the PKC inhibitor chelerythrine did not prevent doxazosin-induced galectin-3 and collagen expression. Consistently, doxazosin treatment did not alter total and phosphorylated PKC. These results suggest that doxazosin-stimulated galectin-3 is independent of PKC pathway. To determine if the α1-adrenergic pathway is involved, we pretreated the cells with the irreversible α-adrenergic receptor blocker phenoxybenzamine and found that doxazosin-stimulated galectin-3 and collagen expression was similar to controls, suggesting that doxazosin acts independently of α1-adrenergic receptor blockade. Collectively, we show a novel effect of doxazosin on cardiomycytes by stimulating heart fibrosis factor galectin-3 expression. The mechanism of action of doxazosin is not mediated through either activation of the PKC pathway or antagonism of α1-adrenergic receptors.

  1. Intracellular calcium overloading and oxidative stress in cardiomyocyte necrosis via a mitochondriocentric signal-transducer-effector pathway

    Science.gov (United States)

    Shaheen, Mazen; Cheema, Yaser; Shahbaz, Atta U; Bhattacharya, Syamal K; Weber, Karl T

    2011-01-01

    Congestive heart failure (CHF), a common clinical syndrome, has reached epidemic proportions. Its disabling symptoms account for frequent hospitalizations and readmissions. Pathophysiological mechanisms that lead to CHF and account for its progressive nature are of considerable interest. Important scientific observations obtained from Dr Pawan K Singal’s laboratory in Winnipeg, Manitoba, have provided crucial insights to our understanding of the pathophysiological factors that contribute to cardiomyocyte necrosis (the heart is a postmitotic organ incapable of tolerating an ongoing loss of these cells without adverse functional consequences). This increment in knowledge and the mechanistic insights afforded by Dr Singal and his colleagues have highlighted the role of excessive intracellular calcium accumulation and the appearance of oxidative stress in CHF, in which the rate of reactive oxygen species generation overwhelms their rate of detoxification by antioxidant defenses. They have shown that this common pathophysiological scenario applies to diverse entities such as ischemia/reperfusion and hypoxia/reoxygenation forms of injury, myocardial infarction and the cardiomyopathies that accompany diabetes and excess levels of catecholamines and adriamycin. The authors are honoured to be invited to contribute to the present focus issue of Experimental & Clinical Cardiology in recognizing Dr Singal’s numerous scholarly accomplishments. The present article reviews the authors’ recent work on a mitochondriocentric signal-transducer-effector pathway to cardiomyocyte necrosis found in rats with either an acute stressor state that accompanies isoproterenol administration or a chronic stressor state manifested after four weeks of aldosterone/salt treatment. PMID:22131852

  2. Use of Human Induced Pluripotent Stem Cell-Derived Cardiomyocytes (hiPSC-CMs) to Monitor Compound Effects on Cardiac Myocyte Signaling Pathways.

    Science.gov (United States)

    Guo, Liang; Eldridge, Sandy; Furniss, Mike; Mussio, Jodie; Davis, Myrtle

    2015-09-01

    There is a need to develop mechanism-based assays to better inform risk of cardiotoxicity. Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) are rapidly gaining acceptance as a biologically relevant in vitro model for use in drug discovery and cardiotoxicity screens. Utilization of hiPSC-CMs for mechanistic investigations would benefit from confirmation of the expression and activity of cellular pathways that are known to regulate cardiac myocyte viability and function. This unit describes an approach to demonstrate the presence and function of signaling pathways in hiPSC-CMs and the effects of treatments on these pathways. We present a workflow that employs protocols to demonstrate protein expression and functional integrity of signaling pathway(s) of interest and to characterize biological consequences of signaling modulation. These protocols utilize a unique combination of structural, functional, and biochemical endpoints to interrogate compound effects on cardiomyocytes. Copyright © 2015 John Wiley & Sons, Inc.

  3. Cardioprotective effect of breviscapine: inhibition of apoptosis in H9c2 cardiomyocytes via the PI3K/Akt/eNOS pathway following simulated ischemia/reperfusion injury.

    Science.gov (United States)

    Wang, Jun; Ji, Shu-Yun; Liu, Si-Zhu; Jing, Rui; Lou, Wei-Juan

    2015-09-01

    Breviscapine (BE) is a standardized Chinese herbal medicine extracted from Erigeron breviscapus (Vant.) Hand.-Mazz. It has been widely used to treat cardiovascular and cerebrovascular diseases. However, there are no reports on the protective effects and underlying molecular mechanisms of BE action on myocardial ischemia/reperfusion (MI/R)-induced cardiomyocyte apoptosis. In the present study, we aimed to confirm the cardioprotective effect of BE from MI/R injury in vivo, and investigate the potential molecular mechanisms against simulated ischemia/reperfusion (SI/R)-induced cardiomyocyte apoptosis in vitro. The rat model of MI/R injury was induced by 30 min of transient vessel occlusion followed by 3 h of reperfusion. BE significantly reduced the myocardium infarct size and production of cardiac troponin (cTnl) in serum. In an in vitro experiment, H9c2 cardiomyocytes were incubated with vehicle or ischemic buffer during hypoxia; then, they were reoxygenated with or without BE. BE markedly improved the cell viability and decreased lactate dehydrogenase (LDH) release. We confirmed the anti-apoptotic effect of BE with the Hoechst 33258 staining assay, and this effect was associated with an increase in Bcl-2 and a decrease in active caspase-3 expression. Western blot analysis also showed that BE increased the phosphorylation of Akt and eNOS in H9c2 cells, and the protective effects of BE were partially inhibited by the phosphatidylinositol 3'-kinase (PI3K) specific inhibitor LY294002. Our results suggested that BE could provide significant cardioprotection against MI/R injury, and the potential mechanisms might involve suppression of cardiomyocyte apoptosis through activating the PI3K/Akt/eNOS signaling pathway.

  4. Microtubular stability affects pVHL-mediated regulation of HIF-1alpha via the p38/MAPK pathway in hypoxic cardiomyocytes.

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    Miao Teng

    Full Text Available BACKGROUND: Our previous research found that structural changes of the microtubule network influence glycolysis in cardiomyocytes by regulating the hypoxia-inducible factor (HIF-1α during the early stages of hypoxia. However, little is known about the underlying regulatory mechanism of the changes of HIF-1α caused by microtubule network alternation. The von Hippel-Lindau tumor suppressor protein (pVHL, as a ubiquitin ligase, is best understood as a negative regulator of HIF-1α. METHODOLOGY/PRINCIPAL FINDINGS: In primary rat cardiomyocytes and H9c2 cardiac cells, microtubule-stabilization was achieved by pretreating with paclitaxel or transfection of microtubule-associated protein 4 (MAP4 overexpression plasmids and microtubule-depolymerization was achieved by pretreating with colchicine or transfection of MAP4 siRNA before hypoxia treatment. Recombinant adenovirus vectors for overexpressing pVHL or silencing of pVHL expression were constructed and transfected in primary rat cardiomyocytes and H9c2 cells. With different microtubule-stabilizing and -depolymerizing treaments, we demonstrated that the protein levels of HIF-1α were down-regulated through overexpression of pVHL and were up-regulated through knockdown of pVHL in hypoxic cardiomyocytes. Importantly, microtubular structure breakdown activated p38/MAPK pathway, accompanied with the upregulation of pVHL. In coincidence, we found that SB203580, a p38/MAPK inhibitor decreased pVHL while MKK6 (Glu overexpression increased pVHL in the microtubule network altered-hypoxic cardiomyocytes and H9c2 cells. CONCLUSIONS/SIGNIFICANCE: This study suggests that pVHL plays an important role in the regulation of HIF-1α caused by the changes of microtubular structure and the p38/MAPK pathway participates in the process of pVHL change following microtubule network alteration in hypoxic cardiomyocytes.

  5. The endoplasmic reticulum stress-autophagy pathway is involved in apelin-13-induced cardiomyocyte hypertrophy in vitro

    Institute of Scientific and Technical Information of China (English)

    Feng XIE; Di WU; Shi-fang HUANG; Jian-gang CAO; He-ning LI; Lu HE; Mei-qing LIU; Lan-fang LI; Lin-xi CHEN

    2017-01-01

    Apelin is the endogenous ligand for the G protein-coupled receptor APJ,and plays important roles in the cardiovascular system.Our previous studies showed that apelin-13 promotes the hypertrophy of H9c2 rat cardiomyocytes through the PI3K-autophagy pathway.The aim of this study was to explore what roles ER stress and autophagy played in apelin-13-induced hypertrophy of cardiomyocytes in vitro.Treatment of H9c2 cells with apelin-13 (0.001-2 μJmol/L) dose-dependently increased the production of ROS and the expression levels of NADPH oxidase 4 (NOX4).Knockdown of Nox4 with siRNAs effectively prevented the reduction of GSH/GSSG ratio in apelin-13-treated cells.Furthermore,apelin-13 treatment dose-dependently increased the expression of Bip and CHOP,two ER stress markers,in the cells.Knockdown of APJ or Nox4 with the corresponding siRNAs,or application of NADPH inhibitor DPI blocked apelin-13-induced increases in Bip and CHOP expression.Moreover,apelin-13 treatment increased the formation of autophagosome and ER fragments and the LC3 puncta in the ER of the cells.Knockdown of APJ,Nox4,Bip or CHOP with the corresponding siRNAs,or application of DPI or salubrinal attenuated apelin-13-induced overexpression of LC3-Ⅱ/Ⅰ and beclin 1.Finally,knockdown of Nox4,Bip or CHOP with the corresponding siRNAs,or application of salubrinal significantly suppressed apelin-13-induced increases in the cell diameter,volume and protein contents.Our results demonstrate that ER stress-autophagy is involved in apelin-13-induced H9c2 cell hypertrophy.

  6. Pharmacological activation of mitochondrial BKCa channels protects isolated cardiomyocytes against simulated reperfusion-induced injury

    Czech Academy of Sciences Publication Activity Database

    Borchert, Gudrun H.; Hlaváčková, Markéta; Kolář, František

    2013-01-01

    Roč. 238, č. 2 (2013), s. 233-241 ISSN 1535-3702 R&D Projects: GA AV ČR(CZ) IAA500110804; GA ČR(CZ) GAP303/12/1162 Institutional research plan: CEZ:AV0Z50110509 Institutional support: RVO:67985823 Keywords : potassium channels * cardiomyocytes * mitochondria * ischemia/reperfusion * cytoprotection * reactive oxygen species Subject RIV: FA - Cardiovascular Diseases incl. Cardiotharic Surgery Impact factor: 2.226, year: 2013

  7. Cardiomyocyte Regeneration

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    Toshio Nakanishi

    2013-01-01

    Full Text Available The heart was initially believed to be a terminally differentiated organ; once the cardiomyocytes died, no recovery could be made to replace the dead cells. However, around a decade ago, the concept of cardiac stem cells (CSCs in adult hearts was proposed. CSCs differentiate into cardiomyocytes, keeping the heart functioning. Studies have proved the existence of stem cells in the heart. These somatic stem cells have been studied for use in cardiac regeneration. Moreover, recently, induced pluripotent stem cells (iPSCs were invented, and methodologies have now been developed to induce stable cardiomyocyte differentiation and purification of mature cardiomyocytes. A reprogramming method has also been applied to direct reprogramming using cardiac fibroblasts into cardiomyocytes. Here, we address cardiomyocyte differentiation of CSCs and iPSCs. Furthermore, we describe the potential of CSCs in regenerative biology and regenerative medicine.

  8. Adrenergic Stress Protection of Human iPS Cell-Derived Cardiomyocytes by Fast Kv7.1 Recycling

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    Ilaria Piccini

    2017-09-01

    Full Text Available The fight-or-flight response (FFR, a physiological acute stress reaction, involves positive chronotropic and inotropic effects on heart muscle cells mediated through β-adrenoceptor activation. Increased systolic calcium is required to enable stronger heart contractions whereas elevated potassium currents are to limit the duration of the action potentials and prevent arrhythmia. The latter effect is accomplished by an increased functional activity of the Kv7.1 channel encoded by KCNQ1. Current knowledge, however, does not sufficiently explain the full extent of rapid Kv7.1 activation and may hence be incomplete. Using inducible genetic KCNQ1 complementation in KCNQ1-deficient human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs, we here reinvestigate the functional role of Kv7.1 in adapting human CMs to adrenergic stress. Under baseline conditions, Kv7.1 was barely detectable at the plasma membrane of hiPSC-CMs, yet it fully protected these from adrenergic stress-induced beat-to-beat variability of repolarization and torsade des pointes-like arrhythmia. Furthermore, isoprenaline treatment increased field potential durations specifically in KCNQ1-deficient CMs to cause these adverse macroscopic effects. Mechanistically, we find that the protective action by Kv7.1 resides in a rapid translocation of channel proteins from intracellular stores to the plasma membrane, induced by adrenergic signaling. Gene silencing experiments targeting RAB GTPases, mediators of intracellular vesicle trafficking, showed that fast Kv7.1 recycling under acute stress conditions is RAB4A-dependent.Our data reveal a key mechanism underlying the rapid adaptation of human cardiomyocytes to adrenergic stress. These findings moreover aid to the understanding of disease pathology in long QT syndrome and bear important implications for safety pharmacological screening.

  9. Examining the protective role of ErbB2 modulation in human-induced pluripotent stem cell-derived cardiomyocytes.

    Science.gov (United States)

    Eldridge, Sandy; Guo, Liang; Mussio, Jodie; Furniss, Mike; Hamre, John; Davis, Myrtle

    2014-10-01

    Human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) are being used as an in vitro model system in cardiac biology and in drug discovery (e.g., cardiotoxicity testing). Qualification of these cells for use in mechanistic investigations will require detailed evaluations of cardiomyocyte signaling pathways and cellular responses. ErbB signaling and the ligand neuregulin play critical roles in survival and functional integrity of cardiac myocytes. As such, we sought to characterize the expression and activity of the ErbB family of receptors. Antibody microarray analysis performed on cell lysates derived from maturing hiPSC-CMs detected expression of ∼570 signaling proteins. EGFR/ErbB1, HER2/ErbB2, and ErbB4, but not ErbB3 receptors, of the epidermal growth factor receptor family were confirmed by Western blot. Activation of ErbB signaling by neuregulin-1β (NRG, a natural ligand for ErbB4) and its modulation by trastuzumab (a monoclonal anti-ErbB2 antibody) and lapatinib (a small molecule ErbB2 tyrosine kinase inhibitor) were evaluated through assessing phosphorylation of AKT and Erk1/2, two major downstream kinases of ErbB signaling, using nanofluidic proteomic immunoassay. Downregulation of ErbB2 expression by siRNA silencing attenuated NRG-induced AKT and Erk1/2 phosphorylation. Activation of ErbB signaling with NRG, or inhibition with trastuzumab, alleviated or aggravated doxorubicin-induced cardiomyocyte damage, respectively, as assessed by a real-time cellular impedance analysis and ATP measurement. Collectively, these results support the expanded use of hiPSC-CMs to examine mechanisms of cardiotoxicity and support the value of using these cells in early assessments of cardiotoxicity or efficacy. Published by Oxford University Press on behalf of Toxicological Sciences 2014. This work is written by (a) US Government employee(s) and is in the public domain in the US.

  10. Similar to spironolactone, oxymatrine is protective in aldosterone-induced cardiomyocyte injury via inhibition of calpain and apoptosis-inducing factor signaling.

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    Ting-Ting Xiao

    Full Text Available Accumulating evidence indicates that oxymatrine (OMT possesses variously pharmacological properties, especially on the cardiovascular system. We previously demonstrated that activated calpain/apoptosis-inducing factor (AIF-mediated pathway was the key molecular mechanism in aldosterone (ALD induces cardiomyocytes apoptosis. In the present study, we extended the experimentation by investigating the effect of OMT on cardiomyocytes exposed to ALD, as compared to spironolactone (Spiro, a classical ALD receptor antagonist. Cardiomyocytes were pre-incubated with OMT, Spiro or vehicle for 1 h, and then, cardiomyocytes were exposed to ALD 24 h. The cell injury was evaluated by 3-(4,5-dimethylthiazol-2-yl-2,5-diphenyltetrazolium bromide (MTT assay and lactate dehydrogenase (LDH leakage ratio. Apoptosis was determined by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL assay, annexin V/PI staining, and relative caspase-3 activity assay. Furthermore, expression of pro-apoptotic proteins including truncated Bid (tBid, calpain and AIF were evaluated by western blot analysis. ALD stimulation increased cardiomyocytes apoptosis, caspase-3 activity and protein expression of calpain, tBid and AIF in the cytosol (p<0.05. Pre-incubated with cardiomyocytes injury and increased caspase-3 activity were significantly attenuated (p<0.05. Furthermore, OMT suppressed ALD-induced high expression of calpain and AIF. And these effects of OMT could be comparable to Spiro. These findings indicated that OMT might be a potential cardioprotective-agent against excessive ALD-induced cardiotoxicity, at least in part, mediated through inhibition of calpain/AIF signaling.

  11. Pi3kcb links Hippo-YAP and PI3K-AKT signaling pathways to promote cardiomyocyte proliferation and survival.

    Science.gov (United States)

    Lin, Zhiqiang; Zhou, Pingzhu; von Gise, Alexander; Gu, Fei; Ma, Qing; Chen, Jinghai; Guo, Haidong; van Gorp, Pim R R; Wang, Da-Zhi; Pu, William T

    2015-01-02

    Yes-associated protein (YAP), the nuclear effector of Hippo signaling, regulates cellular growth and survival in multiple organs, including the heart, by interacting with TEA (transcriptional enhancer activator)-domain sequence-specific DNA-binding proteins. Recent studies showed that YAP stimulates cardiomyocyte proliferation and survival. However, the direct transcriptional targets through which YAP exerts its effects are poorly defined. To identify direct YAP targets that mediate its mitogenic and antiapoptotic effects in the heart. We identified direct YAP targets by combining differential gene expression analysis in YAP gain- and loss-of-function with genome-wide identification of YAP-bound loci using chromatin immunoprecipitation and high throughput sequencing. This screen identified Pik3cb, encoding p110β, a catalytic subunit of phosphoinositol-3-kinase, as a candidate YAP effector that promotes cardiomyocyte proliferation and survival. YAP and TEA-domain occupied a conserved enhancer within the first intron of Pik3cb, and this enhancer drove YAP-dependent reporter gene expression. Yap gain- and loss-of-function studies indicated that YAP is necessary and sufficient to activate the phosphoinositol-3-kinase-Akt pathway. Like Yap, Pik3cb gain-of-function stimulated cardiomyocyte proliferation, and Pik3cb knockdown dampened YAP mitogenic activity. Reciprocally, impaired heart function in Yap loss-of-function was significantly rescued by adeno-associated virus-mediated Pik3cb expression. Pik3cb is a crucial direct target of YAP, through which the YAP activates phosphoinositol-3-kinase-AKT pathway and regulates cardiomyocyte proliferation and survival. © 2014 American Heart Association, Inc.

  12. RhNRG-1β Protects the Myocardium against Irradiation-Induced Damage via the ErbB2-ERK-SIRT1 Signaling Pathway.

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    Anxin Gu

    Full Text Available Radiation-induced heart disease (RIHD, which is a serious side effect of the radiotherapy applied for various tumors due to the inevitable irradiation of the heart, cannot be treated effectively using current clinical therapies. Here, we demonstrated that rhNRG-1β, an epidermal growth factor (EGF-like protein, protects myocardium tissue against irradiation-induced damage and preserves cardiac function. rhNRG-1β effectively ameliorated irradiation-induced myocardial nuclear damage in both cultured adult rat-derived cardiomyocytes and rat myocardium tissue via NRG/ErbB2 signaling. By activating ErbB2, rhNRG-1β maintained mitochondrial integrity, ATP production, respiratory chain function and the Krebs cycle status in irradiated cardiomyocytes. Moreover, the protection of irradiated cardiomyocytes and myocardium tissue by rhNRG-1β was at least partly mediated by the activation of the ErbB2-ERK-SIRT1 signaling pathway. Long-term observations further showed that rhNRG-1β administered in the peri-irradiation period exerts continuous protective effects on cardiac pump function, the myocardial energy metabolism, cardiomyocyte volume and interstitial fibrosis in the rats receiving radiation via NRG/ErbB2 signaling. Our findings indicate that rhNRG-1β can protect the myocardium against irradiation-induced damage and preserve cardiac function via the ErbB2-ERK-SIRT1 signaling pathway.

  13. Growth hormone secretagogues protect mouse cardiomyocytes from in vitro ischemia/reperfusion injury through regulation of intracellular calcium.

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    Yi Ma

    Full Text Available BACKGROUND: Ischemic heart disease is a leading cause of mortality. To study this disease, ischemia/reperfusion (I/R models are widely used to mimic the process of transient blockage and subsequent recovery of cardiac coronary blood supply. We aimed to determine whether the presence of the growth hormone secretagogues, ghrelin and hexarelin, would protect/improve the function of heart from I/R injury and to examine the underlying mechanisms. METHODOLOGY/PRINCIPAL FINDINGS: Isolated hearts from adult male mice underwent 20 min global ischemia and 30 min reperfusion using a Langendorff apparatus. Ghrelin (10 nM or hexarelin (1 nM was introduced into the perfusion system either 10 min before or after ischemia, termed pre- and post-treatments. In freshly isolated cardiomyocytes from these hearts, single cell shortening, intracellular calcium ([Ca(2+](i transients and caffeine-releasable sarcoplasmic reticulum (SR Ca(2+ were measured. In addition, RT-PCR and Western blots were used to examine the expression level of GHS receptor type 1a (GHS-R1a, and phosphorylated phospholamban (p-PLB, respectively. Ghrelin and hexarelin pre- or post-treatments prevented the significant reduction in the cell shortening, [Ca(2+](i transient amplitude and caffeine-releasable SR Ca(2+ content after I/R through recovery of p-PLB. GHS-R1a antagonists, [D-Lys3]-GHRP-6 (200 nM and BIM28163 (100 nM, completely blocked the effects of GHS on both cell shortening and [Ca(2+](i transients. CONCLUSION/SIGNIFICANCE: Through activation of GHS-R1a, ghrelin and hexarelin produced a positive inotropic effect on ischemic cardiomyocytes and protected them from I/R injury probably by protecting or recovering p-PLB (and therefore SR Ca(2+ content to allow the maintenance or recovery of normal cardiac contractility. These observations provide supporting evidence for the potential therapeutic application of ghrelin and hexarelin in patients with cardiac I/R injury.

  14. Fluoride induces apoptosis in H9c2 cardiomyocytes via the mitochondrial pathway.

    Science.gov (United States)

    Yan, Xiaoyan; Wang, Lu; Yang, Xia; Qiu, Yulan; Tian, Xiaolin; Lv, Yi; Tian, Fengjie; Song, Guohua; Wang, Tong

    2017-09-01

    Numerous studies have shown that chronic excessive fluoride intake can adversely affect different organ systems. In particular, the cardiovascular system is susceptible to disruption by a high concentration of fluoride. The objectives of this study were to explore the mechanism of apoptosis by detecting the toxic effects of different concentrations of sodium fluoride (NaF) in H9c2 cells exposed for up to 96 h. NaF not only inhibited H9c2 cell proliferation but also induced apoptosis and morphological damage. With increasing NaF concentrations, early apoptosis of H9c2 cells was increased while the mitochondrial membrane potential was decreased. Compared with the control group, the mRNA levels of caspase-3, caspase-9, and cytochrome c all increased with increasing concentrations of NaF. In summary, these data suggest that apoptosis is involved in NaF-induced H9c2 cell toxicity and that activation of the mitochondrial pathway may occur. Copyright © 2017 Elsevier Ltd. All rights reserved.

  15. Neomysin inhibits Ca2+-stimulated phosphatidylinositol hydrolysis and protects cultured rat cardiomyocytes from Ca2+-dependent cell injury

    International Nuclear Information System (INIS)

    Babson, J.R.; Dougherty, J.M.

    1991-01-01

    Exposure of cultured rat cardiomyocytes to ionomycin and extracellular Ca 2+ leads to a rapid, sustained increase in intracellular free Ca 2+ as monitored by Ca 2+ -dependent phosphorylase a activation and to a subsequent loss of cardiomyocyte viability as determined by lactate dehydrogenase (LDH) leakage. The intracellular free Ca 2+ increase coincided with a rapid hydrolysis of phosphatidylinositol that preceded cell death. Phosphatidylinositol hydrolysis was monitored by the release of radiolabeled phosphoinositides from cardiomyocytes prelabeled with [2- 3 H]-myo-inositol. Neomycin, a known inhibitor of phospholipase C, inhibited the phosphatidylinositol hydrolysis and markedly reduced the extent of cell injury. Inhibitors of other Ca 2+ -activated processes, including intracellular proteases and phospholipase A 2 , had no effect on ionomycin-mediated cell injury. These data suggest that ionomycin-induced Ca 2+ -dependent cell injury in cultured cardiomyocytes may be due in part to the stimulation of phosphatidylinositol hydrolysis, presumably catalyzed by a Ca 2+ -dependent phospholipase C

  16. Role of JAK-STAT pathway in reducing cardiomyocytes hypoxia/reoxygenation injury induced by S1P postconditioning.

    Science.gov (United States)

    Wang, Yuqing; Wang, Dongfei; Zhang, Lizhi; Ye, Fangyu; Li, Mengmeng; Wen, Ke

    2016-08-05

    This experiment was designed to explore the protection of sphingosine1-phosphate (S1P) postconditioning on rat myocardial cells injured by hypoxia/reoxygenation acting via the Janus kinase-signal transducer and activator of transcription (JAK-STAT) signal pathway. The data showed that S1P could significantly increase cell viability, lower the rate of apoptosis, decrease the content of lactate dehydrogenase (LDH) and caspase3 activity in the culture medium, increase the activity of total superoxide dismutase (T-SOD) and manganese superoxide dismutase (Mn-SOD), reduce the loss of mitochondrial membrane potential and the fluorescence intensity of intracellular calcium, as well as increase the phosphorylation of JAK2 and STAT3 in comparison with the H/R group. When the JAK inhibitor AG490 or the STAT inhibitor stattic were added, the effects of S1P were inhibited. Our date shows that S1P protects H9c2 cells from hypoxia/reoxygenation injury and that the protection by S1P was inhibited by AG490 and stattic. Therefore S1P protects H9c2 cells against hypoxia/reoxygenation injury via the JAK-STAT pathway. Copyright © 2016 Elsevier B.V. All rights reserved.

  17. Cardiomyocyte Survival Pathways

    NARCIS (Netherlands)

    Lips, Daniel Jozef

    2004-01-01

    In the present thesis, the link between the genotype of the mouse and the concurrent phenotype is investigated employing sophisticated molecular and cellular techniques combined with in vivo cardiac performance measurements. In chapter 1 we focus on the characteristics of cardiac remodeling

  18. Dystrophin is required for the normal function of the cardio-protective K(ATP channel in cardiomyocytes.

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    Laura Graciotti

    Full Text Available Duchenne and Becker muscular dystrophy patients often develop a cardiomyopathy for which the pathogenesis is still unknown. We have employed the murine animal model of Duchenne muscular dystrophy (mdx, which develops a cardiomyopathy that includes some characteristics of the human disease, to study the molecular basis of this pathology. Here we show that the mdx mouse heart has defects consistent with alteration in compounds that regulate energy homeostasis including a marked decrease in creatine-phosphate (PC. In addition, the mdx heart is more susceptible to anoxia than controls. Since the cardio-protective ATP sensitive potassium channel (K(ATP complex and PC have been shown to interact we investigated whether deficits in PC levels correlate with other molecular events including K(ATP ion channel complex presence, its functionality and interaction with dystrophin. We found that this channel complex is present in the dystrophic cardiac cell membrane but its ability to sense a drop in the intracellular ATP concentration and consequently open is compromised by the absence of dystrophin. We further demonstrate that the creatine kinase muscle isoform (CKm is displaced from the plasma membrane of the mdx cardiac cells. Considering that CKm is a determinant of K(ATP channel complex function we hypothesize that dystrophin acts as a scaffolding protein organizing the K(ATP channel complex and the enzymes necessary for its correct functioning. Therefore, the lack of proper functioning of the cardio-protective K(ATP system in the mdx cardiomyocytes may be part of the mechanism contributing to development of cardiac disease in dystrophic patients.

  19. Protective effect of pioglitazone on cardiomyocyte apoptosis in low-dose streptozotocin & high-fat diet-induced type-2 diabetes in rats

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    Uma Bhandari

    2015-01-01

    Full Text Available Background & objectives: Cardiomyocyte apoptosis is one of the pathologic phenomena associated with diabetes and related conditions including obesity, insulin resistance and hyperlipidaemia. In the present study, the protective effects of pioglitazone on cardiomyocyte apoptosis was evaluated in experimental diabetes induced by low dose of streptozoticin (STZ combined with high fat diet (HFD in rats. Methods: Male Wistar rats (150-200 g were injected with low-dose STZ (45 mg/kg, i.v., single dose and orally fed with a HFD (20 g/day/rat for a period of 28 days and simultaneously treated with pioglitazone (20 mg/kg/p.o. for a period of 21 days (from 8 th day to 28 th day. On 29 th day blood was collected, serum separated and used for biochemical parameters. Heart tissue was used for cardiomyocyte apoptosis measurement and also for histopathological examination. Results: Pioglitazone treatment resulted in a decrease in cardiomyocyte apoptosis as revealed by a decrease in cardiac caspase-3, lactate dehydrogenase (LDH levels and DNA fragmentation, and an increase in Na+K+ATPase levels in diabetic rats. Cardiac histology of diabetic control rats showed dense focal fatty infiltration in the myocardial cells whereas normal architecture with regular morphology and well preserved cytoplasm was observed with pioglitazone treatment. Pioglitazone treatment significantly reduced the heart rate, mean arterial blood pressure, body mass index (BMI and levels of serum glucose, leptin, insulin, HOMA-IR, total cholesterol (TC and triglycerides (TGs, apoliproprotein-B glycosylated haemoglobin (HbA1c levels and atherogenic index, and increased the levels of serum high density lipoprotein cholesterol (HDL-C and cardiac antioxidant enzymes. Interpretation & conclusions: The present study results suggest that pioglitazone possesses cardiac anti-apoptotic potential in diabetic rat model and can be further explored for its use for treatment of diabetic cardiomyopathy.

  20. Protective effect of guggulsterone against cardiomyocyte injury induced by doxorubicin in vitro

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    Wang Wen-Ching

    2012-08-01

    Full Text Available Abstract Background Doxorubicin (DOX is an effective antineoplastic drug; however, clinical use of DOX is limited by its dose-dependent cardiotoxicity. It is well known that reactive oxygen species (ROS play a vital role in the pathological process of DOX-induced cardiotoxicity. For this study, we evaluated the protective effects of guggulsterone (GS, a steroid obtained from myrrh, to determine its preliminary mechanisms in defending against DOX-induced cytotoxicity in H9C2 cells. Methods In this study, we used a 3-(4,5-dimethylthiazol-2-yl-2,5-diphenyl-2H-tetrazolium bromide (MTT assay, lactate dehydrogenase (LDH release measurements, and Hoechst 33258 staining to evaluate the protective effect of GS against DOX-induced cytotoxicity in H9C2 cells. In addition, we observed the immunofluorescence of intracellular ROS and measured lipid peroxidation, caspase-3 activity, and apoptosis-related proteins by using Western blotting. Results The MTT assay and LDH release showed that treatment using GS (1–30 μM did not cause cytotoxicity. Furthermore, GS inhibited DOX (1 μM-induced cytotoxicity in a concentration-dependent manner. Hoechst 33258 staining showed that GS significantly reduced DOX-induced apoptosis and cell death. Using GS at a dose of 10–30 μM significantly reduced intracellular ROS and the formation of MDA in the supernatant of DOX-treated H9C2 cells and suppressed caspase-3 activity to reference levels. In immunoblot analysis, pretreatment using GS significantly reversed DOX-induced decrease of PARP, caspase-3 and bcl-2, and increase of bax, cytochrome C release, cleaved-PARP and cleaved-caspase-3. In addition, the properties of DOX-induced cancer cell (DLD-1 cells death did not interfere when combined GS and DOX. Conclusion These data provide considerable evidence that GS could serve as a novel cardioprotective agent against DOX-induced cardiotoxicity.

  1. Overexpression of glutaredoxin protects cardiomyocytes against nitric oxide-induced apoptosis with suppressing the S-nitrosylation of proteins and nuclear translocation of GAPDH

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    Inadomi, Chiaki, E-mail: inadomic@nagasaki-u.ac.jp [Department of Anesthesiology, Nagasaki University School of Medicine, Nagasaki 852-8501 (Japan); Department of Biochemistry and Molecular Biology in Disease, Atomic Bomb Disease Institute, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki 852-8523 (Japan); Murata, Hiroaki [Department of Anesthesiology, Nagasaki University School of Medicine, Nagasaki 852-8501 (Japan); Department of Biochemistry and Molecular Biology in Disease, Atomic Bomb Disease Institute, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki 852-8523 (Japan); Ihara, Yoshito [Department of Biochemistry and Molecular Biology in Disease, Atomic Bomb Disease Institute, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki 852-8523 (Japan); Department of Biochemistry, Wakayama Medical University, Wakayama 641-8509 (Japan); Goto, Shinji; Urata, Yoshishige [Department of Biochemistry and Molecular Biology in Disease, Atomic Bomb Disease Institute, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki 852-8523 (Japan); Department of Stem Cell Biology, Atomic Bomb Disease Institute, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki 852-8523 (Japan); Yodoi, Junji [Department of Biological Responses, Institute for Virus Research, Kyoto University, Kyoto 606-8507 (Japan); Kondo, Takahito [Department of Biochemistry and Molecular Biology in Disease, Atomic Bomb Disease Institute, Nagasaki University Graduate School of Biomedical Sciences, Nagasaki 852-8523 (Japan); Sumikawa, Koji [Department of Anesthesiology, Nagasaki University School of Medicine, Nagasaki 852-8501 (Japan)

    2012-08-31

    Highlights: Black-Right-Pointing-Pointer GRX1 overexpression protects myocardiac H9c2 cells against NO-induced apoptosis. Black-Right-Pointing-Pointer NO-induced nuclear translocation of GAPDH is suppressed in GRX overexpressors. Black-Right-Pointing-Pointer Oxidation of GAPDH by NO is less in GRX overexpressors than in controls. -- Abstract: There is increasing evidence demonstrating that glutaredoxin 1 (GRX1), a cytosolic enzyme responsible for the catalysis of protein deglutathionylation, plays distinct roles in inflammation and apoptosis by inducing changes in the cellular redox system. In this study, we investigated whether and how the overexpression of GRX1 protects cardiomyocytes against nitric oxide (NO)-induced apoptosis. Cardiomyocytes (H9c2 cells) were transfected with the expression vector for mouse GRX1 cDNA, and mock-transfected cells were used as a control. Compared with the mock-transfected cells, the GRX1-transfected cells were more resistant to NO-induced apoptosis. Stimulation with NO significantly increased the nuclear translocation of glyceraldehyde-3-phosphate dehydrogenase (GAPDH), a pro-apoptotic protein, in the mock-transfected cells, but did not change GAPDH localization in the GRX1-transfected cells. Furthermore, we found that NO stimulation clearly induced the oxidative modification of GAPDH in the mock-transfected cells, whereas less modification of GAPDH was observed in the GRX1-transfected cells. These data suggest that the overexpression of GRX1 could protect cardiomyocytes against NO-induced apoptosis, likely through the inhibition of the oxidative modification and the nuclear translocation of GAPDH.

  2. Overexpression of glutaredoxin protects cardiomyocytes against nitric oxide-induced apoptosis with suppressing the S-nitrosylation of proteins and nuclear translocation of GAPDH

    International Nuclear Information System (INIS)

    Inadomi, Chiaki; Murata, Hiroaki; Ihara, Yoshito; Goto, Shinji; Urata, Yoshishige; Yodoi, Junji; Kondo, Takahito; Sumikawa, Koji

    2012-01-01

    Highlights: ► GRX1 overexpression protects myocardiac H9c2 cells against NO-induced apoptosis. ► NO-induced nuclear translocation of GAPDH is suppressed in GRX overexpressors. ► Oxidation of GAPDH by NO is less in GRX overexpressors than in controls. -- Abstract: There is increasing evidence demonstrating that glutaredoxin 1 (GRX1), a cytosolic enzyme responsible for the catalysis of protein deglutathionylation, plays distinct roles in inflammation and apoptosis by inducing changes in the cellular redox system. In this study, we investigated whether and how the overexpression of GRX1 protects cardiomyocytes against nitric oxide (NO)-induced apoptosis. Cardiomyocytes (H9c2 cells) were transfected with the expression vector for mouse GRX1 cDNA, and mock-transfected cells were used as a control. Compared with the mock-transfected cells, the GRX1-transfected cells were more resistant to NO-induced apoptosis. Stimulation with NO significantly increased the nuclear translocation of glyceraldehyde-3-phosphate dehydrogenase (GAPDH), a pro-apoptotic protein, in the mock-transfected cells, but did not change GAPDH localization in the GRX1-transfected cells. Furthermore, we found that NO stimulation clearly induced the oxidative modification of GAPDH in the mock-transfected cells, whereas less modification of GAPDH was observed in the GRX1-transfected cells. These data suggest that the overexpression of GRX1 could protect cardiomyocytes against NO-induced apoptosis, likely through the inhibition of the oxidative modification and the nuclear translocation of GAPDH.

  3. Acacetin protects against cardiac remodeling after myocardial infarction by mediating MAPK and PI3K/Akt signal pathway

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    Wei Chang

    2017-12-01

    Full Text Available Since inhibiting cardiac remodeling is a critical treatment goal after myocardial infarction (MI, many drugs have been evaluated for this purpose. Acacetin is a flavonoid compound that has been shown to have anti-cancer, anti-mutagenic, anti-inflammatory and anti-peroxidative effects. In this study, we investigated whether acacetin is able to exert a protective effect against MI. One week after anterior wall standard MI surgeries or sham surgeries were performed in mice, acacetin was administered via gavage for two weeks. The results of echocardiographic and hemodynamic evaluation revealed that cardiac dysfunction significantly improved after acacetin treatment. H&E staining indicated that the ratio of the infarct size and the cardiomyocyte cross-sectional area was decreased by acacetin. Masson's staining detected that the fibrotic area ratio was evidently lower in the acacetin-treated MI group. TUNEL assays showed that acacetin ameliorated cardiomyocyte apoptosis after MI. RT-qPCR analysis showed that levels of hypertrophic and fibrotic markers were significantly decreased after acacetin treatment. Western blot analysis of various signaling pathway proteins showed that acacetin targets the MAPK and PI3K/Akt signaling pathways. Collectively, acacetin improves mouse left ventricular function and attenuates cardiac remodeling by inhibiting of the MAPK and PI3K/Akt signaling pathway.

  4. HIV-1 subtype C unproductively infects human cardiomyocytes in vitro and induces apoptosis mitigated by an anti-Gp120 aptamer.

    Science.gov (United States)

    Lopes de Campos, Walter R; Chirwa, Nthato; London, Grace; Rotherham, Lia S; Morris, Lynn; Mayosi, Bongani M; Khati, Makobetsa

    2014-01-01

    HIV-associated cardiomyopathy (HIVCM) is of clinical concern in developing countries because of a high HIV-1 prevalence, especially subtype C, and limited access to highly active antiretroviral therapy (HAART). For these reasons, we investigated the direct and indirect effects of HIV-1 subtype C infection of cultured human cardiomyocytes and the mechanisms leading to cardiomyocytes damage; as well as a way to mitigate the damage. We evaluated a novel approach to mitigate HIVCM using a previously reported gp120 binding and HIV-1 neutralizing aptamer called UCLA1. We established a cell-based model of HIVCM by infecting human cardiomyocytes with cell-free HIV-1 or co-culturing human cardiomyocytes with HIV-infected monocyte derived macrophages (MDM). We discovered that HIV-1 subtype C unproductively (i.e. its life cycle is arrested after reverse transcription) infects cardiomyocytes. Furthermore, we found that HIV-1 initiates apoptosis of cardiomyocytes through caspase-9 activation, preferentially via the intrinsic or mitochondrial initiated pathway. CXCR4 receptor-using viruses were stronger inducers of apoptosis than CCR5 utilizing variants. Importantly, we discovered that HIV-1 induced apoptosis of cardiomyocytes was mitigated by UCLA1. However, UCLA1 had no protective effective on cardiomyocytes when apoptosis was triggered by HIV-infected MDM. When HIV-1 was treated with UCLA1 prior to infection of MDM, it failed to induce apoptosis of cardiomyocytes. These data suggest that HIV-1 causes a mitochondrial initiated apoptotic cascade, which signal through caspase-9, whereas HIV-1 infected MDM causes apoptosis predominantly via the death-receptor pathway, mediated by caspase-8. Furthermore the data suggest that UCLA1 protects cardiomyocytes from caspase-mediated apoptosis, directly by binding to HIV-1 and indirectly by preventing infection of MDM.

  5. Plant Natural Product Formononetin Protects Rat Cardiomyocyte H9c2 Cells against Oxygen Glucose Deprivation and Reoxygenation via Inhibiting ROS Formation and Promoting GSK-3β Phosphorylation

    Directory of Open Access Journals (Sweden)

    Yuanyuan Cheng

    2016-01-01

    Full Text Available The opening of mitochondrial permeability transition pore (mPTP is a major cause of cell death in ischemia reperfusion injury. Based on our pilot experiments, plant natural product formononetin enhanced the survival of rat cardiomyocyte H9c2 cells during oxygen glucose deprivation (OGD and reoxygenation. For mechanistic studies, we focused on two major cellular factors, namely, reactive oxygen species (ROS and glycogen synthase kinase 3β (GSK-3β, in the regulation of mPTP opening. We found that formononetin suppressed the formation of ROS and superoxide in a concentration-dependent manner. Formononetin also rescued OGD/reoxygenation-induced loss of mitochondrial membrane integrity. Further studies suggested that formononetin induced Akt activation and GSK-3β (Ser9 phosphorylation, thereby reducing GSK-3β activity towards mPTP opening. PI3K and PKC inhibitors abolished the effects of formononetin on mPTP opening and GSK-3β phosphorylation. Immunoprecipitation experiments further revealed that formononetin increased the binding of phosphor-GSK-3β to adenine nucleotide translocase (ANT while it disrupted the complex of ANT with cyclophilin D. Moreover, immunofluorescence revealed that phospho-GSK-3β (Ser9 was mainly deposited in the space between mitochondria and cell nucleus. Collectively, these results indicated that formononetin protected cardiomyocytes from OGD/reoxygenation injury via inhibiting ROS formation and promoting GSK-3β phosphorylation.

  6. The use of microelectrode array (MEA) to study the protective effects of potassium channel openers on metabolically compromised HL-1 cardiomyocytes

    International Nuclear Information System (INIS)

    Law, J K Y; Chan, M; Yeung, C K; Rudd, J A; Hofmann, B; Ingebrandt, S; Offenhäusser, A

    2009-01-01

    The microelectrode array (MEA) was used to evaluate the cardioprotective effects of adenosine triphosphate sensitive potassium (K ATP ) channel activation using potassium channel openers (KCOs) on HL-1 cardiomyocytes subjected to acute chemically induced metabolic inhibition. Beat frequency and extracellular action potential (exAP) amplitude were measured in the presence of metabolic inhibitors (sodium azide (NaN 3 ) or 2-deoxyglucose (2-DG)) or KCOs (pinacidil (PIN, a cyanoguanidine derivative, activates sarcolemmal K ATP channels) or SDZ PCO400 (SDZ, a benzopyran derivative, activates mitochondrial K ATP channels)). The protective effects of these KCOs on metabolically inhibited HL-1 cells were subsequently investigated. Signal shapes indicated that NaN 3 and 2-DG reduced the rate of the sodium (Na + ) influx signal as reflected by a reduction in beat frequency. PIN and SDZ appeared to reduce both rate of depolarization and extent of the Na + influx signals. Pre-treating cardiomyocytes with PIN (0.1 mM), but not SDZ, prevented the reduction of beat frequency associated with NaN 3 - or 2-DG-induced metabolic inhibition. The exAP amplitude was not affected by either KCO. The cardioprotective effect of PIN relative to SDZ may be due to the opening of different K ATP channels. This metabolic inhibition model on the MEA may provide a stable platform for the study of cardiac pathophysiology in the future

  7. Epigenetic telomere protection by Drosophila DNA damage response pathways.

    Science.gov (United States)

    Oikemus, Sarah R; Queiroz-Machado, Joana; Lai, KuanJu; McGinnis, Nadine; Sunkel, Claudio; Brodsky, Michael H

    2006-05-01

    Analysis of terminal deletion chromosomes indicates that a sequence-independent mechanism regulates protection of Drosophila telomeres. Mutations in Drosophila DNA damage response genes such as atm/tefu, mre11, or rad50 disrupt telomere protection and localization of the telomere-associated proteins HP1 and HOAP, suggesting that recognition of chromosome ends contributes to telomere protection. However, the partial telomere protection phenotype of these mutations limits the ability to test if they act in the epigenetic telomere protection mechanism. We examined the roles of the Drosophila atm and atr-atrip DNA damage response pathways and the nbs homolog in DNA damage responses and telomere protection. As in other organisms, the atm and atr-atrip pathways act in parallel to promote telomere protection. Cells lacking both pathways exhibit severe defects in telomere protection and fail to localize the protection protein HOAP to telomeres. Drosophila nbs is required for both atm- and atr-dependent DNA damage responses and acts in these pathways during DNA repair. The telomere fusion phenotype of nbs is consistent with defects in each of these activities. Cells defective in both the atm and atr pathways were used to examine if DNA damage response pathways regulate telomere protection without affecting telomere specific sequences. In these cells, chromosome fusion sites retain telomere-specific sequences, demonstrating that loss of these sequences is not responsible for loss of protection. Furthermore, terminally deleted chromosomes also fuse in these cells, directly implicating DNA damage response pathways in the epigenetic protection of telomeres. We propose that recognition of chromosome ends and recruitment of HP1 and HOAP by DNA damage response proteins is essential for the epigenetic protection of Drosophila telomeres. Given the conserved roles of DNA damage response proteins in telomere function, related mechanisms may act at the telomeres of other organisms.

  8. 5-AIQ inhibits H{sub 2}O{sub 2}-induced apoptosis through reactive oxygen species scavenging and Akt/GSK-3β signaling pathway in H9c2 cardiomyocytes

    Energy Technology Data Exchange (ETDEWEB)

    Park, Eun-Seok; Kang, Jun Chul; Kang, Do-Hyun; Jang, Yong Chang [Department of Applied Biochemistry, Konkuk University, Chungju, Chungbuk, 380-701 (Korea, Republic of); Yi, Kyu Yang [Bio-Organic Science Division, Korea Research Institute of Chemical Technology, Daejeon, Chungnam, 305-600 (Korea, Republic of); Chung, Hun-Jong [Industrial Medicine Department, Chungju Hospital, Konkuk Medical School, Konkuk University, Chungju, Chungbuk, 380-701 (Korea, Republic of); Park, Jong Seok [Department of Biomedical Laboratory Science, Taegu Health College, Taegu 702-722 (Korea, Republic of); Kim, Bokyung [Department of Physiology, Konkuk Medical School, Konkuk University, Chungju, Chungbuk, 380-701 (Korea, Republic of); Feng, Zhong-Ping [Department of Physiology, College of Medicine, University of Toronto, Toronto, Ont., Canada M5S 1A8 (Canada); Shin, Hwa-Sup, E-mail: hsshin@kku.ac.kr [Department of Applied Biochemistry, Konkuk University, Chungju, Chungbuk, 380-701 (Korea, Republic of)

    2013-04-01

    Poly(adenosine 5′-diphosphate ribose) polymerase (PARP) is a nuclear enzyme activated by DNA strand breaks and plays an important role in the tissue injury associated with ischemia and reperfusion. The aim of the present study was to investigate the protective effect of 5-aminoisoquinolinone (5-AIQ), a PARP inhibitor, against oxidative stress-induced apoptosis in H9c2 cardiomyocytes. 5-AIQ pretreatment significantly protected against H{sub 2}O{sub 2}-induced cell death, as determined by the XTT assay, cell counting, terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling assay, and Western blot analysis of apoptosis-related proteins such as caspase-3, Bax, and Bcl-2. Upregulation of antioxidant enzymes such as manganese superoxide dismutase and catalase accompanied the protective effect of 5-AIQ on H{sub 2}O{sub 2}-induced cell death. Our data also showed that 5-AIQ pretreatment protected H9c2 cells from H{sub 2}O{sub 2}-induced apoptosis by triggering activation of Akt and glycogen synthase kinase-3β (GSK-3β), and that the protective effect of 5-AIQ was diminished by the PI3K inhibitor LY294002 at a concentration that effectively abolished 5-AIQ-induced Akt and GSK-3β activation. In addition, inhibiting the Akt/GSK-3β pathway by LY294002 significantly attenuated the 5-AIQ-mediated decrease in cleaved caspase-3 and Bax activation and H9c2 cell apoptosis induction. Taken together, these results demonstrate that 5-AIQ prevents H{sub 2}O{sub 2}-induced apoptosis in H9c2 cells by reducing intracellular reactive oxygen species production, regulating apoptosis-related proteins, and activating the Akt/GSK-3β pathway. - Highlights: ► 5-AIQ, a PARP inhibitor, decreased H{sub 2}O{sub 2}-induced H9c2 cell death and apoptosis. ► 5-AIQ upregulated antioxidant Mn-SOD and catalase, while decreasing ROS production. ► 5-AIQ decreased H{sub 2}O{sub 2}-induced increase in cleaved caspase-3 and Bax and decrease in Bcl2. ► 5-AIQ activated Akt and GSK-3

  9. Taurine ameliorated homocysteine-induced H9C2 cardiomyocyte apoptosis by modulating endoplasmic reticulum stress.

    Science.gov (United States)

    Zhang, Zhimin; Zhao, Lianyou; Zhou, Yanfen; Lu, Xuanhao; Wang, Zhengqiang; Wang, Jipeng; Li, Wei

    2017-05-01

    Homocysteine (Hcy)-triggered endoplasmic reticulum (ER) stress-mediated endothelial cell apoptosis has been suggested as a cause of Hcy-dependent vascular injury. However, whether ER stress is the molecular mechanism linking Hcy and cardiomyocytes death is unclear. Taurine has been reported to exert cardioprotective effects via various mechanisms. However, whether taurine protects against Hcy-induced cardiomyocyte death by attenuating ER stress is unknown. This study aimed to evaluate the opposite effects of taurine on Hcy-induced cardiomyocyte apoptosis and their underlying mechanisms. Our results demonstrated that low-dose or short-term Hcy treatment increased the expression of glucose-regulated protein 78 (GRP78) and activated protein kinase RNA-like ER kinase (PERK), inositol-requiring enzyme 1 (IRE1), and activating transcription factor 6 (ATF6), which in turn prevented apoptotic cell death. High-dose Hcy or prolonged Hcy treatment duration significantly up-regulated levels of C/EBP homologous protein (CHOP), cleaved caspase-12, p-c-Jun N-terminal kinase (JNK), and then triggered apoptotic events. High-dose Hcy also resulted in a decrease in mitochondrial membrane potential (Δψm) and an increase in cytoplasmic cytochrome C and the expression of cleaved caspase-9. Pretreatment of cardiomyocytes with sodium 4-phenylbutyric acid (an ER stress inhibitor) significantly inhibited Hcy-induced apoptosis. Furthermore, blocking the PERK pathway partly alleviated Hcy-induced ER stress-modulated cardiomyocyte apoptosis, and down-regulated the levels of Bax and cleaved caspase-3. Experimental taurine pretreatment inhibited the expression of ER stress-related proteins, and protected against apoptotic events triggered by Hcy-induced ER stress. Taken together, our results suggest that Hcy triggered ER stress in cardiomyocytes, which was the crucial molecular mechanism mediating Hcy-induced cardiomyocyte apoptosis, and the adverse effect of Hcy could be prevented by taurine.

  10. The protective effect of lipid emulsion in preventing bupivacaine-induced mitochondrial injury and apoptosis of H9C2 cardiomyocytes.

    Science.gov (United States)

    Chen, Zhe; Jin, Zhousheng; Xia, Yun; Zhao, Shishi; Xu, Xuzhong; Papadimos, Thomas J; Wang, Quanguang

    2017-11-01

    Lipid emulsion (LE) has been shown to be effective in the resuscitation of bupivacaine-induced cardiac arrest, but the precise mechanism of this action has not been fully elucidated. Pursuant to this lack of information on the mechanism in which LE protects the myocardium during bupivacaine-induced toxicity, we explored mitochondrial function and cell apoptosis. H9C2 cardiomyocytes were used in study. Cells were randomly divided in different groups and were cultivated 6 h, 12 h, and 24 h. The mitochondria were extracted and mitochondrial ATP content was measured, as was mitochondrial membrane potential, the concentration of calcium ion (Ca2+), and the activity of Ca2+-ATP enzyme (Ca2+-ATPase). Cells from groups Bup1000, LE group, and Bup1000LE were collected to determine cell viability, cell apoptosis, and electron microscopy scanning of mitochondrial ultrastructure (after 24 h). We found that LE can reverse the inhibition of the mitochondrial function induced by bupivacaine, regulate the concentration of calcium ion in mitochondria, resulting in the protection of myocardial cells from toxicity induced by bupivacaine.

  11. Analysis of cardiomyocyte movement in the developing murine heart

    Energy Technology Data Exchange (ETDEWEB)

    Hashimoto, Hisayuki [Department of Cardiology, Keio University School of Medicine, Tokyo (Japan); Yuasa, Shinsuke, E-mail: yuasa@a8.keio.jp [Department of Cardiology, Keio University School of Medicine, Tokyo (Japan); Tabata, Hidenori [Department of Anatomy, Keio University School of Medicine, Tokyo (Japan); Tohyama, Shugo; Seki, Tomohisa; Egashira, Toru; Hayashiji, Nozomi; Hattori, Fumiyuki; Kusumoto, Dai; Kunitomi, Akira; Takei, Makoto; Kashimura, Shin; Yozu, Gakuto; Shimojima, Masaya; Motoda, Chikaaki; Muraoka, Naoto [Department of Cardiology, Keio University School of Medicine, Tokyo (Japan); Nakajima, Kazunori [Department of Anatomy, Keio University School of Medicine, Tokyo (Japan); Sakaue-Sawano, Asako; Miyawaki, Atsushi [Life Function and Dynamics, ERATO, JST, 2-1 Hirosawa, Wako-city, Saitama 351-0198 (Japan); Laboratory for Cell Function and Dynamics, Advanced Technology Development Group, Brain Science Institute, RIKEN, 2-1 Hirosawa, Wako-city, Saitama 351-0198 (Japan); Fukuda, Keiichi [Department of Cardiology, Keio University School of Medicine, Tokyo (Japan)

    2015-09-04

    The precise assemblage of several types of cardiac precursors controls heart organogenesis. The cardiac precursors show dynamic movement during early development and then form the complicated heart structure. However, cardiomyocyte movements inside the newly organized mammalian heart remain unclear. We previously established the method of ex vivo time-lapse imaging of the murine heart to study cardiomyocyte behavior by using the Fucci (fluorescent ubiquitination-based cell cycle indicator) system, which can effectively label individual G1, S/G2/M, and G1/S-transition phase nuclei in living cardiomyocytes as red, green, and yellow, respectively. Global analysis of gene expression in Fucci green positive ventricular cardiomyocytes confirmed that cell cycle regulatory genes expressed in G1/S, S, G2/M, and M phase transitions were upregulated. Interestingly, pathway analysis revealed that many genes related to the cell cycle were significantly upregulated in the Fucci green positive ventricular cardiomyocytes, while only a small number of genes related to cell motility were upregulated. Time-lapse imaging showed that murine proliferating cardiomyocytes did not exhibit dynamic movement inside the heart, but stayed on site after entering the cell cycle. - Highlights: • We directly visualized cardiomyocyte movement inside the developing murine heart. • Cell cycle related genes were upregulated in the proliferating cardiomyocytes. • Time-lapse imaging revealed that proliferating murine cardiomyocytes stayed in place. • Murine ventricular cardiomyocytes proliferate on site during development.

  12. 8,9-Epoxyeicosatrienoic acid analog protects pulmonary artery smooth muscle cells from apoptosis via ROCK pathway

    International Nuclear Information System (INIS)

    Ma, Jun; Zhang, Lei; Li, Shanshan; Liu, Shulin; Ma, Cui; Li, Weiyang; Falck, J.R.; Manthati, Vijay L.; Reddy, D. Sudarshan; Medhora, Meetha; Jacobs, Elizabeth R.; Zhu, Daling

    2010-01-01

    Epoxyeicosatrienoic acids (EETs), metabolites of arachidonic acid (AA) catalyzed by cytochrome P450 (CYP), have many essential biologic roles in the cardiovascular system including inhibition of apoptosis in cardiomyocytes. In the present study, we tested the potential of 8,9-EET and derivatives to protect pulmonary artery smooth muscle cells (PASMCs) from starvation induced apoptosis. We found 8,9-epoxy-eicos-11(Z)-enoic acid (8,9-EET analog (214)), but not 8,9-EET, increased cell viability, decreased activation of caspase-3 and caspase-9, and decreased TUNEL-positive cells or nuclear condensation induced by serum deprivation (SD) in PASMCs. These effects were reversed after blocking the Rho-kinase (ROCK) pathway with Y-27632 or HA-1077. Therefore, 8,9-EET analog (214) protects PASMC from serum deprivation-induced apoptosis, mediated at least in part via the ROCK pathway. Serum deprivation of PASMCs resulted in mitochondrial membrane depolarization, decreased expression of Bcl-2 and enhanced expression of Bax, all effects were reversed by 8,9-EET analog (214) in a ROCK dependent manner. Because 8,9-EET and not the 8,9-EET analog (214) protects pulmonary artery endothelial cells (PAECs), these observations suggest the potential to differentially promote apoptosis or survival with 8,9-EET or analogs in pulmonary arteries.

  13. 8,9-Epoxyeicosatrienoic acid analog protects pulmonary artery smooth muscle cells from apoptosis via ROCK pathway

    Energy Technology Data Exchange (ETDEWEB)

    Ma, Jun; Zhang, Lei; Li, Shanshan [Department of Biopharmaceutical Sciences, College of Pharmacy, Harbin Medical University, 157 Baojian Road, Nangang District, Harbin, Heilongjiang 150081 (China); Liu, Shulin [Department of Biopharmaceutical Sciences, College of Pharmacy, Harbin Medical University, 157 Baojian Road, Nangang District, Harbin, Heilongjiang 150081 (China); Bio-pharmaceutical Key Laboratory of Heilongjiang Province, Harbin 150081 (China); Ma, Cui [Department of Biopharmaceutical Sciences, College of Pharmacy, Harbin Medical University, 157 Baojian Road, Nangang District, Harbin, Heilongjiang 150081 (China); Li, Weiyang [Mudanjiang Medical College, Mudanjiang 157011 (China); Falck, J.R.; Manthati, Vijay L.; Reddy, D. Sudarshan [University of Texas Southwestern Medical Center, Dallas, TX 75390 (United States); Medhora, Meetha; Jacobs, Elizabeth R. [Division of Pulmonary and Critical Care, Department of Medicine, Cardiovascular Center, Medical College of Wisconsin, Milwaukee, WI 53226 (United States); Zhu, Daling, E-mail: dalingz@yahoo.com [Department of Biopharmaceutical Sciences, College of Pharmacy, Harbin Medical University, 157 Baojian Road, Nangang District, Harbin, Heilongjiang 150081 (China); Bio-pharmaceutical Key Laboratory of Heilongjiang Province, Harbin 150081 (China)

    2010-08-15

    Epoxyeicosatrienoic acids (EETs), metabolites of arachidonic acid (AA) catalyzed by cytochrome P450 (CYP), have many essential biologic roles in the cardiovascular system including inhibition of apoptosis in cardiomyocytes. In the present study, we tested the potential of 8,9-EET and derivatives to protect pulmonary artery smooth muscle cells (PASMCs) from starvation induced apoptosis. We found 8,9-epoxy-eicos-11(Z)-enoic acid (8,9-EET analog (214)), but not 8,9-EET, increased cell viability, decreased activation of caspase-3 and caspase-9, and decreased TUNEL-positive cells or nuclear condensation induced by serum deprivation (SD) in PASMCs. These effects were reversed after blocking the Rho-kinase (ROCK) pathway with Y-27632 or HA-1077. Therefore, 8,9-EET analog (214) protects PASMC from serum deprivation-induced apoptosis, mediated at least in part via the ROCK pathway. Serum deprivation of PASMCs resulted in mitochondrial membrane depolarization, decreased expression of Bcl-2 and enhanced expression of Bax, all effects were reversed by 8,9-EET analog (214) in a ROCK dependent manner. Because 8,9-EET and not the 8,9-EET analog (214) protects pulmonary artery endothelial cells (PAECs), these observations suggest the potential to differentially promote apoptosis or survival with 8,9-EET or analogs in pulmonary arteries.

  14. SOX6 and PDCD4 enhance cardiomyocyte apoptosis through LPS-induced miR-499 inhibition.

    Science.gov (United States)

    Jia, Zhuqing; Wang, Jiaji; Shi, Qiong; Liu, Siyu; Wang, Weiping; Tian, Yuyao; Lu, Qin; Chen, Ping; Ma, Kangtao; Zhou, Chunyan

    2016-02-01

    Sepsis-induced cardiac apoptosis is one of the major pathogenic factors in myocardial dysfunction. As it enhances numerous proinflammatory factors, lipopolysaccharide (LPS) is considered the principal mediator in this pathological process. However, the detailed mechanisms involved are unclear. In this study, we attempted to explore the mechanisms involved in LPS-induced cardiomyocyte apoptosis. We found that LPS stimulation inhibited microRNA (miR)-499 expression and thereby upregulated the expression of SOX6 and PDCD4 in neonatal rat cardiomyocytes. We demonstrate that SOX6 and PDCD4 are target genes of miR-499, and they enhance LPS-induced cardiomyocyte apoptosis by activating the BCL-2 family pathway. The apoptosis process enhanced by overexpression of SOX6 or PDCD4, was rescued by the cardiac-abundant miR-499. Overexpression of miR-499 protected the cardiomyocytes against LPS-induced apoptosis. In brief, our results demonstrate the existence of a miR-499-SOX6/PDCD4-BCL-2 family pathway in cardiomyocytes in response to LPS stimulation.

  15. Shock Wave Therapy Promotes Cardiomyocyte Autophagy and Survival during Hypoxia

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    Ling Du

    2017-06-01

    Full Text Available Background: Autophagy plays an important role in cardiovascular disease. Controversy still exists regarding the effect of autophagy on ischemic/hypoxic myocardium. Cardiac shock wave therapy (CSWT is an effective alternative treatment for refractory ischemic heart disease. Whether CSWT can regulate cardiomyocyte autophagy under hypoxic conditions is not clear. We established a myocardial hypoxia model using the H9c2 cell line and performed shock waves (SWs treatment to evaluate the effect of SW on autophagy. Methods: The H9c2 cells were incubated under hypoxic conditions, and SW treatment was then performed at energies of 0.02, 0.05, or 0.10 mJ/mm2. The cell viability and intracellular ATP level were examined. Western blot analysis was used to assess the expression of LC3B, AMPK, mTOR, Beclin-1, Sirt1, and HIF-1α. Autophagic vacuoles were visualized by monodansylcadaverine staining. Results: After the 24-hour hypoxic period, cardiomyocyte viability and ATP levels were decreased and autophagy was significantly increased in H9c2 cells. SW treatment with an energy of 0.05 mJ/mm2 significantly increased the cellular viability, ATP level, LC3B-II/I, and number of autophagic vacuoles. In addition, phosphorylated AMPK and Sirt1 were increased and phosphorylated mTOR and HIF-1α were decreased after SW treatment. Conclusion: SW treatment can potentially promote cardiomyocyte autophagy during hypoxia and protect cardiomyocyte function by regulating the AMPK/mTOR pathway.

  16. Protective effect of Dendrobium officinale polysaccharides on H2O2-induced injury in H9c2 cardiomyocytes.

    Science.gov (United States)

    Zhao, Xiaoyan; Dou, Mengmeng; Zhang, Zhihao; Zhang, Duoduo; Huang, Chengzhi

    2017-10-01

    The preliminary studies have shown that Dendrobium officinale possessed therapeutic effects on hypertension and atherosclerosis. Studies also reported that Dendrobium officinale polysaccharides showed antioxidant capabilities. However, little is known about its effects on myocardial cells under oxidative stress. The present study was designed to study the protective effect of Dendrobium officinale polysaccharides against H 2 O 2 -induced oxidative stress in H9c2 cells. MTT assay was carried out to determine the cell viability of H9c2 cells when pretreated with Dendrobium officinale polysaccharides. Fluorescent microscopy measurements were performed for evaluating the apoptosis in H9c2 cells. Furthermore, effects of Dendrobium officinale polysaccharides on the activities of antioxidative indicators (malondialdehyde, superoxide dismutase), reactive oxygen species (ROS) production and mitochondrial membrane potential (MMP) levels were analyzed. Dendrobium officinale polysaccharides attenuated H 2 O 2 -induced cell death, as determined by the MTT assay. Dendrobium officinale polysaccharides decreased malondialdehyde levels, increased superoxide dismutase activities, and inhibited the generation of intracellular ROS. Moreover, pretreatment with Dendrobium officinale polysaccharides also inhibited apoptosis and increased the MMP levels in H9c2 cells. These results suggested the protective effects of Dendrobium officinale polysaccharides against H 2 O 2 -induced injury in H9c2 cells. The results also indicated the anti-oxidative capability of Dendrobium officinale polysaccharides. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  17. Endolysosomal pathway activity protects cells from neurotoxic TDP-43

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    Christine Leibiger

    2018-03-01

    Full Text Available The accumulation of protein aggregates in neurons is a typical pathological hallmark of the motor neuron disease amyotrophic lateral sclerosis (ALS and of frontotemporal dementia (FTD. In many cases, these aggregates are composed of the 43 kDa TAR DNA-binding protein (TDP‑43. Using a yeast model for TDP‑43 proteinopathies, we observed that the vacuole (the yeast equivalent of lysosomes markedly contributed to the degradation of TDP‑43. This clearance occurred via TDP‑43-containing vesicles fusing with the vacuole through the concerted action of the endosomal-vacuolar (or endolysosomal pathway and autophagy. In line with its dominant role in the clearance of TDP‑43, endosomal-vacuolar pathway activity protected cells from the detrimental effects of TDP‑43. In contrast, enhanced autophagy contributed to TDP‑43 cytotoxicity, despite being involved in TDP‑43 degradation. TDP‑43’s interference with endosomal-vacuolar pathway activity may have two deleterious consequences. First, it interferes with its own degradation via this pathway, resulting in TDP‑43 accumulation. Second, it affects vacuolar proteolytic activity, which requires endosomal-vacuolar trafficking. We speculate that the latter contributes to aberrant autophagy. In sum, we propose that ameliorating endolysosomal pathway activity enhances cell survival in TDP‑43-associated diseases.

  18. H- ras deletion protects against angiotensin II-induced arterial hypertension and cardiac remodeling through protein kinase G-Iβ pathway activation.

    Science.gov (United States)

    Martín-Sánchez, Paloma; Luengo, Alicia; Griera, Mercedes; Orea, María Jesús; López-Olañeta, Marina; Chiloeches, Antonio; Lara-Pezzi, Enrique; de Frutos, Sergio; Rodríguez-Puyol, Manuel; Calleros, Laura; Rodríguez-Puyol, Diego

    2018-02-01

    Ras proteins regulate cell survival, growth, differentiation, blood pressure, and fibrosis in some organs. We have demonstrated that H- ras gene deletion produces mice hypotension via a soluble guanylate cyclase-protein kinase G (PKG)-dependent mechanism. In this study, we analyzed the consequences of H- ras deletion on cardiac remodeling induced by continuous angiotensin II (AngII) infusion and the molecular mechanisms implied. Left ventricular posterior wall thickness and mass and cardiomyocyte cross-sectional area were similar between AngII-treated H-Ras knockout (H -ras -/- ) and control wild-type (H -ras +/+ ) mice, as were extracellular matrix protein expression. Increased cardiac PKG-Iβ protein expression in H -ras -/- mice suggests the involvement of this protein in heart protection. Ex vivo experiments on cardiac explants could support this mechanism, as PKG blockade blunted protection against AngII-induced cardiac hypertrophy and fibrosis markers in H -ras -/- mice. Genetic modulation studies in cardiomyocytes and cardiac and embryonic fibroblasts revealed that the lack of H-Ras down-regulates the B-RAF/MEK/ERK pathway, which induces the glycogen synthase kinase-3β-dependent activation of the transcription factor, cAMP response element-binding protein, which is responsible for PKG-Iβ overexpression in H -ras -/- mouse embryonic fibroblasts. This study demonstrates that H- ras deletion protects against AngII-induced cardiac remodeling, possibly via a mechanism in which PKG-Iβ overexpression could play a partial role, and points to H-Ras and/or downstream proteins as potential therapeutic targets in cardiovascular disease.-Martín-Sánchez, P., Luengo, A., Griera, M., Orea, M. J., López-Olañeta, M., Chiloeches, A., Lara-Pezzi, E., de Frutos, S., Rodríguez-Puyol, M., Calleros, L., Rodríguez-Puyol, D. H- ras deletion protects against angiotensin II-induced arterial hypertension and cardiac remodeling through protein kinase G-Iβ pathway activation.

  19. Low level tumor necrosis factor-alpha protects cardiomyocytes against high level tumor necrosis factor-alpha: brief insight into a beneficial paradox.

    Science.gov (United States)

    Cacciapaglia, Fabio; Salvatorelli, Emanuela; Minotti, Giorgio; Afeltra, Antonella; Menna, Pierantonio

    2014-12-01

    Whether tumor necrosis factor-alpha (TNFα) caused beneficial or detrimental cardiovascular effects remains poorly defined. Anti-TNFα agents improved cardiac end points in chronic rheumatic diseases characterized by progressive deterioration of cardiac function. In contrast, anti-TNFα agents did not always improve but actually worsened cardiac function in non-rheumatic patients with heart failure (HF), in spite of that HF usually accompanies with high circulating levels of TNFα. To shed light on these mixed findings, we characterized the effects of TNFα in H9c2 cardiomyocytes. Cells were incubated for 24 h with increasing concentrations of TNFα, hydrogen peroxide, aminotriazole, or etoposide. Posttreatment cell viability was assessed by antimycin A-inhibitable reduction of 3-(4,dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide, and the IC50 value of each test compound was defined. H9c2 cells were also preconditioned with a low non-toxic concentration of TNFα and then re-challenged with increasing concentrations of TNFα and other stressor agents. In re-challenge experiments, all of the IC50 values increased significantly, with the IC50 value of TNFα increasing approximately 16-fold. TNFα preconditioning increased cardiomyocytes shedding of the external portion of transmembrane type 1 and type 2 TNFα receptors [(soluble TNFα receptors (sTNFR)]. Levels of survival-oriented soluble TNFR2 (sTNFR2) always exceeded those of death-oriented sTNFR1. When exposed to TNFα at its IC50 value, preconditioned cardiomyocytes showed an increased release of sTNFR2 but not sTNFR1. These results denoted that preconditioning by "low TNFα" helped cardiomyocyte to withstand toxicity from "high TNFα" or other agents. These results also suggested that beneficial or detrimental effects of anti-TNFα agents might well depend on whether these agents spared or intercepted discrete amounts of TNFα that preconditioned cardiomyocytes and made them more resistant to high

  20. Cathepsin B is involved in the heat shock induced cardiomyocytes apoptosis as well as the anti-apoptosis effect of HSP-70.

    Science.gov (United States)

    Hsu, Shu-Fen; Hsu, Chuan-Chih; Cheng, Bor-Chih; Lin, Cheng-Hsien

    2014-11-01

    Cathepsin B is one of the major lysosomal cysteine proteases that plays an important role in apoptosis. Herein, we investigated whether Cathepsin B is involved in cardiomyocyte apoptosis caused by hyperthermic injury (HI) and heat shock protein (HSP)-70 protects these cells from HI-induced apoptosis mediated by Cathepsin. HI was produced in H9C2 cells by putting them in a circulating 43 °C water bath for 120 min, whereas preinduction of HSP-70 was produced in H9C2 cells by mild heat preconditioning (or putting them in 42 °C water bath for 30 min) 8 h before the start of HI. It was found that HI caused both cardiomyocyte apoptosis and increased Cathepsin B activity in H9C2 cells. E-64-c, in addition to reducing Cathepsin B activity, significantly attenuated HI-induced cardiomyocyte apoptosis (evidenced by increased apoptotic cell numbers, increased tuncated Bid (t-Bid), increased cytochrome C, increased caspase-9/-3, and decreased Bcl-2/Bax) in H9C2 cells. In addition, preinduction of HSP-70 by mild heat preconditioning or inhibition of HSP-70 by Tripolide significantly attenuated or exacerbated respectively both the cardiomyocyte apoptosis and increased Cathepsin B activity in H9C2 cells. Furthermore, the beneficial effects of pre-induction of HSP-70 by mild heat production in reducing both cardiomyocyte apoptosis and increased Cathepsin B activity caused by HI can be significantly reduced by Triptolide preconditioning. These results indicate that Cathepsin B is involved in HI-induced cardiomyocyte apoptosis in H9C2 cells and HSP-70 protects these cells from HI-induced cardiomyocyte apoptosis through Cathepsin B pathways.

  1. Ghrelin protects the heart against ischemia/reperfusion injury via inhibition of TLR4/NLRP3 inflammasome pathway.

    Science.gov (United States)

    Wang, Qin; Lin, Ping; Li, Peng; Feng, Li; Ren, Qian; Xie, Xiaofeng; Xu, Jing

    2017-10-01

    The aim of this study was to investigate the cardioprotective effects of ghrelin against myocardial ischemia/reperfusion (I/R) injury and the underlying mechanism. Sprague-Dawley rats were randomized into Sham, I/R and I/R+ghrelin groups. After 30 minutes ischemia, ghrelin (8nmol/kg) was injected intraperitoneally at the time of reperfusion in the I/R+ghrelin group. Then hemodynamic parameters were observed at 24h after reperfusion. Ghrelin exhibited dramatic improvement in cardiac functions, as manifested by increased LVSP and ±dP/dt max and decreased LVDP. At 24h after reperfusion, ghrelin significantly attenuated the myocardial infarction area and apoptosis, accompanied with a decrease in the levels of the myocyte injury marker enzymes. Oxidative stress injury and inflammatory response were also relieved by ghrelin. Western blot showed that the expression of TLR4, NLRP3, and caspase-1 were obviously increased in I/R group, while ghrelin significantly inhibited the I/R-induced TLR4, NLRP3, and caspase-1 expression. Ghrelin could inhibit the increased protein levels of NLRP3, caspase-1, and IL-1β induced by lipopolysacharide in primary cultured cardiomyocytes of neonatal rats. Ghrelin protected the heart against I/R injury by inhibiting oxidative stress and inflammation via TLR4/NLRP3 signaling pathway. Our results might provide new strategy and target for treatment of myocardial ischemia/reperfusion injury. Copyright © 2017 Elsevier Inc. All rights reserved.

  2. Fractalkine depresses cardiomyocyte contractility.

    Directory of Open Access Journals (Sweden)

    David Taube

    Full Text Available Our laboratory reported that male mice with cardiomyocyte-selective knockout of the prostaglandin E2 EP4 receptor sub-type (EP4 KO exhibit reduced cardiac function. Gene array on left ventricles (LV showed increased fractalkine, a chemokine implicated in heart failure. We therefore hypothesized that fractalkine is regulated by PGE2 and contributes to depressed contractility via alterations in intracellular calcium.Fractalkine was measured in LV of 28-32 week old male EP4 KO and wild type controls (WT by ELISA and the effect of PGE2 on fractalkine secretion was measured in cultured neonatal cardiomyocytes and fibroblasts. The effect of fractalkine on contractility and intracellular calcium was determined in Fura-2 AM-loaded, electrical field-paced cardiomyocytes. Cardiomyocytes (AVM from male C57Bl/6 mice were treated with fractalkine and responses measured under basal conditions and after isoproterenol (Iso stimulation.LV fractalkine was increased in EP4 KO mice but surprisingly, PGE2 regulated fractalkine secretion only in fibroblasts. Fractalkine treatment of AVM decreased both the speed of contraction and relaxation under basal conditions and after Iso stimulation. Despite reducing contractility after Iso stimulation, fractalkine increased the Ca(2+ transient amplitude but decreased phosphorylation of cardiac troponin I, suggesting direct effects on the contractile machinery.Fractalkine depresses myocyte contractility by mechanisms downstream of intracellular calcium.

  3. Protective effect of resveratrol against nigrostriatal pathway injury in striatum via JNK pathway.

    Science.gov (United States)

    Li, Dan; Liu, Nan; Zhao, Liang; Tong, Lei; Kawano, Hitoshi; Yan, Hong-Jing; Li, Hong-Peng

    2017-01-01

    Nigrostriatal pathway injury is one of the traumatic brain injury models that usually lead to neurological dysfunction or neuron necrosis. Resveratrol-induced benefits have recently been demonstrated in several models of neuronal degeneration diseases. However, the protective properties of resveratrol against neurodegeneration have not been explored definitely. Thus, we employ the nigrostriatal pathway injury model to mimic the insults on the brain. Resveratrol decreased the p-ERK expression and increased the p-JNK expression compared to the DMSO group, but not alter the p38 MAPK proteins around the lesion site by Western blot. Prior to the injury, mice were infused with resveratrol intracerebroventricularly with or without JNK-IN-8, a specific c-JNK pathway inhibitor for JNK1, JNK2 and JNK4. The study assessed modified improved neurological function score (mNSS) and beam/walking test, the level of inflammatory cytokines IL-1β, IL-6 and TNF-α, and striatal expression of Bax and Bcl-2 proteins associated with neuronal apoptosis. The results revealed that resveratrol exerted a neuroprotective effect as shown by the improved mNSS and beam latency, anti-inflammatory effects as indicated by the decreased level of IL-1β, TNF-α and IL-6. Furthermore, resveratrol up-regulated the protein expression of p-JNK and Bcl-2, down-regulated the expression of Bax and the number of Fluoro-Jade C (FJC) positive neurons. However, these advantages of resveratrol were abolished by JNK-IN-8 treatment. Overall, we demonstrated that resveratrol treatment attenuates the nigrostriatal pathway injury-induced neuronal apoptosis and inflammation via activation of c-JNK signaling. Copyright © 2016 Elsevier B.V. All rights reserved.

  4. Inhibition of CD38/Cyclic ADP-ribose Pathway Protects Rats against Ropivacaine-induced Convulsion

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    Yu Zou

    2017-01-01

    Conclusions: The CD38/cADPR pathway is activated in ropivacaine-induced convulsion. Inhibiting this pathway alleviates ropivacaine-induced convulsion and protects the brain from apoptosis and oxidative stress.

  5. The anti-apoptotic and cardioprotective effects of salvianolic acid a on rat cardiomyocytes following ischemia/reperfusion by DUSP-mediated regulation of the ERK1/2/JNK pathway.

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    Tongda Xu

    Full Text Available The purpose of this study was to observe the effects of salvianolic acid A (SAA pretreatment on the myocardium during ischemia/reperfusion (I/R and to illuminate the interrelationships among dual specificity protein phosphatase (DUSP 2/4/16, ERK1/2 and JNK pathways during myocardial I/R, with the ultimate goal of elucidating how SAA exerts cardioprotection against I/R injury (IRI. Wistar rats were divided into the following six groups: control group (CON, I/R group, SAA+I/R group, ERK1/2 inhibitor PD098059+I/R group (PD+I/R, PD+SAA+I/R group, and JNK inhibitor SP600125+I/R group (SP+I/R. The cardioprotective effects of SAA on the myocardium during I/R were investigated with a Langendorff device. Heart rate (HR, left ventricular systolic pressure (LVSP, left ventricular end-diastolic pressure (LVEDP, maximum rate of ventricular pressure rise and fall (±dp/dtmax, myocardial infarction areas (MIA, lactate dehydrogenase (LDH, and cardiomyocytes apoptosis were monitored. To determine the crosstalk betwee JNK and ERK1/2 via DUSP2/4/16 with SAA pretreatment, siRNA-DUSP2/4/16 were performed. The expression levels of Bcl-2, Bax, caspase 3, p-JNK, p-ERK1/2 and DUSP2/4/16 in cardiomyocytes were assayed by Western blot. Our results showed that LDH, MIA and cell apoptosis were decreased, and various parameters of heart function were improved by SAA pretreatment and SP application. In the I/R group, the expression levels of p-ERK1/2 and DUSP4/16 were not significantly different compared with the CON group, however, the protein expression levels of p-ERK1/2, Bcl-2 and DUSP4/16 were higher, while p-JNK, Bax, caspase 3 and DUSP2 levels were reduced among the SAA+I/R, PD+SAA+I/R and SP+I/R groups. The above indices were not significantly different between the SAA+I/R and SP+I/R groups. Compared with the SAA+I/R group, p-ERK1/2 was increased and p-JNK was decreased in the SAA+si-DUSP2+I/R, however, p-ERK was downregulated and p-JNK was upregulated in SAA+si-DUSP4

  6. Naphthazarin protects against glutamate-induced neuronal death via activation of the Nrf2/ARE pathway.

    Science.gov (United States)

    Son, Tae Gen; Kawamoto, Elisa M; Yu, Qian-Sheng; Greig, Nigel H; Mattson, Mark P; Camandola, Simonetta

    2013-04-19

    Nuclear factor E2-related factor 2 (Nrf2)/antioxidant response element (ARE) pathway is an important cellular stress response pathway involved in neuroprotection. We previously screened several natural phytochemicals and identified plumbagin as a novel activator of the Nrf2/ARE pathway that can protect neurons against ischemic injury. Here we extended our studies to natural and synthetic derivatives of plumbagin. We found that 5,8-dimethoxy-1,4-naphthoquinone (naphthazarin) is a potent activator of the Nrf2/ARE pathway, up-regulates the expression of Nrf2-driven genes in primary neuronal and glial cultures, and protects neurons against glutamate-induced excitotoxicity. Published by Elsevier Inc.

  7. Generation of Cardiomyocytes from Pluripotent Stem Cells.

    Science.gov (United States)

    Nakahama, Hiroko; Di Pasquale, Elisa

    2016-01-01

    The advent of pluripotent stem cells (PSCs) enabled a multitude of studies for modeling the development of diseases and testing pharmaceutical therapeutic potential in vitro. These PSCs have been differentiated to multiple cell types to demonstrate its pluripotent potential, including cardiomyocytes (CMs). However, the efficiency and efficacy of differentiation vary greatly between different cell lines and methods. Here, we describe two different methods for acquiring CMs from human pluripotent lines. One method involves the generation of embryoid bodies, which emulates the natural developmental process, while the other method chemically activates the canonical Wnt signaling pathway to induce a monolayer of cardiac differentiation.

  8. Mena/VASP and αII-Spectrin complexes regulate cytoplasmic actin networks in cardiomyocytes and protect from conduction abnormalities and dilated cardiomyopathy.

    Science.gov (United States)

    Benz, Peter M; Merkel, Carla J; Offner, Kristin; Abeßer, Marco; Ullrich, Melanie; Fischer, Tobias; Bayer, Barbara; Wagner, Helga; Gambaryan, Stepan; Ursitti, Jeanine A; Adham, Ibrahim M; Linke, Wolfgang A; Feller, Stephan M; Fleming, Ingrid; Renné, Thomas; Frantz, Stefan; Unger, Andreas; Schuh, Kai

    2013-08-12

    In the heart, cytoplasmic actin networks are thought to have important roles in mechanical support, myofibrillogenesis, and ion channel function. However, subcellular localization of cytoplasmic actin isoforms and proteins involved in the modulation of the cytoplasmic actin networks are elusive. Mena and VASP are important regulators of actin dynamics. Due to the lethal phenotype of mice with combined deficiency in Mena and VASP, however, distinct cardiac roles of the proteins remain speculative. In the present study, we analyzed the physiological functions of Mena and VASP in the heart and also investigated the role of the proteins in the organization of cytoplasmic actin networks. We generated a mouse model, which simultaneously lacks Mena and VASP in the heart. Mena/VASP double-deficiency induced dilated cardiomyopathy and conduction abnormalities. In wild-type mice, Mena and VASP specifically interacted with a distinct αII-Spectrin splice variant (SH3i), which is in cardiomyocytes exclusively localized at Z- and intercalated discs. At Z- and intercalated discs, Mena and β-actin localized to the edges of the sarcomeres, where the thin filaments are anchored. In Mena/VASP double-deficient mice, β-actin networks were disrupted and the integrity of Z- and intercalated discs was markedly impaired. Together, our data suggest that Mena, VASP, and αII-Spectrin assemble cardiac multi-protein complexes, which regulate cytoplasmic actin networks. Conversely, Mena/VASP deficiency results in disrupted β-actin assembly, Z- and intercalated disc malformation, and induces dilated cardiomyopathy and conduction abnormalities.

  9. Naturally Engineered Maturation of Cardiomyocytes

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    Gaetano J. Scuderi

    2017-05-01

    Full Text Available Ischemic heart disease remains one of the most prominent causes of mortalities worldwide with heart transplantation being the gold-standard treatment option. However, due to the major limitations associated with heart transplants, such as an inadequate supply and heart rejection, there remains a significant clinical need for a viable cardiac regenerative therapy to restore native myocardial function. Over the course of the previous several decades, researchers have made prominent advances in the field of cardiac regeneration with the creation of in vitro human pluripotent stem cell-derived cardiomyocyte tissue engineered constructs. However, these engineered constructs exhibit a functionally immature, disorganized, fetal-like phenotype that is not equivalent physiologically to native adult cardiac tissue. Due to this major limitation, many recent studies have investigated approaches to improve pluripotent stem cell-derived cardiomyocyte maturation to close this large functionality gap between engineered and native cardiac tissue. This review integrates the natural developmental mechanisms of cardiomyocyte structural and functional maturation. The variety of ways researchers have attempted to improve cardiomyocyte maturation in vitro by mimicking natural development, known as natural engineering, is readily discussed. The main focus of this review involves the synergistic role of electrical and mechanical stimulation, extracellular matrix interactions, and non-cardiomyocyte interactions in facilitating cardiomyocyte maturation. Overall, even with these current natural engineering approaches, pluripotent stem cell-derived cardiomyocytes within three-dimensional engineered heart tissue still remain mostly within the early to late fetal stages of cardiomyocyte maturity. Therefore, although the end goal is to achieve adult phenotypic maturity, more emphasis must be placed on elucidating how the in vivo fetal microenvironment drives cardiomyocyte

  10. Naphthazarin protects against glutamate-induced neuronal death via activation of the Nrf2/ARE pathway

    Energy Technology Data Exchange (ETDEWEB)

    Son, Tae Gen; Kawamoto, Elisa M.; Yu, Qian-Sheng; Greig, Nigel H. [Laboratory of Neurosciences, National Institute on Aging, Intramural Research Program, 251 Bayview Blvd., Baltimore, MD 21224 (United States); Mattson, Mark P. [Laboratory of Neurosciences, National Institute on Aging, Intramural Research Program, 251 Bayview Blvd., Baltimore, MD 21224 (United States); Department of Neuroscience, Johns Hopkins University School of Medicine, Baltimore, MD (United States); Camandola, Simonetta, E-mail: camandolasi@mail.nih.gov [Laboratory of Neurosciences, National Institute on Aging, Intramural Research Program, 251 Bayview Blvd., Baltimore, MD 21224 (United States)

    2013-04-19

    Highlights: •Naphthazarin activates the Nrf2/ARE pathway. •Naphthazarin induces Nrf2-driven genes in neurons and astrocytes. •Naphthazarin protects neurons against excitotoxicity. -- Abstract: Nuclear factor E2-related factor 2 (Nrf2)/antioxidant response element (ARE) pathway is an important cellular stress response pathway involved in neuroprotection. We previously screened several natural phytochemicals and identified plumbagin as a novel activator of the Nrf2/ARE pathway that can protect neurons against ischemic injury. Here we extended our studies to natural and synthetic derivatives of plumbagin. We found that 5,8-dimethoxy-1,4-naphthoquinone (naphthazarin) is a potent activator of the Nrf2/ARE pathway, up-regulates the expression of Nrf2-driven genes in primary neuronal and glial cultures, and protects neurons against glutamate-induced excitotoxicity.

  11. Naphthazarin protects against glutamate-induced neuronal death via activation of the Nrf2/ARE pathway

    International Nuclear Information System (INIS)

    Son, Tae Gen; Kawamoto, Elisa M.; Yu, Qian-Sheng; Greig, Nigel H.; Mattson, Mark P.; Camandola, Simonetta

    2013-01-01

    Highlights: •Naphthazarin activates the Nrf2/ARE pathway. •Naphthazarin induces Nrf2-driven genes in neurons and astrocytes. •Naphthazarin protects neurons against excitotoxicity. -- Abstract: Nuclear factor E2-related factor 2 (Nrf2)/antioxidant response element (ARE) pathway is an important cellular stress response pathway involved in neuroprotection. We previously screened several natural phytochemicals and identified plumbagin as a novel activator of the Nrf2/ARE pathway that can protect neurons against ischemic injury. Here we extended our studies to natural and synthetic derivatives of plumbagin. We found that 5,8-dimethoxy-1,4-naphthoquinone (naphthazarin) is a potent activator of the Nrf2/ARE pathway, up-regulates the expression of Nrf2-driven genes in primary neuronal and glial cultures, and protects neurons against glutamate-induced excitotoxicity

  12. c-kitpos GATA-4 high rat cardiac stem cells foster adult cardiomyocyte survival through IGF-1 paracrine signalling.

    Directory of Open Access Journals (Sweden)

    Nanako Kawaguchi

    2010-12-01

    Full Text Available Resident c-kit positive (c-kitpos cardiac stem cells (CSCs could be considered the most appropriate cell type for myocardial regeneration therapies. However, much is still unknown regarding their biological properties and potential.We produced clones of high and low expressing GATA-4 CSCs from long-term bulk-cultured c-kitpos CSCs isolated from adult rat hearts. When c-kitpos GATA-4 high expressing clonal CSCs (cCSCs were co-cultured with adult rat ventricular cardiomyocytes, we observed increased survival and contractility of the cardiomyocytes, compared to cardiomyocytes cultured alone, co-cultured with fibroblasts or c-kitpos GATA-4 low expressing cCSCs. When analysed by ELISA, the concentration of IGF-1 was significantly increased in the c-kitpos GATA-4 high cCSC/cardiomyocyte co-cultures and there was a significant correlation between IGF-1 concentration and cardiomyocyte survival. We showed the activation of the IGF-1 receptor and its downstream molecular targets in cardiomyocytes co-cultured with c-kitpos GATA-4 high cCSCs but not in cardiomyocytes that were cultured alone, co-cultured with fibroblasts or c-kitpos GATA-4 low cCSCs. Addition of a blocking antibody specific to the IGF-1 receptor inhibited the survival of cardiomyocytes and prevented the activation of its signalling in cardiomyocytes in the c-kitpos GATA-4 high cCSC/cardiomyocyte co-culture system. IGF-1 supplementation or IGF-1 high conditioned medium taken from the co-culture of c-kitpos GATA-4 high cCSCs plus cardiomyocytes did extend the survival and contractility of cardiomyocytes cultured alone and cardiomyocytes co-cultured with c-kitpos GATA-4 low cCSCs.c-kitpos GATA-4 high cCSCs exert a paracrine survival effect on cardiomyocytes through induction of the IGF-1R and signalling pathway.

  13. Synergistic role of 5-azacytidine and ascorbic acid in directing cardiosphere derived cells to cardiomyocytes in vitro by downregulating Wnt signaling pathway via phosphorylation of β-catenin.

    Directory of Open Access Journals (Sweden)

    Reddy Sailaja Mundre

    Full Text Available Cardiosphere derived cells (CDCs represent a valuable source in stem cell based therapy for cardiovascular diseases, yet poor differentiation rate hinders the transplantation efficiency. The aim of this study is to check the ability of 5-Azacytidine (Aza alone and in combination with ascorbic acid (Aza+AA in delineating CDCs to cardiomyogenesis and the underlying Wnt signaling mechanism in induced differentiation.CDCs were treated with Aza and Aza+AA for a period of 14 days to examine the expression of cardiac specific markers and Wnt downstream regulators by immunofluorescence, real time PCR and western blot.Results revealed that Aza+AA induced efficient commitment of CDCs to cardiomyogenic lineage. Immunofluorescence analysis showed significant augment for Nkx 2.5, GATA 4 and α-Sarcomeric actinin markers in Aza+AA group than control group (p = 0.0118, p = 0.009 and p = 0.0091, respectively. Relative upregulation of cardiac markers, Nkx 2.5 (p = 0.0156, GATA 4 (p = 0.0087 and down regulation of Wnt markers, β-catenin (p = 0.0107 and Cyclin D1 (p = 0. 0116 in Aza+AA group was revealed by RNA expression analysis. Moreover, the Aza+AA induced prominent expression of GATA 4, α-Sarcomeric actinin and phospho β-catenin while non phospho β-catenin and Cyclin D1 expression was significantly suppressed as displayed in protein expression analysis. Generation of spontaneous beating in Aza+AA treated CDCs further reinforced that Aza+AA accelerates the cardiomyogenic potential of CDCs.Combined treatment of Aza along with AA implicit in inducing cardiomyogenic potential of CDCs and is associated with down regulating Wnt signaling pathway. Altogether, CDCs represent a valuable tool for the treatment of cardiovascular disorders.

  14. Hexarelin Protects Rodent Pancreatic Β-Cells Function from Cytotoxic Effects of Streptozotocin Involving Mitochondrial Signalling Pathways In Vivo and In Vitro.

    Directory of Open Access Journals (Sweden)

    Yan Zhao

    Full Text Available Mitochondrial functions are crucial for pancreatic β-cell survival and glucose-induced insulin secretion. Hexarelin (Hex is a synthetic small peptide ghrelin analogue, which has been shown to protect cardiomyocytes from the ischemia-reperfusion process. In this study, we used in vitro and in vivo models of streptozotocin (STZ-induced β-cell damage to study the protective effect of Hex and the associated mechanisms. We found that STZ produced a cytotoxic effect in a dose- and time-dependent manner in MIN6 cells (a mouse β-cell line. Hex (1.0 μM decreased the STZ-induced damage in β-cells. Rhodamine 123 assay and superoxide DHE production assay revealed that Hex ameliorated STZ-induced mitochondrial damage and excessive superoxide activity in β-cells. In addition, Hex significantly reduced STZ-induced expression of cleaved Caspases-3, Caspases-9 and the ratio of pro-apoptotic protein Bax to anti-apoptotic protein Bcl-2 in MIN6 cells. We further examined the in vivo effect of Hex in a rat model of type 1 diabetes induced by STZ injection. Hex ameliorated STZ-induced decrease in plasma insulin and protected the structure of islets from STZ-induced disruption. Hex also ameliorated STZ-induced expression of cleaved Caspase-9 and the Bax in β-cells. In conclusion, our data indicate that Hex is able to protects β-cell mass from STZ-caused cytotoxic effects involving mitochondrial pathways in vitro and in vivo. Hex may serve as a potential protective agent for the management of diabetes.

  15. [Over-expression of BDNF inhibits angiotensin II-induced apoptosis of cardiomyocytes in SD rats].

    Science.gov (United States)

    Cao, Jingli; Wu, Yingfeng; Liu, Geming; Li, Zhenlong

    2018-03-01

    Objective To investigate the role and molecular mechanism of brain-derived neurotrophic factor (BDNF) against the process of cardiomyocyte hypertrophy and apoptosis. Methods Cardiomyocyte hypertrophy were estabolished by angiotensin II (Ang II) in neonatal cardiomyocytes in vitro and incomplete ligature of abdominal aorta of SD rats in vivo. BDNF over-expressing recombinant vector pcDNA5-BDNF was transfected into cardiomyocytes by liposomes. Immunofluorescence staining was used to detect the effect of BDNF transfection on the surface area of myocardial cells. The effect of BDNF transfection on the apoptosis of cardiomyocytes was assayed by flow cytometry. Real-time fluorescent quantitative PCR was performed to detect the effect of over-expression of BDNF on the expressions of atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP) mRNAs in cardiomyocytes. Western blot assay was used to observe the changes of BDNF, ANP and BNP, calmodulin kinase 2 (CaMK2) and phosphorylated calmodulin kinase 2 (p-CaMK2), calcineurin (CaN), p-CaN, nuclear factor of activated T cells 3 (NFATC3) and p-NFATC3 protein expressions in the myocardial tissues and cardiomyocytes. Results The expression of BDNF protein increased significantly in cardiac hypertrophy animal and cell models in a time-dependent manner. Compared with the untransfected control cardiomyocytes, the surface area of cardiomyocytes, the rate of apoptosis, the levels of ANP and BNP mRNA and protein expression, the levels of p-CaMK2 and CaN protein in the BDNF over-expressed cardiomyocytes were remarkably reduced, while the level of p-NFATC3 protein rose significantly. Conclusion BDNF inhibits the apoptosis of cardiomyocytes induced by Ang II, and it plays the role by inhibiting CaMK2 and CaN signaling pathways.

  16. KLF5 overexpression attenuates cardiomyocyte inflammation induced by oxygen-glucose deprivation/reperfusion through the PPARγ/PGC-1α/TNF-α signaling pathway.

    Science.gov (United States)

    Li, Yang; Li, Jian; Hou, Zhiwen; Yu, Yang; Yu, Bo

    2016-12-01

    The primary physiological function of Krüppel-like zinc-finger transcription factor (KLF5) is the regulation of cardiovascular remodeling. Vascular remodeling is closely related to the amelioration of various ischemic diseases. However, the underlying correlation of KLF5 and ischemia is not clear. In this study, we aim to investigate the role of KLF5 in myocardial ischemia reperfusion (IR) injury and the potential mechanisms involved. Cultured H9C2 cells were subjected to oxygen-glucose deprivation/reperfusion (OGD/Rep) to mimic myocardial IR injury in vivo. Expressions of KLF5 and PPARγ were distinctly inhibited, and PGC-1α expression was activated at 24h after myocardial OGD/Rep injury. After myocardial OGD/Rep injury, we found that KLF5 overexpression down-regulated levels of TNF-α, IL-1β, IL-6 and IL-8. Through the analysis of lactate dehydrogenase (LDH) release, we demonstrate that KLF5 overexpression reduced the release of OGD/Rep-induced LDH. KLF5 overexpression significantly enhanced cell activity and decreased cell apoptosis during OGD/Rep injury. Compared with the OGD/Rep group, cells overexpressing KLF5 showed anti-apoptotic effects, such as decreased expression of Bax and cleaved caspase-3 as well as increased Bcl-2 expression. KLF5 overexpression activated PPARγ, a protein involved in OGD/Rep injury, and increased levels of PGC-1α, while TNF-α expression was remarkably inhibited. In addition, GW9662, a PPARγ receptor antagonist, reversed the expression of PPARγ/PGC-1α/TNF-α and cell activity induced by KLF5 overexpression. The effects of KLF5 overexpression on PPARγ/PGC-1α/TNF-α and cell activity were abolished by co-treatment with GW9662. Taken together, these results suggest that KLF5 can efficiently alleviate OGD/Rep-induced myocardial injury, perhaps through regulation of the PPARγ/PGC-1α/TNF-α pathway. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  17. New Jersey's experience with implementing Protective Action Guides during the 1988 Salem ingestion pathway exercise

    International Nuclear Information System (INIS)

    White, Duncan

    1989-01-01

    On November 30 and December 1, 1988, the New Jersey Department of Environmental Protection (DEP) and three other State agencies (Health, Agriculture and State Police) participated in the ingestion pathway portion of the 1988 Salem Nuclear Generating Station Emergency Exercise. The purpose of this phase of the exercise was to demonstrate the ingestion pathway components of the State's Radiological Emergency Response Plan (RERP) to the Federal Emergency Management Agency (FEMA). The intent of this paper is to provide a summary of difficulties and some lessons learned in implementing the DEP's ingestion pathway Protective Action Guides (PAGs) during the exercise as well as during the preparation of a total population dose estimate (TPDE)

  18. New Jersey's experience with implementing Protective Action Guides during the 1988 Salem ingestion pathway exercise

    Energy Technology Data Exchange (ETDEWEB)

    White, Duncan [New Jersey Department of Environmental Protection, Trenton, NJ (United States)

    1989-09-01

    On November 30 and December 1, 1988, the New Jersey Department of Environmental Protection (DEP) and three other State agencies (Health, Agriculture and State Police) participated in the ingestion pathway portion of the 1988 Salem Nuclear Generating Station Emergency Exercise. The purpose of this phase of the exercise was to demonstrate the ingestion pathway components of the State's Radiological Emergency Response Plan (RERP) to the Federal Emergency Management Agency (FEMA). The intent of this paper is to provide a summary of difficulties and some lessons learned in implementing the DEP's ingestion pathway Protective Action Guides (PAGs) during the exercise as well as during the preparation of a total population dose estimate (TPDE)

  19. Pathways to Institutionalizing and Extending Social Protection in Asia

    International Development Research Centre (IDRC) Digital Library (Canada)

    In the aftermath of the Asian financial crisis (1999), the Ford Foundation initiated a research program on Social Protection in Asia. This program contributed to a growing ... to improve lives and livelihoods. Five world-class research teams are working to develop vaccines for neglected livestock diseases in the Global South.

  20. Glucocorticoid Induced Leucine Zipper inhibits apoptosis of cardiomyocytes by doxorubicin

    International Nuclear Information System (INIS)

    Aguilar, David; Strom, Joshua; Chen, Qin M.

    2014-01-01

    Doxorubicin (Dox) is an indispensable chemotherapeutic agent for the treatment of various forms of neoplasia such as lung, breast, ovarian, and bladder cancers. Cardiotoxicity is a major concern for patients receiving Dox therapy. Previous work from our laboratory indicated that glucocorticoids (GCs) alleviate Dox-induced apoptosis in cardiomyocytes. Here we have found Glucocorticoid-Induced Leucine Zipper (GILZ) to be a mediator of GC-induced cytoprotection. GILZ was found to be induced in cardiomyocytes by GC treatment. Knocking down of GILZ using siRNA resulted in cancelation of GC-induced cytoprotection against apoptosis by Dox treatment. Overexpressing GILZ by transfection was able to protect cells from apoptosis induced by Dox as measured by caspase activation, Annexin V binding and morphologic changes. Western blot analyses indicate that GILZ overexpression prevented cytochrome c release from mitochondria and cleavage of caspase-3. When bcl-2 family proteins were examined, we found that GILZ overexpression causes induction of the pro-survival protein Bcl-xL. Since siRNA against Bcl-xL reverses GC induced cytoprotection, Bcl-xL induction represents an important event in GILZ-induced cytoprotection. Our data suggest that GILZ functions as a cytoprotective gene in cardiomyocytes. - Highlights: • Corticosteroids act as a cytoprotective agent in cardiomyocytes • Corticosteroids induce GILZ expression in cardiomyocytes • Elevated GILZ results in resistance against apoptosis induced by doxorubicin • GILZ induces Bcl-xL protein without inducing Bcl-xL mRNA

  1. Protecting Critical Infrastructure by Identifying Pathways of Exposure to Risk

    Directory of Open Access Journals (Sweden)

    Philip O’Neill

    2013-08-01

    Full Text Available Increasingly, our critical infrastructure is managed and controlled by computers and the information networks that connect them. Cyber-terrorists and other malicious actors understand the economic and social impact that a successful attack on these systems could have. While it is imperative that we defend against such attacks, it is equally imperative that we realize how best to react to them. This article presents the strongest-path method of analyzing all potential pathways of exposure to risk – no matter how indirect or circuitous they may be – in a network model of infrastructure and operations. The method makes direct use of expert knowledge about entities and dependency relationships without the need for any simulation or any other models. By using path analysis in a directed graph model of critical infrastructure, planners can model and assess the effects of a potential attack and develop resilient responses.

  2. Coping capacities for improving adaptation pathways for flood protection in Can Tho, Vietnam

    Science.gov (United States)

    Pathirana, A.; Radhakrishnan, M.; Quan, N. H.; Gersonius, B.; Ashley, R.; Zevenbergen, C.

    2016-12-01

    Studying the evolution of coping and adaptation capacities is a prerequisite for preparing an effective flood management plan for the future, especially in the dynamic and fast changing cities of developing countries. The objectives, requirements, targets, design and performance of flood protection measures will have to be determined after taking into account, or in conjunction with, the coping capacities. A methodology is presented based on adaptation pathways to account for coping capacities and to assess the effect on flood protection measures. The adaptation pathways method determines the point of failure of a particular strategy based on the change in an external driver, a point in time or a socio economic situation where / at which the strategy can no longer meet its objective. Pathways arrived at based on this methodology reflect future reality by considering changing engineering standards along with future uncertainties, risk taking abilities and adaptation capacities. This pathways based methodology determines the Adaptation tipping points (ATP), `time of occurrence of ATP' of flood protection measures after accounting for coping capacities, evaluates the measures and then provides the means to determine the adaptation pathways. Application of this methodology for flood protection measures in Can Tho city in the Mekong delta reveals the effect of coping capacity on the usefulness of flood protection measures and the delay in occurrence of tipping points. Consideration of coping capacity in the system owing to elevated property floor levels lead to the postponement of tipping points and improved the adaptation pathways comprising flood protection measures such as dikes. This information is useful to decision makers for planning and phasing of investments in flood protection.

  3. Oxidative stress and cardiomyocyte necrosis with elevated serum troponins: pathophysiologic mechanisms.

    Science.gov (United States)

    Robinson, Antwon D; Ramanathan, Kodangudi B; McGee, Jesse E; Newman, Kevin P; Weber, Karl T

    2011-08-01

    The progressive nature of heart failure is linked to multiple factors, including an ongoing loss of cardiomyocytes and necrosis. Necrotic cardiomyocytes leave behind several footprints: the spillage of their contents leading to elevations in serum troponins; and morphologic evidence of tissue repair with scarring. The pathophysiologic origins of cardiomyocyte necrosis relates to neurohormonal activation, including the adrenergic nervous system. Catecholamine-initiated excessive intracellular Ca accumulation and mitochondria Ca overloading in particular initiate a mitochondriocentric signal-transducer-effector pathway to necrosis and which includes the induction of oxidative stress and opening of their inner membrane permeability transition pore. Hypokalemia, ionized hypocalcemia and hypomagnesemia, where consequent elevations in parathyroid hormone further account for excessive intracellular Ca accumulation, hypozincemia and hyposelenemia each compromise metalloenzyme-based antioxidant defenses. The necrotic loss of cardiomyocytes and adverse structural remodeling of myocardium is related to the central role played by a mitochondriocentric pathway initiated by neurohormonal activation.

  4. Doxorubicin Blocks Cardiomyocyte Autophagic Flux by Inhibiting Lysosome Acidification.

    Science.gov (United States)

    Li, Dan L; Wang, Zhao V; Ding, Guanqiao; Tan, Wei; Luo, Xiang; Criollo, Alfredo; Xie, Min; Jiang, Nan; May, Herman; Kyrychenko, Viktoriia; Schneider, Jay W; Gillette, Thomas G; Hill, Joseph A

    2016-04-26

    The clinical use of doxorubicin is limited by cardiotoxicity. Histopathological changes include interstitial myocardial fibrosis and the appearance of vacuolated cardiomyocytes. Whereas dysregulation of autophagy in the myocardium has been implicated in a variety of cardiovascular diseases, the role of autophagy in doxorubicin cardiomyopathy remains poorly defined. Most models of doxorubicin cardiotoxicity involve intraperitoneal injection of high-dose drug, which elicits lethargy, anorexia, weight loss, and peritoneal fibrosis, all of which confound the interpretation of autophagy. Given this, we first established a model that provokes modest and progressive cardiotoxicity without constitutional symptoms, reminiscent of the effects seen in patients. We report that doxorubicin blocks cardiomyocyte autophagic flux in vivo and in cardiomyocytes in culture. This block was accompanied by robust accumulation of undegraded autolysosomes. We go on to localize the site of block as a defect in lysosome acidification. To test the functional relevance of doxorubicin-triggered autolysosome accumulation, we studied animals with diminished autophagic activity resulting from haploinsufficiency for Beclin 1. Beclin 1(+/-) mice exposed to doxorubicin were protected in terms of structural and functional changes within the myocardium. Conversely, animals overexpressing Beclin 1 manifested an amplified cardiotoxic response. Doxorubicin blocks autophagic flux in cardiomyocytes by impairing lysosome acidification and lysosomal function. Reducing autophagy initiation protects against doxorubicin cardiotoxicity. © 2016 American Heart Association, Inc.

  5. Cation dyshomeostasis and cardiomyocyte necrosis: the Fleckenstein hypothesis revisited

    Science.gov (United States)

    Borkowski, Brian J.; Cheema, Yaser; Shahbaz, Atta U.; Bhattacharya, Syamal K.; Weber, Karl T.

    2011-01-01

    An ongoing loss of cardiomyocytes to apoptotic and necrotic cell death pathways contributes to the progressive nature of heart failure. The pathophysiological origins of necrotic cell loss relate to the neurohormonal activation that accompanies acute and chronic stressor states and which includes effector hormones of the adrenergic nervous system. Fifty years ago, Albrecht Fleckenstein and coworkers hypothesized the hyperadrenergic state, which accompanies such stressors, causes cardiomyocyte necrosis based on catecholamine-initiated excessive intracellular Ca2+ accumulation (EICA), and mitochondrial Ca2+ overloading in particular, in which the ensuing dysfunction and structural degeneration of these organelles leads to necrosis. In recent years, two downstream factors have been identified which, together with EICA, constitute a signal–transducer–effector pathway: (i) mitochondria-based induction of oxidative stress, in which the rate of reactive oxygen metabolite generation exceeds their rate of detoxification by endogenous antioxidant defences; and (ii) the opening of the mitochondrial inner membrane permeability transition pore (mPTP) followed by organellar swelling and degeneration. The pathogenesis of stress-related cardiomyopathy syndromes is likely related to this pathway. Other factors which can account for cytotoxicity in stressor states include: hypokalaemia; ionized hypocalcaemia and hypomagnesaemia with resultant elevations in parathyroid hormone serving as a potent mediator of EICA; and hypozincaemia with hyposelenaemia, which compromise antioxidant defences. Herein, we revisit the Fleckenstein hypothesis of EICA in leading to cardiomyocyte necrosis and the central role played by mitochondria. PMID:21398641

  6. Sirtuin 6 protects the heart from hypoxic damage

    International Nuclear Information System (INIS)

    Maksin-Matveev, Anna; Kanfi, Yariv; Hochhauser, Edith; Isak, Ahuva; Cohen, Haim Y.; Shainberg, Asher

    2015-01-01

    Sirtuin 6 (SIRT6) is a protein associated with prolonged life expectancy. We investigated whether life extension is associated with cardioprotection against hypoxia. The proposed study is to develop approaches to reduce hypoxic damage through the use of the sirtuin pathway and to elucidate the mechanism involved. For that purpose we subjected cardiomyocytes from transgenic mice (TG) with over-expression of SIRT6, to hypoxic stress in cell cultures. We hypothesized that cardiomyocytes from transgenic mice subjected to prolonged hypoxia may release survival factors or fewer damage markers to protect them from hypoxic stress compared with wild type (WT) mice. Lactate dehydrogenase (LDH) and creatine kinase (CK) released to the medium and propidium iodide (PI) binding, were markedly decreased following hypoxia in TG cardiomyocytes. The protective mechanism of SIRT6 over-expression includes the activation of pAMPKα pathway, the increased protein level of B-cell lymphoma 2 (Bcl2), the inhibition of nuclear factor kappa-light-chain-enhancer of activated B cells (NFκB), the decrease of reactive oxygen species (ROS) and the reduction in the protein level of phospho-protein kinase B (pAkt) during hypoxia. Together, all these processes impede the necrosis/apoptosis pathways leading to the improved survival of cardiomyocytes following hypoxia, which might explain life extension. - Highlights: • Sirtuin 6 is a protein associated with prolonged life expectancy. • Over-expression of sirtuin 6 protects cardiocytes from hypoxia and oxidative stress. • Over-expression of sirtuin 6 activates the pAMPKα pathway and the Bcl2 expression. • Over-expression of sirtuin 6 decreases ROS formation and pAkt level during hypoxia. • These pathways protect cardiocytes from hypoxia and might explain lifespan extension

  7. Sirtuin 6 protects the heart from hypoxic damage

    Energy Technology Data Exchange (ETDEWEB)

    Maksin-Matveev, Anna; Kanfi, Yariv [The Mina and Everard Goodman, Faculty of Life Sciences, Bar-Ilan University, Ramat Gan 52900 (Israel); Hochhauser, Edith [The Laboratory of the Department of Cardiothoracic Surgery, Felsenstein Medical Research Center, Rabin Medical Center, Petach Tikva (Israel); Isak, Ahuva; Cohen, Haim Y. [The Mina and Everard Goodman, Faculty of Life Sciences, Bar-Ilan University, Ramat Gan 52900 (Israel); Shainberg, Asher, E-mail: asher.shainberg@gmail.com [The Mina and Everard Goodman, Faculty of Life Sciences, Bar-Ilan University, Ramat Gan 52900 (Israel)

    2015-01-01

    Sirtuin 6 (SIRT6) is a protein associated with prolonged life expectancy. We investigated whether life extension is associated with cardioprotection against hypoxia. The proposed study is to develop approaches to reduce hypoxic damage through the use of the sirtuin pathway and to elucidate the mechanism involved. For that purpose we subjected cardiomyocytes from transgenic mice (TG) with over-expression of SIRT6, to hypoxic stress in cell cultures. We hypothesized that cardiomyocytes from transgenic mice subjected to prolonged hypoxia may release survival factors or fewer damage markers to protect them from hypoxic stress compared with wild type (WT) mice. Lactate dehydrogenase (LDH) and creatine kinase (CK) released to the medium and propidium iodide (PI) binding, were markedly decreased following hypoxia in TG cardiomyocytes. The protective mechanism of SIRT6 over-expression includes the activation of pAMPKα pathway, the increased protein level of B-cell lymphoma 2 (Bcl2), the inhibition of nuclear factor kappa-light-chain-enhancer of activated B cells (NFκB), the decrease of reactive oxygen species (ROS) and the reduction in the protein level of phospho-protein kinase B (pAkt) during hypoxia. Together, all these processes impede the necrosis/apoptosis pathways leading to the improved survival of cardiomyocytes following hypoxia, which might explain life extension. - Highlights: • Sirtuin 6 is a protein associated with prolonged life expectancy. • Over-expression of sirtuin 6 protects cardiocytes from hypoxia and oxidative stress. • Over-expression of sirtuin 6 activates the pAMPKα pathway and the Bcl2 expression. • Over-expression of sirtuin 6 decreases ROS formation and pAkt level during hypoxia. • These pathways protect cardiocytes from hypoxia and might explain lifespan extension.

  8. Modelling plant invasion pathways in protected areas under climate change: implication for invasion management

    Directory of Open Access Journals (Sweden)

    C.-J. Wang

    2017-12-01

    Full Text Available Global climate change may enable invasive plant species (IPS to invade protected areas (PAs, but plant invasion on a global scale has not yet been explicitly addressed. Here, we mapped the potential invasion pathways for IPS in PAs across the globe and explored potential factors determining the pathways of plant invasion under climate change. We used species distribution modelling to estimate the suitable habitats of 386 IPS and applied a corridor analysis to compute the potential pathways of IPS in PAs under climate change. Subsequently, we analysed the potential factors affecting the pathways in PAs. According to our results, the main potential pathways of IPS in PAs are in Europe, eastern Australia, New Zealand, southern Africa, and eastern regions of South America and are strongly influenced by changes in temperature and precipitation. Protected areas can play an important role in preventing and controlling the spread of IPS under climate change. This is due to the fact that measures are taken to monitor climate change in detail, to provide effective management near or inside PAs, and to control the introduction of IPS with a high capacity for natural dispersal. A review of conservation policies in PAs is urgently needed.

  9. Cardiomyocyte specific expression of Acyl-coA thioesterase 1 attenuates sepsis induced cardiac dysfunction and mortality

    Energy Technology Data Exchange (ETDEWEB)

    Xia, Congying [Departments of Internal Medicine and Institute of Hypertension, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan (China); Dong, Ruolan [Department of Geriatric Medicine, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030 (China); Chen, Chen [Departments of Internal Medicine and Institute of Hypertension, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan (China); Wang, Hong, E-mail: hong.wang1988@yahoo.com [Departments of Internal Medicine and Institute of Hypertension, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan (China); Wang, Dao Wen, E-mail: dwwang@tjh.tjmu.edu.cn [Departments of Internal Medicine and Institute of Hypertension, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan (China)

    2015-12-25

    Compromised cardiac fatty acid oxidation (FAO) induced energy deprivation is a critical cause of cardiac dysfunction in sepsis. Acyl-CoA thioesterase 1 (ACOT1) is involved in regulating cardiac energy production via altering substrate metabolism. This study aims to clarify whether ACOT1 has a potency to ameliorate septic myocardial dysfunction via enhancing cardiac FAO. Transgenic mice with cardiomyocyte specific expression of ACOT1 (αMHC-ACOT1) and their wild type (WT) littermates were challenged with Escherichia coli lipopolysaccharide (LPS; 5 mg/kg i.p.) and myocardial function was assessed 6 h later using echocardiography and hemodynamics. Deteriorated cardiac function evidenced by reduction of the percentage of left ventricular ejection fraction and fractional shortening after LPS administration was significantly attenuated by cardiomyocyte specific expression of ACOT1. αMHC-ACOT1 mice exhibited a markedly increase in glucose utilization and cardiac FAO compared with LPS-treated WT mice. Suppression of cardiac peroxisome proliferator activated receptor alpha (PPARa) and PPARγ-coactivator-1α (PGC1a) signaling observed in LPS-challenged WT mice was activated by the presence of ACOT1. These results suggest that ACOT1 has potential therapeutic values to protect heart from sepsis mediated dysfunction, possibly through activating PPARa/PGC1a signaling. - Highlights: • ACOT1 has potential therapeutic values to protect heart from sepsis mediated dysfunction. • ACOT1 can regulate PPARa/PGC1a signaling pathway. • We first generate the transgenic mice with cardiomyocyte specific expression of ACOT1.

  10. Cardiomyocyte specific expression of Acyl-coA thioesterase 1 attenuates sepsis induced cardiac dysfunction and mortality

    International Nuclear Information System (INIS)

    Xia, Congying; Dong, Ruolan; Chen, Chen; Wang, Hong; Wang, Dao Wen

    2015-01-01

    Compromised cardiac fatty acid oxidation (FAO) induced energy deprivation is a critical cause of cardiac dysfunction in sepsis. Acyl-CoA thioesterase 1 (ACOT1) is involved in regulating cardiac energy production via altering substrate metabolism. This study aims to clarify whether ACOT1 has a potency to ameliorate septic myocardial dysfunction via enhancing cardiac FAO. Transgenic mice with cardiomyocyte specific expression of ACOT1 (αMHC-ACOT1) and their wild type (WT) littermates were challenged with Escherichia coli lipopolysaccharide (LPS; 5 mg/kg i.p.) and myocardial function was assessed 6 h later using echocardiography and hemodynamics. Deteriorated cardiac function evidenced by reduction of the percentage of left ventricular ejection fraction and fractional shortening after LPS administration was significantly attenuated by cardiomyocyte specific expression of ACOT1. αMHC-ACOT1 mice exhibited a markedly increase in glucose utilization and cardiac FAO compared with LPS-treated WT mice. Suppression of cardiac peroxisome proliferator activated receptor alpha (PPARa) and PPARγ-coactivator-1α (PGC1a) signaling observed in LPS-challenged WT mice was activated by the presence of ACOT1. These results suggest that ACOT1 has potential therapeutic values to protect heart from sepsis mediated dysfunction, possibly through activating PPARa/PGC1a signaling. - Highlights: • ACOT1 has potential therapeutic values to protect heart from sepsis mediated dysfunction. • ACOT1 can regulate PPARa/PGC1a signaling pathway. • We first generate the transgenic mice with cardiomyocyte specific expression of ACOT1.

  11. Measuring Fast Calcium Fluxes in Cardiomyocytes

    Science.gov (United States)

    Golebiewska, Urszula; Scarlata, Suzanne

    2011-01-01

    Cardiomyocytes have multiple Ca2+ fluxes of varying duration that work together to optimize function 1,2. Changes in Ca2+ activity in response to extracellular agents is predominantly regulated by the phospholipase Cβ- Gαq pathway localized on the plasma membrane which is stimulated by agents such as acetylcholine 3,4. We have recently found that plasma membrane protein domains called caveolae5,6 can entrap activated Gαq7. This entrapment has the effect of stabilizing the activated state of Gαq and resulting in prolonged Ca2+ signals in cardiomyocytes and other cell types8. We uncovered this surprising result by measuring dynamic calcium responses on a fast scale in living cardiomyocytes. Briefly, cells are loaded with a fluorescent Ca2+ indicator. In our studies, we used Ca2+ Green (Invitrogen, Inc.) which exhibits an increase in fluorescence emission intensity upon binding of calcium ions. The fluorescence intensity is then recorded for using a line-scan mode of a laser scanning confocal microscope. This method allows rapid acquisition of the time course of fluorescence intensity in pixels along a selected line, producing several hundreds of time traces on the microsecond time scale. These very fast traces are transferred into excel and then into Sigmaplot for analysis, and are compared to traces obtained for electronic noise, free dye, and other controls. To dissect Ca2+ responses of different flux rates, we performed a histogram analysis that binned pixel intensities with time. Binning allows us to group over 500 traces of scans and visualize the compiled results spatially and temporally on a single plot. Thus, the slow Ca2+ waves that are difficult to discern when the scans are overlaid due to different peak placement and noise, can be readily seen in the binned histograms. Very fast fluxes in the time scale of the measurement show a narrow distribution of intensities in the very short time bins whereas longer Ca2+ waves show binned data with a broad

  12. SIRT1 Suppresses Doxorubicin-Induced Cardiotoxicity by Regulating the Oxidative Stress and p38MAPK Pathways

    Directory of Open Access Journals (Sweden)

    Yang Ruan

    2015-02-01

    Full Text Available Background: SIRT1, which belongs to the Sirtuin family of NAD-dependent enzymes, plays diverse roles in aging, metabolism, and disease biology. It could regulate cell survival and has been shown to be a protective factor in heart function. Hence, we verified the mechanism by which SIRT1 regulates doxorubicin induced cardiomyocyte injury in vivo and in vitro. Methods: We analyzed SIRT1 expression in doxorubicin-induced neonatal rat cardiomyocyte injury model and adult mouse heart failure model. SIRT1 was over-expressed in cultured neonatal rat cardiomyocyte by adenovirus mediated gene transfer. SIRT1 agonist resveratrol was used to treat the doxorubicin-induced heart failure mouse model. Echocardiography, reactive oxygen species (ROS production, TUNEL, qRT-PCR, and Western blotting were performed to analyze cell survival, oxidative stress, and inflammatory signal pathways in cardiomyocytes. Results: SIRT1 expression was down-regulated in doxorubicin induced cardiomocyte injury, accompanied by elevated oxidative stress and cell apoptosis. SIRT1 over-expression reduced doxorubicin induced cardiomyocyte apoptosis with the attenuated ROS production. SIRT1 also reduced cell apoptosis by inhibition of p38MAPK phosphorylation and caspase-3 activation. The SIRT1 agonist resveratrol was able to prevent doxorubicin-induced heart function loss. Moreover, the SIRT1 inhibitor niacinamide could reverse SIRT1's protective effect in cultured neonatal rat cardiomyocytes. Conclusions: These results support the role of SIRT1 as an important regulator of cardiomyocyte apoptosis during doxorubicin-induced heart injury, which may represent a potential therapeutic target for doxorubicin-induced cardiomyopathy.

  13. Distinct functions for the Drosophila piRNA pathway in genome maintenance and telomere protection.

    Directory of Open Access Journals (Sweden)

    Jaspreet S Khurana

    2010-12-01

    Full Text Available Transposons and other selfish DNA elements can be found in all phyla, and mobilization of these elements can compromise genome integrity. The piRNA (PIWI-interacting RNA pathway silences transposons in the germline, but it is unclear if this pathway has additional functions during development. Here we show that mutations in the Drosophila piRNA pathway genes, armi, aub, ago3, and rhi, lead to extensive fragmentation of the zygotic genome during the cleavage stage of embryonic divisions. Additionally, aub and armi show defects in telomere resolution during meiosis and the cleavage divisions; and mutations in lig-IV, which disrupt non-homologous end joining, suppress these fusions. By contrast, lig-IV mutations enhance chromosome fragmentation. Chromatin immunoprecipitation studies show that aub and armi mutations disrupt telomere binding of HOAP, which is a component of the telomere protection complex, and reduce expression of a subpopulation of 19- to 22-nt telomere-specific piRNAs. Mutations in rhi and ago3, by contrast, do not block HOAP binding or production of these piRNAs. These findings uncover genetically separable functions for the Drosophila piRNA pathway. The aub, armi, rhi, and ago3 genes silence transposons and maintain chromosome integrity during cleavage-stage embryonic divisions. However, the aub and armi genes have an additional function in assembly of the telomere protection complex.

  14. Identification of signaling pathways associated with cancer protection in Laron syndrome.

    Science.gov (United States)

    Lapkina-Gendler, Lena; Rotem, Itai; Pasmanik-Chor, Metsada; Gurwitz, David; Sarfstein, Rive; Laron, Zvi; Werner, Haim

    2016-05-01

    The growth hormone (GH)-insulin-like growth factor-1 (IGF1) pathway emerged in recent years as a critical player in cancer biology. Enhanced expression or activation of specific components of the GH-IGF1 axis, including the IGF1 receptor (IGF1R), is consistently associated with a transformed phenotype. Recent epidemiological studies have shown that patients with Laron syndrome (LS), the best-characterized entity among the congenital IGF1 deficiencies, seem to be protected from cancer development. To identify IGF1-dependent genes and signaling pathways associated with cancer protection in LS, we conducted a genome-wide analysis using immortalized lymphoblastoid cells derived from LS patients and healthy controls of the same gender, age range, and ethnic origin. Our analyses identified a collection of genes that are either over- or under-represented in LS-derived lymphoblastoids. Gene differential expression occurs in several gene families, including cell cycle, metabolic control, cytokine-cytokine receptor interaction, Jak-STAT signaling, and PI3K-AKT signaling. Major differences between LS and healthy controls were also noticed in pathways associated with cell cycle distribution, apoptosis, and autophagy. Our results highlight the key role of the GH-IGF1 axis in the initiation and progression of cancer. Furthermore, data are consistent with the concept that homozygous congenital IGF1 deficiency may confer protection against future tumor development. © 2016 Society for Endocrinology.

  15. Caffeine exposure alters cardiac gene expression in embryonic cardiomyocytes

    Science.gov (United States)

    Fang, Xiefan; Mei, Wenbin; Barbazuk, William B.; Rivkees, Scott A.

    2014-01-01

    Previous studies demonstrated that in utero caffeine treatment at embryonic day (E) 8.5 alters DNA methylation patterns, gene expression, and cardiac function in adult mice. To provide insight into the mechanisms, we examined cardiac gene and microRNA (miRNA) expression in cardiomyocytes shortly after exposure to physiologically relevant doses of caffeine. In HL-1 and primary embryonic cardiomyocytes, caffeine treatment for 48 h significantly altered the expression of cardiac structural genes (Myh6, Myh7, Myh7b, Tnni3), hormonal genes (Anp and BnP), cardiac transcription factors (Gata4, Mef2c, Mef2d, Nfatc1), and microRNAs (miRNAs; miR208a, miR208b, miR499). In addition, expressions of these genes were significantly altered in embryonic hearts exposed to in utero caffeine. For in utero experiments, pregnant CD-1 dams were treated with 20–60 mg/kg of caffeine, which resulted in maternal circulation levels of 37.3–65.3 μM 2 h after treatment. RNA sequencing was performed on embryonic ventricles treated with vehicle or 20 mg/kg of caffeine daily from E6.5-9.5. Differential expression (DE) analysis revealed that 124 genes and 849 transcripts were significantly altered, and differential exon usage (DEU) analysis identified 597 exons that were changed in response to prenatal caffeine exposure. Among the DE genes identified by RNA sequencing were several cardiac structural genes and genes that control DNA methylation and histone modification. Pathway analysis revealed that pathways related to cardiovascular development and diseases were significantly affected by caffeine. In addition, global cardiac DNA methylation was reduced in caffeine-treated cardiomyocytes. Collectively, these data demonstrate that caffeine exposure alters gene expression and DNA methylation in embryonic cardiomyocytes. PMID:25354728

  16. A novel compound DT-010 protects against doxorubicin-induced cardiotoxicity in zebrafish and H9c2 cells by inhibiting reactive oxygen species-mediated apoptotic and autophagic pathways.

    Science.gov (United States)

    Tang, Fan; Zhou, Xinhua; Wang, Liang; Shan, Luchen; Li, Chuwen; Zhou, Hefeng; Lee, Simon Ming-Yuen; Hoi, Maggie Pui-Man

    2018-02-05

    Doxorubicin (Dox) is an effective anti-cancer agent but limited by its cardiotoxicity, thus the search for pharmacological agents for enhancing anti-cancer activities and protecting against cardiotoxicity has been a subject of great interest. We have previously reported the synergistic anti-cancer effects of a novel compound DT-010. In the present study, we further investigated the cardioprotective effects of DT-010 in zebrafish embryos in vivo and the molecular underlying mechanisms in H9c2 cardiomyocytes in vitro. We showed that DT-010 prevented the Dox-induced morphological distortions in the zebrafish heart and the associated cardiac impairments, and especially improved ventricular functions. By using H9c2 cells model, we showed that DT-010 directly inhibited the generation of reactive oxygen species by Dox and protected cell death and cellular damage. We further observed that DT-010 protected against Dox-induced myocardiopathy via inhibiting downstream molecular pathways in response to oxidative stress, including reactive oxygen species-mediated MAPK signaling pathways ERK and JNK, and apoptotic pathways involving the activation of caspase 3, caspase 7, and PARP signaling. Recent studies also suggest the importance of alterations in cardiac autophagy in Dox cardiotoxicity. We further showed that DT-010 could inhibit the induction of autophagosomes formation by Dox via regulating the upstream Akt/AMPK/mTOR signaling. Since Dox-induced cardiotoxicity is multifactorial, our results suggest that multi-functional agent such as DT-010 might be an effective therapeutic agent for combating cardiotoxicity associated with chemotherapeutic agents such as Dox. Copyright © 2017 Elsevier B.V. All rights reserved.

  17. Dedifferentiation, Proliferation, and Redifferentiation of Adult Mammalian Cardiomyocytes After Ischemic Injury.

    Science.gov (United States)

    Wang, Wei Eric; Li, Liangpeng; Xia, Xuewei; Fu, Wenbin; Liao, Qiao; Lan, Cong; Yang, Dezhong; Chen, Hongmei; Yue, Rongchuan; Zeng, Cindy; Zhou, Lin; Zhou, Bin; Duan, Dayue Darrel; Chen, Xiongwen; Houser, Steven R; Zeng, Chunyu

    2017-08-29

    Adult mammalian hearts have a limited ability to generate new cardiomyocytes. Proliferation of existing adult cardiomyocytes (ACMs) is a potential source of new cardiomyocytes. Understanding the fundamental biology of ACM proliferation could be of great clinical significance for treating myocardial infarction (MI). We aim to understand the process and regulation of ACM proliferation and its role in new cardiomyocyte formation of post-MI mouse hearts. β-Actin-green fluorescent protein transgenic mice and fate-mapping Myh6-MerCreMer-tdTomato/lacZ mice were used to trace the fate of ACMs. In a coculture system with neonatal rat ventricular myocytes, ACM proliferation was documented with clear evidence of cytokinesis observed with time-lapse imaging. Cardiomyocyte proliferation in the adult mouse post-MI heart was detected by cell cycle markers and 5-ethynyl-2-deoxyuridine incorporation analysis. Echocardiography was used to measure cardiac function, and histology was performed to determine infarction size. In vitro, mononucleated and bi/multinucleated ACMs were able to proliferate at a similar rate (7.0%) in the coculture. Dedifferentiation proceeded ACM proliferation, which was followed by redifferentiation. Redifferentiation was essential to endow the daughter cells with cardiomyocyte contractile function. Intercellular propagation of Ca 2+ from contracting neonatal rat ventricular myocytes into ACM daughter cells was required to activate the Ca 2+ -dependent calcineurin-nuclear factor of activated T-cell signaling pathway to induce ACM redifferentiation. The properties of neonatal rat ventricular myocyte Ca 2+ transients influenced the rate of ACM redifferentiation. Hypoxia impaired the function of gap junctions by dephosphorylating its component protein connexin 43, the major mediator of intercellular Ca 2+ propagation between cardiomyocytes, thereby impairing ACM redifferentiation. In vivo, ACM proliferation was found primarily in the MI border zone. An ischemia

  18. Genetic enrichment of cardiomyocytes derived from mouse ...

    African Journals Online (AJOL)

    Genetic enrichment of cardiomyocytes derived from mouse embryonic stem cells. WJ He, SC Li, LL Ye, H Liu, QW Wang, WD Han, XB Fu, ZL Chen. Abstract. Pluripotent embryonic stem cells (ESC) have the ability to differentiate into a variety of cell lineages in vitro, including cardiomyocytes. Successful applications of ...

  19. Blueberry polyphenols prevent cardiomyocyte death by preventing calpain activation and oxidative stress.

    Science.gov (United States)

    Louis, Xavier Lieben; Thandapilly, Sijo Joseph; Kalt, Wilhelmina; Vinqvist-Tymchuk, Melinda; Aloud, Basma Milad; Raj, Pema; Yu, Liping; Le, Hoa; Netticadan, Thomas

    2014-08-01

    The purpose of this study was to examine the efficacy of an aqueous wild blueberry extract and five wild blueberry polyphenol fractions on an in vitro model of heart disease. Adult rat cardiomyocytes were pretreated with extract and fractions, and then exposed to norepinephrine (NE). Cardiomyocyte hypertrophy, cell death, oxidative stress, apoptosis and cardiomyocyte contractile function as well as the activities of calpain, superoxide dismutase (SOD) and catalase (CAT) were measured in cardiomyocytes treated with and without NE and blueberry fraction (BF). Four of five blueberry fractions prevented cell death and cardiomyocyte hypertrophy induced by NE. Total phenolic fraction was used for all further analysis. The NE-induced increase in oxidative stress, nuclear condensation, calpain activity and lowering of SOD and CAT activities were prevented upon pretreatment with BF. Reduced contractile function was also significantly improved with BF pretreatment. Blueberry polyphenols prevent NE-induced adult cardiomyocyte hypertrophy and cell death. The protective effects of BF may be in part attributed to a reduction in calpain activity and oxidative stress.

  20. Evidence for Cardiomyocyte Renewal in Humans

    Energy Technology Data Exchange (ETDEWEB)

    Bergmann, O; Bhardwaj, R D; Bernard, S; Zdunek, S; Barnabe-Heider, F; Walsh, S; Zupicich, J; Alkass, K; Buchholz, B A; Druid, H; Jovinge, S; Frisen, J

    2008-10-14

    It has been difficult to establish whether we are limited to the heart muscle cells we are born with or if cardiomyocytes are generated also later in life. We have taken advantage of the integration of {sup 14}C, generated by nuclear bomb tests during the Cold War, into DNA to establish the age of cardiomyocytes in humans. We report that cardiomyocytes renew, with a gradual decrease from 1% turning over annually at the age of 20 to 0.3% at the age of 75. Less than 50% of cardiomyocytes are exchanged during a normal lifespan. The capacity to generate cardiomyocytes in the adult human heart suggests that it may be rational to work towards the development of therapeutic strategies aiming to stimulate this process in cardiac pathologies.

  1. An Ixodes ricinus Tick Salivary Lectin Pathway Inhibitor Protects Borrelia burgdorferi sensu lato from Human Complement.

    Science.gov (United States)

    Wagemakers, Alex; Coumou, Jeroen; Schuijt, Tim J; Oei, Anneke; Nijhof, Ard M; van 't Veer, Cornelis; van der Poll, Tom; Bins, Adriaan D; Hovius, Joppe W R

    2016-04-01

    We previously identified tick salivary lectin pathway inhibitor (TSLPI) in Ixodes scapularis, a vector for Borrelia burgdorferi sensu stricto (s.s.) in North America. TSLPI is a salivary protein facilitating B. burgdorferi s.s. transmission and acquisition by inhibiting the host lectin complement pathway through interference with mannose binding lectin (MBL) activity. Since Ixodes ricinus is the predominant vector for Lyme borreliosis in Europe and transmits several complement sensitive B. burgdorferi sensu lato (s.l.) strains, we aimed to identify, describe, and characterize the I. ricinus ortholog of TSLPI. We performed (q)PCRs on I. ricinus salivary gland cDNA to identify a TSLPI ortholog. Next, we generated recombinant (r)TSLPI in a Drosophila expression system and examined inhibition of the MBL complement pathway and complement-mediated killing of B. burgdorferi s.l. in vitro. We identified a TSLPI ortholog in I. ricinus salivary glands with 93% homology at the RNA and 89% at the protein level compared to I. scapularis TSLPI, which was upregulated during tick feeding. In silico analysis revealed that TSLPI appears to be part of a larger family of Ixodes salivary proteins among which I. persulcatus basic tail salivary proteins and I. scapularis TSLPI and Salp14. I. ricinus rTSLPI inhibited the MBL complement pathway and protected B. burgdorferi s.s. and Borrelia garinii from complement-mediated killing. We have identified a TSLPI ortholog, which protects B. burgdorferi s.l. from complement-mediated killing in I. ricinus, the major vector for tick-borne diseases in Europe.

  2. Cardiomyocyte mitochondrial respiration is reduced by receptor for advanced glycation end-product signaling in a ceramide-dependent manner.

    Science.gov (United States)

    Nelson, Michael B; Swensen, Adam C; Winden, Duane R; Bodine, Jared S; Bikman, Benjamin T; Reynolds, Paul R

    2015-07-01

    Cigarette smoke exposure is associated with an increased risk of cardiovascular complications. The role of advanced glycation end products (AGEs) is already well established in numerous comorbidities, including cardiomyopathy. Given the role of AGEs and their receptor, RAGE, in activating inflammatory pathways, we sought to determine whether ceramides could be a mediator of RAGE-induced altered heart mitochondrial function. Using an in vitro model, we treated H9C2 cardiomyocytes with the AGE carboxy-methyllysine before mitochondrial respiration assessment. We discovered that mitochondrial respiration was significantly impaired in AGE-treated cells, but not when cotreated with myriocin, an inhibitor of de novo ceramide biosynthesis. Moreover, we exposed wild-type and RAGE knockout mice to secondhand cigarette smoke and found reduced mitochondrial respiration in the left ventricular myocardium from wild-type mice, but RAGE knockout mice were protected from this effect. Finally, conditional overexpression of RAGE in the lungs of transgenic mice elicited a robust increase in left ventricular ceramides in the absence of smoke exposure. Taken together, these findings suggest a RAGE-ceramide axis as an important contributor to AGE-mediated disrupted cardiomyocyte mitochondrial function. Copyright © 2015 the American Physiological Society.

  3. Resveratrol, an antioxidant, protects spinal cord injury in rats by suppressing MAPK pathway

    Directory of Open Access Journals (Sweden)

    Song Fu

    2018-02-01

    Full Text Available Resveratrol, a polyphenol found in various plants, including grapes, plums and peanuts has shown various medIRInal properties, including antioxidant, protection of cardiovascular disease and cancer risk. However, the effects of resveratrol on spinal cord reperfusion injury have not been investigated. Hence, the present study was designed to evaluate the effect of resveratrol on nitric oxide synthase (iNOS/p38MAPK signaling pathway and to elucidate its regulating effect on the protection of spinal cord injury. Spinal cord ischemia–reperfusion injury (IRI was performed by the infrarenal abdominal aorta with mini aneurysm clip model. The expressions of iNOS and p38MAPK and the levels of biochemical parameters, including nitrite/nitrate, malondialdehyde (MDA, advanced oxidation products (AOPP, reduced glutathione (GSH, superoxide dismutase (SOD and catalase (CAT were measured in control and experimental groups. IRI-induced rats treated with 10 mg/kg resveratrol protected spinal cord from ischemia injury as supported by improved biological parameters measured in spinal cord tissue homogenates. The resveratrol treatment significantly decreased the levels of plasma nitrite/nitrate, iNOS mRNA and protein expressions and phosphorylation of p38MAPK in IRI-induced rats. Further, IRI-produced free radicals were reduced by resveratrol treatment by increasing enzymatic and non-enzymatic antioxidant levels such as GSH, SOD and CAT. Taken together, administration of resveratrol protects the damage caused by spinal cord ischemia with potential mechanism of suppressing the activation of iNOS/p38MAPK pathway and subsequent reduction of oxidative stress due to IRI.

  4. Nicotine protects kidney from renal ischemia/reperfusion injury through the cholinergic anti-inflammatory pathway.

    Directory of Open Access Journals (Sweden)

    Claude Sadis

    Full Text Available Kidney ischemia/reperfusion injury (I/R is characterized by renal dysfunction and tubular damages resulting from an early activation of innate immunity. Recently, nicotine administration has been shown to be a powerful inhibitor of a variety of innate immune responses, including LPS-induced toxaemia. This cholinergic anti-inflammatory pathway acts via the alpha7 nicotinic acetylcholine receptor (alpha7nAChR. Herein, we tested the potential protective effect of nicotine administration in a mouse model of renal I/R injury induced by bilateral clamping of kidney arteries. Renal function, tubular damages and inflammatory response were compared between control animals and mice receiving nicotine at the time of ischemia. Nicotine pretreatment protected mice from renal dysfunction in a dose-dependent manner and through the alpha7nAChR, as attested by the absence of protection in alpha7nAChR-deficient mice. Additionally, nicotine significantly reduced tubular damages, prevented neutrophil infiltration and decreased productions of the CXC-chemokine KC, TNF-alpha and the proinflammatory high-mobility group box 1 protein. Reduced tubular damage in nicotine pre-treated mice was associated with a decrease in tubular cell apoptosis and proliferative response as attested by the reduction of caspase-3 and Ki67 positive cells, respectively. All together, these data highlight that nicotine exerts a protective anti-inflammatory effect during kidney I/R through the cholinergic alpha7nAChR pathway. In addition, this could provide an opportunity to overcome the effect of surgical cholinergic denervation during kidney transplantation.

  5. Britanin Ameliorates Cerebral Ischemia-Reperfusion Injury by Inducing the Nrf2 Protective Pathway.

    Science.gov (United States)

    Wu, Guozhen; Zhu, Lili; Yuan, Xing; Chen, Hao; Xiong, Rui; Zhang, Shoude; Cheng, Hao; Shen, Yunheng; An, Huazhang; Li, Tiejun; Li, Honglin; Zhang, Weidong

    2017-10-10

    Oxidative stress is considered the major cause of tissue injury after cerebral ischemia. The nuclear factor erythroid 2-related factor 2 (Nrf2) pathway is one of the most important defensive mechanisms against oxidative stresses and has been confirmed as a target for stroke treatment. Thus, we desired to find new Nrf2 activators and test their neuronal protective activity both in vivo and in vitro. The herb-derived compound, Britanin, is a potent inducer of the Nrf2 system. Britanin can induce the expression of protective enzymes and reverse oxygen-glucose deprivation, followed by reperfusion (OGD-R)-induced neuronal injury in primary cortical neurons in vitro. Furthermore, the administration of Britanin significantly ameliorated middle cerebral artery occlusion-reperfusion (MCAO-R) insult in vivo. We report here the crystal structure of the complex of Britanin and the BTB domain of Keap1. Britanin selectively binds to a conserved cysteine residue, cysteine 151, of Keap1 and inhibits Keap1-mediated ubiquitination of Nrf2, leading to induction of the Nrf2 pathway. Britanin is a potent inducer of Nrf2. The complex crystal structure of Britanin and the BTB domain of Keap1 help clarify the mechanism of Nrf2 induction. Britanin was proven to protect primary cortical neurons against OGD-R-induced injury in an Nrf2-dependant way. Additionally, Britanin had excellent cerebroprotective effect in an MCAO-R model. Our results demonstrate that the natural product Britanin with potent Nrf2-activating and neural protective activities both in vitro and in vivo could be developed into a cerebroprotective therapeutic agent. Antioxid. Redox Signal. 27, 754-768.

  6. PGC-1α and Reactive Oxygen Species Regulate Human Embryonic Stem Cell-Derived Cardiomyocyte Function

    NARCIS (Netherlands)

    Birket, Matthew J.; Casini, Simona; Kosmidis, Georgios; Elliott, David A.; Gerencser, Akos A.; Baartscheer, Antonius; Schumacher, Cees; Mastroberardino, Pier G.; Elefanty, Andrew G.; Stanley, Ed G.; Mummery, Christine L.

    2013-01-01

    Diminished mitochondrial function is causally related to some heart diseases. Here, we developed a human disease model based on cardiomyocytes from human embryonic stem cells (hESCs), in which an important pathway of mitochondrial gene expression was inactivated. Repression of PGC-1α, which is

  7. Mitochondria Play a Central Role in Nonischemic Cardiomyocyte Necrosis: Common to Acute and Chronic Stressor States

    Science.gov (United States)

    Khan, M. Usman; Cheema, Yaser; Shahbaz, Atta U.; Ahokas, Robert A.; Sun, Yao; Gerling, Ivan C.; Bhattacharya, Syamal K.; Weber, Karl T.

    2012-01-01

    The survival of cardiomyocytes must be ensured as the myocardium adjusts to a myriad of competing physiologic and pathophysiologic demands. A significant loss of these contractile cells, together with their replacement by stiff fibrillar collagen in the form of fibrous tissue accounts for a transition from a usually efficient muscular pump into one that is failing. Cellular and subcellular mechanisms involved in the pathogenic origins of cardiomyocyte cell death have long been of interest. This includes programmed molecular pathways to either necrosis or apoptosis which are initiated from ischemic or nonischemic origins. Herein we focus on the central role played by a mitochondriocentric signal-transducer-effector pathway to nonischemic cardiomyocyte necrosis which is common to acute and chronic stressor states. We begin by building upon the hypothesis advanced by Albrecht Fleckenstein and coworkers some 40 years ago based on the importance of calcitropic hormone- mediated intracellular Ca2+ overloading which predominantly involves subsarcolemmal mitochondria and is the signal to pathway activation. Other pathway components, which came to be recognized in subsequent years, include the induction of oxidative stress and opening of the mitochondrial inner membrane permeability transition pore. The ensuing loss of cardiomyocytes and consequent replacement fibrosis, or scarring, represents a disease of adaptation and a classic example of when homeostasis begets dyshomeostasis. PMID:22328074

  8. Dnmt1-dependent Chk1 pathway suppression is protective against neuron division.

    Science.gov (United States)

    Oshikawa, Mio; Okada, Kei; Tabata, Hidenori; Nagata, Koh-Ichi; Ajioka, Itsuki

    2017-09-15

    Neuronal differentiation and cell-cycle exit are tightly coordinated, even in pathological situations. When pathological neurons re-enter the cell cycle and progress through the S phase, they undergo cell death instead of division. However, the mechanisms underlying mitotic resistance are mostly unknown. Here, we have found that acute inactivation of retinoblastoma (Rb) family proteins (Rb, p107 and p130) in mouse postmitotic neurons leads to cell death after S-phase progression. Checkpoint kinase 1 (Chk1) pathway activation during the S phase prevented the cell death, and allowed the division of cortical neurons that had undergone acute Rb family inactivation, oxygen-glucose deprivation (OGD) or in vivo hypoxia-ischemia. During neurogenesis, cortical neurons became protected from S-phase Chk1 pathway activation by the DNA methyltransferase Dnmt1, and underwent cell death after S-phase progression. Our results indicate that Chk1 pathway activation overrides mitotic safeguards and uncouples neuronal differentiation from mitotic resistance. © 2017. Published by The Company of Biologists Ltd.

  9. Human Stem Cell Derived Cardiomyocytes: An Alternative ...

    Science.gov (United States)

    Chemical spills and associated deaths in the US has increased 2.6-fold and 16-fold from 1983 to 2012, respectfully. In addition, the number of chemicals to which humans are exposed to in the environment has increased almost 10-fold from 2001 to 2013 within the US. Internationally, a WHO report on the global composite impact of chemicals on health reported that 16% of the total burden of cardiovascular disease was attributed to environmental chemical exposure with 2.5 million deaths per year. Clearly, the cardiovascular system, at all its various developmental and life stages, represents a critical target organ system that can be adversely affected by existing and emerging chemicals (e.g., engineered nanomaterials) in a variety of environmental media. The ability to assess chemical cardiac risk and safety is critically needed but extremely challenging due to the number and categories of chemicals in commerce, as indicated. This presentation\\session will evaluate the use of adult human stem cell derived cardiomyocytes, and existing platforms, as an alternative model to evaluate environmental chemical cardiac toxicity as well as provide key information for the development of predictive adverse outcomes pathways associated with environmental chemical exposures. (This abstract does not represent EPA policy) Rapid and translatable chemical safety screening models for cardiotoxicity current status for informing regulatory decisions, a workshop sponsored by the Society

  10. Dedifferentiation and proliferation of mammalian cardiomyocytes.

    Directory of Open Access Journals (Sweden)

    Yiqiang Zhang

    2010-09-01

    Full Text Available It has long been thought that mammalian cardiomyocytes are terminally-differentiated and unable to proliferate. However, myocytes in more primitive animals such as zebrafish are able to dedifferentiate and proliferate to regenerate amputated cardiac muscle.Here we test the hypothesis that mature mammalian cardiomyocytes retain substantial cellular plasticity, including the ability to dedifferentiate, proliferate, and acquire progenitor cell phenotypes. Two complementary methods were used: 1 cardiomyocyte purification from rat hearts, and 2 genetic fate mapping in cardiac explants from bi-transgenic mice. Cardiomyocytes isolated from rodent hearts were purified by multiple centrifugation and Percoll gradient separation steps, and the purity verified by immunostaining and RT-PCR. Within days in culture, purified cardiomyocytes lost their characteristic electrophysiological properties and striations, flattened and began to divide, as confirmed by proliferation markers and BrdU incorporation. Many dedifferentiated cardiomyocytes went on to express the stem cell antigen c-kit, and the early cardiac transcription factors GATA4 and Nkx2.5. Underlying these changes, inhibitory cell cycle molecules were suppressed in myocyte-derived cells (MDCs, while microRNAs known to orchestrate proliferation and pluripotency increased dramatically. Some, but not all, MDCs self-organized into spheres and re-differentiated into myocytes and endothelial cells in vitro. Cell fate tracking of cardiomyocytes from 4-OH-Tamoxifen-treated double-transgenic MerCreMer/ZEG mouse hearts revealed that green fluorescent protein (GFP continues to be expressed in dedifferentiated cardiomyocytes, two-thirds of which were also c-kit(+.Contradicting the prevailing view that they are terminally-differentiated, postnatal mammalian cardiomyocytes are instead capable of substantial plasticity. Dedifferentiation of myocytes facilitates proliferation and confers a degree of stemness

  11. Regulation of cardiomyocyte autophagy by calcium.

    Science.gov (United States)

    Shaikh, Soni; Troncoso, Rodrigo; Criollo, Alfredo; Bravo-Sagua, Roberto; García, Lorena; Morselli, Eugenia; Cifuentes, Mariana; Quest, Andrew F G; Hill, Joseph A; Lavandero, Sergio

    2016-04-15

    Calcium signaling plays a crucial role in a multitude of events within the cardiomyocyte, including cell cycle control, growth, apoptosis, and autophagy. With respect to calcium-dependent regulation of autophagy, ion channels and exchangers, receptors, and intracellular mediators play fundamental roles. In this review, we discuss calcium-dependent regulation of cardiomyocyte autophagy, a lysosomal mechanism that is often cytoprotective, serving to defend against disease-related stress and nutrient insufficiency. We also highlight the importance of the subcellular distribution of calcium and related proteins, interorganelle communication, and other key signaling events that govern cardiomyocyte autophagy. Copyright © 2016 the American Physiological Society.

  12. Protective Effects of Olive Leaf Extract on Acrolein-Exacerbated Myocardial Infarction via an Endoplasmic Reticulum Stress Pathway

    Directory of Open Access Journals (Sweden)

    Yuyu Xu

    2018-02-01

    Full Text Available Many studies reported that air pollution particulate matter (PM exposure was associated with myocardial infarction (MI. Acrolein representing the unsaturated aldehydes, the main component of PM, derives from the incomplete combustion of wood, plastic, fossil fuels and the main constitute of cigarette smoking. However, the effect of acrolein on MI remains not that clear. In the current study, the effect of acrolein-exacerbated MI was investigated. In vivo, male Sprague–Dawley rats received olive leaf extract (OLE followed by acrolein, then isoprenaline (ISO was received by subcutaneous injection to induce MI. Results showed that the expression levels of GRP78 and CHOP, two major components of endoplasmic reticulum (ER stress were higher in the combination of acrolein and ISO than those in ISO treatment. The apoptosis marker, Bax, was also higher while the anti-apoptosis indicator, Bcl2 expression was lower both at protein and mRNA levels in the combination group. Also, the acrolein-protein adducts and myocardial pathological damage increased in the combination of acrolein and ISO relative to the ISO treatment. Besides, cardiac parameters, ejection fraction (EF and fractional shortening (FS were reduced more significantly when acrolein was added than in ISO treatment. Interestingly, all the changes were able to be ameliorated by OLE. Since hydroxytyrosol (HT and oleuropein (OP were the main components in OLE, we next investigated the effect of HT and OP on cardiomyocyte H9c2 cell apoptosis induced by acrolein through ER stress and Bax pathway. Results showed that GRP78, CHOP and Bax expression were upregulated, while Bcl2 expression was downregulated both at the protein and mRNA levels, when the H9c2 cells were treated with acrolein. In addition, pretreatment with HT can reverse the expression of GRP78, CHOP, Bax and Bcl2 on the protein and mRNA levels, while there was no effect of OP on the expression of GRP78 and CHOP on the mRNA levels

  13. Hsp60 and p70S6K form a complex in human cardiomyocytes

    Directory of Open Access Journals (Sweden)

    Kroupskaya I. V.

    2011-02-01

    Full Text Available Molecular chaperon Hsp60 and protein kinase p70S6K play an important functional role in the regulation of cardiomyocytes vital function or apoptosis. Aim. To study a possibility of in vivo complex formation between Hsp60 and p70S6K in cardiomyocytes. Methods. Co-immunoprecipitation, Western-blot analysis. Results. We have identified in vivo interaction between molecular chaperone Hsp60 and two isoforms of proteinkinase p70S6K in human myocardium, normal and affected by cardiomyopathy. Conclusions. The results obtained suggest a possible participation of molecular chaperon Hsp60 in regulation of p70S6K activity in stressinduced apoptotic signaling pathway in cardiomyocytes.

  14. Genetic enrichment of cardiomyocytes derived from mouse ...

    African Journals Online (AJOL)

    Jane

    2011-06-22

    Jun 22, 2011 ... Pluripotent embryonic stem cells (ESC) have the ability to differentiate into a ... We describe a simple method to generate relatively pure cardiomyocytes from mouse ... In this study, we described the generation of transgenic.

  15. Polydatin Protects Bone Marrow Stem Cells against Oxidative Injury: Involvement of Nrf 2/ARE Pathways

    Directory of Open Access Journals (Sweden)

    Meihui Chen

    2016-01-01

    Full Text Available Polydatin, a glucoside of resveratrol, has been reported to possess potent antioxidative effects. In the present study, we aimed to investigate the effects of polydatin in bone marrow-derived mesenchymal stem cells (BMSCs death caused by hydrogen peroxide (H2O2, imitating the microenvironment surrounding transplanted cells in the injured spinal cord in vitro. In our study, MTT results showed that polydatin effectively prevented the decrease of cell viability caused by H2O2. Hochest 33258, Annexin V-PI, and Western blot assay showed H2O2-induced apoptosis in BMSCs, which was attenuated by polydatin. Further studies indicated that polydatin significantly protects BMSCs against apoptosis due to its antioxidative effects and the regulation of Nrf 2/ARE pathway. Taken together, our results indicate that polydatin could be used in combination with BMSCs for the treatment of spinal cord injury by improving the cell survival and oxidative stress microenvironments.

  16. Identification, Selection, and Enrichment of Cardiomyocyte Precursors

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    Bianca Ferrarini Zanetti

    2013-01-01

    Full Text Available The large-scale production of cardiomyocytes is a key step in the development of cell therapy and tissue engineering to treat cardiovascular diseases, particularly those caused by ischemia. The main objective of this study was to establish a procedure for the efficient production of cardiomyocytes by reprogramming mesenchymal stem cells from adipose tissue. First, lentiviral vectors expressing neoR and GFP under the control of promoters expressed specifically during cardiomyogenesis were constructed to monitor cell reprogramming into precardiomyocytes and to select cells for amplification and characterization. Cellular reprogramming was performed using 5′-azacytidine followed by electroporation with plasmid pOKS2a, which expressed Oct4, Sox2, and Klf4. Under these conditions, GFP expression began only after transfection with pOKS2a, and less than 0.015% of cells were GFP+. These GFP+ cells were selected for G418 resistance to find molecular markers of cardiomyocytes by RT-PCR and immunocytochemistry. Both genetic and protein markers of cardiomyocytes were present in the selected cells, with some variations among them. Cell doubling time did not change after selection. Together, these results indicate that enrichment with vectors expressing GFP and neoR under cardiomyocyte-specific promoters can produce large numbers of cardiomyocyte precursors (CMPs, which can then be differentiated terminally for cell therapy and tissue engineering.

  17. Induction of HO-1 by carbon monoxide releasing molecule-2 attenuates thrombin-induced COX-2 expression and hypertrophy in primary human cardiomyocytes

    International Nuclear Information System (INIS)

    Chien, Peter Tzu-Yu; Lin, Chih-Chung; Hsiao, Li-Der; Yang, Chuen-Mao

    2015-01-01

    Carbon monoxide (CO) is one of the cytoprotective byproducts of heme oxygenase (HO)-1 and exerts anti-inflammatory action in various models. However, the detailed mechanisms underlying CO-induced HO-1 expression in primary human cardiomyocytes remain largely unidentified. We used primary left ventricle myocytes as a model and applied CO releasing molecule (CORM)-2 to investigate the relationship of CO and HO-1 expression. We herein used Western blot, real-time PCR, promoter activity and EIA to investigate the role of HO-1 expression protecting against thrombin-mediated responses. We found that thrombin-induced COX-2 expression, PGE 2 release and cardiomyocyte hypertrophy markers (increase in ANF/BNP, α-actin expression and cell surface area) was attenuated by pretreatment with CORM-2 which was partially reversed by hemoglobin (Hb) or ZnPP (an inhibitor of HO-1 activity), suggesting that HO-1/CO system may be of clinical importance to ameliorate heart failure through inhibition of inflammatory responses. CORM-2-induced HO-1 protein expression, mRNA and promoter was attenuated by pretreatment with the inhibitors of Pyk2 (PF431396), PDGFR (AG1296), PI3K (LY294002), Akt (SH-5), p38 (SB202530), JNK1/2 (SP600125), FoxO1 (AS1842856) and Sp1 (mithramycin A). The involvement of these signaling components was further confirmed by transfection with respective siRNAs, consistent with those of pharmacological inhibitors. These results suggested that CORM-2-induced HO-1 expression is mediated through a Pyk2/PDGFR/PI3K/Akt/FoxO1/Sp1-dependent manner and exerts a cytoprotective effect in human cardiomyocytes. - Graphical abstract: In summary, CORM-2 treatment induces Pyk2 transactivated PDGFR, which induces PI3K/Akt/MAPK activation, and then recruits Sp1/Foxo1 transcriptional factors to regulate HO-1 gene expression in primary human cardiomyocytes. - Highlights: • CORM-2 induces HO-1 expression. • Pyk2-dependent PDGFR activates PI3K/Akt/MAPK pathway in CORM-2-induced HO-1

  18. Induction of HO-1 by carbon monoxide releasing molecule-2 attenuates thrombin-induced COX-2 expression and hypertrophy in primary human cardiomyocytes

    Energy Technology Data Exchange (ETDEWEB)

    Chien, Peter Tzu-Yu [Department of Physiology and Pharmacology and Health Ageing Research Center, Chang Gung University, Kwei-Shan, Tao-Yuan, Taiwan (China); Graduate Institute of Biomedical Sciences, College of Medicine, Chang Gung University, Kwei-Shan, Tao-Yuan, Taiwan (China); Lin, Chih-Chung; Hsiao, Li-Der [Department of Anesthetics, Chang Gung Memorial Hospital at Lin-Kou and College of Medicine, Chang Gung University, Kwei-San, Tao-Yuan, Taiwan (China); Yang, Chuen-Mao, E-mail: chuenmao@mail.cgu.edu.tw [Department of Physiology and Pharmacology and Health Ageing Research Center, Chang Gung University, Kwei-Shan, Tao-Yuan, Taiwan (China); Graduate Institute of Biomedical Sciences, College of Medicine, Chang Gung University, Kwei-Shan, Tao-Yuan, Taiwan (China); Research Center for Industry of Human Ecology and Graduate Institute of Health Industry Technology, Chang Gung University of Science and Technology, Tao-Yuan, Taiwan (China)

    2015-12-01

    Carbon monoxide (CO) is one of the cytoprotective byproducts of heme oxygenase (HO)-1 and exerts anti-inflammatory action in various models. However, the detailed mechanisms underlying CO-induced HO-1 expression in primary human cardiomyocytes remain largely unidentified. We used primary left ventricle myocytes as a model and applied CO releasing molecule (CORM)-2 to investigate the relationship of CO and HO-1 expression. We herein used Western blot, real-time PCR, promoter activity and EIA to investigate the role of HO-1 expression protecting against thrombin-mediated responses. We found that thrombin-induced COX-2 expression, PGE{sub 2} release and cardiomyocyte hypertrophy markers (increase in ANF/BNP, α-actin expression and cell surface area) was attenuated by pretreatment with CORM-2 which was partially reversed by hemoglobin (Hb) or ZnPP (an inhibitor of HO-1 activity), suggesting that HO-1/CO system may be of clinical importance to ameliorate heart failure through inhibition of inflammatory responses. CORM-2-induced HO-1 protein expression, mRNA and promoter was attenuated by pretreatment with the inhibitors of Pyk2 (PF431396), PDGFR (AG1296), PI3K (LY294002), Akt (SH-5), p38 (SB202530), JNK1/2 (SP600125), FoxO1 (AS1842856) and Sp1 (mithramycin A). The involvement of these signaling components was further confirmed by transfection with respective siRNAs, consistent with those of pharmacological inhibitors. These results suggested that CORM-2-induced HO-1 expression is mediated through a Pyk2/PDGFR/PI3K/Akt/FoxO1/Sp1-dependent manner and exerts a cytoprotective effect in human cardiomyocytes. - Graphical abstract: In summary, CORM-2 treatment induces Pyk2 transactivated PDGFR, which induces PI3K/Akt/MAPK activation, and then recruits Sp1/Foxo1 transcriptional factors to regulate HO-1 gene expression in primary human cardiomyocytes. - Highlights: • CORM-2 induces HO-1 expression. • Pyk2-dependent PDGFR activates PI3K/Akt/MAPK pathway in CORM-2-induced HO

  19. Pathways from marine protected area design and management to ecological success

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    Murray A. Rudd

    2015-11-01

    Full Text Available Using an international dataset compiled from 121 sites in 87 marine protected areas (MPAs globally (Edgar et al., 2014, I assessed how various configurations of design and management conditions affected MPA ecological performance, measured in terms of fish species richness and biomass. The set-theoretic approach used Boolean algebra to identify pathways that combined up to five ‘NEOLI’ (No-take, Enforced, Old, Large, Isolated conditions and that were sufficient for achieving positive, and negative, ecological outcomes. Ecological isolation was overwhelming the most important condition affecting ecological outcomes but Old and Large were also conditions important for achieving high levels of biomass among large fishes (jacks, groupers, sharks. Solution coverage was uniformly low (0.50 for negative results (i.e., the absence of high biomass among the large commercially-exploited fishes, implying asymmetries in how MPAs may rebuild populations on the one hand and, on the other, protect against further decline. The results revealed complex interactions involving MPA design, implementation, and management conditions that affect MPA ecological performance. In general terms, the presence of no-take regulations and effective enforcement were insufficient to ensure MPA effectiveness on their own. Given the central role of ecological isolation in securing ecological benefits from MPAs, site selection in the design phase appears critical for success.

  20. Dendritic cells and hepatocytes use distinct pathways to process protective antigen from plasmodium in vivo.

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    Ian A Cockburn

    2011-03-01

    Full Text Available Malaria-protective CD8+ T cells specific for the circumsporozoite (CS protein are primed by dendritic cells (DCs after sporozoite injection by infected mosquitoes. The primed cells then eliminate parasite liver stages after recognizing the CS epitopes presented by hepatocytes. To define the in vivo processing of CS by DCs and hepatocytes, we generated parasites carrying a mutant CS protein containing the H-2K(b epitope SIINFEKL, and evaluated the T cell response using transgenic and mutant mice. We determined that in both DCs and hepatocytes CS epitopes must reach the cytosol and use the TAP transporters to access the ER. Furthermore, we used endosomal mutant (3d and cytochrome c treated mice to address the role of cross-presentation in the priming and effector phases of the T cell response. We determined that in DCs, CS is cross-presented via endosomes while, conversely, in hepatocytes protein must be secreted directly into the cytosol. This suggests that the main targets of protective CD8+ T cells are parasite proteins exported to the hepatocyte cytosol. Surprisingly, however, secretion of the CS protein into hepatocytes was not dependent upon parasite-export (Pexel/VTS motifs in this protein. Together, these results indicate that the presentation of epitopes to CD8+ T cells follows distinct pathways in DCs when the immune response is induced and in hepatocytes during the effector phase.

  1. Allicin protects against cisplatin-induced vestibular dysfunction by inhibiting the apoptotic pathway.

    Science.gov (United States)

    Wu, Xianmin; Cai, Jing; Li, Xiaofei; Li, He; Li, Jianfeng; Bai, Xiaohui; Liu, Wenwen; Han, Yuechen; Xu, Lei; Zhang, Daogong; Wang, Haibo; Fan, Zhaomin

    2017-06-15

    Cisplatin is an anticancer drug that causes the impairment of inner ear function as side effects, including hearing loss and balance dysfunction. The purpose of this study was to investigate the effects of allicin against cisplatin-induced vestibular dysfunction in mice and to make clear the mechanism underlying the protective effects of allicin on oto-vestibulotoxicity. Mice intraperitoneally injected with cisplatin exhibited vestibular dysfunction in swimming test, which agreed with impairment in vestibule. However, these impairments were significantly prevented by pre-treatment with allicin. Allicin markedly reduced cisplatin-activated expression of cleaved-caspase-3 in hair cells and vascular layer cells of utricule, saccule and ampulla, but also decreased AIF nuclear translocation of hair cells in utricule, saccule and ampulla. These results showed that allicin played an effective role in protecting vestibular dysfunction induced by cisplatin via inhibiting caspase-dependent and caspase-independent apoptotic pathways. Therefore, allicin may be useful in preventing oto-vestibulotoxicity mediated by cisplatin. Copyright © 2017. Published by Elsevier B.V.

  2. Vitamin D receptor pathway is required for probiotic protection in colitis.

    Science.gov (United States)

    Wu, Shaoping; Yoon, Sonia; Zhang, Yong-Guo; Lu, Rong; Xia, Yinglin; Wan, Jiandi; Petrof, Elaine O; Claud, Erika C; Chen, Di; Sun, Jun

    2015-09-01

    Low expression of vitamin D receptor (VDR) and dysfunction of vitamin D/VDR signaling are reported in patients with inflammatory bowel disease (IBD); therefore, restoration of VDR function to control inflammation in IBD is desirable. Probiotics have been used in the treatment of IBD. However, the role of probiotics in the modulation of VDR signaling to effectively reduce inflammation is unknown. We identified a novel role of probiotics in activating VDR activity, thus inhibiting inflammation, using cell models and VDR knockout mice. We found that the probiotics Lactobacillus rhamnosus strain GG (LGG) and Lactobacillus plantarum (LP) increased VDR protein expression in both mouse and human intestinal epithelial cells. Using the VDR luciferase reporter vector, we detected increased transcriptional activity of VDR after probiotic treatment. Probiotics increased the expression of the VDR target genes, such as antimicrobial peptide cathelicidin, at the transcriptional level. Furthermore, the role of probiotics in regulating VDR signaling was tested in vivo using a Salmonella-colitis model in VDR knockout mice. Probiotic treatment conferred physiological and histologic protection from Salmonella-induced colitis in VDR(+/+) mice, whereas probiotics had no effects in the VDR(-/-) mice. Probiotic treatment also enhanced numbers of Paneth cells, which secrete AMPs for host defense. These data indicate that the VDR pathway is required for probiotic protection in colitis. Understanding how probiotics enhance VDR signaling and inhibit inflammation will allow probiotics to be used effectively, resulting in innovative approaches to the prevention and treatment of chronic inflammation. Copyright © 2015 the American Physiological Society.

  3. Simultaneous Assessment of Cardiomyocyte DNA Synthesis and Ploidy: A Method to Assist Quantification of Cardiomyocyte Regeneration and Turnover.

    Science.gov (United States)

    Richardson, Gavin D

    2016-05-23

    Although it is accepted that the heart has a limited potential to regenerate cardiomyocytes following injury and that low levels of cardiomyocyte turnover occur during normal ageing, quantification of these events remains challenging. This is in part due to the rarity of the process and the fact that multiple cellular sources contribute to myocardial maintenance. Furthermore, DNA duplication within cardiomyocytes often leads to a polyploid cardiomyocyte and only rarely leads to new cardiomyocytes by cellular division. In order to accurately quantify cardiomyocyte turnover discrimination between these processes is essential. The protocol described here employs long term nucleoside labeling in order to label all nuclei which have arisen as a result of DNA replication and cardiomyocyte nuclei identified by utilizing nuclei isolation and subsequent PCM1 immunolabeling. Together this allows the accurate and sensitive identification of the nucleoside labeling of the cardiomyocyte nuclei population. Furthermore, 4',6-diamidino-2-phenylindole labeling and analysis of nuclei ploidy, enables the discrimination of neo-cardiomyocyte nuclei from nuclei which have incorporated nucleoside during polyploidization. Although this method cannot control for cardiomyocyte binucleation, it allows a rapid and robust quantification of neo-cardiomyocyte nuclei while accounting for polyploidization. This method has a number of downstream applications including assessing the potential therapeutics to enhance cardiomyocyte regeneration or investigating the effects of cardiac disease on cardiomyocyte turnover and ploidy. This technique is also compatible with additional downstream immunohistological techniques, allowing quantification of nucleoside incorporation in all cardiac cell types.

  4. The Adipokine Chemerin Induces Apoptosis in Cardiomyocytes

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    Diego Rodríguez-Penas

    2015-08-01

    Full Text Available Background: The adipokine chemerin has been associated with cardiovascular disease. We investigated the effects of chemerin on viability and intracellular signalling in murine cardiomyocytes, and the effects of insulin and TNF-α on cardiomyocyte chemerin production. Methods: Hoechst dye vital staining and cell cycle analysis were used to analyse the viability of murine cardiac cells in culture. Western blot was used to explore the phosphorylation of AKT and caspase-9 activity in neonatal rat cardiomyocytes and HL-1 cells. Finally, RT-qPCR, ELISA and western blot were performed to examine chemerin and CMKLR1 expression after insulin and TNF-α treatment in cardiac cells. Results: Chemerin treatment increased apoptosis, reduced phosphorylation of AKT at Thr308 and increased caspase-9 activity in murine cardiomyocytes. Insulin treatment lowered chemerin and CMKLR1 mRNA and protein levels, and the amount of chemerin in the cell media, while TNF-α treatment increased chemerin mRNA and protein levels but decreased expression of the CMKLR1 gene. Conclusion: Chemerin induces apoptosis, reduces AKT phosphorylation and increases the cleavage of caspase-9 in murine cardiomyocytes. The expression of chemerin is regulated by important metabolic (insulin and inflammatory (TNF-α mediators at cardiac level. Our results suggest that chemerin could play a role in the physiopathology of cardiac diseases.

  5. Adiponectin protects palmitic acid induced endothelial inflammation and insulin resistance via regulating ROS/IKKβ pathways.

    Science.gov (United States)

    Zhao, Wenwen; Wu, Chuanhong; Li, Shaojing; Chen, Xiuping

    2016-12-01

    Endothelial inflammation and insulin resistance (IR) has been closely associated with endothelial dysfunction. Adiponectin (APN), an adipocyte-secreted hormone from adipose tissues, showed cardioprotective effects. Here, the protective effect of APN on palmitic acid (PA)-induced endothelial inflammation and IR was investigated. Cultured human umbilical vein endothelial cells (HUVECs) were treated with PA without or without APN pretreatment. The expression of inflammatory cytokines TNF-α, IL-6, adhesion molecule ICAM-1 were determined by western blotting, ELISA, and real-time PCR. The protein expression and protein-protein interaction were determined by western blotting and immunoprecipitation. The intracellular reactive oxygen species (ROS) and nitric oxide (NO) production were monitored with fluorescence probes. PA-induced secretion of TNF-α, IL-6, and expression of ICAM-1 at protein and mRNA levels, which was significantly inhibited by APN. PA treatment caused increase of ROS generation, NOX2, p-IKKβ, p-IκBα, p-p65 expression, and p-IκBα-IKKβ interaction, which were all partly reversed by APN. ROS scavenger N-acetylcysteine (NAC) and NF-κB inhibitor PDTC showed similar effect on PA-induced secretion of TNF-α, IL-6, and expression of ICAM-1. Furthermore, APN and NAC pretreatment restored PA-induced increase of p-IRS-1(S307), decrease of p-IRS-1(Tyr). In addition, insulin-triggered expression of p-IRS-1(Tyr), p-PI3K, p-AKT, p-eNOS and NO generation were inhibited by PA, which were also restored by both APN and NAC. These results suggested that APN ameliorated endothelial inflammation and IR through ROS/IKKβ pathway. This study shed new insights into the mechanisms of APN's cardiovascular protective effect. Copyright © 2016 Elsevier Ltd. All rights reserved.

  6. Acute Pre-/Post-Treatment with 8th Day SOD-Like Supreme (a Free Radical Scavenging Health Product Protects against Oxidant-Induced Injury in Cultured Cardiomyocytes and Hepatocytes In Vitro as Well as in Mouse Myocardium and Liver In Vivo

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    Pou Kuan Leong

    2017-04-01

    Full Text Available 8th Day superoxide dismutase (SOD-Like Supreme (SOD-Like Supreme, a free radical scavenging health product is an antioxidant-enriched fermentation preparation with free radical scavenging properties. In the present study, the cellular/tissue protective actions of SOD-Like Supreme against menadione toxicity in cultured H9c2 cardiomyocytes and in AML12 hepatocytes as well as oxidant-induced injury in the mouse myocardium and liver were investigated. SOD-Like Supreme was found to possess potent free radical scavenging activity in vitro as assessed by an oxygen radical absorbance capacity assay. Incubation with SOD-Like Supreme (0.5–3% (v/v was shown to protect against menadione-induced toxicity in H9c2 and AML12 cells, as evidenced by increases in cell viability. The ability of SOD-Like Supreme to protect against menadione cytotoxicity was associated with an elevation in the cellular reduced glutathione (GSH/oxidized glutathione (GSSG ratio in menadione-challenged cells. Consistent with the cell-based studies, pre-/post-treatment with SOD-Like Supreme (0.69 and 2.06 mL/kg, three intermittent doses per day for two consecutive days was found to protect against isoproterenol-induced myocardial injury and carbon tetrachloride hepatotoxicity in mice. The cardio/hepatoprotection afforded by SOD-Like Supreme was also paralleled by increases in myocardial/hepatic mitochondrial GSH/GSSG ratios in the SOD-Like Supreme-treated/oxidant-challenged mice. In conclusion, incubation/treatment with SOD-Like Supreme was found to protect against oxidant-induced injury in vitro and in vivo, presumably by virtue of its free radical scavenging activity.

  7. The Protective Effect of Gangliosides on Lead (Pb)-Induced Neurotoxicity Is Mediated by Autophagic Pathways.

    Science.gov (United States)

    Meng, Hongtao; Wang, Lan; He, Junhong; Wang, Zhufeng

    2016-03-25

    Lead (Pb) is a ubiquitous environmental and industrial pollutant and can affect intelligence development and the learning ability and memory of children. Therefore, necessary measures should be taken to protect the central nervous system (CNS) from Pb toxicity. Gangliosides are sialic acid-containing glycosphingolipids that are constituents of mammalian cell membranes and are more abundantly expressed in the CNS. Studies have shown that gangliosides constitute a useful tool in the attempt to promote functional recovery of CNS and can reverse Pb-induced impairments of synaptic plasticity in rats. However, the detailed mechanisms have yet to be fully understood. In our present study, we tried to investigate the role of gangliosides in Pb-induced injury in hippocampus neurons and to further confirm the detailed mechanism. Our results show that Pb-induced injuries in the spatial reference memory were associated with a reduction of cell viability and cell apoptosis, and treatment with gangliosides markedly ameliorated the Pb-induced injury by inhibition of apoptosis action. Gangliosides further attenuated Pb-induced the abnormal autophagic process by regulation of mTOR pathways. In summary, our study establishes the efficacy of gangliosides as neuroprotective agents and provides a strong rationale for further studies on the underlying mechanisms of their neuroprotective functions.

  8. The Protective Effect of Gangliosides on Lead (Pb-Induced Neurotoxicity Is Mediated by Autophagic Pathways

    Directory of Open Access Journals (Sweden)

    Hongtao Meng

    2016-03-01

    Full Text Available Lead (Pb is a ubiquitous environmental and industrial pollutant and can affect intelligence development and the learning ability and memory of children. Therefore, necessary measures should be taken to protect the central nervous system (CNS from Pb toxicity. Gangliosides are sialic acid-containing glycosphingolipids that are constituents of mammalian cell membranes and are more abundantly expressed in the CNS. Studies have shown that gangliosides constitute a useful tool in the attempt to promote functional recovery of CNS and can reverse Pb-induced impairments of synaptic plasticity in rats. However, the detailed mechanisms have yet to be fully understood. In our present study, we tried to investigate the role of gangliosides in Pb-induced injury in hippocampus neurons and to further confirm the detailed mechanism. Our results show that Pb-induced injuries in the spatial reference memory were associated with a reduction of cell viability and cell apoptosis, and treatment with gangliosides markedly ameliorated the Pb-induced injury by inhibition of apoptosis action. Gangliosides further attenuated Pb-induced the abnormal autophagic process by regulation of mTOR pathways. In summary, our study establishes the efficacy of gangliosides as neuroprotective agents and provides a strong rationale for further studies on the underlying mechanisms of their neuroprotective functions.

  9. Resveratrol Protects PC12 Cell against 6-OHDA Damage via CXCR4 Signaling Pathway

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    Jing Zhang

    2015-01-01

    Full Text Available Resveratrol, herbal nonflavonoid polyphenolic compound naturally derived from grapes, has long been acknowledged to possess extensive biological and pharmacological properties including antioxidant and anti-inflammatory ones and may exert a neuroprotective effect on neuronal damage in neurodegenerative diseases. However, the underlying molecular mechanisms remain undefined. In the present study, we intended to investigate the neuroprotective effects of resveratrol against 6-OHDA-induced neurotoxicity of PC12 cells and further explore the possible mechanisms involved. For this purpose, PC12 cells were exposed to 6-OHDA in the presence of resveratrol (0, 12.5, 25, and 50 μM. The results showed that resveratrol increased cell viability, alleviated the MMP reduction, and reduced the number of apoptotic cells as measured by MTT assay, JC-1 staining, and Hoechst/PI double staining (all p<0.01. Immunofluorescent staining and Western blotting revealed that resveratrol averts 6-OHDA induced CXCR4 upregulation (p<0.01. Our results demonstrated that resveratrol could effectively protect PC12 cells from 6-OHDA-induced oxidative stress and apoptosis via CXCR4 signaling pathway.

  10. Serotonin transporter protects the placental cells against apoptosis in caspase 3-independent pathway.

    Science.gov (United States)

    Hadden, Coedy; Fahmi, Tariq; Cooper, Anthonya; Savenka, Alena V; Lupashin, Vladimir V; Roberts, Drucilla J; Maroteaux, Luc; Hauguel-de Mouzon, Sylvie; Kilic, Fusun

    2017-12-01

    Serotonin (5-HT) and its specific transporter, SERT play important roles in pregnancy. Using placentas dissected from 18d gestational SERT-knock out (KO), peripheral 5-HT (TPH1)-KO, and wild-type (WT) mice, we explored the role of 5-HT and SERT in placental functions in detail. An abnormal thick band of fibrosis and necrosis under the giant cell layer in SERT-KO placentas appeared only moderately in TPH1-KO and minimally present in WT placentas. The majority of the changes were located at the junctional zone of the placentas in SERT. The etiology of these findings was tested with TUNEL assays. The placentas from SERT-KO and TPH1-KO showed 49- and 8-fold increase in TUNEL-positive cells without a concurrent change in the DNA repair or cell proliferation compared to WT placentas. While the proliferation rate in the embryos of TPH1-KO mice was 16-fold lower than the rate in gestational age matched embryos of WT or SERT-KO mice. These findings highlight an important role of continuous 5-HT signaling on trophoblast cell viability. SERT may contribute to protecting trophoblast cells against cell death via terminating the 5-HT signaling which changes cell death ratio in trophoblast as well as proliferation rate in embryos. However, the cell death in SERT-KO placentas is in caspase 3-independent pathway. © 2017 Wiley Periodicals, Inc.

  11. A TIGAR-regulated metabolic pathway is critical for protection of brain ischemia.

    Science.gov (United States)

    Li, Mei; Sun, Meiling; Cao, Lijuan; Gu, Jin-hua; Ge, Jianbin; Chen, Jieyu; Han, Rong; Qin, Yuan-Yuan; Zhou, Zhi-Peng; Ding, Yuqiang; Qin, Zheng-Hong

    2014-05-28

    TP53-induced glycolysis and apoptosis regulator (TIGAR) inhibits glycolysis and increases the flow of pentose phosphate pathway (PPP), which generates NADPH and pentose. We hypothesized that TIGAR plays a neuroprotective role in brain ischemia as neurons do not rely on glycolysis but are vulnerable to oxidative stress. We found that TIGAR was highly expressed in brain neurons and was rapidly upregulated in response to ischemia/reperfusion insult in a TP53-independent manner. Overexpression of TIGAR in normal mice with lentivirus reduced ischemic neuronal injury, whereas lentivirus-mediated TIGAR knockdown aggravated it. In cultured primary neurons, increasing TIGAR expression reduced oxygen and glucose deprivation (OGD)/reoxygenation-induced injury, whereas decreasing its expression worsened the injury. The glucose 6-phosphate dehydrogenase was upregulated in mouse and cellular models of stroke, and its upregulation was further enhanced by overexpression of TIGAR. Supplementation of NADPH also reduced ischemia/reperfusion brain injury and alleviated TIGAR knockdown-induced aggravation of ischemic injury. In animal and cellular stroke models, ischemia/reperfusion increased mitochondrial localization of TIGAR. OGD/reoxygenation-induced elevation of ROS, reduction of GSH, dysfunction of mitochondria, and activation of caspase-3 were rescued by overexpression of TIGAR or supplementation of NADPH, while knockdown of TIGAR aggravated these changes. Together, our results show that TIGAR protects ischemic brain injury via enhancing PPP flux and preserving mitochondria function, and thus may be a valuable therapeutic target for ischemic brain injury. Copyright © 2014 the authors 0270-6474/14/347458-14$15.00/0.

  12. AKIP1 expression modulates mitochondrial function in rat neonatal cardiomyocytes.

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    Hongjuan Yu

    Full Text Available A kinase interacting protein 1 (AKIP1 is a molecular regulator of protein kinase A and nuclear factor kappa B signalling. Recent evidence suggests AKIP1 is increased in response to cardiac stress, modulates acute ischemic stress response, and is localized to mitochondria in cardiomyocytes. The mitochondrial function of AKIP1 is, however, still elusive. Here, we investigated the mitochondrial function of AKIP1 in a neonatal cardiomyocyte model of phenylephrine (PE-induced hypertrophy. Using a seahorse flux analyzer we show that PE stimulated the mitochondrial oxygen consumption rate (OCR in cardiomyocytes. This was partially dependent on PE mediated AKIP1 induction, since silencing of AKIP1 attenuated the increase in OCR. Interestingly, AKIP1 overexpression alone was sufficient to stimulate mitochondrial OCR and in particular ATP-linked OCR. This was also true when pyruvate was used as a substrate, indicating that it was independent of glycolytic flux. The increase in OCR was independent of mitochondrial biogenesis, changes in ETC density or altered mitochondrial membrane potential. In fact, the respiratory flux was elevated per amount of ETC, possibly through enhanced ETC coupling. Furthermore, overexpression of AKIP1 reduced and silencing of AKIP1 increased mitochondrial superoxide production, suggesting that AKIP1 modulates the efficiency of electron flux through the ETC. Together, this suggests that AKIP1 overexpression improves mitochondrial function to enhance respiration without excess superoxide generation, thereby implicating a role for AKIP1 in mitochondrial stress adaptation. Upregulation of AKIP1 during different forms of cardiac stress may therefore be an adaptive mechanism to protect the heart.

  13. Cardiomyocyte hypertrophy induced by Endonuclease G deficiency requires reactive oxygen radicals accumulation and is inhibitable by the micropeptide humanin.

    Science.gov (United States)

    Blasco, Natividad; Cámara, Yolanda; Núñez, Estefanía; Beà, Aida; Barés, Gisel; Forné, Carles; Ruíz-Meana, Marisol; Girón, Cristina; Barba, Ignasi; García-Arumí, Elena; García-Dorado, David; Vázquez, Jesús; Martí, Ramon; Llovera, Marta; Sanchis, Daniel

    2018-06-01

    The endonuclease G gene (Endog), which codes for a mitochondrial nuclease, was identified as a determinant of cardiac hypertrophy. How ENDOG controls cardiomyocyte growth is still unknown. Thus, we aimed at finding the link between ENDOG activity and cardiomyocyte growth. Endog deficiency induced reactive oxygen species (ROS) accumulation and abnormal growth in neonatal rodent cardiomyocytes, altering the AKT-GSK3β and Class-II histone deacethylases (HDAC) signal transduction pathways. These effects were blocked by ROS scavengers. Lack of ENDOG reduced mitochondrial DNA (mtDNA) replication independently of ROS accumulation. Because mtDNA encodes several subunits of the mitochondrial electron transport chain, whose activity is an important source of cellular ROS, we investigated whether Endog deficiency compromised the expression and activity of the respiratory chain complexes but found no changes in these parameters nor in ATP content. MtDNA also codes for humanin, a micropeptide with possible metabolic functions. Nanomolar concentrations of synthetic humanin restored normal ROS levels and cell size in Endog-deficient cardiomyocytes. These results support the involvement of redox signaling in the control of cardiomyocyte growth by ENDOG and suggest a pathway relating mtDNA content to the regulation of cell growth probably involving humanin, which prevents reactive oxygen radicals accumulation and hypertrophy induced by Endog deficiency. Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.

  14. MicroRNA-1 Regulates the Differentiation of Adipose-Derived Stem Cells into Cardiomyocyte-Like Cells

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    Can Chen

    2018-01-01

    Full Text Available Stem cell transplantation is one of most valuable methods in the treatment of myocardial infarction, and adipose-derived stem cells (ASCs are becoming a hot topic in medical research. Previous studies have shown that ASCs can be differentiated into cardiomyocyte-like cells, but the efficiency and survival rates are low. We investigated the role and mechanism of microRNA-1 (miR-1 in the differentiation of ASCs into cardiomyocyte-like cells. ASCs and cardiomyocytes were isolated from neonatal rats. We constructed lentivirus for overexpressing miR-1 and used DAPT, an antagonist of the Notch1 pathway, for in vitro analyses. We performed cocultures with ASCs and cardiomyocytes. The differentiation efficiency of ASCs was detected by cell-specific surface antigens. Our results showed that miR-1 can promote the expression of Notch1 and reduce the expression of Hes1, a Notch pathway factor, and overexpression of miR-1 can promote the differentiation of ASCs into cardiomyocyte-like cells, which may occur by regulating Notch1 and Hes1.

  15. Heme Oxygenase-1/Carbon Monoxide System and Embryonic Stem Cell Differentiation and Maturation into Cardiomyocytes

    Science.gov (United States)

    Suliman, Hagir B.; Zobi, Fabio

    2016-01-01

    Abstract Aims: The differentiation of embryonic stem (ES) cells into energetically efficient cardiomyocytes contributes to functional cardiac repair and is envisioned to ameliorate progressive degenerative cardiac diseases. Advanced cell maturation strategies are therefore needed to create abundant mature cardiomyocytes. In this study, we tested whether the redox-sensitive heme oxygenase-1/carbon monoxide (HO-1/CO) system, operating through mitochondrial biogenesis, acts as a mechanism for ES cell differentiation and cardiomyocyte maturation. Results: Manipulation of HO-1/CO to enhance mitochondrial biogenesis demonstrates a direct pathway to ES cell differentiation and maturation into beating cardiomyocytes that express adult structural markers. Targeted HO-1/CO interventions up- and downregulate specific cardiogenic transcription factors, transcription factor Gata4, homeobox protein Nkx-2.5, heart- and neural crest derivatives-expressed protein 1, and MEF2C. HO-1/CO overexpression increases cardiac gene expression for myosin regulatory light chain 2, atrial isoform, MLC2v, ANP, MHC-β, and sarcomere α-actinin and the major mitochondrial fusion regulators, mitofusin 2 and MICOS complex subunit Mic60. This promotes structural mitochondrial network expansion and maturation, thereby supporting energy provision for beating embryoid bodies. These effects are prevented by silencing HO-1 and by mitochondrial reactive oxygen species scavenging, while disruption of mitochondrial biogenesis and mitochondrial DNA depletion by loss of mitochondrial transcription factor A compromise infrastructure. This leads to failure of cardiomyocyte differentiation and maturation and contractile dysfunction. Innovation: The capacity to augment cardiomyogenesis via a defined mitochondrial pathway has unique therapeutic potential for targeting ES cell maturation in cardiac disease. Conclusion: Our findings establish the HO-1/CO system and redox regulation of mitochondrial biogenesis as

  16. Cardiomyocyte Overexpression of FABP4 Aggravates Pressure Overload-Induced Heart Hypertrophy.

    Directory of Open Access Journals (Sweden)

    Ji Zhang

    Full Text Available Fatty acid binding protein 4 (FABP4 is a member of the intracellular lipid-binding protein family, responsible for the transportation of fatty acids. It is considered to express mainly in adipose tissues, and be strongly associated with inflammation, obesity, diabetes and cardiovasculardiseases. Here we report that FABP4 is also expressed in cardiomyocytes and plays an important role in regulating heart function under pressure overload. We generated heart-specific transgenic FABP4 (FABP4-TG mice using α myosin-heavy chain (α-MHC promoter and human FABP4 sequence, resulting in over-expression of FABP4 in cardiomyocytes. The FABP4-TG mice displayed normal cardiac morphology and contractile function. When they were subjected to the transverse aorta constriction (TAC procedure, the FABP4-TG mice developed more cardiac hypertrophy correlated with significantly increased ERK phosphorylation, compared with wild type controls. FABP4 over-expression in cardiomyocytes activated phosphor-ERK signal and up-regulate the expression of cardiac hypertrophic marker genes. Conversely, FABP4 induced phosphor-ERK signal and hypertrophic gene expressions can be markedly inhibited by an ERK inhibitor PD098059 as well as the FABP4 inhibitor BMS309403. These results suggest that FABP4 over-expression in cardiomyocytes can aggravate the development of cardiac hypertrophy through the activation of ERK signal pathway.

  17. Distinctive Roles of Canonical and Noncanonical Wnt Signaling in Human Embryonic Cardiomyocyte Development

    Directory of Open Access Journals (Sweden)

    Silvia Mazzotta

    2016-10-01

    Full Text Available Wnt signaling is a key regulator of vertebrate heart development; however, specific roles for human cardiomyocyte development remain uncertain. Here we use human embryonic stem cells (hESCs to analyze systematically in human cardiomyocyte development the expression of endogenous Wnt signaling components, monitor pathway activity, and dissect stage-specific requirements for canonical and noncanonical Wnt signaling mechanisms using small-molecule inhibitors. Our analysis suggests that WNT3 and WNT8A, via FZD7 and canonical signaling, regulate BRACHYURY expression and mesoderm induction; that WNT5A/5B, via ROR2 and noncanonical signaling, regulate MESP1 expression and cardiovascular development; and that later in development WNT2, WNT5A/5B, and WNT11, via FZD4 and FZD6, regulate functional cardiomyocyte differentiation via noncanonical Wnt signaling. Our findings confirm in human development previously proposed roles for canonical Wnt signaling in sequential stages of vertebrate cardiomyogenesis, and identify more precise roles for noncanonical signaling and for individual Wnt signal and Wnt receptor genes in human cardiomyocyte development.

  18. Lipopolysaccharide preconditioning protects hepatocytes from ischemia/reperfusion injury (IRI through inhibiting ATF4-CHOP pathway in mice.

    Directory of Open Access Journals (Sweden)

    Jianhua Rao

    Full Text Available BACKGROUND: Low-dose lipopolysaccharide (LPS preconditioning-induced liver protection has been demonstrated during ischemia-reperfusion injury (IRI in several organs but has not been sufficiently elucidated underlying causal mechanism. This study investigated the role of low-dose LPS preconditioning on ATF4-CHOP pathway as well as the effects of the pathway on tissue injury and inflammation in a mouse model of liver partial-warm IRI. METHODS: LPS (100 µg/kg/d was injected intraperitoneally two days before ischemia. Hepatic injury was evaluated based on serum alanine aminotransferase levels, histopathology, and caspase-3 activity. The ATF4-CHOP pathway and its related apoptotic molecules were investigated after reperfusion. The role of LPS preconditioning on apoptosis and ATF4-CHOP pathway was examined in vitro. Moreover, the effects of the ATF4-CHOP pathway on apoptosis, Caspase-12, and Caspase-3 were determined with ATF4 small interfering RNA (siRNA. Inflammatory cytokine expression was also checked after reperfusion. Inflammatory cytokines and related signaling pathways were analyzed in vitro in macrophages treated by LPS preconditioning or ATF4 siRNA. RESULTS: LPS preconditioning significantly attenuated liver injury after IRI. As demonstrated by in vitro experiments, LPS preconditioning significantly reduced the upregulation of the ATF4-CHOP pathway and inhibited Caspase-12 and Caspase-3 activation after IRI. Later experiments showed that ATF4 knockdown significantly suppressed CHOP, cleaved caspase-12 and caspase-3 expression, as well as inhibited hepatocellular apoptosis. In addition, in mice pretreated with LPS, TNF-α and IL-6 were inhibited after reperfusion, whereas IL-10 was upregulated. Similarly, low-dose LPS significantly inhibited TNF-α, IL-6, ATF4-CHOP pathway, NF-κB pathway, and ERK1/2 in high-dose LPS-stimulated macrophages, whereas IL-10 and cytokine signaling (SOCS-3 suppressor were induced. Importantly, ATF4 siRNA is

  19. Axin1 up-regulated 1 accelerates stress-induced cardiomyocytes apoptosis through activating Wnt/β-catenin signaling.

    Science.gov (United States)

    Ye, Xing; Lin, Junyi; Lin, Zebin; Xue, Aimin; Li, Liliang; Zhao, Ziqin; Liu, Li; Shen, Yiwen; Cong, Bin

    2017-10-15

    Stress-induced cardiomyocyte apoptosis contributes to the pathogenesis of a variety of cardiovascular diseases, but how stress induces cardiomyocyte apoptosis remains largely unclear. The present study aims to investigate the effects of Axin1 up-regulated 1 (Axud1), a novel pro-apoptotic protein, on the cardiomyocyte survival and the underlying mechanisms. To this end, a rat model under restraint stress (RS) was established and in vitro stress-induced cardiomyocytes culture was achieved. Our data showed that Axud1 was upregulated in the rat myocardia after exposure to RS. Anti-apoptotic Bcl-2 was decreased, whereas pro-apoptotic Bax and Cleaved caspase-3 (Cc3) were increased in a time-dependent manner. The Wnt/β-catenin signaling was observed to be interestingly activated in heart undergoing RS. In addition, the treatment of norepinephrine (NE) to in vitro cardiomyocytes increased Axud1 level and induced cell apoptosis. Wnt/β-catenin signaling was consistently activated. Knockdown of Axud1 using specific siRNA blunted NE-induced cardiomyocytes apoptosis and also inactivated the Wnt/β-catenin signaling. XAV-939, an inhibitor of Wnt/β-catenin signaling, partially reversed the pro-apoptotic effect of NE. In conclusion, Axud1 accelerated stress-induced cardiomyocytes apoptosis through activation of Wnt/β-catenin signaling pathway. Our data provided novel evidence that therapeutic strategies against Axud1 or Wnt/β-catenin signaling might be promising in relation to RS-induced myocardial injury. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

  20. Coconut oil protects cortical neurons from amyloid beta toxicity by enhancing signaling of cell survival pathways.

    Science.gov (United States)

    Nafar, F; Clarke, J P; Mearow, K M

    2017-05-01

    Alzheimer's disease is a progressive neurodegenerative disease that has links with other conditions that can often be modified by dietary and life-style interventions. In particular, coconut oil has received attention as having potentially having benefits in lessening the cognitive deficits associated with Alzheimer's disease. In a recent report, we showed that neuron survival in cultures co-treated with coconut oil and Aβ was rescued compared to cultures exposed only to Aβ. Here we investigated treatment with Aβ for 1, 6 or 24 h followed by addition of coconut oil for a further 24 h, or treatment with coconut oil for 24 h followed by Aβ exposure for various periods. Neuronal survival and several cellular parameters (cleaved caspase 3, synaptophysin labeling and ROS) were assessed. In addition, the influence of these treatments on relevant signaling pathways was investigated with Western blotting. In terms of the treatment timing, our data indicated that coconut oil rescues cells pre-exposed to Aβ for 1 or 6 h, but is less effective when the pre-exposure has been 24 h. However, pretreatment with coconut oil prior to Aβ exposure showed the best outcomes. Treatment with octanoic or lauric acid also provided protection against Aβ, but was not as effective as the complete oil. The coconut oil treatment reduced the number of cells with cleaved caspase and ROS labeling, as well as rescuing the loss of synaptophysin labeling observed with Aβ treatment. Treatment with coconut oil, as well as octanoic, decanoic and lauric acids, resulted in a modest increase in ketone bodies compared to controls. The biochemical data suggest that Akt and ERK activation may contribute to the survival promoting influence of coconut oil. This was supported by observations that a PI3-Kinase inhibitor blocked the rescue effect of CoOil on Aβ amyloid toxicity. Further studies into the mechanisms of action of coconut oil and its constituent medium chain fatty acids are warranted

  1. Overexpression of BAG3 Attenuates Hypoxia-Induced Cardiomyocyte Apoptosis by Inducing Autophagy.

    Science.gov (United States)

    Zhang, Jiankai; He, Zhangyou; Xiao, Wenjian; Na, Qingqing; Wu, Tianxiu; Su, Kaixin; Cui, Xiaojun

    2016-01-01

    Hypoxia is a well-known factor in the promotion of apoptosis, which contributes to the development of numerous cardiac diseases, such as heart failure and myocardial infarction. Inhibiting apoptosis is an important therapeutic strategy for the treatment of related heart diseases caused by ischemia/hypoxic injury. Previous studies have demonstrated that BAG3 plays an important role in cardiomyocyte apoptosis and survival. However, the role of BAG3 in hypoxia-induced cardiomyocyte apoptosis remains to be clarified. Here, we demonstrate that BAG3 is induced by hypoxia stimuli in cultured cardiomyocytes. BAG3 expression level was measured in H9c2 cells treated with hypoxia for 48 h. Cell proliferation and apoptosis were tested using MTT assay and Annexin V FITC-PI staining assay, respectively. The mRNA or protein expression level of BAG3, LC3-I, LC3-II, Atg5, NF-x03BA;B p65 and phosphorylated NF-x03BA;B p65 were assessed by qRT-PCR and western blot assay, respectively. Resluts: Overexpression of BAG3 inhibited cell apoptosis and promoted proliferation in hypoxia-injured H9c2 cells. Furthermore, autophagy and NF-x03BA;B were activated by BAG3 overexpression, and the NF-x03BA;B inhibitor PDTC could inhibit the activation of autophagy induced by BAG3 overexpression. In addition, the autophagy inhibitor 3-MA partly impeded the inhibitory effect of BAG3 on hypoxia-induced cardiomyocyte apoptosis. these results suggested that overexpression of BAG3 promoted cell proliferation and inhibited apoptosis by activating autophagy though the NF-x03BA;B signaling pathway in hypoxia-injured cardiomyocytes. © 2016 The Author(s) Published by S. Karger AG, Basel.

  2. Overexpression of BAG3 Attenuates Hypoxia-Induced Cardiomyocyte Apoptosis by Inducing Autophagy

    Directory of Open Access Journals (Sweden)

    Jiankai Zhang

    2016-07-01

    Full Text Available Background: Hypoxia is a well-known factor in the promotion of apoptosis, which contributes to the development of numerous cardiac diseases, such as heart failure and myocardial infarction. Inhibiting apoptosis is an important therapeutic strategy for the treatment of related heart diseases caused by ischemia/hypoxic injury. Previous studies have demonstrated that BAG3 plays an important role in cardiomyocyte apoptosis and survival. However, the role of BAG3 in hypoxia-induced cardiomyocyte apoptosis remains to be clarified. Here, we demonstrate that BAG3 is induced by hypoxia stimuli in cultured cardiomyocytes. Methods: BAG3 expression level was measured in H9c2 cells treated with hypoxia for 48 h. Cell proliferation and apoptosis were tested using MTT assay and Annexin V FITC-PI staining assay, respectively. The mRNA or protein expression level of BAG3, LC3-I, LC3-II, Atg5, NF-κB p65 and phosphorylated NF-κB p65 were assessed by qRT-PCR and western blot assay, respectively. Resluts: Overexpression of BAG3 inhibited cell apoptosis and promoted proliferation in hypoxia-injured H9c2 cells. Furthermore, autophagy and NF-κB were activated by BAG3 overexpression, and the NF-κB inhibitor PDTC could inhibit the activation of autophagy induced by BAG3 overexpression. In addition, the autophagy inhibitor 3-MA partly impeded the inhibitory effect of BAG3 on hypoxia-induced cardiomyocyte apoptosis. Conclusion: these results suggested that overexpression of BAG3 promoted cell proliferation and inhibited apoptosis by activating autophagy though the NF-κB signaling pathway in hypoxia-injured cardiomyocytes.

  3. Role of alpha- and beta-adrenergic receptors in cardiomyocyte differentiation from murine-induced pluripotent stem cells.

    Science.gov (United States)

    Li, Xiao-Li; Zeng, Di; Chen, Yan; Ding, Lu; Li, Wen-Ju; Wei, Ting; Ou, Dong-Bo; Yan, Song; Wang, Bin; Zheng, Qiang-Sun

    2017-02-01

    Induced pluripotent stem cell (iPSC)-derived cardiomyocytes are a promising source of cells for regenerative heart disease therapies, but progress towards their use has been limited by their low differentiation efficiency and high cellular heterogeneity. Previous studies have demonstrated expression of adrenergic receptors (ARs) in stem cells after differentiation; however, roles of ARs in fate specification of stem cells, particularly in cardiomyocyte differentiation and development, have not been characterized. Murine-induced pluripotent stem cells (miPSCs) were cultured in hanging drops to form embryoid bodies, cells of which were then differentiated into cardiomyocytes. To determine whether ARs regulated miPSC differentiation into cardiac lineages, effects of the AR agonist, epinephrine (EPI), on miPSC differentiation and underlying signalling mechanisms, were evaluated. Treatment with EPI, robustly enhanced miPSC cardiac differentiation, as indicated by increased expression levels of cardiac-specific markers, GATA4, Nkx2.5 and Tnnt2. Although β-AR signalling is the foremost signalling pathway in cardiomyocytes, EPI-enhanced cardiac differentiation depended more on α-AR signalling than β-AR signalling. In addition, selective activation of α 1 -AR signalling with specific agonists induced vigorous cardiomyocyte differentiation, whereas selective activation of α 2 - or β-AR signalling induced no or less differentiation, respectively. EPI- and α 1 -AR-dependent cardiomyocyte differentiation from miPSCs occurred through specific promotion of CPC proliferation via the MEK-ERK1/2 pathway and regulation of miPS cell-cycle progression. These results demonstrate that activation of ARs, particularly of α 1 -ARs, promoted miPSC differentiation into cardiac lineages via MEK-ERK1/2 signalling. © 2016 John Wiley & Sons Ltd.

  4. The Association of German Radiation Protection Physicians: Its origins, pathways and objectives

    International Nuclear Information System (INIS)

    Beck, H.R.

    1985-01-01

    The paper highlights three principles which will hold good for future development, too: 1) In terms of the protection of the health of personnel, patients and total population, radiation protection constitutes a primary task for the physician; 2) The association endeavours to make a realistic assessment of the radiation hazard and dismisses both its overestimation and its underestimation; 3) Training and vocational qualification appear to be more important for radiation protection than perfectionist-bureaucratic provisions. (orig./HSCH) [de

  5. Directed Differentiation of Zebrafish Pluripotent Embryonic Cells to Functional Cardiomyocytes

    Directory of Open Access Journals (Sweden)

    Yao Xiao

    2016-09-01

    Full Text Available A cardiomyocyte differentiation in vitro system from zebrafish embryos remains to be established. Here, we have determined pluripotency window of zebrafish embryos by analyzing their gene-expression patterns of pluripotency factors together with markers of three germ layers, and have found that zebrafish undergoes a very narrow period of pluripotency maintenance from zygotic genome activation to a brief moment after oblong stage. Based on the pluripotency and a combination of appropriate conditions, we established a rapid and efficient method for cardiomyocyte generation in vitro from primary embryonic cells. The induced cardiomyocytes differentiated into functional and specific cardiomyocyte subtypes. Notably, these in vitro generated cardiomyocytes exhibited typical contractile kinetics and electrophysiological features. The system provides a new paradigm of cardiomyocyte differentiation from primary embryonic cells in zebrafish. The technology provides a new platform for the study of heart development and regeneration, in addition to drug discovery, disease modeling, and assessment of cardiotoxic agents.

  6. The soluble guanylyl cyclase activator bay 58-2667 selectively limits cardiomyocyte hypertrophy.

    Directory of Open Access Journals (Sweden)

    Jennifer C Irvine

    Full Text Available Although evidence now suggests cGMP is a negative regulator of cardiac hypertrophy, the direct consequences of the soluble guanylyl cyclase (sGC activator BAY 58-2667 on cardiac remodeling, independent of changes in hemodynamic load, has not been investigated. In the present study, we tested the hypothesis that the NO(•-independent sGC activator BAY 58-2667 inhibits cardiomyocyte hypertrophy in vitro. Concomitant impact of BAY 58-2667 on cardiac fibroblast proliferation, and insights into potential mechanisms of action, were also sought. Results were compared to the sGC stimulator BAY 41-2272.Neonatal rat cardiomyocytes were incubated with endothelin-1 (ET(1, 60nmol/L in the presence and absence of BAY 41-2272 and BAY 58-2667 (0.01-0.3 µmol/L. Hypertrophic responses and its triggers, as well as cGMP signaling, were determined. The impact of both sGC ligands on basal and stimulated cardiac fibroblast proliferation in vitro was also determined.We now demonstrate that BAY 58-2667 (0.01-0.3 µmol/L elicited concentration-dependent antihypertrophic actions, inhibiting ET(1-mediated increases in cardiomyocyte 2D area and de novo protein synthesis, as well as suppressing ET(1-induced cardiomyocyte superoxide generation. This was accompanied by potent increases in cardiomyocyte cGMP accumulation and activity of its downstream signal, vasodilator-stimulated phosphoprotein (VASP, without elevating cardiomyocyte cAMP. In contrast, submicromolar concentrations of BAY 58-2667 had no effect on basal or stimulated cardiac fibroblast proliferation. Indeed, only at concentrations ≥10 µmol/L was inhibition of cardiac fibrosis seen in vitro. The effects of BAY 58-2667 in both cell types were mimicked by BAY 41-2272.Our results demonstrate that BAY 58-2667 elicits protective, cardiomyocyte-selective effects in vitro. These actions are associated with sGC activation and are evident in the absence of confounding hemodynamic factors, at low (submicromolar

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  1. File list: NoD.CDV.50.AllAg.Cardiomyocytes [Chip-atlas[Archive

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  2. Differentiation and characterization of rhesus monkey atrial and ventricular cardiomyocytes from induced pluripotent stem cells.

    Science.gov (United States)

    Zhang, Xiaoqian; Cao, Henghua; Bai, Shuyun; Huo, Weibang; Ma, Yue

    2017-04-01

    The combination of non-human primate animals and their induced pluripotent stem cell derived cardiomyocytes (iPSC-CMs) provides not only transplantation models for cell-based therapy of heart diseases, but also opportunities for heart-related drug research on both cellular and animal levels. However, the subtypes and electrophysiology properties of non-human primate iPSC-CMs hadn't been detailed characterized. In this study, we generated rhesus monkey induced pluripotent stem cells (riPSCs), and efficiently differentiated them into ventricular or atrial cardiomyocytes by modulating retinoic acid (RA) pathways. Our results revealed that the electrophysiological characteristics and response to canonical drugs of riPSC-CMs were similar with those of human pluripotent stem cell derived CMs. Therefore, rhesus monkeys and their iPSC-CMs provide a powerful and practicable system for heart related biomedical research. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

  3. (Re-)programming of subtype specific cardiomyocytes.

    Science.gov (United States)

    Hausburg, Frauke; Jung, Julia Jeannine; Hoch, Matti; Wolfien, Markus; Yavari, Arash; Rimmbach, Christian; David, Robert

    2017-10-01

    Adult cardiomyocytes (CMs) possess a highly restricted intrinsic regenerative potential - a major barrier to the effective treatment of a range of chronic degenerative cardiac disorders characterized by cellular loss and/or irreversible dysfunction and which underlies the majority of deaths in developed countries. Both stem cell programming and direct cell reprogramming hold promise as novel, potentially curative approaches to address this therapeutic challenge. The advent of induced pluripotent stem cells (iPSCs) has introduced a second pluripotent stem cell source besides embryonic stem cells (ESCs), enabling even autologous cardiomyocyte production. In addition, the recent achievement of directly reprogramming somatic cells into cardiomyocytes is likely to become of great importance. In either case, different clinical scenarios will require the generation of highly pure, specific cardiac cellular-subtypes. In this review, we discuss these themes as related to the cardiovascular stem cell and programming field, including a focus on the emergent topic of pacemaker cell generation for the development of biological pacemakers and in vitro drug testing. Copyright © 2017 Elsevier B.V. All rights reserved.

  4. Blockage of NOX2/MAPK/NF-κB Pathway Protects Photoreceptors against Glucose Deprivation-Induced Cell Death

    Directory of Open Access Journals (Sweden)

    Bin Fan

    2017-01-01

    Full Text Available Acute energy failure is one of the critical factors contributing to the pathogenic mechanisms of retinal ischemia. Our previous study demonstrated that glucose deprivation can lead to a caspase-dependent cell death of photoreceptors. The aim of this study was to decipher the upstream signal pathway in glucose deprivation- (GD- induced cell death. We mimicked acute energy failure by using glucose deprivation in photoreceptor cells (661W cells. GD-induced oxidative stress was evaluated by measuring ROS with the DCFH-DA assay and HO-1 expression by Western blot analysis. The activation of NOX2/MAPK/NF-κB signal was assessed by Western blot and immunohistochemical assays. The roles of these signals in GD-induced cell death were measured by using their specific inhibitors. Inhibition of Rac-1 and NOX2 suppressed GD-induced oxidative stress and protected photoreceptors against GD-induced cell death. NOX2 was an upstream signal in the caspase-dependent cell death cascade, yet the downstream MAPK pathways were activated and blocking MAPK signals rescued 661W cells from GD-induced death. In addition, GD caused the activation of NF-κB signal and inhibiting NF-κB significantly protected 661W cells. These observations may provide insights for treating retinal ischemic diseases and protecting retinal neurons from ischemia-induced cell death.

  5. EGb 761 Protects Cardiac Microvascular Endothelial Cells against Hypoxia/Reoxygenation Injury and Exerts Inhibitory Effect on the ATM Pathway.

    Science.gov (United States)

    Zhang, Chao; Wang, Deng-Feng; Zhang, Zhuang; Han, Dong; Yang, Kan

    2017-03-28

    Ginkgo bilob a extract (EGb 761) has been widely used clinically to reduce myocardial ischemia reperfusion injury (MIRI). Microvascular endothelial cells (MVECs) may be a proper cellular model in vitro for the effect and mechanism study against MIRI. However, the protective effect of EGb 761 on MVECs resisting hypoxia/reoxygenation (H/R) injury is little reported. In this study, H/R-injured MVECs were treated with EGb 761, and then the cell viability, apoptosis, ROS production, SOD activity, caspase-3 activity, and protein level of ATM, γ-H2AX, p53, and Bax were measured. ATM siRNA was transfected to study the changes of protein in the ATM pathway. EGb 761 presented protective effect on H/R-injured MVECs, with decreasing cell death, apoptosis, and ROS, and elevated SOD activity. Next, EGb 761 could inhibit H/R-induced ATM, γ-H2AX, p53, and Bax in a dose-dependent manner. Moreover, ATM siRNA also could inhibit H/R-induced ATM, γ-H2AX, p53, and Bax. Overall, these findings verify that EGb 761 protects cardiac MVECs from H/R injury, and for the first time, illustrate the influence on the ATM pathway and apoptosis by EGb 761 via dampening ROS.

  6. An Ixodes ricinus Tick Salivary Lectin Pathway Inhibitor Protects Borrelia burgdorferi sensu lato from Human Complement

    NARCIS (Netherlands)

    Wagemakers, Alex; Coumou, Jeroen; Schuijt, Tim J.; Oei, Anneke; Nijhof, Ard M.; van 't Veer, Cornelis; van der Poll, Tom; Bins, Adriaan D.; Hovius, Joppe W. R.

    2016-01-01

    We previously identified tick salivary lectin pathway inhibitor (TSLPI) in Ixodes scapularis, a vector for Borrelia burgdorferi sensu stricto (s.s.) in North America. TSLPI is a salivary protein facilitating B. burgdorferi s.s. transmission and acquisition by inhibiting the host lectin complement

  7. Pathway of protection: Ethnic identity, self-esteem, and substance use among multiracial youth.

    Science.gov (United States)

    Fisher, Sycarah; Zapolski, Tamika C B; Sheehan, Chelsea; Barnes-Najor, Jessica

    2017-09-01

    Fifty percent of adolescents have tried an illicit drug and 70% have tried alcohol by the end of high school, with even higher rates among multiracial youth. Ethnic identity is a protective factor against substance use for minority groups. However, little is known about the mechanisms that facilitate its protective effects, and even less is known about this relationship for multiracial youth. The purpose of the present study was to examine the protective effect of ethnic identity on substance use and to determine whether this relationship operated indirectly through self-esteem, a strong predictor of substance use for among adolescent populations. Participants included 468 multiracial youth in grades six through 12 (53% female). The results found that ethnic identity was indeed related to substance use, partially through changes in self-esteem. Findings from this study contribute to our understanding and development of models of risk and protection for an understudied population. Copyright © 2017. Published by Elsevier Ltd.

  8. Astrocytes protect neurons against methylmercury via ATP/P2Y(1) receptor-mediated pathways in astrocytes.

    Science.gov (United States)

    Noguchi, Yusuke; Shinozaki, Youichi; Fujishita, Kayoko; Shibata, Keisuke; Imura, Yoshio; Morizawa, Yosuke; Gachet, Christian; Koizumi, Schuichi

    2013-01-01

    Methylmercury (MeHg) is a well known environmental pollutant that induces serious neuronal damage. Although MeHg readily crosses the blood-brain barrier, and should affect both neurons and glial cells, how it affects glia or neuron-to-glia interactions has received only limited attention. Here, we report that MeHg triggers ATP/P2Y1 receptor signals in astrocytes, thereby protecting neurons against MeHg via interleukin-6 (IL-6)-mediated pathways. MeHg increased several mRNAs in astrocytes, among which IL-6 was the highest. For this, ATP/P2Y1 receptor-mediated mechanisms were required because the IL-6 production was (i) inhibited by a P2Y1 receptor antagonist, MRS2179, (ii) abolished in astrocytes obtained from P2Y1 receptor-knockout mice, and (iii) mimicked by exogenously applied ATP. In addition, (iv) MeHg released ATP by exocytosis from astrocytes. As for the intracellular mechanisms responsible for IL-6 production, p38 MAP kinase was involved. MeHg-treated astrocyte-conditioned medium (ACM) showed neuro-protective effects against MeHg, which was blocked by anti-IL-6 antibody and was mimicked by the application of recombinant IL-6. As for the mechanism of neuro-protection by IL-6, an adenosine A1 receptor-mediated pathway in neurons seems to be involved. Taken together, when astrocytes sense MeHg, they release ATP that autostimulates P2Y1 receptors to upregulate IL-6, thereby leading to A1 receptor-mediated neuro-protection against MeHg.

  9. Dietary Fiber and Bacterial SCFA Enhance Oral Tolerance and Protect against Food Allergy through Diverse Cellular Pathways.

    Science.gov (United States)

    Tan, Jian; McKenzie, Craig; Vuillermin, Peter J; Goverse, Gera; Vinuesa, Carola G; Mebius, Reina E; Macia, Laurence; Mackay, Charles R

    2016-06-21

    The incidence of food allergies in western countries has increased dramatically in recent decades. Tolerance to food antigens relies on mucosal CD103(+) dendritic cells (DCs), which promote differentiation of regulatory T (Treg) cells. We show that high-fiber feeding in mice improved oral tolerance and protected from food allergy. High-fiber feeding reshaped gut microbial ecology and increased the release of short-chain fatty acids (SCFAs), particularly acetate and butyrate. High-fiber feeding enhanced oral tolerance and protected against food allergy by enhancing retinal dehydrogenase activity in CD103(+) DC. This protection depended on vitamin A in the diet. This feeding regimen also boosted IgA production and enhanced T follicular helper and mucosal germinal center responses. Mice lacking GPR43 or GPR109A, receptors for SCFAs, showed exacerbated food allergy and fewer CD103(+) DCs. Dietary elements, including fiber and vitamin A, therefore regulate numerous protective pathways in the gastrointestinal tract, necessary for immune non-responsiveness to food antigens. Copyright © 2016 The Author(s). Published by Elsevier Inc. All rights reserved.

  10. Virus-like particles activate type I interferon pathways to facilitate post-exposure protection against Ebola virus infection.

    Directory of Open Access Journals (Sweden)

    Natarajan Ayithan

    Full Text Available Ebola virus (EBOV causes a severe hemorrhagic disease with high fatality. Virus-like particles (VLPs are a promising vaccine candidate against EBOV. We recently showed that VLPs protect mice from lethal EBOV infection when given before or after viral infection. To elucidate pathways through which VLPs confer post-exposure protection, we investigated the role of type I interferon (IFN signaling. We found that VLPs lead to accelerated induction of IFN stimulated genes (ISGs in liver and spleen of wild type mice, but not in Ifnar-/- mice. Accordingly, EBOV infected Ifnar-/- mice, unlike wild type mice succumbed to death even after VLP treatment. The ISGs induced in wild type mice included anti-viral proteins and negative feedback factors known to restrict viral replication and excessive inflammatory responses. Importantly, proinflammatory cytokine/chemokine expression was much higher in WT mice without VLPs than mice treated with VLPs. In EBOV infected Ifnar-/- mice, however, uninhibited viral replication and elevated proinflammatory factor expression ensued, irrespective of VLP treatment, supporting the view that type I IFN signaling helps to limit viral replication and attenuate inflammatory responses. Further analyses showed that VLP protection requires the transcription factor, IRF8 known to amplify type I IFN signaling in dendritic cells and macrophages, the probable sites of initial EBOV infection. Together, this study indicates that VLPs afford post-exposure protection by promoting expeditious initiation of type I IFN signaling in the host.

  11. Dietary Fiber and Bacterial SCFA Enhance Oral Tolerance and Protect against Food Allergy through Diverse Cellular Pathways

    Directory of Open Access Journals (Sweden)

    Jian Tan

    2016-06-01

    Full Text Available The incidence of food allergies in western countries has increased dramatically in recent decades. Tolerance to food antigens relies on mucosal CD103+ dendritic cells (DCs, which promote differentiation of regulatory T (Treg cells. We show that high-fiber feeding in mice improved oral tolerance and protected from food allergy. High-fiber feeding reshaped gut microbial ecology and increased the release of short-chain fatty acids (SCFAs, particularly acetate and butyrate. High-fiber feeding enhanced oral tolerance and protected against food allergy by enhancing retinal dehydrogenase activity in CD103+ DC. This protection depended on vitamin A in the diet. This feeding regimen also boosted IgA production and enhanced T follicular helper and mucosal germinal center responses. Mice lacking GPR43 or GPR109A, receptors for SCFAs, showed exacerbated food allergy and fewer CD103+ DCs. Dietary elements, including fiber and vitamin A, therefore regulate numerous protective pathways in the gastrointestinal tract, necessary for immune non-responsiveness to food antigens.

  12. RP105 Protects Against Apoptosis in Ischemia/Reperfusion-Induced Myocardial Damage in Rats by Suppressing TLR4-Mediated Signaling Pathways

    Directory of Open Access Journals (Sweden)

    Jun Yang

    2015-07-01

    Full Text Available Background: Myocardial apoptosis is heavily implicated in the myocardial damage caused by ischemia-reperfusion (I/R. Toll-like receptor 4 (TLR4 is a potent inducer of these apoptotic cascades. In contrast, the radioprotective 105 kDa protein (RP105 is a specific negative regulator of TLR4 signaling pathways. However, the precise mechanisms by which RP105 inhibits myocardium apoptosis via TLR4-associated pathways during I/R is not fully understood. Methods: We utilized a rat model of myocardial ischemic reperfusion injury (MIRI. Animals were pre-treated with Ad-EGFP adenovirus, Ad-EGFP-RP105 adenovirus, saline, or nothing (sham. After three days, rats underwent a 30min left anterior descending coronary artery occlusion and a 4h reperfusion. Mycardial tissue was assessed by immunohistochemistry, TUNEL-staining, Western blot, quantitative RT-PCR, and a morphometric assay. Results: RP105 overexpression resulted in a reduction in infarct size, fewer TUNEL-positive cardiomyocytes, and a reduction in mitochondrial-associated apoptosis cascade activity. Further, RP105 overexpression repressed I/R-induced myocardial injury by attenuating myocardial apoptosis. This was mediated by inhibiting TLR4 activation and the phosphorylation of P38MAPK and the downstream transcription factor AP-1. Conclusion: RP105 overexpression leads to the de-activation of TLR4, P38MAPK, and AP-1 signaling pathways, and subsequently represses apoptotic cascades and ensuing damage of myocardial ischemic reperfusion. These findings may become the basis of a novel therapeutic approach for reducing of cardiac damage caused by MIRI.

  13. Klotho Protects Dopaminergic Neuron Oxidant-Induced Degeneration by Modulating ASK1 and p38 MAPK Signaling Pathways.

    Directory of Open Access Journals (Sweden)

    Reynolds K Brobey

    Full Text Available Klotho transgenic mice exhibit resistance to oxidative stress as measured by their urinal levels of 8-hydroxy-2-deoxyguanosine, albeit this anti-oxidant defense mechanism has not been locally investigated in the brain. Here, we tested the hypothesis that the reactive oxygen species (ROS-sensitive apoptosis signal-regulating kinase 1 (ASK1/p38 MAPK pathway regulates stress levels in the brain of these mice and showed that: 1 the ratio of free ASK1 to thioredoxin (Trx-bound ASK1 is relatively lower in the transgenic brain whereas the reverse is true for the Klotho knockout mice; 2 the reduced p38 activation level in the transgene corresponds to higher level of ASK1-bound Trx, while the KO mice showed elevated p38 activation and lower level of-bound Trx; and 3 that 14-3-3ζ is hyper phosphorylated (Ser-58 in the transgene which correlated with increased monomer forms. In addition, we evaluated the in vivo robustness of the protection by challenging the brains of Klotho transgenic mice with a neurotoxin, MPTP and analyzed for residual neuron numbers and integrity in the substantia nigra pars compacta. Our results show that Klotho overexpression significantly protects dopaminergic neurons against oxidative damage, partly by modulating p38 MAPK activation level. Our data highlight the importance of ASK1/p38 MAPK pathway in the brain and identify Klotho as a possible anti-oxidant effector.

  14. Role of the Antigen Capture Pathway in the Induction of a Neutralizing Antibody Response to Anthrax Protective Antigen

    Directory of Open Access Journals (Sweden)

    Anita Verma

    2018-02-01

    Full Text Available Toxin neutralizing antibodies represent the major mode of protective immunity against a number of toxin-mediated bacterial diseases, including anthrax; however, the cellular mechanisms that lead to optimal neutralizing antibody responses remain ill defined. Here we show that the cellular binding pathway of anthrax protective antigen (PA, the binding component of anthrax toxin, determines the toxin neutralizing antibody response to this antigen. PA, which binds cellular receptors and efficiently enters antigen-presenting cells by receptor-mediated endocytosis, was found to elicit robust anti-PA IgG and toxin neutralizing antibody responses. In contrast, a receptor binding-deficient mutant of PA, which does not bind receptors and only inefficiently enters antigen-presenting cells by macropinocytosis, elicited very poor antibody responses. A chimeric protein consisting of the receptor binding-deficient PA mutant tethered to the binding subunit of cholera toxin, which efficiently enters cells using the cholera toxin receptor rather than the PA receptor, elicited an anti-PA IgG antibody response similar to that elicited by wild-type PA; however, the chimeric protein elicited a poor toxin neutralizing antibody response. Taken together, our results demonstrate that the antigen capture pathway can dictate the magnitudes of the total IgG and toxin neutralizing antibody responses to PA as well as the ratio of the two responses.

  15. Hyperin protects against LPS-induced acute kidney injury by inhibiting TLR4 and NLRP3 signaling pathways

    Science.gov (United States)

    Chunzhi, Gong; Zunfeng, Li; Chengwei, Qin; Xiangmei, Bu; Jingui, Yu

    2016-01-01

    Hyperin is a flavonoid compound derived from Ericaceae, Guttifera, and Celastraceae that has been shown to have various biological effects, such as anti-inflammatory and anti-oxidant effects. However, there is no evidence to show the protective effects of hyperin on lipopolysaccharide (LPS)-induced acute kidney injury (AKI). Therefore, we investigated the protective effects and mechanism of hyperin on LPS-induced AKI in mice. The levels of TNF-α, IL-6, and IL-1β were tested by ELISA. The effects of hyperin on blood urea nitrogen (BUN) and serum creatinine were also detected. In addition, the expression of TLR4, NF-κB, and NLRP3 were detected by western blot analysis. The results showed that hyperin significantly inhibited LPS-induced TNF-α, IL-6, and IL-1β production. The levels of BUN and creatinine were also suppressed by hyperin. Furthermore, LPS-induced TLR4 expression and NF-κB activation were also inhibited by hyperin. In addition, treatment of hyperin dose-dependently inhibited LPS-induced NLRP3 signaling pathway. In conclusion, the results showed that hyperin inhibited LPS-induced inflammatory response by inhibiting TLR4 and NLRP3 signaling pathways. Hyperin has potential application prospects in the treatment of sepsis-induced AKI. PMID:27813491

  16. Pathway Aggregation in the Risk Assessment of Proliferation Resistance and Physical Protection (PR&PP) of Nuclear Energy Systems

    International Nuclear Information System (INIS)

    2015-01-01

    The framework for Proliferation Resistance and Physical Protection (PR & PP) evaluation is to define a set of challenges, to obtain the system responses, and to assess the outcomes. The assessment of outcomes heavily relies on pathways, defined as sequences of events or actions that could potentially be followed by a State or a group of individuals in order to achieve a proliferation objective, with the defined threats as initiating events. There may be large number of segments connecting pathway stages (e.g. acquisition, processing, and fabrication for PR) which can lead to even larger number of pathways or scenarios through possible different combinations of segment connections, each with associated probabilities contributing to the overall risk. Clustering of these scenarios in specified stage attribute intervals is important for their tractable analysis and outcome assessment. A software tool for scenario generation and clustering (OSUPR) is developed that utilizes the PRCALC code developed at the Brookhaven National Laboratory for scenario generation and the K- means, mean shift and adaptive mean shift algorithms as possible clustering schemes. The results of the study using the Example Sodium Fast Breeder as an example system show that clustering facilitates the probabilistic or deterministic analysis of scenarios to identify system vulnerabilities and communication of the major risk contributors to stakeholders. The results of the study also show that the mean shift algorithm has the most potential for assisting the analysis of the scenarios generated by PRCALC.

  17. Pathway Aggregation in the Risk Assessment of Proliferation Resistance and Physical Protection (PR&PP) of Nuclear Energy Systems

    Energy Technology Data Exchange (ETDEWEB)

    Aldemir, Tunc [Ohio State Univ., Columbus, OH (United States); Denning, Richard [Ohio State Univ., Columbus, OH (United States); Catalyurek, Umit [Ohio State Univ., Columbus, OH (United States); Yilmaz, Alper [Ohio State Univ., Columbus, OH (United States); Yue, Meng [Brookhaven National Lab. (BNL), Upton, NY (United States); Cheng, Lap-Yan [Brookhaven National Lab. (BNL), Upton, NY (United States)

    2015-01-23

    The framework for Proliferation Resistance and Physical Protection (PR & PP) evaluation is to define a set of challenges, to obtain the system responses, and to assess the outcomes. The assessment of outcomes heavily relies on pathways, defined as sequences of events or actions that could potentially be followed by a State or a group of individuals in order to achieve a proliferation objective, with the defined threats as initiating events. There may be large number of segments connecting pathway stages (e.g. acquisition, processing, and fabrication for PR) which can lead to even larger number of pathways or scenarios through possible different combinations of segment connections, each with associated probabilities contributing to the overall risk. Clustering of these scenarios in specified stage attribute intervals is important for their tractable analysis and outcome assessment. A software tool for scenario generation and clustering (OSUPR) is developed that utilizes the PRCALC code developed at the Brookhaven National Laboratory for scenario generation and the K- means, mean shift and adaptive mean shift algorithms as possible clustering schemes. The results of the study using the Example Sodium Fast Breeder as an example system show that clustering facilitates the probabilistic or deterministic analysis of scenarios to identify system vulnerabilities and communication of the major risk contributors to stakeholders. The results of the study also show that the mean shift algorithm has the most potential for assisting the analysis of the scenarios generated by PRCALC.

  18. Kaempferol Attenuates Cardiac Hypertrophy via Regulation of ASK1/MAPK Signaling Pathway and Oxidative Stress.

    Science.gov (United States)

    Feng, Hong; Cao, Jianlei; Zhang, Guangyu; Wang, Yanggan

    2017-07-01

    Kaempferol has been demonstrated to provide benefits for the treatment of atherosclerosis, coronary heart disease, hyperlipidemia, and diabetes through its antioxidant and anti-inflammatory properties. However, its role in cardiac hypertrophy remains to be elucidated. The aim of our study was to investigate the effects of kaempferol on cardiac hypertrophy and the underlying mechanism. Mice subjected to aorta banding were treated with or without kaempferol (100 mg/kg/d, p. o.) for 6 weeks. Echocardiography was performed to evaluate cardiac function. Mice hearts were collected for pathological observation and molecular mechanism investigation. H9c2 cardiomyocytes were stimulated with or without phenylephrine for in vitro study. Kaempferol significantly attenuated cardiac hypertrophy induced by aorta banding as evidenced by decreased cardiomyocyte areas and interstitial fibrosis, accompanied with improved cardiac functions and decreased apoptosis. The ASK1/MAPK signaling pathways (JNK1/2 and p38) were markedly activated in the aorta banding mouse heart but inhibited by kaempferol treatment. In in vitro experiments, kaempferol also inhibited the activity of ASK1/JNK1/2/p38 signaling pathway and the enlargement of H9c2 cardiomyocytes. Furthermore, our study revealed that kaempferol could protect the mouse heart and H9c2 cells from pathological oxidative stress. Our investigation indicated that treatment with kaempferol protects against cardiac hypertrophy, and its cardioprotection may be partially explained by the inhibition of the ASK1/MAPK signaling pathway and the regulation of oxidative stress. Georg Thieme Verlag KG Stuttgart · New York.

  19. Acoustical sensing of cardiomyocyte cluster beating

    Energy Technology Data Exchange (ETDEWEB)

    Tymchenko, Nina; Kunze, Angelika [Dept. of Applied Physics, Chalmers University of Technology, 412 96 Göteborg (Sweden); Dahlenborg, Kerstin [Cellectis, 413 46 Göteborg (Sweden); Svedhem, Sofia, E-mail: sofia.svedhem@chalmers.se [Dept. of Applied Physics, Chalmers University of Technology, 412 96 Göteborg (Sweden); Steel, Daniella [Cellectis, 413 46 Göteborg (Sweden)

    2013-06-14

    Highlights: •An example of the application of QCM-D to live cell studies. •Detection of human pluripotent stem cell-derived cardiomyocyte cluster beating. •Clusters were studied in a thin liquid film and in a large liquid volume. •The QCM-D beating profile provides an individual fingerprint of the hPS-CMCs. -- Abstract: Spontaneously beating human pluripotent stem cell-derived cardiomyocytes clusters (CMCs) represent an excellent in vitro tool for studies of human cardiomyocyte function and for pharmacological cardiac safety assessment. Such testing typically requires highly trained operators, precision plating, or large cell quantities, and there is a demand for real-time, label-free monitoring of small cell quantities, especially rare cells and tissue-like structures. Array formats based on sensing of electrical or optical properties of cells are being developed and in use by the pharmaceutical industry. A potential alternative to these techniques is represented by the quartz crystal microbalance with dissipation monitoring (QCM-D) technique, which is an acoustic surface sensitive technique that measures changes in mass and viscoelastic properties close to the sensor surface (from nm to μm). There is an increasing number of studies where QCM-D has successfully been applied to monitor properties of cells and cellular processes. In the present study, we show that spontaneous beating of CMCs on QCM-D sensors can be clearly detected, both in the frequency and the dissipation signals. Beating rates in the range of 66–168 bpm for CMCs were detected and confirmed by simultaneous light microscopy. The QCM-D beating profile was found to provide individual fingerprints of the hPS-CMCs. The presented results point towards acoustical assays for evaluation cardiotoxicity.

  20. Polycystin-2-dependent control of cardiomyocyte autophagy.

    Science.gov (United States)

    Criollo, Alfredo; Altamirano, Francisco; Pedrozo, Zully; Schiattarella, Gabriele G; Li, Dan L; Rivera-Mejías, Pablo; Sotomayor-Flores, Cristian; Parra, Valentina; Villalobos, Elisa; Battiprolu, Pavan K; Jiang, Nan; May, Herman I; Morselli, Eugenia; Somlo, Stefan; de Smedt, Humbert; Gillette, Thomas G; Lavandero, Sergio; Hill, Joseph A

    2018-05-01

    Considerable evidence points to critical roles of intracellular Ca 2+ homeostasis in the modulation and control of autophagic activity. Yet, underlying molecular mechanisms remain unknown. Mutations in the gene (pkd2) encoding polycystin-2 (PC2) are associated with autosomal dominant polycystic kidney disease (ADPKD), the most common inherited nephropathy. PC2 has been associated with impaired Ca 2+ handling in cardiomyocytes and indirect evidence suggests that this protein may be involved in autophagic control. Here, we investigated the role for PC2 as an essential regulator of Ca 2+ homeostasis and autophagy. Activation of autophagic flux triggered by mTOR inhibition either pharmacologically (rapamycin) or by means of nutrient depletion was suppressed in cells depleted of PC2. Moreover, cardiomyocyte-specific PC2 knockout mice (αMhc-cre;Pkd2 F/F mice) manifested impaired autophagic flux in the setting of nutrient deprivation. Stress-induced autophagy was blunted by intracellular Ca 2+ chelation using BAPTA-AM, whereas removal of extracellular Ca 2+ had no effect, pointing to a role of intracellular Ca 2+ homeostasis in stress-induced cardiomyocyte autophagy. To determine the link between stress-induced autophagy and PC2-induced Ca 2+ mobilization, we over-expressed either wild-type PC2 (WT) or a Ca 2+ -channel deficient PC2 mutant (PC2-D509V). PC2 over-expression increased autophagic flux, whereas PC2-D509V expression did not. Importantly, autophagy induction triggered by PC2 over-expression was attenuated by BAPTA-AM, supporting a model of PC2-dependent control of autophagy through intracellular Ca 2+ . Furthermore, PC2 ablation was associated with impaired Ca 2+ handling in cardiomyocytes marked by partial depletion of sarcoplasmic reticulum Ca 2+ stores. Finally, we provide evidence that Ca 2+ -mediated autophagy elicited by PC2 is a mechanism conserved across multiple cell types. Together, this study unveils PC2 as a novel regulator of autophagy acting

  1. Stanniocalcin-1 Protects a Mouse Model from Renal Ischemia-Reperfusion Injury by Affecting ROS-Mediated Multiple Signaling Pathways.

    Science.gov (United States)

    Liu, Dajun; Shang, Huiping; Liu, Ying

    2016-07-12

    Stanniocalcin-1 (STC-1) protects against renal ischemia-reperfusion injury (RIRI). However, the molecular mechanisms remain widely unknown. STC-1 inhibits reactive oxygen species (ROS), whereas most ROS-mediated pathways are associated with ischemic injury. Therefore, to explore the mechanism, the effects of STC-1 on ROS-medicated pathways were studied. Non-traumatic vascular clamps were used to establish RIRI mouse models. The serum levels of STC-1, interleukin-6 (IL-6), interferon (IFN) γ, P53, and capase-3 were measured by ELISA kits. Superoxide dismutase (SOD) and malondialdehyde (MDA) were measured by fluorescence spectrofluorometer. All these molecules changed significantly in a RIRI model mouse when compared with those in a sham control. Kidney cells were isolated from sham and model mice. STC-1 was overexpressed or knockout in these kidney cells. The molecules in ROS-medicated pathways were measured by real-time quantitative PCR and Western blot. The results showed that STC-1 is an effective ROS scavenger. The serum levels of STC-1, MDA and SOD activity were increased while the serum levels of IL-6, iIFN-γ, P53, and capase-3 were decreased in a model group when compared with a sham control (p ROS-mediated molecules. Therefore, STC-1 maybe improve anti-inflammation, anti-oxidant and anti-apoptosis activities by affecting ROS-mediated pathways, especially the phospho-modifications of the respective proteins, resulting in the increase of SOD and reduce of capase-3, p53, IL-6 and IFN-γ.

  2. Progranulin protects vascular endothelium against atherosclerotic inflammatory reaction via Akt/eNOS and nuclear factor-κB pathways.

    Science.gov (United States)

    Hwang, Hwan-Jin; Jung, Tae Woo; Hong, Ho Cheol; Choi, Hae Yoon; Seo, Ji-A; Kim, Sin Gon; Kim, Nan Hee; Choi, Kyung Mook; Choi, Dong Seop; Baik, Sei Hyun; Yoo, Hye Jin

    2013-01-01

    Atherosclerosis is considered a chronic inflammatory disease, initiated by activation and dysfunction of the endothelium. Recently, progranulin has been regarded as an important modulator of inflammatory processes; however, the role for prgranulin in regulating inflammation in vascular endothelial cells has not been described. Signaling pathways mediated by progranulin were analyzed in human umbilical vein endothelial cells (HUVECs) treated with progranulin. Progranulin significantly induced Akt and endothelial nitric oxide synthase (eNOS) phosphorylation in HUVECs, an effect that was blocked with Akt inhibitor. Furthermore, nitric oxide (NO) level, the end product of Akt/eNOS pathway, was significantly upregulated after progranulin treatment. Next, we showed that progranulin efficiently inhibited lipopolysaccharide (LPS)-mediated pro-inflammatory signaling. LPS-induced phosphorylation of IκB and nuclear factor-κB (NF-κB) levels decreased after progranulin treatment. Also, progranulin blocked translocation of NF-κB from the cytosol to the nucleus. In addition, progranulin significantly reduced the expression of vascular cell adhesion molecule-1 (VCAM-1) and intercellular adhesion molecule-1 (ICAM-1) by inhibiting binding of NF- κB to their promoter regions and blocked attachment of monocytes to HUVECs. Progranulin also significantly reduced the expression of tumor necrosis factor receptor-α (TNF-α) and monocyte chemo-attractant protein-1 (MCP-1), the crucial inflammatory molecules known to aggravate atherosclerosis. Progranulin efficiently inhibited LPS-mediated pro-inflammatory signaling in endothelial cells through activation of the Akt/eNOS pathway and attenuation of the NF-κB pathway, suggesting its protective roles in vascular endothelium against inflammatory reaction underlying atherosclerosis.

  3. Progranulin protects vascular endothelium against atherosclerotic inflammatory reaction via Akt/eNOS and nuclear factor-κB pathways.

    Directory of Open Access Journals (Sweden)

    Hwan-Jin Hwang

    Full Text Available OBJECTIVE: Atherosclerosis is considered a chronic inflammatory disease, initiated by activation and dysfunction of the endothelium. Recently, progranulin has been regarded as an important modulator of inflammatory processes; however, the role for prgranulin in regulating inflammation in vascular endothelial cells has not been described. METHOD AND RESULTS: Signaling pathways mediated by progranulin were analyzed in human umbilical vein endothelial cells (HUVECs treated with progranulin. Progranulin significantly induced Akt and endothelial nitric oxide synthase (eNOS phosphorylation in HUVECs, an effect that was blocked with Akt inhibitor. Furthermore, nitric oxide (NO level, the end product of Akt/eNOS pathway, was significantly upregulated after progranulin treatment. Next, we showed that progranulin efficiently inhibited lipopolysaccharide (LPS-mediated pro-inflammatory signaling. LPS-induced phosphorylation of IκB and nuclear factor-κB (NF-κB levels decreased after progranulin treatment. Also, progranulin blocked translocation of NF-κB from the cytosol to the nucleus. In addition, progranulin significantly reduced the expression of vascular cell adhesion molecule-1 (VCAM-1 and intercellular adhesion molecule-1 (ICAM-1 by inhibiting binding of NF- κB to their promoter regions and blocked attachment of monocytes to HUVECs. Progranulin also significantly reduced the expression of tumor necrosis factor receptor-α (TNF-α and monocyte chemo-attractant protein-1 (MCP-1, the crucial inflammatory molecules known to aggravate atherosclerosis. CONCLUSION: Progranulin efficiently inhibited LPS-mediated pro-inflammatory signaling in endothelial cells through activation of the Akt/eNOS pathway and attenuation of the NF-κB pathway, suggesting its protective roles in vascular endothelium against inflammatory reaction underlying atherosclerosis.

  4. Streptozotocin induced activation of oxidative stress responsive splenic cell signaling pathways: Protective role of arjunolic acid

    International Nuclear Information System (INIS)

    Manna, Prasenjit; Ghosh, Jyotirmoy; Das, Joydeep; Sil, Parames C.

    2010-01-01

    Present study investigates the beneficial role of arjunolic acid (AA) against the alteration in the cytokine levels and simultaneous activation of oxidative stress responsive signaling pathways in spleen under hyperglycemic condition. Diabetes was induced by injection of streptozotocin (STZ) (at a dose of 70 mg/kg body weight, injected in the tail vain). STZ administration elevated the levels of IL-2 as well as IFN-γ and attenuated the level of TNF-α in the sera of diabetic animals. In addition, hyperglycemia is also associated with the increased production of intracellular reactive intermediates resulting with the elevation in lipid peroxidation, protein carbonylation and reduction in intracellular antioxidant defense. Investigating the oxidative stress responsive cell signaling pathways, increased expressions (immunoreactive concentrations) of phosphorylated p65 as well as its inhibitor protein phospho IκBα and phosphorylated mitogen activated protein kinases (MAPKs) have been observed in diabetic spleen tissue. Studies on isolated splenocytes revealed that hyperglycemia caused disruption of mitochondrial membrane potential, elevation in the concentration of cytosolic cytochrome c as well as activation of caspase 3 leading to apoptotic cell death. Histological examination revealed that diabetic induction depleted the white pulp scoring which is in agreement with the reduced immunological response. Treatment with AA prevented the hyperglycemia and its associated pathogenesis in spleen tissue. Results suggest that AA might act as an anti-diabetic and immunomodulatory agent against hyperglycemia.

  5. A non-cardiomyocyte autonomous mechanism of cardioprotection involving the SLO1 BK channel

    Directory of Open Access Journals (Sweden)

    Andrew P. Wojtovich

    2013-03-01

    Full Text Available Opening of BK-type Ca2+ activated K+ channels protects the heart against ischemia-reperfusion (IR injury. However, the location of BK channels responsible for cardioprotection is debated. Herein we confirmed that openers of the SLO1 BK channel, NS1619 and NS11021, were protective in a mouse perfused heart model of IR injury. As anticipated, deletion of the Slo1 gene blocked this protection. However, in an isolated cardiomyocyte model of IR injury, protection by NS1619 and NS11021 was insensitive to Slo1 deletion. These data suggest that protection in intact hearts occurs by a non-cardiomyocyte autonomous, SLO1-dependent, mechanism. In this regard, an in-situ assay of intrinsic cardiac neuronal function (tachycardic response to nicotine revealed that NS1619 preserved cardiac neurons following IR injury. Furthermore, blockade of synaptic transmission by hexamethonium suppressed cardioprotection by NS1619 in intact hearts. These results suggest that opening SLO1 protects the heart during IR injury, via a mechanism that involves intrinsic cardiac neurons. Cardiac neuronal ion channels may be useful therapeutic targets for eliciting cardioprotection.

  6. Effect of Berberine on PPARα/NO Activation in High Glucose- and Insulin-Induced Cardiomyocyte Hypertrophy

    Directory of Open Access Journals (Sweden)

    Mingfeng Wang

    2013-01-01

    Full Text Available Rhizoma coptidis, the root of Coptis chinensis Franch, has been used in China as a folk medicine in the treatment of diabetes for thousands of years. Berberine, one of the active ingredients of Rhizoma coptidis, has been reported to improve symptoms of diabetes and to treat experimental cardiac hypertrophy, respectively. The objective of this study was to evaluate the potential effect of berberine on cardiomyocyte hypertrophy in diabetes and its possible influence on peroxisome proliferator-activated receptor-α (PPARα/nitric oxide (NO signaling pathway. The cardiomyocyte hypertrophy induced by high glucose (25.5 mmol/L and insulin (0.1 μmol/L (HGI was characterized in rat primary cardiomyocyte by measuring the cell surface area, protein content, and atrial natriuretic factor mRNA expression level. Protein and mRNA expression were measured by western blot and real-time RT-PCR, respectively. The enzymatic activity of NO synthase (NOS was measured using a spectrophotometric assay, and NO concentration was measured using the Griess assay. HGI significantly induced cardiomyocyte hypertrophy and decreased the expression of PPARα and endothelial NOS at the mRNA and protein levels, which occurred in parallel with declining NOS activity and NO concentration. The effect of HGI was inhibited by berberine (0.1 to 100 μmol/L, fenofibrate (0.3 μmol/L, or L-arginine (100 μmol/L. MK886 (0.3 μmol/L, a selective PPARα antagonist, could abolish the effects of berberine and fenofibrate. NG-nitro-L-arginine-methyl ester (100 μmol/L, a NOS inhibitor, could block the effects of L-arginine, but only partially blocked the effects of berberine. These results suggest that berberine can blunt HGI-induced cardiomyocyte hypertrophy in vitro, through the activation of the PPARα/NO signaling pathway.

  7. Isorhynchophylline Protects PC12 Cells Against Beta-Amyloid-Induced Apoptosis via PI3K/Akt Signaling Pathway

    Science.gov (United States)

    Xian, Yan-Fang; Lin, Zhi-Xiu; Mao, Qing-Qiu; Chen, Jian-Nan; Su, Zi-Ren; Lai, Xiao-Ping; Ip, Paul Siu-Po

    2013-01-01

    The neurotoxicity of amyloid-β (Aβ) has been implicated as a critical cause of Alzheimer's disease. Isorhynchophylline (IRN), an oxindole alkaloid isolated from Uncaria rhynchophylla, exerts neuroprotective effect against Aβ 25–35-induced neurotoxicity in vitro. However, the exact mechanism for its neuroprotective effect is not well understood. The present study aimed to investigate the molecular mechanisms underlying the protective action of IRN against Aβ 25–35-induced neurotoxicity in cultured rat pheochromocytoma (PC12) cells. Pretreatment with IRN significantly increased the cell viability, inhibited the release of lactate dehydrogenase and the extent of DNA fragmentation in Aβ 25–35-treated cells. IRN treatment was able to enhance the protein levels of phosphorylated Akt (p-Akt) and glycogen synthase kinase-3β (p-GSK-3β). Lithium chloride blocked Aβ 25–35-induced cellular apoptosis in a similar manner as IRN, suggesting that GSK-3β inhibition was involved in neuroprotective action of IRN. Pretreatment with LY294002 completely abolished the protective effects of IRN. Furthermore, IRN reversed Aβ 25–35-induced attenuation in the level of phosphorylated cyclic AMP response element binding protein (p-CREB) and the effect of IRN could be blocked by the PI3K inhibitor. These experimental findings unambiguously suggested that the protective effect of IRN against Aβ 25–35-induced apoptosis in PC12 cells was associated with the enhancement of p-CREB expression via PI3K/Akt/GSK-3β signaling pathway. PMID:24319473

  8. Short-term psychosocial stress protects photoreceptors from damage via corticosterone-mediated activation of the AKT pathway.

    Science.gov (United States)

    Forkwa, Tembei K; Neumann, Inga D; Tamm, Ernst R; Ohlmann, Andreas; Reber, Stefan O

    2014-02-01

    Apoptotic death of photoreceptors in hereditary retinal degenerations can be prevented by neuroprotective molecules. Here, we report that adrenal glucocorticoids (GC) released during psychosocial stress protect photoreceptors from apoptosis after light damage. Psychosocial stress is known to be the main type of stressor humans are exposed to and was induced here in mice by 10h of chronic subordinate colony housing (CSC). Photoreceptor damage was generated by subsequent exposure to white light. Short-term psychosocial stress prior to illumination significantly reduced the number of apoptotic photoreceptors, an effect that was absent in adrenalectomized (ADX) mice. The neuroprotective effect was completely restored in ADX mice substituted with GC. Moreover, phosphorylation of retinal AKT increased following CSC or exogenous GC treatment, an effect that was again absent in ADX mice exposed to CSC. Finally, inhibition of AKT signaling with triciribine blocked the stress- and GC-mediated neuroprotective effects on photoreceptors. In summary, we provide evidence that 1) short-term psychosocial stress protects photoreceptors from light-induced damage and 2) the protective effect is most likely mediated by GC-induced activation of the AKT signaling pathway. Copyright © 2013 Elsevier Inc. All rights reserved.

  9. Muscles provide protection during microbial infection by activating innate immune response pathways in Drosophila and zebrafish

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    Arunita Chatterjee

    2016-06-01

    Full Text Available Muscle contraction brings about movement and locomotion in animals. However, muscles have also been implicated in several atypical physiological processes including immune response. The role of muscles in immunity and the mechanism involved has not yet been deciphered. In this paper, using Drosophila indirect flight muscles (IFMs as a model, we show that muscles are immune-responsive tissues. Flies with defective IFMs are incapable of mounting a potent humoral immune response. Upon immune challenge, the IFMs produce anti-microbial peptides (AMPs through the activation of canonical signaling pathways, and these IFM-synthesized AMPs are essential for survival upon infection. The trunk muscles of zebrafish, a vertebrate model system, also possess the capacity to mount an immune response against bacterial infections, thus establishing that immune responsiveness of muscles is evolutionarily conserved. Our results suggest that physiologically fit muscles might boost the innate immune response of an individual.

  10. Salidroside protects against foam cell formation and apoptosis, possibly via the MAPK and AKT signaling pathways.

    Science.gov (United States)

    Ni, Jing; Li, Yuanmin; Li, Weiming; Guo, Rong

    2017-10-10

    Foam cell formation and apoptosis are closely associated with atherosclerosis pathogenesis. We determined the effect of salidroside on oxidized low-density lipoprotein (ox-LDL)-induced foam cell formation and apoptosis in THP1 human acute monocytic leukemia cells and investigated the associated molecular mechanisms. THP1-derived macrophages were incubated with salidroside for 5 h and then exposed to ox-LDL for 24 h to induce foam cell formation. Cytotoxicity, lipid deposition, apoptosis, and the expression of various proteins were tested using the CCK8 kit, Oil Red O staining, flow cytometry, and western blotting, respectively. Ox-LDL treatment alone promoted macrophage-derived foam cell formation, while salidroside treatment alone inhibited it (p foam cell formation and apoptosis, partly by regulating the MAPK and Akt signaling pathways.

  11. Heparan Sulfate Induces Necroptosis in Murine Cardiomyocytes: A Medical-In silico Approach Combining In vitro Experiments and Machine Learning.

    Science.gov (United States)

    Zechendorf, Elisabeth; Vaßen, Phillip; Zhang, Jieyi; Hallawa, Ahmed; Martincuks, Antons; Krenkel, Oliver; Müller-Newen, Gerhard; Schuerholz, Tobias; Simon, Tim-Philipp; Marx, Gernot; Ascheid, Gerd; Schmeink, Anke; Dartmann, Guido; Thiemermann, Christoph; Martin, Lukas

    2018-01-01

    Life-threatening cardiomyopathy is a severe, but common, complication associated with severe trauma or sepsis. Several signaling pathways involved in apoptosis and necroptosis are linked to trauma- or sepsis-associated cardiomyopathy. However, the underling causative factors are still debatable. Heparan sulfate (HS) fragments belong to the class of danger/damage-associated molecular patterns liberated from endothelial-bound proteoglycans by heparanase during tissue injury associated with trauma or sepsis. We hypothesized that HS induces apoptosis or necroptosis in murine cardiomyocytes. By using a novel Medical- In silico approach that combines conventional cell culture experiments with machine learning algorithms, we aimed to reduce a significant part of the expensive and time-consuming cell culture experiments and data generation by using computational intelligence (refinement and replacement). Cardiomyocytes exposed to HS showed an activation of the intrinsic apoptosis signal pathway via cytochrome C and the activation of caspase 3 (both p  machine learning algorithms.

  12. Intracellular diffusion restrictions in isolated cardiomyocytes from rainbow trout

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    Birkedal Rikke

    2009-12-01

    Full Text Available Abstract Background Restriction of intracellular diffusion of adenine nucleotides has been studied intensively on adult rat cardiomyocytes. However, their cause and role in vivo is still uncertain. Intracellular membrane structures have been suggested to play a role. We therefore chose to study cardiomyocytes from rainbow trout (Oncorhynchus mykiss, which are thinner and have fewer intracellular membrane structures than adult rat cardiomyocytes. Previous studies suggest that trout permeabilized cardiac fibers also have diffusion restrictions. However, results from fibers may be affected by incomplete separation of the cells. This is avoided when studying permeabilized, isolated cardiomyocytes. The aim of this study was to verify the existence of diffusion restrictions in trout cardiomyocytes by comparing ADP-kinetics of mitochondrial respiration in permeabilized fibers, permeabilized cardiomyocytes and isolated mitochondria from rainbow trout heart. Experiments were performed at 10, 15 and 20°C in the absence and presence of creatine. Results Trout cardiomyocytes hypercontracted in the solutions used for mammalian cardiomyocytes. We developed a new solution in which they retained their shape and showed stable steady state respiration rates throughout an experiment. The apparent ADP-affinity of permeabilized cardiomyocytes was different from that of fibers. It was higher, independent of temperature and not increased by creatine. However, it was still about ten times lower than in isolated mitochondria. Conclusions The differences between fibers and cardiomyocytes suggest that results from trout heart fibers were affected by incomplete separation of the cells. However, the lower ADP-affinity of cardiomyocytes compared to isolated mitochondria indicate that intracellular diffusion restrictions are still present in trout cardiomyocytes despite their lower density of intracellular membrane structures. The lack of a creatine effect indicates that

  13. Engineered Biomaterials Control Differentiation and Proliferation of Human-Embryonic-Stem-Cell-Derived Cardiomyocytes via Timed Notch Activation

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    Jason C. Tung

    2014-03-01

    Full Text Available For cell-based treatments of myocardial infarction, a better understanding of key developmental signaling pathways and more robust techniques for producing cardiomyocytes are required. Manipulation of Notch signaling has promise as it plays an important role during cardiovascular development, but previous studies presented conflicting results that Notch activation both positively and negatively regulates cardiogenesis. We developed surface- and microparticle-based Notch-signaling biomaterials that function in a time-specific activation-tunable manner, enabling precise investigation of Notch activation at specific developmental stages. Using our technologies, a biphasic effect of Notch activation on cardiac differentiation was found: early activation in undifferentiated human embryonic stem cells (hESCs promotes ectodermal differentiation, activation in specified cardiovascular progenitor cells increases cardiac differentiation. Signaling also induces cardiomyocyte proliferation, and repeated doses of Notch-signaling microparticles further enhance cardiomyocyte population size. These results highlight the diverse effects of Notch activation during cardiac development and provide approaches for generating large quantities of cardiomyocytes.

  14. Lithium protects against methamphetamine-induced neurotoxicity in PC12 cells via Akt/GSK3β/mTOR pathway

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    Wu, Jintao; Zhu, Dexiao; Zhang, Jing; Li, Guibao [Department of Anatomy, School of Medicine, Shandong University, Jinan, Shandong, 250012 (China); Liu, Zengxun [Department of Psychiatry, School of Medicine, Shandong University, Jinan, Shandong, 250012 China (China); Sun, Jinhao, E-mail: sunjinhao@gmail.com [Department of Anatomy, School of Medicine, Shandong University, Jinan, Shandong, 250012 (China)

    2015-09-25

    Methamphetamine (MA) is neurotoxic, especially in dopaminergic neurons. Long-lasting exposure to MA causes psychosis and increases the risk of Parkinson's disease. Lithium (Li) is a known mood stabilizer and has neuroprotective effects. Previous studies suggest that MA exposure decreases the phosphorylation of Akt/GSK3β pathway in vivo, whereas Li facilitates the phosphorylation of Akt/GSK3β pathway. Moreover, GSK3β and mTOR are implicated in the locomotor sensitization induced by psychostimulants and mTOR plays a critical role in MA induced toxicity. However, the effect of MA on Akt/GSK3β/mTOR pathway has not been fully investigated in vitro. Here, we found that MA exposure significantly dephosphorylated Akt/GSK3β/mTOR pathway in PC12 cells. In addition, Li remarkably attenuated the dephosphorylation effect of MA exposure on Akt/GSK3β/mTOR pathway. Furthermore, Li showed obvious protective effects against MA toxicity and LY294002 (Akt inhibitor) suppressed the protective effects of Li. Together, MA exposure dephosphorylates Akt/GSK3β/mTOR pathway in vitro, while lithium protects against MA-induced neurotoxicity via phosphorylation of Akt/GSK3β/mTOR pathway. - Highlights: • Lithium protects against methamphetamine-induced neurotoxicity in vitro. • Methamphetamine exposure dephosphorylates Akt/GSK3β/mTOR pathway. • Lithium attenuates methamphetamine-induced toxicity via phosphorylating Akt/GSK3β/mTOR pathway.

  15. Lithium protects against methamphetamine-induced neurotoxicity in PC12 cells via Akt/GSK3β/mTOR pathway

    International Nuclear Information System (INIS)

    Wu, Jintao; Zhu, Dexiao; Zhang, Jing; Li, Guibao; Liu, Zengxun; Sun, Jinhao

    2015-01-01

    Methamphetamine (MA) is neurotoxic, especially in dopaminergic neurons. Long-lasting exposure to MA causes psychosis and increases the risk of Parkinson's disease. Lithium (Li) is a known mood stabilizer and has neuroprotective effects. Previous studies suggest that MA exposure decreases the phosphorylation of Akt/GSK3β pathway in vivo, whereas Li facilitates the phosphorylation of Akt/GSK3β pathway. Moreover, GSK3β and mTOR are implicated in the locomotor sensitization induced by psychostimulants and mTOR plays a critical role in MA induced toxicity. However, the effect of MA on Akt/GSK3β/mTOR pathway has not been fully investigated in vitro. Here, we found that MA exposure significantly dephosphorylated Akt/GSK3β/mTOR pathway in PC12 cells. In addition, Li remarkably attenuated the dephosphorylation effect of MA exposure on Akt/GSK3β/mTOR pathway. Furthermore, Li showed obvious protective effects against MA toxicity and LY294002 (Akt inhibitor) suppressed the protective effects of Li. Together, MA exposure dephosphorylates Akt/GSK3β/mTOR pathway in vitro, while lithium protects against MA-induced neurotoxicity via phosphorylation of Akt/GSK3β/mTOR pathway. - Highlights: • Lithium protects against methamphetamine-induced neurotoxicity in vitro. • Methamphetamine exposure dephosphorylates Akt/GSK3β/mTOR pathway. • Lithium attenuates methamphetamine-induced toxicity via phosphorylating Akt/GSK3β/mTOR pathway

  16. Vitamin B1 Analog Benfotiamine Prevents Diabetes-Induced Diastolic Dysfunction and Heart Failure Through Akt/Pim-1–Mediated Survival Pathway

    Science.gov (United States)

    Katare, Rajesh G.; Caporali, Andrea; Oikawa, Atsuhiko; Meloni, Marco; Emanueli, Costanza; Madeddu, Paolo

    2010-01-01

    Background The increasing incidence of diabetes mellitus will result in a new epidemic of heart failure unless novel treatments able to halt diabetic cardiomyopathy early in its course are introduced. This study aimed to determine whether the activity of the Akt/Pim-1 signaling pathway is altered at critical stages of diabetic cardiomyopathy and whether supplementation with vitamin B1 analog benfotiamine (BFT) helps to sustain the above prosurvival mechanism, thereby preserving cardiomyocyte viability and function. Methods and Results Untreated streptozotocin-induced type 1 or leptin-receptor mutant type 2 diabetic mice showed diastolic dysfunction evolving to contractile impairment and cardiac dilatation and failure. BFT (70 mg/kg−1/d−1) improved diastolic and systolic function and prevented left ventricular end-diastolic pressure increase and chamber dilatation in both diabetic models. Moreover, BFT improved cardiac perfusion and reduced cardiomyocyte apoptosis and interstitial fibrosis. In hearts of untreated diabetic mice, the expression and activity of Akt/Pim-1 signaling declined along with O-N-acetylglucosamine modification of Akt, inhibition of pentose phosphate pathway, activation of oxidative stress, and accumulation of glycation end products. Furthermore, diabetes reduced signal transducer and activator of transcription 3 phosphorylation independently of Akt. BFT inhibited these effects of diabetes mellitus, thereby conferring cardiomyocytes with improved resistance to high glucose-induced damage. The phosphoinositide-3-kinase inhibitor LY294002 and dominant-negative Akt inhibited antiapoptotic action of BFT and Pim-1 upregulation in high glucose-challenged cardiomyocytes. Conclusions These results show that BFT protects from diabetes mellitus-induced cardiac dysfunction through pleiotropic mechanisms, culminating in the activation of prosurvival signaling pathway. Thus, BFT merits attention for application in clinical practice. PMID:20107192

  17. Vitamin B1 analog benfotiamine prevents diabetes-induced diastolic dysfunction and heart failure through Akt/Pim-1-mediated survival pathway.

    Science.gov (United States)

    Katare, Rajesh G; Caporali, Andrea; Oikawa, Atsuhiko; Meloni, Marco; Emanueli, Costanza; Madeddu, Paolo

    2010-03-01

    The increasing incidence of diabetes mellitus will result in a new epidemic of heart failure unless novel treatments able to halt diabetic cardiomyopathy early in its course are introduced. This study aimed to determine whether the activity of the Akt/Pim-1 signaling pathway is altered at critical stages of diabetic cardiomyopathy and whether supplementation with vitamin B1 analog benfotiamine (BFT) helps to sustain the above prosurvival mechanism, thereby preserving cardiomyocyte viability and function. Untreated streptozotocin-induced type 1 or leptin-receptor mutant type 2 diabetic mice showed diastolic dysfunction evolving to contractile impairment and cardiac dilatation and failure. BFT (70 mg/kg(-1)/d(-1)) improved diastolic and systolic function and prevented left ventricular end-diastolic pressure increase and chamber dilatation in both diabetic models. Moreover, BFT improved cardiac perfusion and reduced cardiomyocyte apoptosis and interstitial fibrosis. In hearts of untreated diabetic mice, the expression and activity of Akt/Pim-1 signaling declined along with O-N-acetylglucosamine modification of Akt, inhibition of pentose phosphate pathway, activation of oxidative stress, and accumulation of glycation end products. Furthermore, diabetes reduced pSTAT3 independently of Akt. BFT inhibited these effects of diabetes mellitus, thereby conferring cardiomyocytes with improved resistance to high glucose-induced damage. The phosphoinositide-3-kinase inhibitor LY294002 and dominant-negative Akt inhibited antiapoptotic action of BFT-induced and Pim-1 upregulation in high glucose-challenged cardiomyocytes. These results show that BFT protects from diabetes mellitus-induced cardiac dysfunction through pleiotropic mechanisms, culminating in the activation of prosurvival signaling pathway. Thus, BFT merits attention for application in clinical practice.

  18. Sulforaphane protects rodent retinas against ischemia-reperfusion injury through the activation of the Nrf2/HO-1 antioxidant pathway.

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    Hong Pan

    Full Text Available Retinal ischemia-reperfusion (I/R injury induces oxidative stress, leukocyte infiltration, and neuronal cell death. Sulforaphane (SF, which can be obtained in cruciferous vegetables such as broccoli, exerts protective effects in response to oxidative stress in various tissues. These effects can be initiated through nuclear factor E2-related factor 2 (Nrf2-mediated induction of heme oxygenase-1 (HO-1. This investigation was designed to elucidate the neural protective mechanisms of SF in the retinal I/R rat model. Animals were intraperitoneally (i.p. injected with SF (12.5 mg/kg or vehicle (corn oil once a day for 7 consecutive days. Then, retinal I/R was made by elevating the intraocular pressure (IOP to 130 mmHg for 1 h. To determine if HO-1 was involved in the Nrf2 antioxidant pathway, rats were subjected to protoporphyrin IX zinc (II (ZnPP, 30 mg/kg, i.p. treatments at 24 h before retinal ischemia. The neuroprotective effects of SF were assessed by determining the morphology of the retina, counting the infiltrating inflammatory cells and the surviving retinal ganglion cells (RGCs and amacrine cells, and measuring apoptosis in the retinal layers. The expression of Nrf2 and HO-1 was studied by immunofluorescence analysis and western blotting. I/R induced a marked increase of ROS generation, caused pronounced inflammation, increased the apoptosis of RGCs and amacrine cells and caused the thinning of the inner retinal layer (IRL, and these effects were diminished or abolished by SF pretreatment. Meanwhile, SF pretreatment significantly elevated the nuclear accumulation of Nrf2 and the level of HO-1 expression in the I/R retinas; however, ZnPP reversed the protective effects of SF on I/R retinas. Together, we offer direct evidence that SF had protective effects on I/R retinas, which could be attributed, at least in part, to the activation of the Nrf2/HO-1 antioxidant pathway.

  19. Protection of Momordica charantia polysaccharide against intracerebral hemorrhage-induced brain injury through JNK3 signaling pathway.

    Science.gov (United States)

    Duan, Zhen-Zhen; Zhou, Xiao-Ling; Li, Yi-Hang; Zhang, Feng; Li, Feng-Ying; Su-Hua, Qi

    2015-01-01

    It has been well documented that Momordica charantia polysaccharide (MCP) has multiple biological effects such as immune enhancement, anti-oxidation and anti-cancer. However, the potential protective effects of MCP on stroke damage and its relative mechanisms remain unclear. Our present study demonstrated that MCP could scavenge reactive oxygen species (ROS) in intra-cerebral hemorrhage damage, significantly attenuating the neuronal death induced by thrombin in primary hippocampal neurons. Furthermore, we found that MCP prevented the activation of the c-Jun N-terminal protein kinase (JNK3), c-Jun and caspase-3, which was caused by the intra-cerebral hemorrhage injury. Taken together, our study demonstrated that MCP had a neuroprotective effect in response to intra-cerebral hemorrhage and its mechanisms involved the inhibition of JNK3 signaling pathway.

  20. Trimetazidine protects retinal ganglion cells from acute glaucoma via the Nrf2/Ho-1 pathway.

    Science.gov (United States)

    Wan, Peixing; Su, Wenru; Zhang, Yingying; Li, Zhidong; Deng, Caibin; Zhuo, Yehong

    2017-09-15

    Acute glaucoma is one of the leading causes of irreversible vision impairment characterized by the rapid elevation of intraocular pressure (IOP) and consequent retinal ganglion cell (RGC) death. Oxidative stress and neuroinflammation have been considered critical for the pathogenesis of RGC death in acute glaucoma. Trimetazidine (TMZ), an anti-ischemic drug, possesses antioxidative and anti-inflammatory properties, contributing to its therapeutic potential in tissue damage. However, the role of TMZ in acute glaucoma and the underlying molecular mechanisms remain elusive. Here, we report that treatment with TMZ significantly attenuated retinal damage and RGC death in mice with acute glaucoma, with a significant decrease in reactive oxygen species (ROS) and inflammatory cytokine production in the retina. Furthermore, TMZ treatment directly decreased ROS production and rebalanced the intracellular redox state, thus contributing to the survival of RGCs in vitro TMZ treatment also reduced the production of inflammatory cytokines in vitro Mechanistically, the TMZ-mediated inhibition of apoptosis and inflammatory cytokine production in RGCs occurred via the regulation of the nuclear factor erythroid 2-related factor 2/heme oxygenase 1/caspase-8 pathway. Moreover, the TMZ-mediated neuroprotection in acute glaucoma was abrogated when an HO-1 inhibitor, SnPP, was used. Our findings identify potential mechanisms of RGC apoptosis and propose a novel therapeutic agent, TMZ, which exerts a precise neuroprotective effect against acute glaucoma. © 2017 The Author(s).

  1. Naringenin protects against 6-OHDA-induced neurotoxicity via activation of the Nrf2/ARE signaling pathway.

    Science.gov (United States)

    Lou, Haiyan; Jing, Xu; Wei, Xinbing; Shi, Huanying; Ren, Dongmei; Zhang, Xiumei

    2014-04-01

    There is increasing evidence that oxidative stress is critically involved in the pathogenesis of Parkinson's disease (PD), suggesting that pharmacological targeting of the antioxidant machinery may have therapeutic value. Naringenin, a natural flavonoid compound, has been reported to possess neuroprotective effect against PD related pathology; however the mechanisms underlying its beneficial effects are poorly defined. Thus, the purpose of the present study was to investigate the potential neuroprotective role of naringenin and to delineate its mechanism of action against 6-hydroxydopamine (6-OHDA)-induced neurotoxicity in models of PD both in vitro and in vivo. Naringenin treatment resulted in an increase in nuclear factor E2-related factor 2 (Nrf2) protein levels and subsequent activation of antioxidant response element (ARE) pathway genes in SH-SY5Y cells and in mice. Exposure of SH-SY5Y cells to naringenin provided protection against 6-OHDA-induced oxidative insults that was dependent on Nrf2, since treatment with Nrf2 siRNA failed to block against 6-OHDA neurotoxicity or induce Nrf2-dependent cytoprotective genes in SH-SY5Y cells. In mice, oral administration of naringenin resulted in significant protection against 6-OHDA-induced nigrostriatal dopaminergic neurodegeneration and oxidative damage. Our results indicate that activation of Nrf2/ARE signaling by naringenin is strongly associated with its neuroprotective effects against 6-OHDA neurotoxicity and suggest that targeting the Nrf2/ARE pathway may be a promising approach for therapeutic intervention in PD. Copyright © 2013 Elsevier Ltd. All rights reserved.

  2. The TPO/c-MPL pathway in the bone marrow may protect leukemia cells from chemotherapy in AML Patients.

    Science.gov (United States)

    Dong-Feng, Zeng; Ting, Liu; Yong, Zhang; Cheng, Chang; Xi, Zhang; Pei-Yan, Kong

    2014-04-01

    Accumulating evidence indicates that the interaction of human LSCs (leukemic stem cells) with the hematopoietic microenvironment, mediated by the thrombopoietin (TPO)/c-MPL pathway, may be an underlying mechanism for resistance to cell cycle-dependent cytotoxic chemotherapy. However, the role of TPO/c-MPL signaling in AML (acute myelogenous leukemia) chemotherapy resistance hasn't been fully understood. The c-MPL and TPO levels in different AML samples were measured by flow cytometry and ELISA. We also assessed the TPO levels in the osteoblasts derived from bone mesenchymal stem cells (BMSCs). The survival rate of an AML cell line that had been co-cultured with different BMSC-derived osteoblasts was measured to determine the IC50 of an AML chemotherapy drug daunorubicin (DNR). The levels of TPO/c-MPL in the initial and relapse AML patients were significantly higher than that in the control (P MPL expression was found in the bone marrow mononuclear cells of the relapse AML patients. More importantly, the IC50 of DNR in the HEL + AML-derived osteoblasts was the highest among all co-culture systems. High level of TPO/c-MPL signaling may protect LSCs from chemotherapy in AML. The effects of inhibition of the TPO/c-MPL pathway on enhancing the chemotherapy sensitivity of AML cells, and on their downstream effector molecules that direct the interactions between patient-derived blasts and leukemia repopulating cells need to be further studied.

  3. Mesenchymal Stem Cells Protect Nucleus Pulposus Cells from Compression-Induced Apoptosis by Inhibiting the Mitochondrial Pathway

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    Sheng Chen

    2017-01-01

    Full Text Available Objective. Excessive apoptosis of nucleus pulposus cells (NPCs induced by various stresses, including compression, contributes to the development of intervertebral disc degeneration (IVDD. Mesenchymal stem cells (MSCs can benefit the regeneration of NPCs and delay IVDD, but the underlying molecular mechanism is poorly understood. This study aimed to evaluate the antiapoptosis effects of bone marrow-derived MSC (BMSC on rat NPCs exposed to compression and investigate whether the mitochondrial pathway was involved. Methods. BMSCs and NPCs were cocultured in the compression apparatus at 1.0 MPa for 36 h. Cell viability, apoptosis, mitochondrial function, and the expression of apoptosis-related proteins were evaluated. Results. The results showed that coculturing with BMSCs increased the cell viability and reduced apoptosis of NPCs exposed to compression. Meanwhile, BMSCs could relieve the compression-induced mitochondrial damage of NPCs by decreasing reactive oxygen species level and maintaining mitochondrial membrane potential as well as mitochondrial integrity. Furthermore, coculturing with BMSCs suppressed the activated caspase-3 and activated caspase-9, decreased the expressions of cytosolic cytochrome c and Bax, and increased the expression of Bcl-2. Conclusions. Our results suggest that BMSCs can protect against compression-induced apoptosis of NPCs by inhibiting the mitochondrial pathway and thus enhance our understanding on the MSC-based therapy for IVDD.

  4. HAX-1 Protects Glioblastoma Cells from Apoptosis through the Akt1 Pathway

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    Xin Deng

    2017-12-01

    Full Text Available Glioblastoma is the most common malignant tumor in central nervous system (CNS, and it is still insurmountable and has a poor prognosis. The proliferation and survival mechanism of glioma cells needs to be explored further for the development of glioma treatment. Hematopoietic-substrate-1 associated protein X-1 (HAX-1 has been reported as an anti-apoptosis protein that plays an important role in several malignant tumors. However, the effect and mechanism of HAX-1 in glioblastomas remains unknown. This study aimed to investigate the effect of HAX-1 in glioblastoma cells and explore the mechanism. The results of clone formation and Edu proliferation assay showed slower multiplication in HAX-1 knock-out cells. Flow cytometry showed cell cycle arrest mainly in G0/G1 phase. Apoptosis due to oxidative stress was increased after HAX-1 was knocked out. Western-blot assay exhibited that the levels of p21, Bax, and p53 proteins were significantly raised, and that the activation of the caspase cascade was enhanced in the absence of HAX-1. The degradation rate and ubiquitination of p53 declined because of the decrease in phosphorylation of proteins MDM2 and Akt1. Co-immunoprecipitation (Co-IP and immunefluorescent co-localization assays were performed to test the influence of HAX-1 on the interaction between Akt1 and Hsp90, which is crucial for the activity of Akt1. In conclusion, this novel study suggested that HAX-1 could affect the Akt1 pathway through Hsp90. The knock-out of HAX-1 leads to the inactivity of the Ak1t/MDM2 axis, which leads to increased levels of p53, and finally generates cell cycle arrest and results in the apoptosis of glioblastoma cells.

  5. A Non-canonical Melanin Biosynthesis Pathway Protects Aspergillus terreus Conidia from Environmental Stress.

    Science.gov (United States)

    Geib, Elena; Gressler, Markus; Viediernikova, Iuliia; Hillmann, Falk; Jacobsen, Ilse D; Nietzsche, Sandor; Hertweck, Christian; Brock, Matthias

    2016-05-19

    Melanins are ubiquitous pigments found in all kingdoms of life. Most organisms use them for protection from environmental stress, although some fungi employ melanins as virulence determinants. The human pathogenic fungus Aspergillus fumigatus and related Ascomycetes produce dihydroxynaphthalene- (DHN) melanin in their spores, the conidia, and use it to inhibit phagolysosome acidification. However, biosynthetic origin of melanin in a related fungus, Aspergillus terreus, has remained a mystery because A. terreus lacks genes for synthesis of DHN-melanin. Here we identify genes coding for an unusual NRPS-like enzyme (MelA) and a tyrosinase (TyrP) that A. terreus expressed under conidiation conditions. We demonstrate that MelA produces aspulvinone E, which is activated for polymerization by TyrP. Functional studies reveal that this new pigment, Asp-melanin, confers resistance against UV light and hampers phagocytosis by soil amoeba. Unexpectedly, Asp-melanin does not inhibit acidification of phagolysosomes, thus likely contributing specifically to survival of A. terreus conidia in acidic environments. Copyright © 2016 Elsevier Ltd. All rights reserved.

  6. Heme oxygenase-1 dependant pathway contributes to protection by tetramethylpyrazine against chronic hypoxic injury on medulla oblongata in rats.

    Science.gov (United States)

    Ding, Yan; Hou, Xuefei; Chen, Li; Zhou, Hua; Gong, Yanju; Dai, Liqun; Zheng, Yu

    2016-02-15

    Tetramethylpyrazine (TMP), one of the active ingredients of the Chinese herb Lingusticum Wallichii (Chuan Xiong) has been proved to protect the medulla oblongata from chronic hypoxia injury. However, the underlying mechanism remains unclear. The purpose of this study was to determine whether the protective effects of TMP are associated with the heme oxygenase-1 (HO-1) dependant pathway in adult rats. The morphological changes of neurons in the hypoglossal nucleus (12N), the nucleus ambiguus (Amb), the nucleus tractus solitarius (NTS), and the pre-Bötzinger complex (pre-BötC) were investigated by Nissl staining; the malondialdehyde (MDA) content and superoxide dismutase (SOD) activity were measured to evaluate the anti-oxidant effect; some apoptosis parameters, Bax mRNA and Bcl-2 mRNA, were tested; and the double immunochemistry staining of active caspase-3/NeuN was performed. Meanwhile, the HO-1 protein expression and heme oxygenase (HO) activity were examined. Tin-protoporphyrin (SnPP), a potent inhibitor of HO, was used to further confirm the effect of HO-1. We found that TMP ameliorated the neuron loss in 12N, Amb and NTS, the decrease in SOD activity and the increase in MDA content, the decrease in Bcl-2 mRNA of medulla oblongata (Pmedulla oblongata in the rats. Copyright © 2015 Elsevier B.V. All rights reserved.

  7. Sulforaphane protects against cytokine- and streptozotocin-induced β-cell damage by suppressing the NF-κB pathway

    International Nuclear Information System (INIS)

    Song, Mi-Young; Kim, Eun-Kyung; Moon, Woo-Sung; Park, Jin-Woo; Kim, Hyung-Jin; So, Hong-Seob; Park, Raekil; Kwon, Kang-Beom; Park, Byung-Hyun

    2009-01-01

    Sulforaphane (SFN) is an indirect antioxidant that protects animal tissues from chemical or biological insults by stimulating the expression of several NF-E2-related factor-2 (Nrf2)-regulated phase 2 enzymes. Treatment of RINm5F insulinoma cells with SFN increases Nrf2 nuclear translocation and expression of phase 2 enzymes. In this study, we investigated whether the activation of Nrf2 by SFN treatment or ectopic overexpression of Nrf2 inhibited cytokine-induced β-cell damage. Treatment of RIN cells with IL-1β and IFN-γ induced β-cell damage through a NF-κB-dependent signaling pathway. Activation of Nrf2 by treatment with SFN and induction of Nrf2 overexpression by transfection with Nrf2 prevented cytokine toxicity. The mechanism by which Nrf2 activation inhibited NF-κB-dependent cell death signals appeared to involve the reduction of oxidative stress, as demonstrated by the inhibition of cytokine-induced H 2 O 2 production. The protective effect of SFN was further demonstrated by the restoration of normal insulin secreting responses to glucose in cytokine-treated rat pancreatic islets. Furthermore, pretreatment with SFN blocked the development of type 1 diabetes in streptozotocin-treated mice

  8. Cyclophilin B protects SH-SY5Y human neuroblastoma cells against MPP(+)-induced neurotoxicity via JNK pathway.

    Science.gov (United States)

    Oh, Yoojung; Jeong, Kwon; Kim, Kiyoon; Lee, Young-Seok; Jeong, Suyun; Kim, Sung Soo; Yoon, Kyung-Sik; Ha, Joohun; Kang, Insug; Choe, Wonchae

    2016-09-23

    Parkinson's disease (PD) is the second most common neurodegenerative disorder of aging. PD involves a progressive loss of dopaminergic neurons in the substantia nigra pars compacta. 1-Methyl-4-phenyl-1, 2, 3, 6-tetrahydropyidine (MPTP) and its toxic metabolite 1-methyl-4-phenylpyridinium ion (MPP+) inhibit the complex I of the mitochondrial electron transport chain, and have been widely used to construct PD models. Cyclophilin B (CypB) is an endoplasmic reticulum protein that binds to cyclosporine A as a cyclophilin family member. CypB has peptidyl-prolyl cis-trans isomerase (PPIase) activity. We investigated the protective effects of overexpressed CypB on MPP+-induced neurocytotoxicity in SH-SY5Y human neuroblastoma cells. Overexpressed CypB decreased MPP(+)-induced oxidative stress through the modulation of antioxidant enzymes including manganese superoxide dismutase and catalase, and prevented neurocytotoxicity via mitogen-activated protein kinase, especially the c-Jun N-terminal kinase pathway. In addition, CypB inhibited the activation of MPP(+)-induced the pro-apoptotic molecules poly (ADP-ribose) polymerase, Bax, and Bcl-2, and attenuated MPP(+)-induced mitochondrial dysfunction. The data suggest that overexpressed CypB protects neuronal cells from MPP+-induced dopaminergic neuronal cell death. Copyright © 2016 Elsevier Inc. All rights reserved.

  9. Melatonin ameliorates oxidative stress, modulates death receptor pathway proteins, and protects the rat cerebrum against bisphenol-A-induced apoptosis.

    Science.gov (United States)

    El-Missiry, Mohamed A; Othman, Azza I; Al-Abdan, Monera A; El-Sayed, Aml A

    2014-12-15

    Epidemiological reports have indicated a correlation between the increasing of bisphenol-A (BPA) levels in the environment and the incidence of neurodegenerative diseases. In the present study, the protective effect of melatonin on oxidative stress and the death receptor apoptotic proteins in the cerebrum of the bisphenol-A-treated rats were examined. Adult male rats were orally administered melatonin (10mg/kg bw) concurrently with BPA (50mg/kg bw) 3 days a week for 6 weeks. BPA exposure resulted in significant elevations of oxidative stress, as evidenced by the increased malondialdehyde level and the decreased glutathione level and superoxide dismutase activity in the cerebrum. BPA caused an upregulation of p53 and CD95-Fas and activation of capsases-3 and 8, resulting in cerebral cell apoptosis. Melatonin significantly attenuated the BPA-evoked brain oxidative stress, modulated apoptotic-regulating proteins and protected against apoptosis. These data suggest that melatonin modulated important steps in the death receptor apoptotic pathway which likely related to its redox control properties. Melatonin is a promising pharmacological agent for preventing the potential neurotoxicity of BPA following occupational or environmental exposures. Copyright © 2014 Elsevier B.V. All rights reserved.

  10. Protective Effect of Tempol on Acute Kidney Injury Through PI3K/Akt/Nrf2 Signaling Pathway

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    Gensheng Zhang

    2016-02-01

    Full Text Available Background/Aims: Tempol is a protective antioxidant against ischemic injury in many animal models. The molecular mechanisms are not well understood. Nuclear factor erythroid 2-related factor (Nrf2 is a master transcription factor during oxidative stress, which is enhanced by activation of protein kinase C (PKC pathway. Another factor, tubular epithelial apoptosis, is mediated by activation of phosphoinositide 3-kinase (PI3K/protein kinase B (PKB, Akt signaling pathway during renal ischemic injury. We tested the hypothesis that tempol activates PKC or PI3K/Akt/Nrf2 pathways to transcribe many genes that coordinate endogenous antioxidant defense. Methods: The right renal pedicle was clamped for 45 minutes and the left kidney was removed to study renal ischemia/reperfusion (I/R injury in C57BL/6 mice. The response was assessed from serum parameters, renal morphology and renal expression of PKC, phosphorylated-PKC (p-PKC, Nrf2, heme oxygenase-1 (HO-1, Akt, phosphorylated-Akt (p-Akt, pro-caspase-3 and cleaved caspase-3 in groups of sham and I/R mice given vehicle, or tempol (50 or 100 mg/kg, intraperitoneal injection. Results: The serum malondialdehyde (MDA, marker of reactive oxygen species doubled and the BUN and creatinine increased 5- to 10-fold after I/R injury. Tempol (50 or 100 mg/kg prevented the increases in MDA but only tempol (50 mg/kg lessened the increases in BUN and creatinine and moderated the acute tubular necrosis. I/R did not change expression of PKC or p-PKC but reduced renal expression of Nrf2, p-Akt, HO-1 and pro-caspase-3 and increased cleaved caspase-3. Tempol (50 mg/kg prevented these changes produced by I/R whereas tempol (100 mg/kg had lesser or inconsistent effects. Conclusion: Tempol (50 mg/kg prevents lipid peroxidation and attenuates renal damage after I/R injury. The beneficial pathway apparently is not dependent on upregulation or phosphorylation of PKC, at lower tempol doses, does implicate upregulation of Akt with

  11. Schisandra chinensis Protects the Skin from Global Pollution by Inflammatory and Redox Balance Pathway Modulations: An In Vitro Study

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    Edwige Ranouille

    2018-06-01

    Full Text Available Epidemiological results show that airborne particulate matter (PM induces health alterations in line with pulmonary and cardiovascular pathologies. Deleterious effects of PM on the skin have also been investigated. A possible approach to prevent Reactive Oxygen Species (ROS-mediated disorders for both preventive and treatment means is based on the use of substances, which can be found in plants. These can act as secondary metabolites, and lignans are a promising candidate. Thus, the objective of this study was firstly to identify reconstructed human epidermis, using a transcriptomic approach, and also to identify the effects of Urban Dust and of Urban Dust and Schisandra chinensis (S.C. extract on the expression of genes that are involved in the response to cellular protection mechanisms. Secondly, we examined the effect of an active extract from S.C. on the protection of human keratinocytes damages that were caused by pollution, through the evaluation of Nrf2 and AhR pathways, NF-kB, and DJ-1. Urban Dust included the over-expression of metalloproteinases MMP-1 and MMP-9 and an increase in Glutathione peroxidase 2 (GPX2. In the presence of Urban Dust, S.C. extract activated the over-expression of several genes that are involved in the antioxidant response and in the detoxification pathway, including Ferritin light chain (FTL and GPX2. Exposure to urban dust activated the cytoplasmic expression of NF-kB and AhR, when compared to the control. Co-treatment of Urban Dust and S.C. extract increased DJ-1 protein levels, Nrf2 expression, and decreased AhR and NF-kB in the cytoplasm. At the same time, this co-treatment increased SOD2 expression (50%: p < 0.001 and catalase activity (120%: p < 0.05, when compared to Urban Dust alone. Thus, S.C. might be able to protect the Normal Human Epidermal Keratinocytes (NHEK from environmental aggression, by fighting the harmful effects of urban pollution.

  12. Interference of silibinin with IGF-1R signalling pathways protects human epidermoid carcinoma A431 cells from UVB-induced apoptosis

    International Nuclear Information System (INIS)

    Liu, Weiwei; Otkur, Wuxiyar; Li, Lingzhi; Wang, Qiong; He, Hao; Zang, Linghe; Hayashi, Toshihiko; Tashiro, Shin-ichi; Onodera, Satoshi; Xia, Mingyu; Ikejima, Takashi

    2013-01-01

    Highlights: ► Silibinin protects A431 cells from UVB irradiation-induced apoptosis. ► Up-regulation of the IGF-1R-JNK/ERK pathways by UVB induces cell apoptosis. ► Silibinin inhibits IGF-1R pathways to repress caspase-8-mediated apoptosis. -- Abstract: Ultraviolet B (UVB) from sunlight is a major cause of cutaneous lesion. Silibinin, a traditional hepatic protectant, elicits protective effects against UVB-induced cellular damage. In A431 cells, the insulin-like growth factor-1 receptor (IGF-1R) was markedly up-regulated by UVB irradiation. The activation of the IGF-1R signalling pathways contributed to apoptosis of the cells rather than rescuing the cells from death. Up-regulated IGF-1R stimulated downstream mitogen-activated protein kinases (MAPKs), such as c-Jun N-terminal kinases (JNK) and extracellular signal-regulated protein kinases 1/2 (ERK1/2). The subsequent activation of caspase-8 and caspase-3 led to apoptosis. The activation of IGF-1R signalling pathways is the cause of A431 cell death. The pharmacological inhibitors and the small interfering RNA (siRNA) targeting IGF-1R suppressed the downstream activation of JNK/ERK-caspases to help the survival of the UVB-irradiated A431 cells. Indeed, silibinin treatment suppressed the IGF-1R-JNK/ERK pathways and thus protected the cells from UVB-induced apoptosis

  13. Unique sex- and age-dependent effects in protective pathways in acute kidney injury.

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    Boddu, Ravindra; Fan, Chunlan; Rangarajan, Sunil; Sunil, Bhuvana; Bolisetty, Subhashini; Curtis, Lisa M

    2017-09-01

    Sex and age influence susceptibility to acute kidney injury (AKI), with young females exhibiting lowest incidence. In these studies, we investigated mechanisms which may underlie the sex/age-based dissimilarities. Cisplatin (Cp)-induced AKI resulted in morphological evidence of injury in all groups. A minimal rise in plasma creatinine (PCr) was seen in Young Females, whereas in Aged Females, PCr rose precipitously. Relative to Young Males, Aged Males showed significantly, but temporally, comparably elevated PCr. Notably, Aged Females showed significantly greater mortality, whereas Young Females exhibited none. Tissue KIM-1 and plasma NGAL were significantly lower in Young Females than all others. IGFBP7 levels were modestly increased in both Young groups. IGFBP7 levels in Aged Females were significantly elevated at baseline relative to Aged Males, and increased linearly through day 3 , when these levels were comparable in both Aged groups. Plasma cytokine levels similarly showed a pattern of protective effects preferentially in Young Females. Expression of the drug transporter MATE2 did not explain the sex/age distinctions. Heme oxygenase-1 (HO-1) levels (~28-kDa species) showed elevation at day 1 in all groups with highest levels seen in Young Males. Exclusively in Young Females, these levels returned to baseline on day 3 , suggestive of a more efficient recovery. In aggregate, we demonstrate, for the first time, a distinctive pattern of response to AKI in Young Females relative to males which appears to be significantly altered in aging. These distinctions may offer novel targets to exploit therapeutically in both females and males in the treatment of AKI.

  14. Isorhynchophylline Protects PC12 Cells Against Beta-Amyloid-Induced Apoptosis via PI3K/Akt Signaling Pathway

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    Yan-Fang Xian

    2013-01-01

    Full Text Available The neurotoxicity of amyloid-β (Aβ has been implicated as a critical cause of Alzheimer’s disease. Isorhynchophylline (IRN, an oxindole alkaloid isolated from Uncaria rhynchophylla, exerts neuroprotective effect against Aβ25–35-induced neurotoxicity in vitro. However, the exact mechanism for its neuroprotective effect is not well understood. The present study aimed to investigate the molecular mechanisms underlying the protective action of IRN against Aβ25–35-induced neurotoxicity in cultured rat pheochromocytoma (PC12 cells. Pretreatment with IRN significantly increased the cell viability, inhibited the release of lactate dehydrogenase and the extent of DNA fragmentation in Aβ25–35-treated cells. IRN treatment was able to enhance the protein levels of phosphorylated Akt (p-Akt and glycogen synthase kinase-3β (p-GSK-3β. Lithium chloride blocked Aβ25–35-induced cellular apoptosis in a similar manner as IRN, suggesting that GSK-3β inhibition was involved in neuroprotective action of IRN. Pretreatment with LY294002 completely abolished the protective effects of IRN. Furthermore, IRN reversed Aβ25–35-induced attenuation in the level of phosphorylated cyclic AMP response element binding protein (p-CREB and the effect of IRN could be blocked by the PI3K inhibitor. These experimental findings unambiguously suggested that the protective effect of IRN against Aβ25–35-induced apoptosis in PC12 cells was associated with the enhancement of p-CREB expression via PI3K/Akt/GSK-3β signaling pathway.

  15. Nerves Regulate Cardiomyocyte Proliferation and Heart Regeneration.

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    Mahmoud, Ahmed I; O'Meara, Caitlin C; Gemberling, Matthew; Zhao, Long; Bryant, Donald M; Zheng, Ruimao; Gannon, Joseph B; Cai, Lei; Choi, Wen-Yee; Egnaczyk, Gregory F; Burns, Caroline E; Burns, C Geoffrey; MacRae, Calum A; Poss, Kenneth D; Lee, Richard T

    2015-08-24

    Some organisms, such as adult zebrafish and newborn mice, have the capacity to regenerate heart tissue following injury. Unraveling the mechanisms of heart regeneration is fundamental to understanding why regeneration fails in adult humans. Numerous studies have revealed that nerves are crucial for organ regeneration, thus we aimed to determine whether nerves guide heart regeneration. Here, we show using transgenic zebrafish that inhibition of cardiac innervation leads to reduction of myocyte proliferation following injury. Specifically, pharmacological inhibition of cholinergic nerve function reduces cardiomyocyte proliferation in the injured hearts of both zebrafish and neonatal mice. Direct mechanical denervation impairs heart regeneration in neonatal mice, which was rescued by the administration of neuregulin 1 (NRG1) and nerve growth factor (NGF) recombinant proteins. Transcriptional analysis of mechanically denervated hearts revealed a blunted inflammatory and immune response following injury. These findings demonstrate that nerve function is required for both zebrafish and mouse heart regeneration. Copyright © 2015 Elsevier Inc. All rights reserved.

  16. Role of heterotrimeric G protein and calcium in cardiomyocyte hypertrophy induced by IGF-1.

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    Carrasco, Loreto; Cea, Paola; Rocco, Paola; Peña-Oyarzún, Daniel; Rivera-Mejias, Pablo; Sotomayor-Flores, Cristian; Quiroga, Clara; Criollo, Alfredo; Ibarra, Cristian; Chiong, Mario; Lavandero, Sergio

    2014-04-01

    In the heart, insulin-like growth factor-1 (IGF-1) is a peptide with pro-hypertrophic and anti-apoptotic actions. The pro-hypertrophic properties of IGF-1 have been attributed to the extracellular regulated kinase (ERK) pathway. Recently, we reported that IGF-1 also increases intracellular Ca(2+) levels through a pertussis toxin (PTX)-sensitive G protein. Here we investigate whether this Ca(2+) signal is involved in IGF-1-induced cardiomyocyte hypertrophy. Our results show that the IGF-1-induced increase in Ca(2+) level is abolished by the IGF-1 receptor tyrosine kinase inhibitor AG538, PTX and the peptide inhibitor of Gβγ signaling, βARKct. Increases in the activities of Ca(2+) -dependent enzymes calcineurin, calmodulin kinase II (CaMKII), and protein kinase Cα (PKCα) were observed at 5 min after IGF-1 exposure. AG538, PTX, βARKct, and the dominant negative PKCα prevented the IGF-1-dependent phosphorylation of ERK1/2. Participation of calcineurin and CaMKII in ERK phosphorylation was discounted. IGF-1-induced cardiomyocyte hypertrophy, determined by cell size and β-myosin heavy chain (β-MHC), was prevented by AG538, PTX, βARKct, dominant negative PKCα, and the MEK1/2 inhibitor PD98059. Inhibition of calcineurin with CAIN did not abolish IGF-1-induced cardiac hypertrophy. We conclude that IGF-1 induces hypertrophy in cultured cardiomyocytes by activation of the receptor tyrosine kinase activity/βγ-subunits of a PTX-sensitive G protein/Ca(2+) /PKCα/ERK pathway without the participation of calcineurin. © 2013 Wiley Periodicals, Inc.

  17. Calcium and mitochondrial metabolism in ceramide-induced cardiomyocyte death.

    Science.gov (United States)

    Parra, Valentina; Moraga, Francisco; Kuzmicic, Jovan; López-Crisosto, Camila; Troncoso, Rodrigo; Torrealba, Natalia; Criollo, Alfredo; Díaz-Elizondo, Jessica; Rothermel, Beverly A; Quest, Andrew F G; Lavandero, Sergio

    2013-08-01

    Ceramides are important intermediates in the biosynthesis and degradation of sphingolipids that regulate numerous cellular processes, including cell cycle progression, cell growth, differentiation and death. In cardiomyocytes, ceramides induce apoptosis by decreasing mitochondrial membrane potential and promoting cytochrome-c release. Ca(2+) overload is a common feature of all types of cell death. The aim of this study was to determine the effect of ceramides on cytoplasmic Ca(2+) levels, mitochondrial function and cardiomyocyte death. Our data show that C2-ceramide induces apoptosis and necrosis in cultured cardiomyocytes by a mechanism involving increased Ca(2+) influx, mitochondrial network fragmentation and loss of the mitochondrial Ca(2+) buffer capacity. These biochemical events increase cytosolic Ca(2+) levels and trigger cardiomyocyte death via the activation of calpains. Copyright © 2013 Elsevier B.V. All rights reserved.

  18. Generation and purification of human stem cell-derived cardiomyocytes

    NARCIS (Netherlands)

    Schwach, Verena; Passier, Robert

    2016-01-01

    © 2016 International Society of Differentiation Efficient and reproducible generation and purification of human stem cell-derived cardiomyocytes (CMs) is crucial for regenerative medicine, disease modeling, drug screening and study of developmental events during cardiac specification. Established

  19. [Analysis on "component-target-pathway" of Paeonia lactiflora in treating cardiac diseases based on data mining].

    Science.gov (United States)

    Liu, Yang; Zhang, Fang-Bo; Tang, Shi-Huan; Wang, Ping; Li, Sen; Su, Jin; Zhou, Rong-Rong; Zhang, Jia-Qi; Sun, Hui-Feng

    2018-04-01

    Based on the literature review and modern application of Paeonia lactiflora in heart diseases, this article would predict the target of drug and disease by intergrative pharmacology platform of traditional Chinese medicine (TCMIP, http://www.tcmip.cn), and then explore the molecular mechanism of P. lactiflora in treatment of heart disease, providing theoretical basis and method for further studies on P. lactiflora. According to the ancient books, P. lactiflora with functions of "removing the vascular obstruction, removing the lumps, relieving pain, diuretic, nutrient qi" and other effects, have been used for many times to treat heart disease. Some prescriptions are also favored by the modern physicians nowadays. With the development of science, the chemical components that play a role in heart disease and the interrelation between these components and the body become the research hotspot. In order to further reveal the pharmacological substance base and molecular mechanism of P. lactiflora for the treatment of such diseases, TCM-IP was used to obtain multiple molecular targets and signaling pathways in treatment of heart disease. ATP1A1, a common target of drug and disease, was related to energy, and HDAC2 mainly regulated cardiomyocyte hypertrophy gene and cardiomyocyte expression. Other main drug targets such as GCK, CHUK and PRKAA2 indirectly regulated heart disease through many pathways; multiple disease-associated signaling pathways interfered with various heart diseases including coronary heart disease, myocardial ischemia and myocardial hypertrophy through influencing energy metabolism, enzyme activity and gene expression. In conclusion, P. lactiflora plays a role in protecting heart function by regulating the gene expression of cardiomyocytes directly. Meanwhile, it can indirectly intervene in other pathways of heart function, and thus participate in the treatment of heart disease. In this paper, the molecular mechanism of P. lactiflora for treatment of

  20. Adenosine A1 Receptor Protects Against Cisplatin Ototoxicity by Suppressing the NOX3/STAT1 Inflammatory Pathway in the Cochlea.

    Science.gov (United States)

    Kaur, Tejbeer; Borse, Vikrant; Sheth, Sandeep; Sheehan, Kelly; Ghosh, Sumana; Tupal, Srinivasan; Jajoo, Sarvesh; Mukherjea, Debashree; Rybak, Leonard P; Ramkumar, Vickram

    2016-04-06

    Cisplatin is a commonly used antineoplastic agent that produces ototoxicity that is mediated in part by increasing levels of reactive oxygen species (ROS) via the NOX3 NADPH oxidase pathway in the cochlea. Recent studies implicate ROS generation in mediating inflammatory and apoptotic processes and hearing loss by activating signal transducer and activator of transcription (STAT1). In this study, we show that the adenosine A1 receptor (A1AR) protects against cisplatin ototoxicity by suppressing an inflammatory response initiated by ROS generation via NOX3 NADPH oxidase, leading to inhibition of STAT1. Trans-tympanic administration of the A1AR agonist R-phenylisopropyladenosine (R-PIA) inhibited cisplatin-induced ototoxicity, as measured by auditory brainstem responses and scanning electron microscopy in male Wistar rats. This was associated with reduced NOX3 expression, STAT1 activation, tumor necrosis factor-α (TNF-α) levels, and apoptosis in the cochlea. In vitro studies in UB/OC-1 cells, an organ of Corti immortalized cell line, showed that R-PIA reduced cisplatin-induced phosphorylation of STAT1 Ser(727) (but not Tyr(701)) and STAT1 luciferase activity by suppressing the ERK1/2, p38, and JNK mitogen-activated protein kinase (MAPK) pathways.R-PIA also decreased the expression of STAT1 target genes, such as TNF-α, inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) and reduced cisplatin-mediated apoptosis. These data suggest that the A1AR provides otoprotection by suppressing NOX3 and inflammation in the cochlea and could serve as an ideal target for otoprotective drug therapy. Cisplatin is a widely used chemotherapeutic agent for the treatment of solid tumors. Its use results in significant and permanent hearing loss, for which no US Food and Drug Administration-approved treatment is currently available. In this study, we targeted the cochlear adenosine A1 receptor (A1AR) by trans-tympanic injections of the agonist R

  1. Adenosine A1 Receptor Protects Against Cisplatin Ototoxicity by Suppressing the NOX3/STAT1 Inflammatory Pathway in the Cochlea

    Science.gov (United States)

    Kaur, Tejbeer; Borse, Vikrant; Sheth, Sandeep; Sheehan, Kelly; Ghosh, Sumana; Tupal, Srinivasan; Jajoo, Sarvesh; Mukherjea, Debashree; Rybak, Leonard P.

    2016-01-01

    Cisplatin is a commonly used antineoplastic agent that produces ototoxicity that is mediated in part by increasing levels of reactive oxygen species (ROS) via the NOX3 NADPH oxidase pathway in the cochlea. Recent studies implicate ROS generation in mediating inflammatory and apoptotic processes and hearing loss by activating signal transducer and activator of transcription (STAT1). In this study, we show that the adenosine A1 receptor (A1AR) protects against cisplatin ototoxicity by suppressing an inflammatory response initiated by ROS generation via NOX3 NADPH oxidase, leading to inhibition of STAT1. Trans-tympanic administration of the A1AR agonist R-phenylisopropyladenosine (R-PIA) inhibited cisplatin-induced ototoxicity, as measured by auditory brainstem responses and scanning electron microscopy in male Wistar rats. This was associated with reduced NOX3 expression, STAT1 activation, tumor necrosis factor-α (TNF-α) levels, and apoptosis in the cochlea. In vitro studies in UB/OC-1 cells, an organ of Corti immortalized cell line, showed that R-PIA reduced cisplatin-induced phosphorylation of STAT1 Ser727 (but not Tyr701) and STAT1 luciferase activity by suppressing the ERK1/2, p38, and JNK mitogen-activated protein kinase (MAPK) pathways. R-PIA also decreased the expression of STAT1 target genes, such as TNF-α, inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) and reduced cisplatin-mediated apoptosis. These data suggest that the A1AR provides otoprotection by suppressing NOX3 and inflammation in the cochlea and could serve as an ideal target for otoprotective drug therapy. SIGNIFICANCE STATEMENT Cisplatin is a widely used chemotherapeutic agent for the treatment of solid tumors. Its use results in significant and permanent hearing loss, for which no US Food and Drug Administration-approved treatment is currently available. In this study, we targeted the cochlear adenosine A1 receptor (A1AR) by trans-tympanic injections of the agonist R

  2. A BAG3 chaperone complex maintains cardiomyocyte function during proteotoxic stress.

    Science.gov (United States)

    Judge, Luke M; Perez-Bermejo, Juan A; Truong, Annie; Ribeiro, Alexandre Js; Yoo, Jennie C; Jensen, Christina L; Mandegar, Mohammad A; Huebsch, Nathaniel; Kaake, Robyn M; So, Po-Lin; Srivastava, Deepak; Pruitt, Beth L; Krogan, Nevan J; Conklin, Bruce R

    2017-07-20

    Molecular chaperones regulate quality control in the human proteome, pathways that have been implicated in many diseases, including heart failure. Mutations in the BAG3 gene, which encodes a co-chaperone protein, have been associated with heart failure due to both inherited and sporadic dilated cardiomyopathy. Familial BAG3 mutations are autosomal dominant and frequently cause truncation of the coding sequence, suggesting a heterozygous loss-of-function mechanism. However, heterozygous knockout of the murine BAG3 gene did not cause a detectable phenotype. To model BAG3 cardiomyopathy in a human system, we generated an isogenic series of human induced pluripotent stem cells (iPSCs) with loss-of-function mutations in BAG3. Heterozygous BAG3 mutations reduced protein expression, disrupted myofibril structure, and compromised contractile function in iPSC-derived cardiomyocytes (iPS-CMs). BAG3-deficient iPS-CMs were particularly sensitive to further myofibril disruption and contractile dysfunction upon exposure to proteasome inhibitors known to cause cardiotoxicity. We performed affinity tagging of the endogenous BAG3 protein and mass spectrometry proteomics to further define the cardioprotective chaperone complex that BAG3 coordinates in the human heart. Our results establish a model for evaluating protein quality control pathways in human cardiomyocytes and their potential as therapeutic targets and susceptibility factors for cardiac drug toxicity.

  3. Polysaccharide from Angelica sinensis protects H9c2 cells against oxidative injury and endoplasmic reticulum stress by activating the ATF6 pathway.

    Science.gov (United States)

    Niu, Xiaowei; Zhang, Jingjing; Ling, Chun; Bai, Ming; Peng, Yu; Sun, Shaobo; Li, Yingdong; Zhang, Zheng

    2018-01-01

    Objectives Angelica sinensis exerts various pharmacological effects, such as antioxidant and anti-apoptotic activity. This study aimed to investigate the active ingredients in A. sinensis with antioxidant properties and whether A. sinensis polysaccharide (ASP) protects H9c2 cells against oxidative and endoplasmic reticulum (ER) stress. Methods The ingredients of A. sinensis and their targets and related pathways were determined using web-based databases. Markers of oxidative stress, cell viability, apoptosis, and ER stress-related signalling pathways were measured in H9c2 cells treated with hydrogen peroxide (H 2 O 2 ) and ASP. Results The ingredient-pathway-disease network showed that A. sinensis exerted protective effects against oxidative injury through its various active ingredients on regulation of multiple pathways. Subsequent experiments showed that ASP pretreatment significantly decreased H 2 O 2 -induced cytotoxicity and apoptosis in H9c2 cells. ASP pretreatment inhibited H 2 O 2 -induced reactive oxygen species generation, lactic dehydrogenase release, and malondialdehyde production. ASP exerted beneficial effects by inducing activating transcription factor 6 (ATF6) and increasing ATF6 target protein levels, which in turn attenuated ER stress and increased antioxidant activity. Conclusions Our findings indicate that ASP, a major water-soluble component of A. sinensis, exerts protective effects against H 2 O 2 -induced injury in H9c2 cells by activating the ATF6 pathway, thus ameliorating ER and oxidative stress.

  4. Allicin protects against H2O2-induced apoptosis of PC12 cells via the mitochondrial pathway.

    Science.gov (United States)

    Lv, Runxiao; Du, Lili; Lu, Chunwen; Wu, Jinhui; Ding, Muchen; Wang, Chao; Mao, Ningfang; Shi, Zhicai

    2017-09-01

    Allicin is a major bioactive ingredient of garlic and has a broad range of biological activities. Allicin has been reported to protect against cell apoptosis induced by H 2 O 2 in human umbilical vein endothelial cells. The present study evaluated the neuroprotective effect of allicin on the H 2 O 2 -induced apoptosis of rat pheochromocytoma PC12 cells in vitro and explored the underlying mechanism involved. PC12 cells were incubated with increasing concentrations of allicin and the toxic effect of allicin was measured by MTT assay. The cells were pretreated for 24 h with low dose (L-), medium dose (M-) and high dose (H-) of allicin, followed by exposure to 200 µM H 2 O 2 for 2 h, and the cell viability was examined by MTT assay. In addition, cell apoptosis rate was analyzed by Annexin V-FITC/PI assay, while intracellular reactive oxygen species (ROS) and mitochondrial transmembrane potential (∆ψm) were measured by flow cytometry. Bcl-2, Bax, cleaved-caspase-3 and cytochrome c (Cyt C) in the mitochondria were also examined by western blotting. The results demonstrated that 0.01 µg/ml (L-allicin), 0.1 µg/ml (M-allicin) and 1 µg/ml (H-allicin) were non-toxic doses of allicin. Furthermore, H 2 O 2 reduced cell viability, promoted cell apoptosis, induced ROS production and decreased ∆ψm. However, allicin treatment reversed the effect of H 2 O 2 in a dose-dependent manner. It was also observed that H 2 O 2 exposure significantly decreased Bcl-2 and mitochondrial Cyt C, while it increased Bax and cleaved-caspase-3, which were attenuated by allicin pretreatment. The results revealed that allicin protected PC12 cells from H 2 O 2 -induced cell apoptosis via the mitochondrial pathway, suggesting the potential neuroprotective effect of allicin against neurological diseases.

  5. Salvianic acid A sodium protects HUVEC cells against tert-butyl hydroperoxide induced oxidative injury via mitochondria-dependent pathway.

    Science.gov (United States)

    Jia, Dan; Li, Tian; Chen, Xiaofei; Ding, Xuan; Chai, Yifeng; Chen, Alex F; Zhu, Zhenyu; Zhang, Chuan

    2018-01-05

    Salvianic acid A (Danshensu) is a major water-soluble component extracted from Salvia miltiorrhiza (Danshen), which has been widely used in clinic in China for treatment of cardiovascular diseases (CVDs). This study aimed to investigate the protective effects of salvianic acid A sodium (SAAS) against tert-butyl hydroperoxide (t-BHP) induced human umbilical vein endothelial cell (HUVEC) oxidative injury and the underlying molecular mechanisms. In the antioxidant activity-assessing model, SAAS pretreatment significantly ameliorated the cell growth inhibition and apoptosis induced by t-BHP. An ultra-performance liquid chromatography/quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS) based-metabolic profiling was developed to investigate the metabolic changes of HUVEC cells in response to t-BHP and SAAS. The results revealed that t-BHP injury upregulated 13 metabolites mainly involved in tryptophan metabolism and phenylalanine metabolism which were highly correlated with mitochondrial function and oxidative stress, and 50 μM SAAS pretreatment effectively reversed these metabolic changes. Further biomedical research indicated that SAAS pretreatment reduced the t-BHP induced increase of lactate dehydrogenase (LDH), intracellular reactive oxygen species (ROS), malondialdehyde (MDA) and mitochondrial membrane potential (MMP), and the decrease of key antioxidant enzymes through mitochondria antioxidative pathways via JAK2/STAT3 and PI3K/Akt/GSK-3β signalings. Taken together, our results suggested that SAAS may protect HUVEC cells against t-BHP induced oxidative injury via mitochondrial antioxidative defense system. Copyright © 2017 Elsevier B.V. All rights reserved.

  6. Emodin Protects against Diabetic Cardiomyopathy by Regulating the AKT/GSK-3β Signaling Pathway in the Rat Model

    Directory of Open Access Journals (Sweden)

    Zhiqin Wu

    2014-09-01

    Full Text Available Diabetes mellitus (DM has been recognized as a major health problem. Emodin (Emo has been reported to exhibit protective effects against diabetic nephropathy. However, little has been known about the effect of Emo on diabetic cardiomyopathy (DCM. A type 2 DM model was induced in rats by low dose streptozotocin (STZ combined with high energy intake. We found that Emo-treated groups displayed significantly higher body weight (BW and lower heart weight (HW/BW. Furthermore, Emo could significantly decrease blood glucose, total cholesterol (TG levels, and triglyceride (TC levels in diabetic rats. Moreover, the Emo-treated group showed a marked increase in heart rate (HR and showed lower left ventricular end-diastolic diameter (LVEDD, left ventricular end-systolic diameter (LVESD, left ventricular posterior wall thickness (LWPWT, and interventricular septal diastolic wall thickness (IVSD. Emo induced a significant increase in phosphorylation of Akt and GSK-3β in myocardium. These results suggest that Emo may have great therapeutic potential in the treatment of DCM by Akt/GSK-3β signaling pathway.

  7. The polyphenol fisetin protects bone by repressing NF-κB and MKP-1-dependent signaling pathways in osteoclasts.

    Directory of Open Access Journals (Sweden)

    Laurent Léotoing

    Full Text Available Osteoporosis is a bone pathology leading to increase fractures risk and challenging quality of life. Since current treatments could exhibit deleterious side effects, the use of food compounds derived from plants represents a promising innovative alternative due to their potential therapeutic and preventive activities against human diseases. In this study, we investigated the ability of the polyphenol fisetin to counter osteoporosis and analyzed the cellular and molecular mechanisms involved. In vivo, fisetin consumption significantly prevented bone loss in estrogen deficiency and inflammation mice osteoporosis models. Indeed, bone mineral density, micro-architecture parameters and bone markers were positively modulated by fisetin. Consistent with in vivo results, we showed that fisetin represses RANKL-induced osteoclast differentiation and activity as demonstrated by an inhibition of multinucleated cells formation, TRAP activity and differentiation genes expression. The signaling pathways NF-κB, p38 MAPK, JNK and the key transcription factors c-Fos and NFATc1 expressions induced by RANKL, were negatively regulated by fisetin. We further showed that fisetin inhibits the constitutive proteasomal degradation of MKP-1, the phosphatase that deactivates p38 and JNK. Consistently, using shRNA stable cell lines, we demonstrated that impairment of MKP-1 decreases fisetin potency. Taken together, these results strongly support that fisetin should be further considered as a bone protective agent.

  8. The Polyphenol Fisetin Protects Bone by Repressing NF-κB and MKP-1-Dependent Signaling Pathways in Osteoclasts

    Science.gov (United States)

    Léotoing, Laurent; Wauquier, Fabien; Guicheux, Jérôme; Miot-Noirault, Elisabeth; Wittrant, Yohann; Coxam, Véronique

    2013-01-01

    Osteoporosis is a bone pathology leading to increase fractures risk and challenging quality of life. Since current treatments could exhibit deleterious side effects, the use of food compounds derived from plants represents a promising innovative alternative due to their potential therapeutic and preventive activities against human diseases. In this study, we investigated the ability of the polyphenol fisetin to counter osteoporosis and analyzed the cellular and molecular mechanisms involved. In vivo, fisetin consumption significantly prevented bone loss in estrogen deficiency and inflammation mice osteoporosis models. Indeed, bone mineral density, micro-architecture parameters and bone markers were positively modulated by fisetin. Consistent with in vivo results, we showed that fisetin represses RANKL-induced osteoclast differentiation and activity as demonstrated by an inhibition of multinucleated cells formation, TRAP activity and differentiation genes expression. The signaling pathways NF-κB, p38 MAPK, JNK and the key transcription factors c-Fos and NFATc1 expressions induced by RANKL, were negatively regulated by fisetin. We further showed that fisetin inhibits the constitutive proteasomal degradation of MKP-1, the phosphatase that deactivates p38 and JNK. Consistently, using shRNA stable cell lines, we demonstrated that impairment of MKP-1 decreases fisetin potency. Taken together, these results strongly support that fisetin should be further considered as a bone protective agent. PMID:23861901

  9. Cardioprotective Effects of Quercetin in Cardiomyocyte under Ischemia/Reperfusion Injury

    Directory of Open Access Journals (Sweden)

    Yi-Wen Chen

    2013-01-01

    Full Text Available Quercetin, a polyphenolic compound existing in many vegetables, fruits, has antiinflammatory, antiproliferation, and antioxidant effect on mammalian cells. Quercetin was evaluated for protecting cardiomyocytes from ischemia/reperfusion injury, but its protective mechanism remains unclear in the current study. The cardioprotective effects of quercetin are achieved by reducing the activity of Src kinase, signal transducer and activator of transcription 3 (STAT3, caspase 9, Bax, intracellular reactive oxygen species production, and inflammatory factor and inducible MnSOD expression. Fluorescence two-dimensional differential gel electrophoresis (2D-DIGE and matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS can reveal the differentially expressed proteins of H9C2 cells treated with H2O2 or quercetin. Although 17 identified proteins were altered in H2O2-induced cells, these proteins such as alpha-soluble NSF attachment protein (α-SNAP, Ena/VASP-like protein (Evl, and isopentenyl-diphosphate delta-isomerase 1 (Idi-1 were reverted by pretreatment with quercetin, which correlates with kinase activation, DNA repair, lipid, and protein metabolism. Quercetin dephosphorylates Src kinase in H2O2-induced H9C2 cells and likely blocks the H2O2-induced inflammatory response through STAT3 kinase modulation. This probably contributes to prevent ischemia/reperfusion injury in cardiomyocytes.

  10. Differential Gene Expression Patterns in Chicken Cardiomyocytes during Hydrogen Peroxide-Induced Apoptosis.

    Science.gov (United States)

    Wan, Chunyun; Xiang, Jinmei; Li, Youwen; Guo, Dingzong

    2016-01-01

    Hydrogen peroxide (H2O2) is both an exogenous and endogenous cytotoxic agent that can reliably induce apoptosis in numerous cell types for studies on apoptosis signaling pathways. However, little is known of these apoptotic processes in myocardial cells of chicken, a species prone to progressive heart failure. Sequencing of mRNA transcripts (RNA-Seq) allows for the identification of differentially expressed genes under various physiological and pathological conditions to elucidate the molecular pathways involved, including cellular responses to exogenous and endogenous toxins. We used RNA-seq to examine genes differentially expressed during H2O2-induced apoptosis in primary cultures of embryonic chicken cardiomyocytes. Following control or H2O2 treatment, RNA was extracted and sequencing performed to identify novel transcripts up- or downregulated in the H2O2 treatment group and construct protein-protein interaction networks. Of the 19,268 known and 2,160 novel transcripts identified in both control and H2O2 treatment groups, 4,650 showed significant differential expression. Among them, 55.63% were upregulated and 44.37% downregulated. Initiation of apoptosis by H2O2 was associated with upregulation of caspase-8, caspase-9, and caspase-3, and downregulation of anti-apoptotic genes API5 and TRIA1. Many other differentially expressed genes were associated with metabolic pathways (including 'Fatty acid metabolism', 'Alanine, aspartate, and glutamate metabolism', and 'Biosynthesis of unsaturated fatty acids') and cell signaling pathways (including 'PPAR signaling pathway', 'Adipocytokine signaling pathway', 'TGF-beta signaling pathway', 'MAPK signaling pathway', and 'p53 signaling pathway'). In chicken cardiomyocytes, H2O2 alters the expression of numerous genes linked to cell signaling and metabolism as well as genes directly associated with apoptosis. In particular, H2O2 also affects the biosynthesis and processing of proteins and unsaturated fatty acids. These

  11. αvβ5 Integrin/FAK/PGC-1α Pathway Confers Protective Effects on Retinal Pigment Epithelium.

    Directory of Open Access Journals (Sweden)

    Murilo F Roggia

    Full Text Available To elucidate the mechanism of the induction of peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α by photoreceptor outer segments (POS and its effects on retinal pigment epithelium (RPE.PGC-1α upregulation by POS was confirmed in ARPE-19 cells and in RPE ex vivo. To elucidate the mechanism, siRNAs against β5 integrin, CD36, Mer tyrosine kinase (MerTK, and Atg5, blocking antibodies against CD36 and MerTK, and a specific inhibitor for focal adhesion kinase (FAK were used. We examined the effect of POS-induced PGC-1α upregulation on the levels of reactive oxygen species (ROS, mitochondrial biogenesis, senescence-associated β-galactosidase (SA-β-gal after H2O2 treatment, and lysosomal activity. Lysosomal activity was evaluated through transcriptional factor EB and its target genes, and the activity of cathepsin D. Lipid metabolism after POS treatment was assessed using Oil Red O and BODIPY C11. RPE phenotypes of PGC-1α-deficient mice were examined.POS-induced PGC-1α upregulation was suppressed by siRNA against β5 integrin and a FAK inhibitor. siRNAs and blocking antibodies against CD36 and MerTK enhanced the effect of POS on PGC-1α. The upregulation of PGC-1α increased the levels of mRNA for antioxidant enzymes and stimulated mitochondrial biogenesis, decreased ROS levels, and reduced SA-β-gal staining in H2O2-treated ARPE-19 cells. PGC-1α was critical for lysosomal activity and lipid metabolism after POS treatment. PGC-1α-deficient mice demonstrated an accumulation of type 2 lysosomes in RPE, thickening of Bruch's membrane, and poor choriocapillaris vasculature.The binding, but not the internalization of POS confers protective effects on RPE cells through the αvβ5 integrin/FAK/PGC-1α pathway.

  12. Inhibition of the kynurenine pathway protects against reactive microglial-associated reductions in the complexity of primary cortical neurons.

    Science.gov (United States)

    O'Farrell, Katherine; Fagan, Eimear; Connor, Thomas J; Harkin, Andrew

    2017-09-05

    Brain glia possess the rate limiting enzyme indoleamine 2, 3-dioxygenase (IDO) which catalyses the conversion of tryptophan to kynurenine. Microglia also express kynurenine monooxygenase (KMO) and kynureninase (KYNU) which lead to the production of the free radical producing metabolites, 3-hydroxykynurenine and 3-hydroxyanthranillic acid respectively and subsequently production of the NMDA receptor agonist quinolinic acid. The aim of this study was to examine the effect of IFNγ-stimulated kynurenine pathway (KP) induction in microglia on neurite outgrowth and complexity, and to determine whether alterations could be abrogated using pharmacological inhibitors of the KP. BV-2 microglia were treated with IFNγ (5ng/ml) for 24h and conditioned media (CM) was placed on primary cortical neurons 3 days in vitro (DIV) for 48h. Neurons were fixed and neurite outgrowth and complexity was assessed using fluorescent immunocytochemistry followed by Sholl analysis. Results show increased mRNA expression of IDO, KMO and KYNU, and increased concentrations of tryptophan, kynurenine, and 3-hydroxykynurenine in the CM of IFNγ-stimulated BV-2 microglia. The IFNγ-stimulated BV-2 microglial CM reduced neurite outgrowth and complexity with reductions in various parameters of neurite outgrowth prevented when BV-2 microglia were pre-treated with either the IDO inhibitor, 1-methyltryptophan (1-MT) (L) (0.5mM; 30min), the KMO inhibitor, Ro 61-8048 (1μM; 30min), the synthetic glucocorticoid, dexamethasone (1μM; 2h) -which suppresses IFNγ-induced IDO - and the N-methyl-D-aspartate (NMDA) receptor antagonist, MK801 (0.1μM; 30min). Overall this study indicates that inhibition of the KP in microglia may be targeted to protect against reactive microglial-associated neuronal atrophy. Copyright © 2017 Elsevier B.V. All rights reserved.

  13. Rcan1-1L overexpression induces mitochondrial autophagy and improves cell survival in angiotensin II-exposed cardiomyocytes

    International Nuclear Information System (INIS)

    Duan, Hongyan; Li, Yongqiang; Yan, Lijie; Yang, Haitao; Wu, Jintao; Qian, Peng; Li, Bing; Wang, Shanling

    2015-01-01

    Mitochondrial autophagy is an important adaptive stress response and can be modulated by various key molecules. A previous study found that the regulator of calcineurin 1-1L (Rcan1-1L) may regulate mitochondrial autophagy and cause mitochondria degradation in neurocytes. However, the effect of Rcan1-1L on cardiomyocytes has not been determined. In the present study, we aimed to investigate the role of Rcan1-1L in angiotensin II (Ang II)-exposed human cardiomyocytes. Above all, Human adult cardiac myocytes (HACMs) were exposed to 200 nmol/L Ang II for 4 days. Enhanced H 2 O 2 production, cytochrome C release and mitochondrial permeability were observed in these cells, which were blocked by valsartan. Consistently, Ang II exposure significantly reduced cardiomyocyte viability. However, transfection of Rcan1-1L vector promoted cell viability and ameliorated the apoptosis caused by Ang II. Rcan1-1L clearly promoted mitochondrial autophagy in HACMs, with elevated autophagy protein (ATG) 5 and light chain 3 (LC3) expression. Transient mitochondrial biogenesis and reduced cytochrome C release was also induced by Rcan1-1L. Additionally, Rcan1-1L significantly inhibited calcineurin/nuclear factor of activated T cells (NFAT) signaling. We thus conclude that Rcan1-1L may play a protective role in Ang II-treated cardiomyocytes through the induction of mitochondrial autophagy, and may be an alternative method of cardiac protection. - Highlights: • Transfection of Rcan1-1L into HACMs promoted cell viability and reduced apoptosis. • Transfection of Rcan1-1L promoted mitochondrial autophagy in HACMs. • Rcan1-1L inhibited the calcineurin/nuclear factor of activated T cells signaling

  14. Rcan1-1L overexpression induces mitochondrial autophagy and improves cell survival in angiotensin II-exposed cardiomyocytes

    Energy Technology Data Exchange (ETDEWEB)

    Duan, Hongyan; Li, Yongqiang; Yan, Lijie; Yang, Haitao; Wu, Jintao; Qian, Peng; Li, Bing; Wang, Shanling, E-mail: shanglingwang@126.com

    2015-07-01

    Mitochondrial autophagy is an important adaptive stress response and can be modulated by various key molecules. A previous study found that the regulator of calcineurin 1-1L (Rcan1-1L) may regulate mitochondrial autophagy and cause mitochondria degradation in neurocytes. However, the effect of Rcan1-1L on cardiomyocytes has not been determined. In the present study, we aimed to investigate the role of Rcan1-1L in angiotensin II (Ang II)-exposed human cardiomyocytes. Above all, Human adult cardiac myocytes (HACMs) were exposed to 200 nmol/L Ang II for 4 days. Enhanced H{sub 2}O{sub 2} production, cytochrome C release and mitochondrial permeability were observed in these cells, which were blocked by valsartan. Consistently, Ang II exposure significantly reduced cardiomyocyte viability. However, transfection of Rcan1-1L vector promoted cell viability and ameliorated the apoptosis caused by Ang II. Rcan1-1L clearly promoted mitochondrial autophagy in HACMs, with elevated autophagy protein (ATG) 5 and light chain 3 (LC3) expression. Transient mitochondrial biogenesis and reduced cytochrome C release was also induced by Rcan1-1L. Additionally, Rcan1-1L significantly inhibited calcineurin/nuclear factor of activated T cells (NFAT) signaling. We thus conclude that Rcan1-1L may play a protective role in Ang II-treated cardiomyocytes through the induction of mitochondrial autophagy, and may be an alternative method of cardiac protection. - Highlights: • Transfection of Rcan1-1L into HACMs promoted cell viability and reduced apoptosis. • Transfection of Rcan1-1L promoted mitochondrial autophagy in HACMs. • Rcan1-1L inhibited the calcineurin/nuclear factor of activated T cells signaling.

  15. Particulate matter exposure exacerbates high glucose-induced cardiomyocyte dysfunction through ROS generation.

    Directory of Open Access Journals (Sweden)

    Li Zuo

    cardiomyocytes exposed to HG-containing media, which is potentially mediated by various ROS generation pathways.

  16. Adrenaline and reactive oxygen species elicit proteome and energetic metabolism modifications in freshly isolated rat cardiomyocytes

    International Nuclear Information System (INIS)

    Costa, Vera Marisa; Silva, Renata; Tavares, Ludgero Canario; Vitorino, Rui; Amado, Francisco; Carvalho, Felix; Bastos, Maria de Lourdes; Carvalho, Marcia; Carvalho, Rui Albuquerque; Remiao, Fernando

    2009-01-01

    The sustained elevation of plasma and interstitial catecholamine levels, namely adrenaline (ADR), and the generation of reactive oxygen species (ROS) are well recognized hallmarks of several cardiopathologic conditions, like cardiac ischemia/reperfusion (I/R) and heart failure (HF). The present work aimed to investigate the proteomics and energetic metabolism of cardiomyocytes incubated with ADR and/or ROS. To mimic pathologic conditions, freshly isolated calcium-tolerant cardiomyocytes from adult rat were incubated with ADR alone or in the presence of a system capable of generating ROS [(xanthine with xanthine oxidase) (XXO)]. Two-dimensional electrophoresis with matrix-assisted laser desorption/ionization and time-of-flight mass spectrometer analysis were used to define protein spot alterations in the cardiomyocytes incubated with ADR and/or ROS. Moreover, the energetic metabolism and the activity of mitochondrial complexes were evaluated by nuclear magnetic resonance and spectrophotometric determinations, respectively. The protein extract was mainly constituted by cardiac mitochondrial proteins and the alterations found were included in five functional classes: (i) structural proteins, notably myosin light chain-2; (ii) redox regulation proteins, in particular superoxide dismutase (SOD); (iii) energetic metabolism proteins, encompassing ATP synthase alpha chain and dihydrolipoyllysine-residue acetyltransferase component of pyruvate dehydrogenase complex; (iv) stress response proteins, like the heat shock proteins; and (v) regulatory proteins, like cytochrome c and voltage-dependent anion channel 1. The XXO system elicited alterations in cardiac contractile proteins, as they showed high levels of cleavage, and also altered energetic metabolism, through increased lactate and alanine levels. The cardiomyocytes incubation with ADR resulted in an accentuated increase in mitochondrial complexes activity and the decrease in alanine/lactate ratio, thus reflecting a high

  17. Mitochondrial p38β and manganese superoxide dismutase interaction mediated by estrogen in cardiomyocytes.

    Directory of Open Access Journals (Sweden)

    Han Liu

    Full Text Available While etiology behind the observed acceleration of ischemic heart disease in postmenopausal women is poorly understood, collective scientific data suggest cardioprotective effects of the endogenous female sex hormone, estrogen. We have previously shown that 17β-estradiol (E2 protects cardiomyocytes exposed to hypoxia-reoxygenation (H/R by inhibiting p38α - p53 signaling in apoptosis and activating pro-survival p38β mitogen activated protein kinase (p38β MAPK, leading to suppression of reactive oxygen species (ROS post H/R. However, little is known about the mechanism behind the antioxidant actions of E2-dependent p38β. The aim of this study is to determine whether the cytoprotection by estrogen involves regulation of manganese superoxide dismutase (MnSOD, a major mitochondrial ROS scavenging enzyme, via cardiac p38β.We identified mitochondrial p38β by immunocytochemistry and by immunoblotting in mitochondria isolated from neonatal cardiomyocytes of Sprague-Dawley rats. E2 facilitated the mitochondrial localization of the active form of the kinase, phosphorylated p38β (p-p38β. E2 also reduced the H/R-induced mitochondrial membrane potential decline, augmented the MnSOD activity and suppressed anion superoxide generation, while the dismutase protein expression remained unaltered. Co-immunoprecipitation studies showed physical association between MnSOD and p38β. p38β phosphorylated MnSOD in an E2-dependent manner in in-vitro kinase assays.This work demonstrates for the first time a mitochondrial pool of active p38β and E2-mediated phosphorylation of MnSOD by the kinase. The results shed light on the mechanism behind the cytoprotective actions of E2 in cardiomyocytes under oxidative stress.

  18. Functional Differences in Engineered Myocardium from Embryonic Stem Cell-Derived versus Neonatal Cardiomyocytes

    NARCIS (Netherlands)

    Feinberg, Adam W.; Ripplinger, Crystal M.; van der Meer, Peter; Sheehy, Sean P.; Domian, Ibrahim; Chien, Kenneth R.; Parker, Kevin Kit

    2013-01-01

    Stem cell-derived cardiomyocytes represent unique tools for cell-and tissue-based regenerative therapies, drug discovery and safety, and studies of fundamental heart-failure mechanisms. However, the degree to which stem cell-derived cardiomyocytes compare to mature cardiomyocytes is often debated.

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  6. Rapid Induction of Aldosterone Synthesis in Cultured Neonatal Rat Cardiomyocytes under High Glucose Conditions

    Directory of Open Access Journals (Sweden)

    Masami Fujisaki

    2013-01-01

    Full Text Available In addition to classical adrenal cortical biosynthetic pathway, there is increasing evidence that aldosterone is produced in extra-adrenal tissues. Although we previously reported aldosterone production in the heart, the concept of cardiac aldosterone synthesis remains controversial. This is partly due to lack of established experimental models representing aldosterone synthase (CYP11B2 expression in robustly reproducible fashion. We herein investigated suitable conditions in neonatal rat cardiomyocytes (NRCMs culture system producing CYP11B2 with considerable efficacy. NRCMs were cultured with various glucose doses for 2–24 hours. CYP11B2 mRNA expression and aldosterone concentrations secreted from NRCMs were determined using real-time PCR and enzyme immunoassay, respectively. We found that suitable conditions for CYP11B2 induction included four-hour incubation with high glucose conditions. Under these particular conditions, CYP11B2 expression, in accordance with aldosterone secretion, was significantly increased compared to those observed in the cells cultured under standard-glucose condition. Angiotensin II receptor blocker partially inhibited this CYP11B2 induction, suggesting that there is local renin-angiotensin-aldosterone system activation under high glucose conditions. The suitable conditions for CYP11B2 induction in NRCMs culture system are now clarified: high-glucose conditions with relatively brief period of culture promote CYP11B2 expression in cardiomyocytes. The current system will help to accelerate further progress in research on cardiac tissue aldosterone synthesis.

  7. Krp1 (Sarcosin) promotes lateral fusion of myofibril assembly intermediates in cultured mouse cardiomyocytes

    International Nuclear Information System (INIS)

    Greenberg, Cynthia C.; Connelly, Patricia S.; Daniels, Mathew P.; Horowits, Robert

    2008-01-01

    Krp1, also called sarcosin, is a cardiac and skeletal muscle kelch repeat protein hypothesized to promote the assembly of myofibrils, the contractile organelles of striated muscles, through interaction with N-RAP and actin. To elucidate its role, endogenous Krp1 was studied in primary embryonic mouse cardiomyocytes. While immunofluorescence showed punctate Krp1 distribution throughout the cell, detergent extraction revealed a significant pool of Krp1 associated with cytoskeletal elements. Reduction of Krp1 expression with siRNA resulted in specific inhibition of myofibril accumulation with no effect on cell spreading. Immunostaining analysis and electron microscopy revealed that cardiomyocytes lacking Krp1 contained sarcomeric proteins with longitudinal periodicities similar to mature myofibrils, but fibrils remained thin and separated. These thin myofibrils were degraded by a scission mechanism distinct from the myofibril disassembly pathway observed during cell division in the developing heart. The data are consistent with a model in which Krp1 promotes lateral fusion of adjacent thin fibrils into mature, wide myofibrils and contribute insight into mechanisms of myofibrillogenesis and disassembly

  8. Effects of Mechanical Coupling Between Cardiomyocytes and Cardiac Fibroblasts on Myocardium

    Science.gov (United States)

    Zorlutuna, Pinar; Nguyen, Trung Dung; Nagarajan, Neerajha

    Cardiomyocytes show excitatory responses to stimulation solely by mechanical forces through their stretch-activated ion channels, and can fire action potentials upon mechanical stimulation through a pathway known as mechano-electric feedback. Furthermore, cardiomyocyte (CM) - cardiac fibroblasts (CF) can couple mechanically through cell-cell junctions. Here we investigated the effects of CM and CF mechanical coupling on myocardial physiology and pathology using a bio-nanoindentered coupled with fast calcium imaging and microelectrode arrays. In order to study mechanical signal transmission, we measured the contractile forces generated by CMs, as well as by CFs that were coupled to the CMs. We observed that CFs were beating with the same frequency but at smaller magnitude compared to CMs, and their contractility was dependent on the substrate stiffness. Our results showed that beating CMs actively stretched neighbouring CFs through the deformation of the substrate the cells were seeded on, which promoted the myocardial contractility through mechanical coupling. The results also revealed that CM contractility was propagated greater on soft substrates than stiff ones. Results of this study could help identify the role of the infarcted tissue stiffness and size on heart failure. This study is supported by NSF Grant No: 1530884.

  9. Bauhinia championii Flavone Attenuates Hypoxia-Reoxygenation Induced Apoptosis in H9c2 Cardiomyocytes by Improving Mitochondrial Dysfunction.

    Science.gov (United States)

    Liao, Ping; Sun, Guibo; Zhang, Chan; Wang, Min; Sun, Yao; Zhou, Yuehan; Sun, Xiaobo; Jian, Jie

    2016-11-04

    This study aimed to determine the effects of Bauhinia championii flavone (BCF) on hypoxia-reoxygenation (H/R) induced apoptosis in H9c2 cardiomyocytes and to explore potential mechanisms. The H/R model in H9c2 cardiomyocytes was established by 6 h of hypoxia and 12 h of reoxygenation. Cell viability was detected by CCK-8 assay. Apoptotic rate was measured by Annexin V/PI staining. Levels of mitochondria-associated ROS, mitochondrial transmembrane potential (∆Ψm) and mitochondrial permeability transition pores (MPTP) opening were assessed by fluorescent probes. ATP production was measured by ATP assay kit. The release of cytochrome c, translocation of Bax, and related proteins were measured by western blotting. Our results showed that pretreatment with BCF significantly improved cell viability and attenuated the cardiomyocyte apoptosis caused by H/R. Furthermore, BCF increased ATP production and inhibited ROS-generating mitochondria, depolarization of ΔΨm, and MPTP opening. Moreover, BCF pretreatment decreased Bax mitochondrial translocation, cytochrome c release, and activation of caspase-3, as well as increased the expression of p-PI3K, p-Akt, and the ratio of Bcl-2 to Bax. Interestingly, a specific inhibitor of phosphatidylinositol 3-kinase, LY294002, partly reversed the anti-apoptotic effect of BCF. These observations indicated that BCF pretreatment attenuates H/R-induced myocardial apoptosis strength by improving mitochondrial dysfunction via PI3K/Akt signaling pathway.

  10. Altered calcium handling and increased contraction force in human embryonic stem cell derived cardiomyocytes following short term dexamethasone exposure

    Energy Technology Data Exchange (ETDEWEB)

    Kosmidis, Georgios; Bellin, Milena; Ribeiro, Marcelo C.; Meer, Berend van; Ward-van Oostwaard, Dorien [Department of Anatomy and Embryology, Leiden University Medical Center, Leiden (Netherlands); Passier, Robert [Department of Anatomy and Embryology, Leiden University Medical Center, Leiden (Netherlands); MIRA, University of Twente (Netherlands); Tertoolen, Leon G.J.; Mummery, Christine L. [Department of Anatomy and Embryology, Leiden University Medical Center, Leiden (Netherlands); Casini, Simona, E-mail: s.casini@amc.uva.nl [Department of Anatomy and Embryology, Leiden University Medical Center, Leiden (Netherlands)

    2015-11-27

    One limitation in using human pluripotent stem cell derived cardiomyocytes (hPSC-CMs) for disease modeling and cardiac safety pharmacology is their immature functional phenotype compared with adult cardiomyocytes. Here, we report that treatment of human embryonic stem cell derived cardiomyocytes (hESC-CMs) with dexamethasone, a synthetic glucocorticoid, activated glucocorticoid signaling which in turn improved their calcium handling properties and contractility. L-type calcium current and action potential properties were not affected by dexamethasone but significantly faster calcium decay, increased forces of contraction and sarcomeric lengths, were observed in hESC-CMs after dexamethasone exposure. Activating the glucocorticoid pathway can thus contribute to mediating hPSC-CMs maturation. - Highlights: • Dexamethasone accelerates Ca{sup 2+} transient decay in hESC-CMs. • Dexamethasone enhances SERCA and NCX function in hESC-CMs. • Dexamethasone increases force of contraction and sarcomere length in hESC-CMs. • Dexamethasone does not alter I{sub Ca,L} and action potential characteristics in hESC-CMs.

  11. Altered calcium handling and increased contraction force in human embryonic stem cell derived cardiomyocytes following short term dexamethasone exposure

    International Nuclear Information System (INIS)

    Kosmidis, Georgios; Bellin, Milena; Ribeiro, Marcelo C.; Meer, Berend van; Ward-van Oostwaard, Dorien; Passier, Robert; Tertoolen, Leon G.J.; Mummery, Christine L.; Casini, Simona

    2015-01-01

    One limitation in using human pluripotent stem cell derived cardiomyocytes (hPSC-CMs) for disease modeling and cardiac safety pharmacology is their immature functional phenotype compared with adult cardiomyocytes. Here, we report that treatment of human embryonic stem cell derived cardiomyocytes (hESC-CMs) with dexamethasone, a synthetic glucocorticoid, activated glucocorticoid signaling which in turn improved their calcium handling properties and contractility. L-type calcium current and action potential properties were not affected by dexamethasone but significantly faster calcium decay, increased forces of contraction and sarcomeric lengths, were observed in hESC-CMs after dexamethasone exposure. Activating the glucocorticoid pathway can thus contribute to mediating hPSC-CMs maturation. - Highlights: • Dexamethasone accelerates Ca"2"+ transient decay in hESC-CMs. • Dexamethasone enhances SERCA and NCX function in hESC-CMs. • Dexamethasone increases force of contraction and sarcomere length in hESC-CMs. • Dexamethasone does not alter I_C_a_,_L and action potential characteristics in hESC-CMs.

  12. Cardiomyocyte-specific deletion of the G protein-coupled estrogen receptor (GPER) leads to left ventricular dysfunction and adverse remodeling: A sex-specific gene profiling analysis.

    Science.gov (United States)

    Wang, Hao; Sun, Xuming; Chou, Jeff; Lin, Marina; Ferrario, Carlos M; Zapata-Sudo, Gisele; Groban, Leanne

    2017-08-01

    Activation of G protein-coupled estrogen receptor (GPER) by its agonist, G1, protects the heart from stressors such as pressure-overload, ischemia, a high-salt diet, estrogen loss, and aging, in various male and female animal models. Due to nonspecific effects of G1, the exact functions of cardiac GPER cannot be concluded from studies using systemic G1 administration. Moreover, global knockdown of GPER affects glucose homeostasis, blood pressure, and many other cardiovascular-related systems, thereby confounding interpretation of its direct cardiac actions. We generated a cardiomyocyte-specific GPER knockout (KO) mouse model to specifically investigate the functions of GPER in cardiomyocytes. Compared to wild type mice, cardiomyocyte-specific GPER KO mice exhibited adverse alterations in cardiac structure and impaired systolic and diastolic function, as measured by echocardiography. Gene deletion effects on left ventricular dimensions were more profound in male KO mice compared to female KO mice. Analysis of DNA microarray data from isolated cardiomyocytes of wild type and KO mice revealed sex-based differences in gene expression profiles affecting multiple transcriptional networks. Gene Set Enrichment Analysis (GSEA) revealed that mitochondrial genes are enriched in GPER KO females, whereas inflammatory response genes are enriched in GPER KO males, compared to their wild type counterparts of the same sex. The cardiomyocyte-specific GPER KO mouse model provides us with a powerful tool to study the functions of GPER in cardiomyocytes. The gene expression profiles of the GPER KO mice provide foundational information for further study of the mechanisms underlying sex-specific cardioprotection by GPER. Copyright © 2016 Elsevier B.V. All rights reserved.

  13. The dectin-1/inflammasome pathway is responsible for the induction of protective T-helper 17 responses that discriminate between yeasts and hyphae of Candida albicans.

    Science.gov (United States)

    Cheng, Shih-Chin; van de Veerdonk, Frank L; Lenardon, Megan; Stoffels, Monique; Plantinga, Theo; Smeekens, Sanne; Rizzetto, Lisa; Mukaremera, Liliane; Preechasuth, Kanya; Cavalieri, Duccio; Kanneganti, Thirumala Devi; van der Meer, Jos W M; Kullberg, Bart Jan; Joosten, Leo A B; Gow, Neil A R; Netea, Mihai G

    2011-08-01

    In the mucosa, the immune pathways discriminating between colonizing and invasive Candida, thus inducing tolerance or inflammation, are poorly understood. Th17 responses induced by Candida albicans hyphae are central for the activation of mucosal antifungal immunity. An essential step for the discrimination between yeasts and hyphae and induction of Th17 responses is the activation of the inflammasome by C. albicans hyphae and the subsequent release of active IL-1β in macrophages. Inflammasome activation in macrophages results from differences in cell-wall architecture between yeasts and hyphae and is partly mediated by the dectin-1/Syk pathway. These results define the dectin-1/inflammasome pathway as the mechanism that enables the host immune system to mount a protective Th17 response and distinguish between colonization and tissue invasion by C. albicans.

  14. 1,4 Naphthoquinone protects radiation induced cell death and DNA damage in lymphocytes by activation Nrf2/are pathway and enhancing DNA repair

    Energy Technology Data Exchange (ETDEWEB)

    Khan, Nazir M; Sandur, Santosh K; Checker, Rahul; Sharma, Deepak; Poduval, T.B., E-mail: nazirbiotech@rediffmail.com [Radiation Biology and Health Sciences Division, Bhabha Atomic Research Centre, Mumbai (India)

    2012-07-01

    1,4-Naphthoquinone (NQ) is the parent molecule of many clinically approved anticancer, anti-infective, and antiparasitic drugs such as anthracycline, mitomycin, daunorubicin, doxorubicin, diospyrin, and malarone. Presence of NQ during a-irradiation (4Gy) significantly reduced the death of irradiated murine splenic lymphocytes in a dose dependent manner (0.05-liM), with complete protection at liM as assessed by PI staining. Radioprotection by NQ was further confirmed by inhibition of caspase activation, decrease in cell size, DNA-fragmentation, nuclear-blebbing and clonogenic assay. All trans retinoic acid which is inhibitor of Nrf-2 pathway, completely abrogated the radioprotective effect of NQ, suggesting that radioprotective activity of NQ may be due to activation of Nrf-2 signaling pathways. Further, addition of NQ to lymphocytes activated Nrf-2 in time dependent manner as shown by confocal microscopy, electrophoretic mobility shift assay and quantitative real time PCR. It also increased the expression of Nrf-2 dependent cytoprotective genes like hemeoxygenase-1, MnSOD, catalse as demonstrated by real time PCR and flowcytometry. NQ protected lymphocytes significantly against radiation-induced cell death even when added after irradiation. Complete protection was observed by addition of NQ up to 2 h after irradiation. However, percentage protection decreased with increasing time interval. These results suggested that NQ may offer protection to lymphocytes activating repair pathways. Repair of radiation induced DNA strand breaks was studied by comet assay. Pretreatment of lymphocytes with NQ induced single strand breaks up to 6h but not double strand breaks in DNA. However, NQ mediated single strand breaks were repaired completely at longer time intervals. Addition of NQ to lymphocytes prior to 4 Gy a-radiation exposure showed decrease in the yield of DNA double strand breaks. The observed time-dependent decrease in the DNA strand breaks could be attributed to

  15. Structural phenotyping of stem cell-derived cardiomyocytes.

    Science.gov (United States)

    Pasqualini, Francesco Silvio; Sheehy, Sean Paul; Agarwal, Ashutosh; Aratyn-Schaus, Yvonne; Parker, Kevin Kit

    2015-03-10

    Structural phenotyping based on classical image feature detection has been adopted to elucidate the molecular mechanisms behind genetically or pharmacologically induced changes in cell morphology. Here, we developed a set of 11 metrics to capture the increasing sarcomere organization that occurs intracellularly during striated muscle cell development. To test our metrics, we analyzed the localization of the contractile protein α-actinin in a variety of primary and stem-cell derived cardiomyocytes. Further, we combined these metrics with data mining algorithms to unbiasedly score the phenotypic maturity of human-induced pluripotent stem cell-derived cardiomyocytes. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

  16. Anti-apoptosis effect of polysaccharide isolated from the seeds of Cuscuta chinensis Lam on cardiomyocytes in aging rats.

    Science.gov (United States)

    Sun, Shou-Li; Guo, Li; Ren, Ya-Chao; Wang, Bing; Li, Rong-Hui; Qi, Yu-Shan; Yu, Hui; Chang, Nai-Dan; Li, Ming-Hui; Peng, Hai-Sheng

    2014-09-01

    To investigate the mechanism of apoptosis in myocardial cells of aging rats induced by D-galactose and to study the effect of the Polysaccharide isolated from the seeds of Cuscuta chinensis Lam (PCCL) on apoptosis of cardiomyocytes and its corresponding machinasim in aging rat model. Fifty male SD rats were randomly divided into 5 groups. Normal control group (NC). D-galactose (100 mg · kg(-1)d(-1) for 56 day) indued aging group (MC), D-galactose plus 100 mg kg(-1) d(-1) PCCL group (ML), D-galactose plus 200 mg kg(-1) d(-1) PCCL group (MM), and D-galactose plus 400 mg kg(-1) d(-1) PCCL group (MH). Same volume of solution (water, or PCCL aqueous solution) was given by gavage for 56 days. Then the hearts were collected and apoptosis parameters were evaluated. Caspase-3 and Cyt c were determined by fluorescence spectrometer, the apoptosis rate was assessed by AnnexinV-FITC method by Flow-Cytometry, [Ca(2+)]i and [Ca(2+)]i overloaded by KCL were observed by laser scanning confocal microscopy (LSCM); Bcl-2 and Bax were examined by immunohistochemistry. The content of Cyt C, [Ca(2+)]i of cardiomyocytes, the activity of Caspase-3, Bax expression level in D-galactose induced aging group were higher than NC (p < 0.05). The ratio of Bcl-2/Bax was decreased in D-galactose induced aging group compared to NC. On the other hand, the content of Cyt C, [Ca(2+)]i of cardiomyocytes, the activity of Caspase-3 and apoptosis rate, as well as Bax expression level in all three PCCL groups were decreased compared to galactose induced group (p < 0.05). Bcl-2/Bax ratio was increased in all PCCL groups compared to galactose induced aging group. PCCL could decrease the apoptosis of cardiomyocytes by the mitochondria apoptosis pathway.

  17. MicroRNA-145 suppresses ROS-induced Ca2+ overload of cardiomyocytes by targeting CaMKIIδ

    International Nuclear Information System (INIS)

    Cha, Min-Ji; Jang, Jin-Kyung; Ham, Onju; Song, Byeong-Wook; Lee, Se-Yeon; Lee, Chang Yeon; Park, Jun-Hee; Lee, Jiyun; Seo, Hyang-Hee; Choi, Eunhyun; Jeon, Woo-min; Hwang, Hye Jin; Shin, Hyun-Taek

    2013-01-01

    Highlights: •CaMKIIδ mediates H 2 O 2 -induced Ca 2+ overload in cardiomyocytes. •miR-145 can inhibit Ca 2+ overload. •A luciferase assay confirms that miR-145 functions as a CaMKIIδ-targeting miRNA. •Overexpression of miR-145 regulates CaMKIIδ-related genes and ameliorates apoptosis. -- Abstract: A change in intracellular free calcium (Ca 2+ ) is a common signaling mechanism of reperfusion-induced cardiomyocyte death. Calcium/calmodulin dependent protein kinase II (CaMKII) is a critical regulator of Ca 2+ signaling and mediates signaling pathways responsible for functions in the heart including hypertrophy, apoptosis, arrhythmia, and heart disease. MicroRNAs (miRNA) are involved in the regulation of cell response, including survival, proliferation, apoptosis, and development. However, the roles of miRNAs in Ca 2+ -mediated apoptosis of cardiomyocytes are uncertain. Here, we determined the potential role of miRNA in the regulation of CaMKII dependent apoptosis and explored its underlying mechanism. To determine the potential roles of miRNAs in H 2 O 2 -mediated Ca 2+ overload, we selected and tested 6 putative miRNAs that targeted CaMKIIδ, and showed that miR-145 represses CaMKIIδ protein expression and Ca 2+ overload. We confirmed CaMKIIδ as a direct downstream target of miR-145. Furthermore, miR-145 regulates Ca 2+ -related signals and ameliorates apoptosis. This study demonstrates that miR-145 regulates reactive oxygen species (ROS)-induced Ca 2+ overload in cardiomyocytes. Thus, miR-145 affects ROS-mediated gene regulation and cellular injury responses

  18. MicroRNA-145 suppresses ROS-induced Ca{sup 2+} overload of cardiomyocytes by targeting CaMKIIδ

    Energy Technology Data Exchange (ETDEWEB)

    Cha, Min-Ji [Cardiovascular Research Institute, Yonsei University College of Medicine, 250 Seongsanno, Seodamun-gu, Seoul 120-752 (Korea, Republic of); Brain Korea 21 Project for Medical Science, Yonsei University College of Medicine, 250 Seongsanno, Seodamun-gu, Seoul 120-752 (Korea, Republic of); Jang, Jin-Kyung [College of Pharmacy, Sookmyung Women’s University, 52 HyoChangWon-Gil, Yongsan-ku, Seoul 140-742 (Korea, Republic of); Ham, Onju; Song, Byeong-Wook; Lee, Se-Yeon [Cardiovascular Research Institute, Yonsei University College of Medicine, 250 Seongsanno, Seodamun-gu, Seoul 120-752 (Korea, Republic of); Brain Korea 21 Project for Medical Science, Yonsei University College of Medicine, 250 Seongsanno, Seodamun-gu, Seoul 120-752 (Korea, Republic of); Lee, Chang Yeon; Park, Jun-Hee [Department of Integrated Omics for Biomedical Sciences, Graduate School, Yonsei University, 50 Yonsei-ro, Seodamun-gu, Seoul 120-759 (Korea, Republic of); Lee, Jiyun; Seo, Hyang-Hee [Cardiovascular Research Institute, Yonsei University College of Medicine, 250 Seongsanno, Seodamun-gu, Seoul 120-752 (Korea, Republic of); Brain Korea 21 Project for Medical Science, Yonsei University College of Medicine, 250 Seongsanno, Seodamun-gu, Seoul 120-752 (Korea, Republic of); Choi, Eunhyun [Severance Integrative Research Institute for Cerebral and Cardiovascular Disease, Yonsei University Health System, 250 Seongsanno, Seodamun-gu, Seoul 120-752 (Korea, Republic of); Jeon, Woo-min [Department of Animal Resource, Sahmyook University, Seoul 139-742 (Korea, Republic of); Hwang, Hye Jin [Cardiovascular Research Institute, Yonsei University College of Medicine, 250 Seongsanno, Seodamun-gu, Seoul 120-752 (Korea, Republic of); Shin, Hyun-Taek [College of Pharmacy, Sookmyung Women’s University, 52 HyoChangWon-Gil, Yongsan-ku, Seoul 140-742 (Korea, Republic of); and others

    2013-06-14

    Highlights: •CaMKIIδ mediates H{sub 2}O{sub 2}-induced Ca{sup 2+} overload in cardiomyocytes. •miR-145 can inhibit Ca{sup 2+} overload. •A luciferase assay confirms that miR-145 functions as a CaMKIIδ-targeting miRNA. •Overexpression of miR-145 regulates CaMKIIδ-related genes and ameliorates apoptosis. -- Abstract: A change in intracellular free calcium (Ca{sup 2+}) is a common signaling mechanism of reperfusion-induced cardiomyocyte death. Calcium/calmodulin dependent protein kinase II (CaMKII) is a critical regulator of Ca{sup 2+} signaling and mediates signaling pathways responsible for functions in the heart including hypertrophy, apoptosis, arrhythmia, and heart disease. MicroRNAs (miRNA) are involved in the regulation of cell response, including survival, proliferation, apoptosis, and development. However, the roles of miRNAs in Ca{sup 2+}-mediated apoptosis of cardiomyocytes are uncertain. Here, we determined the potential role of miRNA in the regulation of CaMKII dependent apoptosis and explored its underlying mechanism. To determine the potential roles of miRNAs in H{sub 2}O{sub 2}-mediated Ca{sup 2+} overload, we selected and tested 6 putative miRNAs that targeted CaMKIIδ, and showed that miR-145 represses CaMKIIδ protein expression and Ca{sup 2+} overload. We confirmed CaMKIIδ as a direct downstream target of miR-145. Furthermore, miR-145 regulates Ca{sup 2+}-related signals and ameliorates apoptosis. This study demonstrates that miR-145 regulates reactive oxygen species (ROS)-induced Ca{sup 2+} overload in cardiomyocytes. Thus, miR-145 affects ROS-mediated gene regulation and cellular injury responses.

  19. Edaravone protects rats and human pulmonary alveolar epithelial cells against hyperoxia injury: heme oxygenase-1 and PI3K/Akt pathway may be involved.

    Science.gov (United States)

    Cao, Huifang; Feng, Ying; Ning, Yunye; Zhang, Zinan; Li, Weihao; Li, Qiang

    2015-01-01

    Hyperoxic acute lung injury (HALI) is a clinical syndrome as a result of prolonged supplement of high concentrations of oxygen. As yet, no specific treatment is available for HALI. The present study aims to investigate the effects of edaravone on hyperoxia-induced oxidative injury and the underlying mechanism. We treated rats and human pulmonary alveolar epithelial cells with hyperoxia and different concentration of edaravone, then examined the effects of edaravone on cell viability, cell injury and two oxidative products. The roles of heme oxygenase-1 (HO-1) and PI3K/Akt pathway were explored using Western blot and corresponding inhibitors. The results showed that edaravone reduced lung biochemical alterations induced by hyperoxia and mortality of rats, dose-dependently alleviated cell mortality, cell injury, and peroxidation of cellular lipid and DNA oxidative damage. It upregulated cellular HO-1 expression and activity, which was reversed by PI3K/Akt pathway inhibition. The administration of zinc protoporphyrin-IX, a HO-1 inhibitor, and LY249002, a PI3K/Akt pathway inhibitor, abolished the protective effects of edaravone in cells. This study indicates that edaravone protects rats and human pulmonary alveolar epithelial cells against hyperoxia-induced injury and the antioxidant effect may be related to upregulation of HO-1, which is regulated by PI3K/Akt pathway.

  20. Schisandra sphenanthera extract (Wuzhi Tablet protects against chronic-binge and acute alcohol-induced liver injury by regulating the NRF2-ARE pathway in mice

    Directory of Open Access Journals (Sweden)

    Xuezhen Zeng

    2017-09-01

    Full Text Available Alcohol abuse leads to alcoholic liver disease and no effective therapy is currently available. Wuzhi Tablet (WZ, a preparation of extract from Schisandra sphenanthera that is a traditional hepato-protective herb, exerted a significant protective effect against acetaminophen-induced liver injury in our recent studies, but whether WZ can alleviate alcohol-induced toxicity remains unclear. This study aimed to investigate the contribution of WZ to alcohol-induced liver injury by using chronic-binge and acute models of alcohol feeding. The activities of ALT and AST in serum were assessed as well as the level of GSH and the activity of SOD in the liver. The expression of CYP2E1 and proteins in the NRF2-ARE signaling pathway including NRF2, GCLC, GCLM, HO-1 were measured, and the effect of WZ on NRF2 transcriptional activity was determined. We found that both models resulted in liver steatosis accompanied by increased transaminase activities, but that liver injury was significantly attenuated by WZ. WZ administration also inhibited CYP2E1 expression induced by alcohol, and elevated the level of GSH and the activity of SOD in the liver. Moreover, the NRF2-ARE signaling pathway was activated by WZ and the target genes were all upregulated. Furthermore, WZ significantly activated NRF2 transcriptional activity. Collectively, our study demonstrates that WZ protected against alcohol-induced liver injury by reducing oxidative stress and improving antioxidant defense, possibly by activating the NRF2-ARE pathway.

  1. PI3K/Akt Pathway Contributes to Neurovascular Unit Protection of Xiao-Xu-Ming Decoction against Focal Cerebral Ischemia and Reperfusion Injury in Rats

    Directory of Open Access Journals (Sweden)

    Rui Lan

    2013-01-01

    Full Text Available In the present study, we used a focal cerebral ischemia and reperfusion rat model to investigate the protective effects of Xiao-Xu-Ming decoction (XXMD on neurovascular unit and to examine the role of PI3K (phosphatidylinositol 3-kinase/Akt pathway in this protection. The cerebral ischemia was induced by 90 min of middle cerebral artery occlusion. Cerebral infarct area was measured by tetrazolium staining, and neurological function was observed at 24 h after reperfusion. DNA fragmentation assay, combined with immunofluorescence, was performed to evaluate apoptosis of neuron, astrocyte, and vascular endothelial cell which constitute neurovascular unit. The expression levels of proteins involved in PI3K/Akt pathway were detected by Western blot. The results showed that XXMD improved neurological function, decreased cerebral infarct area and neuronal damage, and attenuated cellular apoptosis in neurovascular unit, while these effects were abolished by inhibition of PI3K/Akt with LY294002. We also found that XXMD upregulated p-PDKl, p-Akt, and p-GSK3β expression levels, which were partly reversed by LY294002. In addition, the increases of p-PTEN and p-c-Raf expression levels on which LY294002 had no effect were also observed in response to XXMD treatment. The data indicated the protective effects of XXMD on neurovascular unit partly through the activation of PI3K/Akt pathway.

  2. The Bone Marrow-Mediated Protection of Myeloproliferative Neoplastic Cells to Vorinostat and Ruxolitinib Relies on the Activation of JNK and PI3K Signalling Pathways.

    Directory of Open Access Journals (Sweden)

    Bruno A Cardoso

    Full Text Available The classical BCR-ABL-negative Myeloproliferative Neoplasms (MPN are a group of heterogeneous haematological diseases characterized by constitutive JAK-STAT pathway activation. Targeted therapy with Ruxolitinib, a JAK1/2-specific inhibitor, achieves symptomatic improvement but does not eliminate the neoplastic clone. Similar effects are seen with histone deacetylase inhibitors (HDACi, albeit with poorer tolerance. Here, we show that bone marrow (BM stromal cells (HS-5 protected MPN-derived cell lines (SET-2; HEL and UKE-1 and MPN patient-derived BM cells from the cytotoxic effects of Ruxolitinib and the HDACi Vorinostat. This protective effect was mediated, at least in part, by the secretion of soluble factors from the BM stroma. In addition, it correlated with the activation of signalling pathways important for cellular homeostasis, such as JAK-STAT, PI3K, JNK, MEK-ERK and NF-κB. Importantly, the pharmacological inhibition of JNK and PI3K pathways completely abrogated the BM protective effect on MPN cell lines and MPN patient samples. Our findings shed light on mechanisms of tumour survival and may indicate novel therapeutic approaches for the treatment of MPN.

  3. The Bone Marrow-Mediated Protection of Myeloproliferative Neoplastic Cells to Vorinostat and Ruxolitinib Relies on the Activation of JNK and PI3K Signalling Pathways

    Science.gov (United States)

    Cardoso, Bruno A.; Belo, Hélio; Barata, João T.; Almeida, António M.

    2015-01-01

    The classical BCR-ABL-negative Myeloproliferative Neoplasms (MPN) are a group of heterogeneous haematological diseases characterized by constitutive JAK-STAT pathway activation. Targeted therapy with Ruxolitinib, a JAK1/2-specific inhibitor, achieves symptomatic improvement but does not eliminate the neoplastic clone. Similar effects are seen with histone deacetylase inhibitors (HDACi), albeit with poorer tolerance. Here, we show that bone marrow (BM) stromal cells (HS-5) protected MPN-derived cell lines (SET-2; HEL and UKE-1) and MPN patient-derived BM cells from the cytotoxic effects of Ruxolitinib and the HDACi Vorinostat. This protective effect was mediated, at least in part, by the secretion of soluble factors from the BM stroma. In addition, it correlated with the activation of signalling pathways important for cellular homeostasis, such as JAK-STAT, PI3K, JNK, MEK-ERK and NF-κB. Importantly, the pharmacological inhibition of JNK and PI3K pathways completely abrogated the BM protective effect on MPN cell lines and MPN patient samples. Our findings shed light on mechanisms of tumour survival and may indicate novel therapeutic approaches for the treatment of MPN. PMID:26623653

  4. Apelin and APJ orchestrate complex tissue-specific control of cardiomyocyte hypertrophy and contractility in the hypertrophy-heart failure transition.

    Science.gov (United States)

    Parikh, Victoria Nicole; Liu, Jing; Shang, Ching; Woods, Christopher; Chang, Alex Chia Yu; Zhao, Mingming; Charo, David N; Grunwald, Zachary; Huang, Yong; Seo, Kinya; Tsao, Philip S; Bernstein, Daniel; Ruiz-Lozano, Pilar; Quertermous, Thomas; Ashley, Euan A

    2018-05-18

    The G protein coupled receptor APJ is a promising therapeutic target for heart failure. Constitutive deletion of APJ in the mouse is protective against the hypertrophy-heart failure transition via elimination of ligand-independent, β-arrestin dependent stretch transduction. However, the cellular origin of this stretch transduction and the details of its interaction with apelin signaling remain unknown. We generated mice with conditional elimination of APJ in the endothelium (APJ endo-/- ) and myocardium (APJ myo-/- ). No baseline difference was observed in LV function in APJ endo-/- , APJ myo-/- or controls (APJ endo+/+ , APJ myo+/+ ). After exposure to transaortic constriction (TAC), APJ endo-/- animals developed left ventricular failure while APJ myo-/- were protected. At the cellular level, carbon fiber stretch of freshly isolated single cardiomyocytes demonstrated decreased contractile response to stretch in APJ -/- cardiomyocytes compared to APJ +/+ cardiomyocytes. Calcium transient did not change with stretch in either APJ -/- or APJ +/+ cardiomyocytes. Application of apelin to APJ +/+ cardiomyocytes resulted in decreased calcium transient. Further, hearts of mice treated with apelin exhibited decreased phosphorylation at Troponin I (cTnI) N-terminal residues (Ser 22,23), consistent with increased calcium sensitivity. These data establish that APJ stretch transduction is mediated specifically by myocardial APJ, that APJ is necessary for stretch-induced increases in contractility, and that apelin opposes APJ's stretch-mediated hypertrophy signaling by lowering calcium transient while maintaining contractility through myofilament calcium sensitization. These findings underscore apelin's unique potential as a therapeutic agent that can simultaneously support cardiac function and protect against the hypertrophy-heart failure transition.

  5. Hydrogen sulfide postconditioning protects isolated rat hearts against ischemia and reperfusion injury mediated by the JAK2/STAT3 survival pathway

    Directory of Open Access Journals (Sweden)

    Heng-Fei Luan

    2012-10-01

    Full Text Available The JAK2/STAT3 signal pathway is an important component of survivor activating factor enhancement (SAFE pathway. The objective of the present study was to determine whether the JAK2/STAT3 signaling pathway participates in hydrogen sulfide (H2S postconditioning, protecting isolated rat hearts from ischemic-reperfusion injury. Male Sprague-Dawley rats (230-270 g were divided into 6 groups (N = 14 per group: time-matched perfusion (Sham group, ischemia/reperfusion (I/R group, NaHS postconditioning group, NaHS with AG-490 group, AG-490 (5 µM group, and dimethyl sulfoxide (DMSO; <0.2% group. Langendorff-perfused rat hearts, with the exception of the Sham group, were subjected to 30 min of ischemia followed by 90 min of reperfusion after 20 min of equilibrium. Heart rate, left ventricular developed pressure (LVDP, left ventricular end-diastolic pressure (LVEDP, and the maximum rate of increase or decrease of left ventricular pressure (± dp/dt max were recorded. Infarct size was determined using triphenyltetrazolium chloride (TTC staining. Myocardial TUNEL staining was used as the in situ cell death detection method and the percentage of TUNEL-positive nuclei to all nuclei counted was used as the apoptotic index. The expression of STAT3, bcl-2 and bax was determined by Western blotting. After reperfusion, compared to the I/R group, H2S significantly improved functional recovery and decreased infarct size (23.3 ± 3.8 vs 41.2 ± 4.7%, P < 0.05 and apoptotic index (22.1 ± 3.6 vs 43.0 ± 4.8%, P < 0.05. However, H2S-mediated protection was abolished by AG-490, the JAK2 inhibitor. In conclusion, H2S postconditioning effectively protects isolated I/R rat hearts via activation of the JAK2/STAT3 signaling pathway.

  6. Intermittent Hypoxia Inhibits Na+-H+ Exchange-Mediated Acid Extrusion Via Intracellular Na+ Accumulation in Cardiomyocytes

    Directory of Open Access Journals (Sweden)

    Huai-Ren Chang

    2018-04-01

    Full Text Available Background/Aims: Intermittent hypoxia (IH has been shown to exert preconditioning-like cardioprotective effects. It also has been reported that IH preserves intracellular pH (pHi during ischemia and protects cardiomyocytes against ischemic reperfusion injury. However, the exact mechanism is still unclear. Methods: In this study, we used proton indicator BCECF-AM to analyze the rate of pHi recovery from acidosis in the IH model of rat neonatal cardiomyocytes. Neonatal cardiomyocytes were first treated with repetitive hypoxia-normoxia cycles for 1-4 days. Cells were then acid loaded with NH4Cl, and the rate of pHi recovery from acidosis was measured. Results: We found that the pHi recovery rate from acidosis was much slower in the IH group than in the room air (RA group. When we treated cardiomyocytes with Na+-H+ exchange (NHE inhibitors (Amiloride and HOE642 or Na+-free Tyrode solution during the recovery, there was no difference between RA and IH groups. We also found intracellular Na+ concentration ([Na+]i significantly increased after IH exposure for 4 days. However, the phenomenon could be abolished by pretreatment with ROS inhibitors (SOD and phenanathroline, intracellular calcium chelator or Na+-Ca2+ exchange (NCX inhibitor. Furthermore, the pHi recovery rate from acidosis became faster in the IH group than in the RA group when inhibition of NCX activity. Conclusions: These results suggest that IH would induce the elevation of ROS production. ROS then activates Ca2+-efflux mode of NCX and results in intracellular Na+ accumulation. The rise of [Na+]i further inhibits the activity of NHE-mediated acid extrusion and retards the rate of pHi recovery from acidosis during IH.

  7. Excitation model of pacemaker cardiomyocytes of cardiac conduction system

    Science.gov (United States)

    Grigoriev, M.; Babich, L.

    2015-11-01

    Myocardium includes typical and atypical cardiomyocytes - pacemakers, which form the cardiac conduction system. Excitation from the atrioventricular node in normal conditions is possible only in one direction. Retrograde direction of pulses is impossible. The most important prerequisite for the work of cardiomyocytes is the anatomical integrity of the conduction system. Changes in contractile force of the cardiomyocytes, which appear periodically, are due to two mechanisms of self-regulation - heterometric and homeometric. Graphic course of the excitation pulse propagation along the heart muscle more accurately reveals the understanding of the arrhythmia mechanism. These models have the ability to visualize the essence of excitation dynamics. However, they do not have the proper forecasting function for result estimation. Integrative mathematical model enables further investigation of general laws of the myocardium active behavior, allows for determination of the violation mechanism of electrical and contractile function of cardiomyocytes. Currently, there is no full understanding of the topography of pacemakers and ionic mechanisms. There is a need for the development of direction of mathematical modeling and comparative studies of the electrophysiological arrangement of cells of atrioventricular connection and ventricular conduction system.

  8. AKIP1 expression modulates mitochondrial function in rat neonatal cardiomyocytes

    NARCIS (Netherlands)

    Yu, Hongjuan; Tigchelaar, Wardit; Koonen, Debby P. Y.; Patel, Hemal H.; de Boer, Rudolf A.; van Gilst, Wiek H.; Westenbrink, B. Daan; Sillje, Herman H. W.

    2013-01-01

    A kinase interacting protein 1 (AKIP1) is a molecular regulator of protein kinase A and nuclear factor kappa B signalling. Recent evidence suggests AKIP1 is increased in response to cardiac stress, modulates acute ischemic stress response, and is localized to mitochondria in cardiomyocytes. The

  9. Common marmoset embryonic stem cell can differentiate into cardiomyocytes

    International Nuclear Information System (INIS)

    Chen Hao; Hattori, Fumiyuki; Murata, Mitsushige; Li Weizhen; Yuasa, Shinsuke; Onizuka, Takeshi; Shimoji, Kenichiro; Ohno, Yohei; Sasaki, Erika; Kimura, Kensuke; Hakuno, Daihiko

    2008-01-01

    Common marmoset monkeys have recently attracted much attention as a primate research model, and are preferred to rhesus and cynomolgus monkeys due to their small bodies, easy handling and efficient breeding. We recently reported the establishment of common marmoset embryonic stem cell (CMESC) lines that could differentiate into three germ layers. Here, we report that our CMESC can also differentiate into cardiomyocytes and investigated their characteristics. After induction, FOG-2 was expressed, followed by GATA4 and Tbx20, then Nkx2.5 and Tbx5. Spontaneous beating could be detected at days 12-15. Immunofluorescent staining and ultrastructural analyses revealed that they possessed characteristics typical of functional cardiomyocytes. They showed sinus node-like action potentials, and the beating rate was augmented by isoproterenol stimulation. The BrdU incorporation assay revealed that CMESC-derived cardiomyocytes retained a high proliferative potential for up to 24 weeks. We believe that CMESC-derived cardiomyocytes will advance preclinical studies in cardiovascular regenerative medicine

  10. Reverse engineering life: physical and chemical mimetics for controlled stem cell differentiation into cardiomyocytes.

    Science.gov (United States)

    Skuse, Gary R; Lamkin-Kennard, Kathleen A

    2013-01-01

    Our ability to manipulate stem cells in order to induce differentiation along a desired developmental pathway has improved immeasurably in recent years. That is in part because we have a better understanding of the intracellular and extracellular signals that regulate differentiation. However, there has also been a realization that stem cell differentiation is not regulated only by chemical signals but also by the physical milieu in which a particular stem cell exists. In this regard we are challenged to mimic both chemical and physical environments. Herein we describe a method to induce stem cell differentiation into cardiomyocytes using a combination of chemical and physical cues. This method can be applied to produce differentiated cells for research and potentially for cell-based therapy of cardiomyopathies.

  11. The primary cilium coordinates early cardiogenesis and hedgehog signaling in cardiomyocyte differentiation

    DEFF Research Database (Denmark)

    Clement, Christian A; Kristensen, Stine G; Møllgård, Kjeld

    2009-01-01

    Defects in the assembly or function of primary cilia, which are sensory organelles, are tightly coupled to developmental defects and diseases in mammals. Here, we investigated the function of the primary cilium in regulating hedgehog signaling and early cardiogenesis. We report that the pluripotent...... P19.CL6 mouse stem cell line, which can differentiate into beating cardiomyocytes, forms primary cilia that contain essential components of the hedgehog pathway, including Smoothened, Patched-1 and Gli2. Knockdown of the primary cilium by Ift88 and Ift20 siRNA or treatment with cyclopamine...... development. These data support the conclusion that cardiac primary cilia are crucial in early heart development, where they partly coordinate hedgehog signaling....

  12. 1,5-Disubstituted benzimidazoles that direct cardiomyocyte differentiation from mouse embryonic stem cells.

    Science.gov (United States)

    Okolotowicz, Karl J; Bushway, Paul; Lanier, Marion; Gilley, Cynthia; Mercola, Mark; Cashman, John R

    2015-09-01

    Cardiomyopathy is the leading cause of death worldwide. Despite progress in medical treatments, heart transplantation is one of the only current options for those with infarcted heart muscle. Stem cell differentiation technology may afford cell-based therapeutics that may lead to the generation of new, healthy heart muscle cells from undifferentiated stem cells. Our approach is to use small molecules to stimulate stem cell differentiation. Herein, we describe a novel class of 1,5-disubstituted benzimidazoles that induce differentiation of stem cells into cardiac cells. We report on the evaluation in vitro for cardiomyocyte differentiation and describe structure-activity relationship results that led to molecules with drug-like properties. The results of this study show the promise of small molecules to direct stem cell lineage commitment, to probe signaling pathways and to develop compounds for the stimulation of stem cells to repair damaged heart tissue. Copyright © 2015 Elsevier Ltd. All rights reserved.

  13. IGF-1 protects SH-SY5Y cells against MPP+-induced apoptosis via PI3K/PDK-1/Akt pathway.

    Science.gov (United States)

    Kim, Chanyang; Park, Seungjoon

    2018-03-01

    Insulin-like growth factor (IGF)-1 is a well-known anti-apoptotic pro-survival factor and phosphatidylinositol-3-kinase (PI3K)/Akt pathway is linked to cell survival induced by IGF-1. It is also reported that Akt signaling is modulated by 3-phosphoinositide-dependent kinase-1 (PDK1). In the current study, we investigated whether the anti-apoptotic effect of IGF-1 in SH-SY5Y cells exposed to 1-methyl-4-phenylpyridinium (MPP + ) is associated with the activity of PI3K/PDK1/Akt pathway. Treatment of cells with IGF-1 inhibited MPP + -induced apoptotic cell death. IGF-1-induced activation of Akt and the protective effect of IGF-1 on MPP + -induced apoptosis were abolished by chemical inhibition of PDK1 (GSK2334470) or PI3K (LY294002). The phosphorylated levels of Akt and PDK1 were significantly suppressed after MPP + exposure, while IGF-1 treatment completely restored MPP+-induced reductions in phosphorylation. IGF-1 protected cells from MPP + insult by suppressing intracellular reactive oxygen species (ROS) production and malondialdehyde levels and increasing superoxide dismutase activity. Mitochondrial ROS levels were also increased during MPP + exposure, which were attenuated by IGF-1 treatment. In addition, IGF-1-treated cells showed increased activities of succinate dehydrogenase and citrate synthase, stabilization of mitochondrial transmembrane potential, increased ratio of Bcl-2 to Bax, prevention of cytochrome c release and inhibition of caspase-3 activation with PARP cleavage. Furthermore, the protective effects of IGF-1 on oxidative stress and mitochondrial dysfunction were attenuated when cells were preincubated with GSK2334470 or LY294002. Our data suggest that IGF-1 protects SH-SY5Y cells against MPP + -associated oxidative stress by preserving mitochondrial integrity and inhibiting mitochondrial apoptotic cascades via the activation of PI3K/PDK1/Akt pathway. © 2018 The authors.

  14. Atrial natriuretic peptide regulates Ca channel in early developmental cardiomyocytes.

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    Lin Miao

    Full Text Available BACKGROUND: Cardiomyocytes derived from murine embryonic stem (ES cells possess various membrane currents and signaling cascades link to that of embryonic hearts. The role of atrial natriuretic peptide (ANP in regulation of membrane potentials and Ca(2+ currents has not been investigated in developmental cardiomyocytes. METHODOLOGY/PRINCIPAL FINDINGS: We investigated the role of ANP in regulating L-type Ca(2+ channel current (I(CaL in different developmental stages of cardiomyocytes derived from ES cells. ANP decreased the frequency of action potentials (APs in early developmental stage (EDS cardiomyocytes, embryonic bodies (EB as well as whole embryo hearts. ANP exerted an inhibitory effect on basal I(CaL in about 70% EDS cardiomyocytes tested but only in about 30% late developmental stage (LDS cells. However, after stimulation of I(CaL by isoproterenol (ISO in LDS cells, ANP inhibited the response in about 70% cells. The depression of I(CaL induced by ANP was not affected by either Nomega, Nitro-L-Arginine methyl ester (L-NAME, a nitric oxide synthetase (NOS inhibitor, or KT5823, a cGMP-dependent protein kinase (PKG selective inhibitor, in either EDS and LDS cells; whereas depression of I(CaL by ANP was entirely abolished by erythro-9-(2-Hydroxy-3-nonyl adenine (EHNA, a selective inhibitor of type 2 phosphodiesterase(PDE2 in most cells tested. CONCLUSION/SIGNIFICANCES: Taken together, these results indicate that ANP induced depression of action potentials and I(CaL is due to activation of particulate guanylyl cyclase (GC, cGMP production and cGMP-activation of PDE2 mediated depression of adenosine 3', 5'-cyclic monophophate (cAMP-cAMP-dependent protein kinase (PKA in early cardiomyogenesis.

  15. Serotonin and dopamine protect from hypothermia/rewarming damage through the CBS/H2S pathway

    NARCIS (Netherlands)

    Talaei, Fatemeh; Bouma, Hjalmar; Van der Graaf, Adrianus; Strijkstra, Arjen; Schmidt, Martina; Henning, Robert

    2011-01-01

    Biogenic amines have been demonstrated to protect cells from apoptotic cell death. Herein we show for the first time that serotonin and dopamine increase H2S production by the endogenous enzyme cystathionine-beta-synthase (CBS) and protect cells against hypothermia/rewarming induced reactive oxygen

  16. Protective

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    Wessam M. Abdel-Wahab

    2013-10-01

    Full Text Available Many active ingredients extracted from herbal and medicinal plants are extensively studied for their beneficial effects. Antioxidant activity and free radical scavenging properties of thymoquinone (TQ have been reported. The present study evaluated the possible protective effects of TQ against the toxicity and oxidative stress of sodium fluoride (NaF in the liver of rats. Rats were divided into four groups, the first group served as the control group and was administered distilled water whereas the NaF group received NaF orally at a dose of 10 mg/kg for 4 weeks, TQ group was administered TQ orally at a dose of 10 mg/kg for 5 weeks, and the NaF-TQ group was first given TQ for 1 week and was secondly administered 10 mg/kg/day NaF in association with 10 mg/kg TQ for 4 weeks. Rats intoxicated with NaF showed a significant increase in lipid peroxidation whereas the level of reduced glutathione (GSH and the activity of superoxide dismutase (SOD, catalase (CAT, glutathione S-transferase (GST and glutathione peroxidase (GPx were reduced in hepatic tissues. The proper functioning of the liver was also disrupted as indicated by alterations in the measured liver function indices and biochemical parameters. TQ supplementation counteracted the NaF-induced hepatotoxicity probably due to its strong antioxidant activity. In conclusion, the results obtained clearly indicated the role of oxidative stress in the induction of NaF toxicity and suggested hepatoprotective effects of TQ against the toxicity of fluoride compounds.

  17. Activation of the Nrf2/HO-1 Antioxidant Pathway Contributes to the Protective Effects of Lycium Barbarum Polysaccharides in the Rodent Retina after Ischemia-Reperfusion-Induced Damage

    Science.gov (United States)

    Chang, Raymond Chuen-Chung; So, Kwok-Fai; Brecha, Nicholas C.; Pu, Mingliang

    2014-01-01

    Lycium barbarum polysaccharides (LBP), extracts from the wolfberries, are protective to retina after ischemia-reperfusion (I/R). The antioxidant response element (ARE)–mediated antioxidant pathway plays an important role in maintaining the redox status of the retina. Heme oxygenase-1 (HO-1), combined with potent AREs in its promoter, is a highly effective therapeutic target for the protection against neurodegenerative diseases, including I/R-induced retinal damage. The aim of our present study was to investigate whether the protective effect of LBP after I/R damage was mediated via activation of the Nrf2/HO-1-antioxidant pathway in the retina. Retinal I/R was induced by an increase in intraocular pressure to 130 mm Hg for 60 minutes. Prior to the induction of ischemia, rats were orally treated with either vehicle (PBS) or LBP (1 mg/kg) once a day for 1 week. For specific experiments, zinc protoporphyrin (ZnPP, 20 mg/kg), an HO-1 inhibitor, was intraperitoneally administered at 24 h prior to ischemia. The protective effects of LBP were evaluated by quantifying ganglion cell and amacrine cell survival, and by measuring cell apoptosis in the retinal layers. In addition, HO-1 expression was examined using Western blotting and immunofluorescence analyses. Cytosolic and nuclear Nrf2 was measured using immunofluorescent staining. LBP treatment significantly increased Nrf2 nuclear accumulation and HO-1 expression in the retina after I/R injury. Increased apoptosis and a decrease in the number of viable cells were observed in the ganglion cell layer (GCL) and inner nuclear layer (INL) in the I/R retina, which were reversed by LBP treatment. The HO-1 inhibitor, ZnPP, diminished the LBP treatment-induced protective effects in the retina after I/R. Taken together, these results suggested that LBP partially exerted its beneficial neuroprotective effects via the activation of Nrf2 and an increase in HO-1 protein expression. PMID:24400114

  18. Hydrogen-rich medium protects mouse embryonic fibroblasts from oxidative stress by activating LKB1-AMPK-FoxO1 signal pathway.

    Science.gov (United States)

    Lee, Jihyun; Yang, Goowon; Kim, Young-Joo; Tran, Quynh Hoa; Choe, Wonchae; Kang, Insug; Kim, Sung Soo; Ha, Joohun

    2017-09-23

    Persistent oxidative stress is recognized as a major cause of many pathological conditions as well as ageing. However, most clinical trials of dietary antioxidants have failed to produce successful outcomes in treating oxidative stress-induced diseases. Molecular hydrogen (H 2 ) has recently received considerable attention as a therapeutic agent owing to its novel antioxidant properties, a selective scavenger of hydroxyl and peroxynitrite radicals. Beyond this, numerous reports support that H 2 can modulate the activity of various cellular signal pathways. However, its effect on AMP-activated protein kinase (AMPK) signal pathway, a central regulator of energy hemostasis, has remained almost elusive. Here, we report that hydrogen-rich medium activated LKB1-AMPK signal pathway without ATP depletion, which in turn induced FoxO1-dependent transcription of manganese superoxide dismutase and catalase in mouse embryonic fibroblasts. Moreover, hydrogen-rich media effectively reduced the level of reactive oxygen species in cells treated with hydrogen peroxide and protected these cells from apoptosis in an AMPK-dependent manner. These results suggest that the LKB1-AMPK-FoxO1 signaling pathway is a critical mediator of the antioxidant properties of H 2 , further supporting the idea that H 2 acts as a signaling molecule to serve various physiological functions. Copyright © 2017 Elsevier Inc. All rights reserved.

  19. Activation of the Nrf2-ARE pathway by siRNA knockdown of Keap1 reduces oxidative stress and provides partial protection from MPTP-mediated neurotoxicity.

    Science.gov (United States)

    Williamson, Tracy P; Johnson, Delinda A; Johnson, Jeffrey A

    2012-06-01

    Nuclear factor erythroid 2-related factor 2 (Nrf2) is a transcription factor that binds to the antioxidant response element, a cis-acting regulatory element that increases expression of detoxifying enzymes and antioxidant proteins. Kelch-like ECH associating protein 1 (Keap1) protein is a negative regulator of Nrf2. Previous work has shown that genetic overexpression of Nrf2 is protective in vitro and in vivo. To modulate the Nrf2-ARE system without overexpressing Nrf2, we used short interfering RNA (siRNA) directed against Keap1. Keap1 siRNA administration in primary astrocytes increased the levels of Nrf2-ARE driven genes and protected against oxidative stress. Moreover, Keap1 siRNA resulted in a persistent upregulation of the Nrf2-ARE pathway and protection against oxidative stress in primary astrocytes. Keap1 siRNA injected into the striatum was also modestly protective against MPTP-induced dopaminergic terminal damage. These data indicate that activation of endogenous intracellular levels of Nrf2 is sufficient to protect in models of oxidative stress and Parkinson's disease. Copyright © 2012 Elsevier Inc. All rights reserved.

  20. Effect of hepatocyte growth factor and angiotensin II on rat cardiomyocyte hypertrophy

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Ai-Lan [Department of Cardiology, The First Affiliated Hospital of Guangzhou Medical University, Guangzhou (China); Ou, Cai-Wen [The Fourth Affiliated Hospital of Guangzhou Medical University, Guangzhou (China); He, Zhao-Chu [Department of Cardiology, The First Affiliated Hospital of Guangzhou Medical University, Guangzhou (China); Liu, Qi-Cai [Experimental Medical Research Center, Guangzhou Medical University, Guangzhou (China); Dong, Qi [Department of Physiology, Guangzhou Medical University, Guangzhou (China); Chen, Min-Sheng [Guangzhou Key Laboratory of Cardiovascular Disease, Guangzhou Institute of Cardiovascular Disease, The Second Affiliated Hospital of Guangzhou Medical University, Guangzhou (China)

    2012-10-15

    Angiotensin II (Ang II) plays an important role in cardiomyocyte hypertrophy. The combined effect of hepatocyte growth factor (HGF) and Ang II on cardiomyocytes is unknown. The present study was designed to determine the effect of HGF on cardiomyocyte hypertrophy and to explore the combined effect of HGF and Ang II on cardiomyocyte hypertrophy. Primary cardiomyocytes were isolated from neonatal rat hearts and cultured in vitro. Cells were treated with Ang II (1 µM) alone, HGF (10 ng/mL) alone, and Ang II (1 µM) plus HGF (10 ng/mL) for 24, 48, and 72 h. The amount of [{sup 3}H]-leucine incorporation was then measured to evaluate protein synthesis. The mRNA levels of β-myosin heavy chain and atrial natriuretic factor were determined by real-time PCR to evaluate the presence of fetal phenotypes of gene expression. The cell size of cardiomyocytes was also studied. Ang II (1 µM) increased cardiomyocyte hypertrophy. Similar to Ang II, treatment with 1 µM HGF promoted cardiomyocyte hypertrophy. Moreover, the combination of 1 µM Ang II and 10 ng/mL HGF clearly induced a combined pro-hypertrophy effect on cardiomyocytes. The present study demonstrates for the first time a novel, combined effect of HGF and Ang II in promoting cardiomyocyte hypertrophy.

  1. Calcitonin protects chondrocytes from lipopolysaccharide-induced apoptosis and inflammatory response through MAPK/Wnt/NF-κB pathways.

    Science.gov (United States)

    Zhang, Lai-Bo; Man, Zhen-Tao; Li, Wei; Zhang, Wei; Wang, Xian-Quan; Sun, Shui

    2017-07-01

    Calcitonin (CT) is an anti-absorbent, which has long been used for treatment of osteoporosis. However, little information is available about the effects of CT on osteoarthritis (OA). This study was mainly aimed to explore the effects of CT on the treatment of OA, as well as the underlying mechanisms. Chondrocytes were isolated from immature mice and then were incubated with lipopolysaccharide (LPS), CT, small interfering (si) RNA against bone morphogenetic protein (BMP)-2, and/or the inhibitors of MAPK/Wnt/NF-κB pathway. Thereafter, cell viability, apoptosis, nitric oxide (NO) and inflammatory factors productions, and expression levels of cartilage synthesis protein key factors, cartilage-derived morphogenetic protein (CDMP) 1, SRY (sex-determining region Y)-box 9 protein (SOX9), and MAPK/Wnt/NF-κB pathways key factors were determined. CT significantly reversed LPS-induced cell viability decrease, apoptosis increase, the inflammatory factors and NO secretion, the abnormally expression of cartilage synthesis proteins and the activation of MAPK/Wnt/NF-κB pathways (Ppathways statistically further increased the levels of CDMP1 and SOX9 (Ppathways, and could partially abolish CT-modulated the expression changes in CDMP1 and SOX9, and MAPK/Wnt/NF-κB pathways key factors (Ppathways. Copyright © 2017 Elsevier Ltd. All rights reserved.

  2. Recombinant adeno-associated virus-delivered hypoxia-inducible stanniocalcin-1 expression effectively inhibits hypoxia-induced cell apoptosis in cardiomyocytes.

    Science.gov (United States)

    Shi, Xin; Wang, Jianzhong; Qin, Yan

    2014-12-01

    Ischemia/hypoxia-induced oxidative stress is detrimental for the survival of cardiomyocytes and cardiac function. Stanniocalcin-1 (STC-1), a glycoprotein, has been found to play an inhibitory role in the production of reactive oxygen species (ROS). Here, we speculated that the overexpression of STC-1 might alleviate oxidative damage in cardiomyocytes under conditions of hypoxia. To control the expression of STC-1 in hypoxia, we constructed a recombinant adeno-associated virus (AAV) carrying the hypoxia-responsive element (HRE) to mediate hypoxia induction. Cardiomyocytes were infected with AAV-HRE-STC-1 and cultured in normoxic or hypoxic conditions, and STC-1 overexpression was only detected in hypoxic cultured cardiomyocytes by using quantitative real-time polymerase chain reaction and Western blot analysis. Using the 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, AAV-HRE-STC-1 infection was shown to significantly enhance cell survival under hypoxia. Hypoxia-induced cell apoptosis was inhibited by AAV-HRE-STC-1 infection by using the Annexin V-fluorescein isothiocyanate (FITC)/propidium iodide apoptosis assay. Moreover, the proapoptotic protein Caspase-3 and anti-apoptotic protein Bcl-2, which were dysregulated by hypoxia, were reversed by AAV-HRE-STC-1 infection. AAV-HRE-STC-1-mediated STC-1 overexpression markedly inhibited ROS production in cardiomyocytes cultured under hypoxic conditions. AAV-HRE-STC-1 infection significantly upregulated uncoupled protein 3 (UCP3), whereas silencing of UCP3 blocked the inhibitory effect of AAV-HRE-STC-1 on ROS production. In contrast, AAV-HRE-STC-1 infection had no effect on UCP2, and knockdown of UCP2 did not block the inhibitory effect of AAV-HRE-STC-1 on ROS production in the cardiomyocytes cultured under hypoxic conditions. Taken together, STC1 activates antioxidant pathway in cardiomyocytes through the induction of UCP3, implying that AAV-HRE-STC-1 has potential in the treatment of ischemic

  3. SIRT1 protects cardiac cells against apoptosis induced by zearalenone or its metabolites α- and β-zearalenol through an autophagy-dependent pathway

    International Nuclear Information System (INIS)

    Ben Salem, Intidhar; Boussabbeh, Manel; Da Silva, Julie Pires; Guilbert, Arnaud; Bacha, Hassen; Abid-Essefi, Salwa; Lemaire, Christophe

    2017-01-01

    Zearalenone (ZEN) is a non-steroidal estrogenic mycotoxin produced by several species of Fusarium in cereals and agricultural products. The major ZEN metabolites are α-zearalenol (α-ZOL) and β-zearalenol (β-ZOL). In the present study, we investigated the underlying mechanism of the toxicity induced by ZEN, α-ZOL and β-ZOL in cardiac cells (H9c2). We show that treatment with ZEN or its metabolites induces the activation of the mitochondrial pathway of apoptosis as characterized by an increase in ROS generation, a loss of mitochondrial transmembrane potential (ΔΨm) and an activation of caspases. Besides, we demonstrate that these mycotoxins promote the activation of autophagy before the onset of apoptosis. Indeed, we observed that a short-time (6 h) treatment with ZEN, α-ZOL or β-ZOL, increased the level of Beclin-1 and LC3-II and induced the accumulation of the CytoID® autophagy detection probe. Moreover, the inhibition of autophagy by Chloroquine significantly increased cell death induced by ZEN, α-ZOL or β-ZOL, suggesting that the activation of autophagy serves as a cardioprotective mechanism against these mycotoxins. In addition, we found that the inhibition (EX527) or the knockdown of SIRT1 (siRNA) significantly increased apoptosis induced by ZEN or its derivatives, whereas SIRT1 activation with RSV greatly prevents the cytotoxic effects of these mycotoxins. By contrast, when autophagy was inhibited by CQ, the activation of SIRT1 by RSV had no protection against the cardiotoxicity of ZEN or its metabolites, suggesting that SIRT1 protects cardiac cells by an autophagy-dependent pathway. - Highlights: • ZEN, α- and β-ZOL induce the mitochondrial pathway of apoptosis in cardiac cells. • Inhibition of autophagy enhanced ZEN-, α-ZOL- and β-ZOL-induced apoptosis. • SIRT1 activates autophagy to protect cells from ZEN, α- and β-ZOL-induced toxicity.

  4. Tetramethylpyrazine Protects Against Oxygen-Glucose Deprivation-Induced Brain Microvascular Endothelial Cells Injury via Rho/Rho-kinase Signaling Pathway.

    Science.gov (United States)

    Yang, Guang; Qian, Chen; Wang, Ning; Lin, Chenyu; Wang, Yan; Wang, Guangyun; Piao, Xinxin

    2017-05-01

    Tetramethylpyrazine (TMP, also known as Ligustrazine), which is isolated from Chinese Herb Medicine Ligustium wollichii Franchat (Chuan Xiong), has been widely used in China for the treatment of ischemic stroke by Chinese herbalists. Brain microvascular endothelial cells (BMECs) are the integral parts of the blood-brain barrier (BBB), protecting BMECs against oxygen-glucose deprivation (OGD) which is important for the treatment of ischemic stroke. Here, we investigated the protective mechanisms of TMP, focusing on OGD-injured BMECs and the Rho/Rho-kinase (Rho-associated kinases, ROCK) signaling pathway. The model of OGD-injured BMECs was established in this study. BMECs were identified by von Willebrand factor III staining and exposed to fasudil, or TMP at different concentrations (14.3, 28.6, 57.3 µM) for 2 h before 24 h of OGD injury. The effect of each treatment was examined by cell viability assays, measurement of intracellular reactive oxygen species (ROS), and transendothelial electric resistance and western blot analysis (caspase-3, endothelial nitric oxide synthase (eNOS), RhoA, Rac1). Our results show that TMP significantly attenuated apoptosis and the permeability of BMECs induced by OGD. In addition, TMP could notably down-regulate the characteristic proteins in Rho/ROCK signaling pathway such as RhoA and Rac1, which triggered abnormal changes of eNOS and ROS, respectively. Altogether, our results show that TMP has a strong protective effect against OGD-induced BMECs injury and suggest that the mechanism might be related to the inhibition of the Rho/ROCK signaling pathway.

  5. Curcumin Protects Skin against UVB-Induced Cytotoxicity via the Keap1-Nrf2 Pathway: The Use of a Microemulsion Delivery System

    Directory of Open Access Journals (Sweden)

    Maya Ben Yehuda Greenwald

    2017-01-01

    Full Text Available Curcumin was found to be beneficial in treating several skin pathologies and diseases, providing antioxidant protection due to its reducing properties and its electrophilic properties (the ability to activate the Nrf2 pathway and induce phase II cytoprotective enzymes. Nevertheless, clinical applications of curcumin are being hampered by its insufficient solubility, chemical instability, and poor absorption, leading to low efficacy in preventing skin pathologies. These limitations can be overcome by using a nanotechnology-based delivery system. Here, we elucidated the possibility of using curcumin encapsulated in a microemulsion preserving its unique chemical structure. We also examined whether curcumin microemulsion would reduce UVB-induced toxicity in skin. A significant curcumin concentration was found in the human skin dermis following topical application of a curcumin microemulsion. Moreover, curcumin microemulsion enhanced the reduction of UV-induced cytotoxicity in epidermal cells, paving the way for other incorporated electrophiles in encapsulated form protecting skin against stress-related diseases.

  6. Curcumin Protects Skin against UVB-Induced Cytotoxicity via the Keap1-Nrf2 Pathway: The Use of a Microemulsion Delivery System

    Science.gov (United States)

    Ben Yehuda Greenwald, Maya; Frušić-Zlotkin, Marina; Soroka, Yoram; Ben Sasson, Shmuel; Bitton, Ronit; Bianco-Peled, Havazelet

    2017-01-01

    Curcumin was found to be beneficial in treating several skin pathologies and diseases, providing antioxidant protection due to its reducing properties and its electrophilic properties (the ability to activate the Nrf2 pathway and induce phase II cytoprotective enzymes). Nevertheless, clinical applications of curcumin are being hampered by its insufficient solubility, chemical instability, and poor absorption, leading to low efficacy in preventing skin pathologies. These limitations can be overcome by using a nanotechnology-based delivery system. Here, we elucidated the possibility of using curcumin encapsulated in a microemulsion preserving its unique chemical structure. We also examined whether curcumin microemulsion would reduce UVB-induced toxicity in skin. A significant curcumin concentration was found in the human skin dermis following topical application of a curcumin microemulsion. Moreover, curcumin microemulsion enhanced the reduction of UV-induced cytotoxicity in epidermal cells, paving the way for other incorporated electrophiles in encapsulated form protecting skin against stress-related diseases. PMID:28757910

  7. Determining the Molecular Pathways Underlying the Protective Effect of Non-Steroidal Anti-Inflammatory Drugs for Alzheimer's Disease: A Bioinformatics Approach

    Directory of Open Access Journals (Sweden)

    Alejo J Nevado-Holgado

    Full Text Available Alzheimer's disease (AD represents a substantial unmet need, due to increasing prevalence in an ageing society and the absence of a disease modifying therapy. Epidemiological evidence shows a protective effect of non steroidal anti inflammatory (NSAID drugs, and genome wide association studies (GWAS show consistent linkage to inflammatory pathways; both observations suggesting anti-inflammatory compounds might be effective in AD therapy although clinical trials to date have not been positive.In this study, we use pathway enrichment and fuzzy logic to identify pathways (KEGG database simultaneously affected in both AD and by NSAIDs (Sulindac, Piroxicam, Paracetamol, Naproxen, Nabumetone, Ketoprofen, Diclofenac and Aspirin. Gene expression signatures were derived for disease from both blood (n = 344 and post-mortem brain (n = 690, and for drugs from immortalised human cell lines exposed to drugs of interest as part of the Connectivity Map platform. Using this novel approach to combine datasets we find striking overlap between AD gene expression in blood and NSAID induced changes in KEGG pathways of Ribosome and Oxidative Phosphorylation. No overlap was found in non NSAID comparison drugs. In brain we find little such overlap, although Oxidative Phosphorylation approaches our pre-specified significance level.These findings suggest that NSAIDs might have a mode of action beyond inflammation and moreover that their therapeutic effects might be mediated in particular by alteration of Oxidative Phosphorylation and possibly the Ribosome pathway. Mining of such datasets might prove increasingly productive as they increase in size and richness. Keywords: Alzheimer's disease, NSAID, Inflammation, Fuzzy logic, Ribosome

  8. Exposure scenarios of the German Federal Soil Protection and Contamination Ordinance - impact pathway from soil to humans

    International Nuclear Information System (INIS)

    Konietzka, R.

    2005-01-01

    The soil uptake values used for calculating soil values have not undergone strict scientific validation by extensive testing. As far as possible be they can and should be based on empirical data. In many cases it is inevitable to resort to conventions, but these should be made plausible through empirical studies. Land use scenarios relevant to the calculation of test values for direct uptake from soil to humans are playgrounds, residential areas, park and leisure facilities and industrial and commercial properties. The relevant exposure pathways are assumed to differ between scenarios depending on certain characteristics of the potentially exposed population groups. In most cases the pathways oral, inhalational (inhalation of contaminated dusts) and dermal uptake from the soil are considered

  9. The coiled-coil domain of MURC/cavin-4 is involved in membrane trafficking of caveolin-3 in cardiomyocytes.

    Science.gov (United States)

    Naito, Daisuke; Ogata, Takehiro; Hamaoka, Tetsuro; Nakanishi, Naohiko; Miyagawa, Kotaro; Maruyama, Naoki; Kasahara, Takeru; Taniguchi, Takuya; Nishi, Masahiro; Matoba, Satoaki; Ueyama, Tomomi

    2015-12-15

    Muscle-restricted coiled-coil protein (MURC), also referred to as cavin-4, is a member of the cavin family that works cooperatively with caveolins in caveola formation and function. Cavins are cytoplasmic proteins with coiled-coil domains and form heteromeric complexes, which are recruited to caveolae in cells expressing caveolins. Among caveolins, caveolin-3 (Cav3) is exclusively expressed in muscle cells, similar to MURC/cavin-4. In the heart, Cav3 overexpression contributes to cardiac protection, and its deficiency leads to progressive cardiomyopathy. Mutations in the MURC/cavin-4 gene have been identified in patients with dilated cardiomyopathy. In the present study, we show the role of MURC/cavin-4 as a caveolar component in the heart. In H9c2 cells, MURC/cavin-4 was localized at the plasma membrane, whereas a MURC/cavin-4 mutant lacking the coiled-coil domain (ΔCC) was primarily localized to the cytoplasm. ΔCC bound to Cav3 and impaired membrane localization of Cav3 in cardiomyocytes. Additionally, although ΔCC did not alter Cav3 mRNA expression, ΔCC decreased the Cav3 protein level. MURC/cavin-4 and ΔCC similarly induced cardiomyocyte hypertrophy; however, ΔCC showed higher hypertrophy-related fetal gene expression than MURC/cavin-4. ΔCC induced ERK activation in cardiomyocytes. Transgenic mice expressing ΔCC in the heart (ΔCC-Tg mice) showed impaired cardiac function accompanied by cardiomyocyte hypertrophy and marked interstitial fibrosis. Hearts from ΔCC-Tg mice showed a reduction of the Cav3 protein level and activation of ERK. These results suggest that MURC/cavin-4 requires its coiled-coil domain to target the plasma membrane and to stabilize Cav3 at the plasma membrane of cardiomyocytes and that MURC/cavin-4 functions as a crucial caveolar component to regulate cardiac function. Copyright © 2015 the American Physiological Society.

  10. Creating a Structurally Realistic Finite Element Geometric Model of a Cardiomyocyte to Study the Role of Cellular Architecture in Cardiomyocyte Systems Biology.

    Science.gov (United States)

    Rajagopal, Vijay; Bass, Gregory; Ghosh, Shouryadipta; Hunt, Hilary; Walker, Cameron; Hanssen, Eric; Crampin, Edmund; Soeller, Christian

    2018-04-18

    With the advent of three-dimensional (3D) imaging technologies such as electron tomography, serial-block-face scanning electron microscopy and confocal microscopy, the scientific community has unprecedented access to large datasets at sub-micrometer resolution that characterize the architectural remodeling that accompanies changes in cardiomyocyte function in health and disease. However, these datasets have been under-utilized for investigating the role of cellular architecture remodeling in cardiomyocyte function. The purpose of this protocol is to outline how to create an accurate finite element model of a cardiomyocyte using high resolution electron microscopy and confocal microscopy images. A detailed and accurate model of cellular architecture has significant potential to provide new insights into cardiomyocyte biology, more than experiments alone can garner. The power of this method lies in its ability to computationally fuse information from two disparate imaging modalities of cardiomyocyte ultrastructure to develop one unified and detailed model of the cardiomyocyte. This protocol outlines steps to integrate electron tomography and confocal microscopy images of adult male Wistar (name for a specific breed of albino rat) rat cardiomyocytes to develop a half-sarcomere finite element model of the cardiomyocyte. The procedure generates a 3D finite element model that contains an accurate, high-resolution depiction (on the order of ~35 nm) of the distribution of mitochondria, myofibrils and ryanodine receptor clusters that release the necessary calcium for cardiomyocyte contraction from the sarcoplasmic reticular network (SR) into the myofibril and cytosolic compartment. The model generated here as an illustration does not incorporate details of the transverse-tubule architecture or the sarcoplasmic reticular network and is therefore a minimal model of the cardiomyocyte. Nevertheless, the model can already be applied in simulation-based investigations into the

  11. Taurine protects against As2O3-induced autophagy in pancreas of rat offsprings through Nrf2/Trx pathway.

    Science.gov (United States)

    Bai, Jie; Yao, Xiaofeng; Jiang, Liping; Qiu, Tianming; Liu, Shuang; Qi, Baoxu; Zheng, Yue; Kong, Yuan; Yang, Guang; Chen, Min; Liu, Xiaofang; Sun, Xiance

    2016-04-01

    Arsenic was increasingly to blame as a risk factor for type 2 diabetes mellitus. In our previous study, we had found iAs stimulated autophagic flux and caused autophagic cell death through ROS pathway in INS-1 cells. Since NF-E2-related factor 2 (Nrf2) and the thioredoxin (Trx) system was a crucial line of defense against ROS, we investigated whether Nrf2/Trx pathway contributed to As2O3-stimulated autophagy and the role of taurine in this study. After treatment with 2 mg/kg BW-8 mg/kg BW As2O3 for 57 d, the expression of Nrf2 protein was decreased significantly in offsprings' pancreas. The expression of Trx gene was decreased significantly in pancreas subsequently. Finally, the generation of reactive oxygen species stimulated autophagy in arsenic-treated pancreas. Taurine could reverse arsenic-inhibited Nrf2 and Trx and inhibit autophagy. In short, inhibition of Nrf2/Trx pathway might play an important role in the pathogenesis of arsenic-related diabetes. Taurine could serve as nutrition supplementation against arsenic-related diabetes in high arsenic exposure area. Copyright © 2016 Elsevier B.V. and Société Française de Biochimie et Biologie Moléculaire (SFBBM). All rights reserved.

  12. Chinese herbal formula Tongluo Jiunao injection protects against cerebral ischemia by activating neurotrophin 3/tropomyosin-related kinase C pathway

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    Peiman Alesheikh

    2015-01-01

    Full Text Available The Chinese herbal formula Tongluo Jiunao, containing the active components Panax notoginseng and Gardenia jasminoides, has recently been patented and is in use clinically. It is known to be neuroprotective in cerebral ischemia, but the underlying pathway remains poorly understood. In the present study, we established a rat model of cerebral ischemia by occlusion of the middle cerebral artery, and administered Tongluo Jiunao, a positive control (Xuesai Tong, containing Panax notoginseng or saline intraperitoneally to investigate the pathway involved in the action of Tongluo Jiunao injection. 2,3,5-Triphenyltetrazolium chloride (TTC staining showed that the cerebral infarct area was significantly smaller in model rats that received Tongluo Jiunao than in those that received saline. Enzyme-linked immunosorbent assay revealed significantly greater expression of neurotrophin 3 and growth-associated protein 43 in ischemic cerebral tissue, and serum levels of neurotrophin 3, in the Tongluo Jiunao group than in the saline group. The reverse transcription polymerase chain reaction and immunohistochemical staining showed that after treatment with Tongluo Jiunao or Xuesai Tong, tropomyosin-related kinase C gene expression and immunoreactivity were significantly elevated compared with saline, with the greatest expression observed after Tongluo Jiunao treatment. These findings suggest that Tongluo Jiunao injection exerts a neuroprotective effect in rats with cerebral ischemia by activating the neurotrophin 3/tropomyosin-related kinase C pathway.

  13. Lycium barbarum Polysaccharides Protect Rat Corneal Epithelial Cells against Ultraviolet B-Induced Apoptosis by Attenuating the Mitochondrial Pathway and Inhibiting JNK Phosphorylation

    Directory of Open Access Journals (Sweden)

    Shaobo Du

    2017-01-01

    Full Text Available Lycium barbarum polysaccharides (LBPs have been shown to play a key role in protecting the eyes by reducing the apoptosis induced by certain types of damage. However, it is not known whether LBPs can protect damaged corneal cells from apoptosis. Moreover, no reports have focused on the role of LBPs in guarding against ultraviolet B- (UVB- induced apoptosis. The present study aimed to investigate the protective effect and underlying mechanism of LBPs against UVB-induced apoptosis in rat corneal epithelial (RCE cells. The results showed that LBPs significantly prevented the loss of cell viability and inhibited cell apoptosis induced by UVB in RCE cells. LBPs also inhibited UVB-induced loss of mitochondrial membrane potential, downregulation of Bcl-2, and upregulation of Bax and caspase-3. Finally, LBPs attenuated the phosphorylation of c-Jun NH2-terminal kinase (JNK triggered by UVB. In summary, LBPs protect RCE cells against UVB-induced damage and apoptosis, and the underlying mechanism involves the attenuation of the mitochondrial apoptosis pathway and the inhibition of JNK phosphorylation.

  14. Araloside C Prevents Hypoxia/Reoxygenation-Induced Endoplasmic Reticulum Stress via Increasing Heat Shock Protein 90 in H9c2 Cardiomyocytes

    Directory of Open Access Journals (Sweden)

    Yuyang Du

    2018-04-01

    Full Text Available Araloside C (AsC is a cardioprotective triterpenoid compound that is mainly isolated from Aralia elata. This study aims to determine the effects of AsC on hypoxia-reoxygenation (H/R-induced apoptosis in H9c2 cardiomyocytes and its underlying mechanisms. Results demonstrated that pretreatment with AsC (12.5 μM for 12 h significantly suppressed the H/R injury in H9c2 cardiomyocytes, including improving cell viability, attenuating the LDH leakage and preventing cardiomyocyte apoptosis. AsC also inhibited H/R-induced ER stress by reducing the activation of ER stress pathways (PERK/eIF2α and ATF6, and decreasing the expression of ER stress-related apoptotic proteins (CHOP and caspase-12. Moreover, AsC greatly improved the expression level of HSP90 compared with that in the H/R group. The use of HSP90 inhibitor 17-AAG and HSP90 siRNA blocked the above suppression effect of AsC on ER stress-related apoptosis caused by H/R. Taken together, AsC could reduce H/R-induced apoptosis possibly because it attenuates ER stress-dependent apoptotic pathways by increasing HSP90 expression.

  15. Doxorubicin impairs the insulin-like growth factor-1 system and causes insulin-like growth factor-1 resistance in cardiomyocytes.

    Directory of Open Access Journals (Sweden)

    Patrizia Fabbi

    Full Text Available Insulin-like growth factor-1 (IGF-1 promotes the survival of cardiomyocytes by activating type 1 IGF receptor (IGF-1R. Within the myocardium, IGF-1 action is modulated by IGF binding protein-3 (IGFBP-3, which sequesters IGF-1 away from IGF-1R. Since cardiomyocyte apoptosis is implicated in anthracycline cardiotoxicity, we investigated the effects of the anthracycline, doxorubicin, on the IGF-1 system in H9c2 cardiomyocytes.Besides inducing apoptosis, concentrations of doxorubicin comparable to those observed in patients after bolus infusion (0.1-1 µM caused a progressive decrease in IGF-1R and increase in IGFBP-3 expression. Exogenous IGF-1 was capable to rescue cardiomyocytes from apoptosis triggered by 0.1 and 0.5 µM, but not 1 µM doxorubicin. The loss of response to IGF-1 was paralleled by a significant reduction in IGF-1 availability and signaling, as assessed by free hormone levels in conditioned media and Akt phosphorylation in cell lysates, respectively. Doxorubicin also dose-dependently induced p53, which is known to repress the transcription of IGF1R and induce that of IGFBP3. Pre-treatment with the p53 inhibitor, pifithrin-α, prevented apoptosis and the changes in IGF-1R and IGFBP-3 elicited by doxorubicin. The decrease in IGF-1R and increase in IGFBP-3, as well as apoptosis, were also antagonized by pre-treatment with the antioxidant agents, N-acetylcysteine, dexrazoxane, and carvedilol.Doxorubicin down-regulates IGF-1R and up-regulates IGFBP-3 via p53 and oxidative stress in H9c2 cells. This leads to resistance to IGF-1 that may contribute to doxorubicin-initiated apoptosis. Further studies are needed to confirm these findings in human cardiomyocytes and explore the possibility of manipulating the IGF-1 axis to protect against anthracycline cardiotoxicity.

  16. Cardiac injury of the newborn mammalian heart accelerates cardiomyocyte terminal differentiation

    DEFF Research Database (Denmark)

    Zebrowski, David C.; Jensen, Charlotte H.; Becker, Robert

    2017-01-01

    exhibited midbody formation consistent with successful abscission, whereas those from 3 day-old cardiomyocytes after apical resection exhibited midbody formation consistent with abscission failure. Lastly, injured hearts failed to fully regenerate as evidenced by persistent scarring and reduced wall motion......After birth cardiomyocytes undergo terminal differentiation, characterized by binucleation and centrosome disassembly, rendering the heart unable to regenerate. Yet, it has been suggested that newborn mammals regenerate their hearts after apical resection by cardiomyocyte proliferation. Thus, we...... increased rate of binucleation there was a nearly 2-fold increase in the number of cardiomyocytes in mitosis indicating that the majority of injury-induced cardiomyocyte cell cycle activity results in binucleation, not proliferation. Concurrently, cardiomyocytes undergoing cytokinesis from embryonic hearts...

  17. Frequency of mononuclear diploid cardiomyocytes underlies natural variation in heart regeneration.

    Science.gov (United States)

    Patterson, Michaela; Barske, Lindsey; Van Handel, Ben; Rau, Christoph D; Gan, Peiheng; Sharma, Avneesh; Parikh, Shan; Denholtz, Matt; Huang, Ying; Yamaguchi, Yukiko; Shen, Hua; Allayee, Hooman; Crump, J Gage; Force, Thomas I; Lien, Ching-Ling; Makita, Takako; Lusis, Aldons J; Kumar, S Ram; Sucov, Henry M

    2017-09-01

    Adult mammalian cardiomyocyte regeneration after injury is thought to be minimal. Mononuclear diploid cardiomyocytes (MNDCMs), a relatively small subpopulation in the adult heart, may account for the observed degree of regeneration, but this has not been tested. We surveyed 120 inbred mouse strains and found that the frequency of adult mononuclear cardiomyocytes was surprisingly variable (>7-fold). Cardiomyocyte proliferation and heart functional recovery after coronary artery ligation both correlated with pre-injury MNDCM content. Using genome-wide association, we identified Tnni3k as one gene that influences variation in this composition and demonstrated that Tnni3k knockout resulted in elevated MNDCM content and increased cardiomyocyte proliferation after injury. Reciprocally, overexpression of Tnni3k in zebrafish promoted cardiomyocyte polyploidization and compromised heart regeneration. Our results corroborate the relevance of MNDCMs in heart regeneration. Moreover, they imply that intrinsic heart regeneration is not limited nor uniform in all individuals, but rather is a variable trait influenced by multiple genes.

  18. Mitochondrial apoptotic pathway activation in the atria of heart failure patients due to mitral and tricuspid regurgitation.

    Science.gov (United States)

    Chang, Jen-Ping; Chen, Mien-Cheng; Liu, Wen-Hao; Lin, Yu-Sheng; Huang, Yao-Kuang; Pan, Kuo-Li; Ho, Wan-Chun; Fang, Chih-Yuan; Chen, Chien-Jen; Chen, Huang-Chung

    2015-08-01

    Apoptosis occurs in atrial cardiomyocytes in mitral and tricuspid valve disease. The purpose of this study was to examine the respective roles of the mitochondrial and tumor necrosis factor-α receptor associated death domain (TRADD)-mediated death receptor pathways for apoptosis in the atrial cardiomyocytes of heart failure patients due to severe mitral and moderate-to-severe tricuspid regurgitation. This study comprised eighteen patients (7 patients with persistent atrial fibrillation and 11 in sinus rhythm). Atrial appendage tissues were obtained during surgery. Three purchased normal human left atrial tissues served as normal controls. Moderately-to-severely myolytic cardiomyocytes comprised 59.7±22.1% of the cardiomyocytes in the right atria and 52.4±12.9% of the cardiomyocytes in the left atria of mitral and tricuspid regurgitation patients with atrial fibrillation group and comprised 58.4±24.8% of the cardiomyocytes in the right atria of mitral and tricuspid regurgitation patients with sinus rhythm. In contrast, no myolysis was observed in the normal human adult left atrial tissue samples. Immunohistochemical analysis showed expression of cleaved caspase-9, an effector of the mitochondrial pathways, in the majority of right atrial cardiomyocytes (87.3±10.0%) of mitral and tricuspid regurgitation patients with sinus rhythm, and right atrial cardiomyocytes (90.6±31.4%) and left atrial cardiomyocytes (70.7±22.0%) of mitral and tricuspid regurgitation patients with atrial fibrillation. In contrast, only 5.7% of cardiomyocytes of the normal left atrial tissues showed strongly positive expression of cleaved caspase-9. Of note, none of the atrial cardiomyocytes in right atrial tissue in sinus rhythm and in the fibrillating right and left atria of mitral and tricuspid regurgitation patients, and in the normal human adult left atrial tissue samples showed cleaved caspase-8 expression, which is a downstream effector of TRADD of the death receptor pathway

  19. Rac1 modulates cardiomyocyte adhesion during mouse embryonic development

    Energy Technology Data Exchange (ETDEWEB)

    Abu-Issa, Radwan, E-mail: rabuissa@umich.edu

    2015-01-24

    Highlights: • Conditional knockout of Rac1 using Nkx2.5 Cre line is lethal at E13.5. • The myocardium of the mutant is thin and disorganized. • The phenotype is not due to cardiomyocyte low proliferation or apoptosis. • The phenotype is due to specific defect in cardiomyocyte adhesion. - Abstract: Rac1, a member of the Rho subfamily of small GTPases, is involved in morphogenesis and differentiation of many cell types. Here we define a role of Rac1 in cardiac development by specifically deleting Rac1 in the pre-cardiac mesoderm using the Nkx2.5-Cre transgenic driver line. Rac1-conditional knockout embryos initiate heart development normally until embryonic day 11.5 (E11.5); their cardiac mesoderm is specified, and the heart tube is formed and looped. However, by E12.5-E13.5 the mutant hearts start failing and embryos develop edema and hemorrhage which is probably the cause for the lethality observed soon after. The hearts of Rac1-cKO embryos exhibit disorganized and thin myocardial walls and defects in outflow tract alignment. No significant differences of cardiomyocyte death or proliferation were found between developing control and mutant embryos. To uncover the role of Rac1 in the heart, E11.5 primary heart cells were cultured and analyzed in vitro. Rac1-deficient cardiomyocytes were less spread, round and loosely attached to the substrate and to each other implying that Rac1-mediated signaling is required for appropriate cell–cell and/or cellmatrix adhesion during cardiac development.

  20. Apoptosis of rats’ cardiomyocytes after chronic energy drinks consumption

    Directory of Open Access Journals (Sweden)

    Slawinski Miroslaw Aleksander

    2018-03-01

    Full Text Available Energy drinks (ED are beverages containing caffeine, taurine, vitamins, herbal extracts, and sugar or sweeteners. They are marketed as capable of improving stamina, athletic performance and concentration, moreover, as serving as a source of energy. Still, there are very few papers describing the impact of ED on cell biology – including cell apoptosis within tissues. Therefore, in our study, we assessed the symptoms of rat cardiomyocytes apoptosis after 8 weeks consumption of ED.

  1. Rac1 modulates cardiomyocyte adhesion during mouse embryonic development

    International Nuclear Information System (INIS)

    Abu-Issa, Radwan

    2015-01-01

    Highlights: • Conditional knockout of Rac1 using Nkx2.5 Cre line is lethal at E13.5. • The myocardium of the mutant is thin and disorganized. • The phenotype is not due to cardiomyocyte low proliferation or apoptosis. • The phenotype is due to specific defect in cardiomyocyte adhesion. - Abstract: Rac1, a member of the Rho subfamily of small GTPases, is involved in morphogenesis and differentiation of many cell types. Here we define a role of Rac1 in cardiac development by specifically deleting Rac1 in the pre-cardiac mesoderm using the Nkx2.5-Cre transgenic driver line. Rac1-conditional knockout embryos initiate heart development normally until embryonic day 11.5 (E11.5); their cardiac mesoderm is specified, and the heart tube is formed and looped. However, by E12.5-E13.5 the mutant hearts start failing and embryos develop edema and hemorrhage which is probably the cause for the lethality observed soon after. The hearts of Rac1-cKO embryos exhibit disorganized and thin myocardial walls and defects in outflow tract alignment. No significant differences of cardiomyocyte death or proliferation were found between developing control and mutant embryos. To uncover the role of Rac1 in the heart, E11.5 primary heart cells were cultured and analyzed in vitro. Rac1-deficient cardiomyocytes were less spread, round and loosely attached to the substrate and to each other implying that Rac1-mediated signaling is required for appropriate cell–cell and/or cellmatrix adhesion during cardiac development

  2. JANEX-1, a JAK3 inhibitor, protects pancreatic islets from cytokine toxicity through downregulation of NF-{kappa}B activation and the JAK/STAT pathway

    Energy Technology Data Exchange (ETDEWEB)

    Lv, Na; Kim, Eun-Kyung; Song, Mi-Young [Department of Biochemistry, Medical School and Diabetes Research Center, Chonbuk National University, Jeonju, Jeonbuk 561-756 (Korea, Republic of); Choi, Ha-Na; Moon, Woo Sung [Department of Pathology, Medical School and Diabetes Research Center, Chonbuk National University, Jeonju, Jeonbuk 561-756 (Korea, Republic of); Park, Sung-Joo [Department of Herbology, School of Oriental Medicine, Wonkwang University, Iksan, Jeonbuk 570-749 (Korea, Republic of); Park, Jin-Woo [Department of Biochemistry, Medical School and Diabetes Research Center, Chonbuk National University, Jeonju, Jeonbuk 561-756 (Korea, Republic of); Kwon, Kang-Beom, E-mail: desson@wonkwang.ac.kr [Department of Physiology, School of Oriental Medicine, Wonkwang University, Iksan, Jeonbuk 570-749 (Korea, Republic of); Park, Byung-Hyun, E-mail: bhpark@chonbuk.ac.kr [Department of Biochemistry, Medical School and Diabetes Research Center, Chonbuk National University, Jeonju, Jeonbuk 561-756 (Korea, Republic of)

    2009-07-15

    JANEX-1/WHI-P131, a selective Janus kinase 3 (JAK3) inhibitor, has been shown to delay the onset of diabetes in the NOD mouse model. However, the molecular mechanism by which JANEX-1 protects pancreatic {beta}-cells is unknown. In the current study, we investigated the role of JANEX-1 on interleukin (IL)-1{beta} and interferon (IFN)-{gamma}-induced {beta}-cell damage using isolated islets. JANEX-1-pretreated islets showed resistance to cytokine toxicity, namely suppressed nitric oxide (NO) production, reduced inducible form of NO synthase (iNOS) expression, and decreased islet destruction. The molecular mechanism by which JANEX-1 inhibits iNOS expression was mediated through suppression of the nuclear factor {kappa}B (NF-{kappa}B) and JAK/signal transducer and activator of transcription (STAT) pathways. Islets treated with the cytokines downregulated the protein levels of suppressor of cytokine signaling (SOCS)-1 and SOCS-3, but pretreatment with JANEX-1 attenuated these decreases. Additionally, islets from JAK3{sup -/-} mice were more resistant to cytokine toxicity than islets from control mice. These results demonstrate that JANEX-1 protects {beta}-cells from cytokine toxicity through suppression of the NF-{kappa}B and JAK/STAT pathways and upregulation of SOCS proteins, suggesting that JANEX-1 may be used to preserve functional {beta}-cell mass.

  3. Urinary and Serum Metabolomics Analyses Uncover That Total Glucosides of Paeony Protect Liver against Acute Injury Potentially via Reprogramming of Multiple Metabolic Pathways

    Directory of Open Access Journals (Sweden)

    Haojie Li

    2017-01-01

    Full Text Available Total glucosides of paeony (TGP have been confirmed to be hepatoprotective. However, the underlying mechanism is largely unclear. In this study, we investigated the metabolic profiles of urine and serum in rats with carbon tetrachloride- (CCl4- induced experimental liver injury and TGP administration by using ultra-performance liquid chromatography-mass spectrometry (UPLC-MS. The vehicle or a single dose of TGP was intragastrically administered to Wistar rats once a day for 14 consecutive days. To induce ALI, 50% CCl4 was injected intraperitoneally into these rats 2 hours after the last time administration of saline of TGP at the 14th day. The results indicated that TGP administration could protect rats from CCl4-induced ALI and alanine aminotransferase (ALT and aspartate aminotransferase (AST elevation, as well as hepatocyte apoptosis and inflammation. Furthermore, metabolomics analysis showed that TGP treatment significantly attenuated CCl4-triggered deregulation of multiple metabolites in both urine and serum, including glycine, alanine, proline, and glutamine. Metabolite set enrichment and pathway analyses demonstrated that amino acid cycling and glutathione metabolism were two main pathways involved in CCl4-induced experimental liver injury and TGP administration. Taken together, these findings revealed that regulation of metabolites potentially plays a pivotal role in the protective effect of TGP on ALI.

  4. Protection by the gross saponins of Tribulus terrestris against cerebral ischemic injury in rats involves the NF-κB pathway

    Directory of Open Access Journals (Sweden)

    En-ping Jiang

    2011-06-01

    Full Text Available The aim of this study was to investigate whether the gross saponins of Tribulus terrestris (GSTT, a traditional Chinese herbal medicine, have neuroprotective effects on rats subjected to middle cerebral artery occlusion (MCAO, through nuclear factor-κB (NF-κB pathway and inflammatory mediators. Cerebral ischemia was produced by MCAO in either untreated (control or GSTT-pretreated rats, and the animals were examined for infarct volume, cerebral edema, neuro-behavioral abnormality and pathological changes. Meanwhile, the expression of NF-κB protein in brain tissue was analyzed on Western blots and the serum levels of TNF-α and IL-1 were determined by ELISA. The experimental results demonstrated that, compared with the control MCAO group, GSTT-pretreated MCAO group had significantly reduced infarct volume, brain edema and neuro-behavioral abnormality, and lesser degree of pathologic changes in the brain, as well as had lower levels of serum TNF-α and IL-1β, and higher levels of brain NF-κB (P<0.05. Furthermore, treatment with an NF-κB inhibitor pyrrolidine dithiocarbamate (PDTC abolished the protective effects of GSTT against MCAO-induced cerebral ischemic injury. These results indicated that GSTT's ability to protect against cerebral ischemic injury was mediated through the NF-κB signaling pathway, and that GSTT may act through inhibition of the production of inflammatory mediators.

  5. Naoxintong Protects Primary Neurons from Oxygen-Glucose Deprivation/Reoxygenation Induced Injury through PI3K-Akt Signaling Pathway

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    Yan Ma

    2016-01-01

    Full Text Available Naoxintong capsule (NXT, developed from Buyang Huanwu Decoction, has shown the neuroprotective effects in cerebrovascular diseases, but the neuroprotection mechanisms of NXT on ischemia/reperfusion injured neurons have not yet been well known. In this study, we established the oxygen-glucose deprivation/reoxygenation (OGD/R induced neurons injury model and treat the neurons with cerebrospinal fluid containing NXT (BNC to investigate the effects of NXT on OGD/R induced neurons injury and potential mechanisms. BNC improved neuron viability and decreased apoptotic rate induced by OGD/R. BNC attenuated OGD/R induced cytosolic and mitochondrial Ca2+ overload, ROS generation, intracellular NO levels and nNOS mRNA increase, and cytochrome-c release when compared with OGD/R group. BNC significantly inhibited both mPTP opening and ΔΨm depolarization. BNC increased Bcl-2 expression and decreased Bax expression, upregulated the Bcl-2/Bax ratio, downregulated caspase-3 mRNA and caspase-9 mRNA expression, and decreased cleaved caspase-3 expression and caspase-3 activity. BNC increased phosphorylation of Akt following OGD/R, while LY294002 attenuated BNC induced increase of phosphorylated Akt expression. Our study demonstrated that NXT protected primary neurons from OGD/R induced injury by inhibiting calcium overload and ROS generation, protecting mitochondria, and inhibiting mitochondrial apoptotic pathway which was mediated partially by PI3K-Akt signaling pathway activation.

  6. A cross-scale impact assessment of European nature protection policies under contrasting future socio-economic pathways

    NARCIS (Netherlands)

    Lotze-Campen, Hermann; Verburg, Peter H.; Popp, Alexander; Lindner, Marcus; Verkerk, Pieter J.; Moiseyev, Alexander; Schrammeijer, Elizabeth; Helming, John; Tabeau, Andrzej; Schulp, Catharina J.E.; Zanden, van der Emma H.; Lavalle, Carlo; E Silva, Filipe Batista; Walz, Ariane; Bodirsky, Benjamin

    2018-01-01

    Protection of natural or semi-natural ecosystems is an important part of societal strategies for maintaining biodiversity, ecosystem services, and achieving overall sustainable development. The assessment of multiple emerging land use trade-offs is complicated by the fact that land use changes occur

  7. Mutations in Alström Protein Impair Terminal Differentiation of Cardiomyocytes

    OpenAIRE

    Shenje, Lincoln T.; Andersen, Peter; Halushka, Marc K.; Lui, Cecillia; Fernandez, Laviel; Collin, Gayle B.; Amat-Alarcon, Nuria; Meschino, Wendy; Cutz, Ernest; Chang, Kenneth; Yonescu, Raluca; Batista, Denise A. S.; Chen, Yan; Chelko, Stephen; Crosson, Jane E.

    2014-01-01

    Cardiomyocyte cell division and replication in mammals proceed through embryonic development and abruptly decline soon after birth. The process governing cardiomyocyte cell cycle arrest is poorly understood. Here we carry out whole exome sequencing in an infant with evidence of persistent postnatal cardiomyocyte replication to determine the genetic risk factors. We identify compound heterozygous ALMS1 mutations in the proband, and confirm their presence in her affected sibling, one copy inher...

  8. Non-homologous end joining pathway is the major route of protection against 4β-hydroxywithanolide E-induced DNA damage in MCF-7 cells.

    Science.gov (United States)

    You, B-J; Wu, Y-C; Lee, C-L; Lee, H-Z

    2014-03-01

    4β-Hydroxywithanolide E is a bioactive withanolide extracted from Physalis peruviana. 4β-Hydroxywithanolide E caused reactive oxygen species production and cell apoptosis in human breast cancer MCF-7 cells. We further found that 4β-hydroxywithanolide E induced DNA damage and regulated the DNA damage signaling in MCF-7 cells. The DNA damage sensors and repair proteins act promptly to remove DNA lesions by 4β-hydroxywithanolide E. The ataxia-telangiectasia mutated protein (ATM)-dependent DNA damage signaling pathway is involved in 4β-hydroxywithanolide E-induced apoptosis of MCF-7 cells. Non-homologous end joining pathway, but not homologous recombination, is the major route of protection of MCF-7 cells against 4β-hydroxywithanolide E-induced DNA damage. 4β-Hydroxywithanolide E had no significant impact on the base excision repair pathway. In this study, we examined the 4β-hydroxywithanolide E-induced DNA damage as a research tool in project investigating the DNA repair signaling in breast cancer cells. We also suggest that 4β-hydroxywithanolide E assert its anti-tumor activity in carcinogenic progression and develop into a dietary chemopreventive agent. Copyright © 2014 Elsevier Ltd. All rights reserved.

  9. Brucella abortus Triggers a cGAS-Independent STING Pathway To Induce Host Protection That Involves Guanylate-Binding Proteins and Inflammasome Activation.

    Science.gov (United States)

    Costa Franco, Miriam M; Marim, Fernanda; Guimarães, Erika S; Assis, Natan R G; Cerqueira, Daiane M; Alves-Silva, Juliana; Harms, Jerome; Splitter, Gary; Smith, Judith; Kanneganti, Thirumala-Devi; de Queiroz, Nina M G P; Gutman, Delia; Barber, Glen N; Oliveira, Sergio C

    2018-01-15

    Immunity against microbes depends on recognition of pathogen-associated molecular patterns by innate receptors. Signaling pathways triggered by Brucella abortus DNA involves TLR9, AIM2, and stimulator of IFN genes (STING). In this study, we observed by microarray analysis that several type I IFN-associated genes, such as IFN-β and guanylate-binding proteins (GBPs), are downregulated in STING knockout (KO) macrophages infected with Brucella or transfected with DNA. Additionally, we determined that STING and cyclic GMP-AMP synthase (cGAS) are important to engage the type I IFN pathway, but only STING is required to induce IL-1β secretion, caspase-1 activation, and GBP2 and GBP3 expression. Furthermore, we determined that STING but not cGAS is critical for host protection against Brucella infection in macrophages and in vivo. This study provides evidence of a cGAS-independent mechanism of STING-mediated protection against an intracellular bacterial infection. Additionally, infected IFN regulatory factor-1 and IFNAR KO macrophages had reduced GBP2 and GBP3 expression and these cells were more permissive to Brucella replication compared with wild-type control macrophages. Because GBPs are critical to target vacuolar bacteria, we determined whether GBP2 and GBP chr3 affect Brucella control in vivo. GBP chr3 but not GBP2 KO mice were more susceptible to bacterial infection, and small interfering RNA treated-macrophages showed reduction in IL-1β secretion and caspase-1 activation. Finally, we also demonstrated that Brucella DNA colocalizes with AIM2, and AIM2 KO mice are less resistant to B. abortus infection. In conclusion, these findings suggest that the STING-dependent type I IFN pathway is critical for the GBP-mediated release of Brucella DNA into the cytosol and subsequent activation of AIM2. Copyright © 2018 by The American Association of Immunologists, Inc.

  10. Qishenyiqi protects ligation-induced left ventricular remodeling by attenuating inflammation and fibrosis via STAT3 and NF-κB signaling pathway.

    Directory of Open Access Journals (Sweden)

    Chun Li

    Full Text Available AIM: Qi-shen-yi-qi (QSYQ, a formula used for the routine treatment of heart failure (HF in China, has been demonstrated to improve cardiac function through down-regulating the activation of the Renin-Angiotensin-Aldosterone System (RAAS. However, the mechanisms governing its therapeutic effects are largely unknown. The present study aims to demonstrate that QSYQ treatment can prevent left ventricular remodeling in heart failure by attenuating oxidative stress and inhabiting inflammation. METHODS: Sprague-Dawley (SD rats were randomly divided into 6 groups: sham group, model group (LAD coronary artery ligation, QSYQ group with high dosage, middle dosage and low dosage (LAD ligation and treated with QSYQ, and captopril group (LAD ligation and treated with captopril as the positive drug. Indicators of fibrosis (Masson, MMPs, and collagens and inflammation factors were detected 28 days after surgery. RESULTS: Results of hemodynamic alterations (dp/dt value in the model group as well as other ventricular remodeling (VR markers, such as MMP-2, MMP-9, collagen I and III elevated compared with sham group. VR was accompanied by activation of RAAS (angiotensin II and NADPHoxidase. Levels of pro-inflammatory cytokines (TNF-α, IL-6 in myocardial tissue were also up-regulated. Treatment of QSYQ improved cardiac remodeling through counter-acting the aforementioned events. The improvement of QSYQ was accompanied with a restoration of angiotensin II-NADPHoxidase-ROS-MMPs pathways. In addition, "therapeutic" QSYQ administration can reduce both TNF-α-NF-B and IL-6-STAT3 pathways, respectively, which further proves the beneficial effects of QSYQ. CONCLUSIONS: Our study demonstrated that QSYQ protected LAD ligation-induced left VR via attenuating AngII -NADPH oxidase pathway and inhabiting inflammation. These findings provide evidence as to the cardiac protective efficacy of QSYQ to HF and explain the beneficial effects of QSYQ in the clinical application for HF.

  11. Caloric restriction protects against electrical kindling of the amygdala by inhibiting the mTOR signaling pathway

    Directory of Open Access Journals (Sweden)

    Bryan Victor Phillips-Farfan

    2015-03-01

    Full Text Available Caloric restriction (CR has been shown to possess antiepileptic properties; however its mechanism of action is poorly understood. CR might inhibit the activity of the mammalian or mechanistic target of rapamycin (mTOR signaling cascade, which seems to participate crucially in the generation of epilepsy. Thus, we investigated the effect of CR on the mTOR pathway and whether CR modified epilepsy generation due to electrical amygdala kindling. The former was studied by analyzing the phosphorylation of adenosine monophosphate-activated protein kinase, protein kinase B and the ribosomal protein S6. The mTOR cascade is regulated by energy and by insulin levels, both of which may be changed by CR; thus we investigated if CR altered the levels of energy substrates in the blood or the level of insulin in plasma. Finally, we studied if CR modified the expression of genes that encode proteins participating in the mTOR pathway. CR increased the after-discharge threshold and tended to reduce the after-discharge duration, indicating an anti-convulsive action. CR diminished the phosphorylation of protein kinase B and ribosomal protein S6, suggesting an inhibition of the mTOR cascade. However, CR did not change glucose, β-hydroxybutyrate or insulin levels; thus the effects of CR were independent from them. Interestingly, CR also did not modify the expression of any investigated gene. The results suggest that the anti-epileptic effect of CR may be partly due to inhibition of the mTOR pathway.

  12. Attenuation of everolimus-induced cytotoxicity by a protective autophagic pathway involving ERK activation in renal cell carcinoma cells

    Science.gov (United States)

    Zeng, Yizhou; Tian, Xiaofang; Wang, Quan; He, Weiyang; Fan, Jing; Gou, Xin

    2018-01-01

    Aim The mammalian target of rapamycin (mTOR) pathway is a critical target for cancer treatment and the mTOR inhibitor everolimus (RAD001) has been approved for treatment of renal cell carcinoma (RCC). However, the limited efficacy of RAD001 has led to the development of drug resistance. Autophagy is closely related to cell survival and death, which may be activated under RAD001 stimulation. The aim of the present study was to identify the underlying mechanisms of RAD001 resistance in RCC cells through cytoprotective autophagy involving activation of the extracellular signal-regulated kinase (ERK) pathway. Methods and results: RAD001 strongly induced autophagy of RCC cells in a dose- and time-dependent manner, as confirmed by Western blot analysis. Importantly, suppression of autophagy by the pharmacological inhibitor chloroquine effectively enhanced RAD001-induced apoptotic cytotoxicity, as demonstrated by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and Western blot analysis, indicating a cytoprotective role for RAD001-induced autophagy. In addition, as was shown by the MTT assay, flow cytometry, and Western blot analysis, RAD001 robustly activated ERK, but not c-Jun N-terminal kinase and p38. Activation of ERK was inhibited by the pharmacological inhibitor selumetinib (AZD6244), which effectively promoted RAD001-induced cell death. Moreover, employing AZD6244 markedly attenuated RAD001-induced autophagy and enhanced RAD001-induced apoptosis, which play a central role in RAD001-induced cell death. Furthermore, RAD001-induced autophagy is regulated by ERK-mediated phosphorylation of Beclin-1 and B-cell lymphoma 2, as confirmed by Western blot analysis. Conclusion These results suggest that RAD001-induced autophagy involves activation of the ERK, which may impair cytotoxicity of RAD001 in RCC cells. Thus, inhibition of the activation of ERK pathway-mediated autophagy may be useful to overcome chemoresistance to RAD001. PMID:29719377

  13. Mutations in Alström protein impair terminal differentiation of cardiomyocytes.

    Science.gov (United States)

    Shenje, Lincoln T; Andersen, Peter; Halushka, Marc K; Lui, Cecillia; Fernandez, Laviel; Collin, Gayle B; Amat-Alarcon, Nuria; Meschino, Wendy; Cutz, Ernest; Chang, Kenneth; Yonescu, Raluca; Batista, Denise A S; Chen, Yan; Chelko, Stephen; Crosson, Jane E; Scheel, Janet; Vricella, Luca; Craig, Brian D; Marosy, Beth A; Mohr, David W; Hetrick, Kurt N; Romm, Jane M; Scott, Alan F; Valle, David; Naggert, Jürgen K; Kwon, Chulan; Doheny, Kimberly F; Judge, Daniel P

    2014-03-04

    Cardiomyocyte cell division and replication in mammals proceed through embryonic development and abruptly decline soon after birth. The process governing cardiomyocyte cell cycle arrest is poorly understood. Here we carry out whole-exome sequencing in an infant with evidence of persistent postnatal cardiomyocyte replication to determine the genetic risk factors. We identify compound heterozygous ALMS1 mutations in the proband, and confirm their presence in her affected sibling, one copy inherited from each heterozygous parent. Next, we recognize homozygous or compound heterozygous truncating mutations in ALMS1 in four other children with high levels of postnatal cardiomyocyte proliferation. Alms1 mRNA knockdown increases multiple markers of proliferation in cardiomyocytes, the percentage of cardiomyocytes in G2/M phases, and the number of cardiomyocytes by 10% in cultured cells. Homozygous Alms1-mutant mice have increased cardiomyocyte proliferation at 2 weeks postnatal compared with wild-type littermates. We conclude that deficiency of Alström protein impairs postnatal cardiomyocyte cell cycle arrest.

  14. Solving the puzzle of pluripotent stem cell-derived cardiomyocyte maturation: piece by piece.

    Science.gov (United States)

    Lundy, David J; Lee, Desy S; Hsieh, Patrick C H

    2017-03-01

    There is a growing need for in vitro models which can serve as platforms for drug screening and basic research. Human adult cardiomyocytes cannot be readily obtained or cultured, and so pluripotent stem cell-derived cardiomyocytes appear to be an attractive option. Unfortunately, these cells are structurally and functionally immature-more comparable to foetal cardiomyocytes than adult. A recent study by Ruan et al ., provides new insights into accelerating the maturation process and takes us a step closer to solving the puzzle of pluripotent stem cell-derived cardiomyocyte maturation.

  15. Protective Effect of Saccharomyces boulardii on Deoxynivalenol-Induced Injury of Porcine Macrophage via Attenuating p38 MAPK Signal Pathway.

    Science.gov (United States)

    Chang, Chao; Wang, Kun; Zhou, Sheng-Nan; Wang, Xue-Dong; Wu, Jin-E

    2017-05-01

    The aims of our study were to evaluate the effects of Saccharomyces boulardii (S. boulardii) on deoxynivalenol (DON)-induced injury in porcine alveolar macrophage cells (PAMCs) and to explore the underlying mechanisms. 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, flow cytometric analysis, ELISA, qRT-PCR, and western blot were performed to assess whether S. boulardii could prevent DON-induced injury by p38 mitogen-activated protein kinase (p38 MAPK) signal pathway. The results showed that pretreatment with 8 μM DON could decrease the viability of PAMC and significantly increase the apoptosis rate of PAMC, whereas S. boulardii could rescue apoptotic PAMC cells induced by DON. Further experiments revealed that S. boulardii effectively reversed DON-induced cytotoxicity via downregulating the expression of TNF-α, IL-6, and IL-lβ. In addition, S. boulardii significantly alleviated DON-induced phosphorylation and mRNA expression of p38 and further increased the expression of apoptosis regulation genes Bcl-xl and Bcl-2 and inhibited the activation of Bax. Our results suggest that S. boulardii could suppress DON-induced p38 MAPK pathway activation and reduce the expression of downstream inflammatory cytokines, as well as promote the expression of anti-apoptotic genes to inhibit apoptosis induced by DON in PAMC.

  16. Ebselen protects mitochondrial function and oxidative stress while inhibiting the mitochondrial apoptosis pathway after acute spinal cord injury.

    Science.gov (United States)

    Jia, Zhi-Qiang; Li, San-Qiang; Qiao, Wei-Qiang; Xu, Wen-Zhong; Xing, Jian-Wu; Liu, Jian-Tao; Song, Hui; Gao, Zhong-Yang; Xing, Bing-Wen; He, Xi-Jing

    2018-05-04

    Ebselen is a fat-soluble small molecule and organic selenium compound that regulates the activity of glutathione peroxidase to alleviate mitochondrial oxidative stress and improve mitochondrial function. In the present study, we aimed to investigate the effects of ebselen on mitochondrial oxidative stress response, mitochondrial apotosis, and motor behaviors after spinal cord injury (SCI). We found that ebselen significantly increased the BBB score in motor behavior, thus suggesting a rescue effect of ebselen on motor function after SCI in rats. Meanwhile, we revealed that ebselen can increase glutathione (GSH) content as well as superoxide dismutase (SOD) and catalase (CAT) activities after SCI-this suggests ebselen has an antioxidant effect. Furthermore, the ATP content and Na + -K + -ATPase activity in mitochondria were increased by ebselen after SCI, while the mitochondrial membrane potential (MMP) was decreased by ebselen. The Cytochrome C and Smac release from mitochondria were reduced by ebselen after SCI, thus indicating improved membrane permeability by ebselen. Moreover, the alterations in caspase-3, Bax and Bcl-2 protein expression, as well as the proportion of cell apoptosis were improved by ebselen treatment, which together suggested that ebselen has an inhibitory effect on mitochondrial apotosis pathways after SCI. Taken together, our results suggest that ebselen can inhibit secondary damage caused by spinal cord injury. Indeed it plays a neuroprotective role in spinal cord injury perhaps by improving mitochondrial function and inhibiting the mitochondrial apoptosis pathway. Copyright © 2018 Elsevier B.V. All rights reserved.

  17. Tetramethylpyrazine analogue CXC195 protects against cerebral ischemia/reperfusion-induced apoptosis through PI3K/Akt/GSK3β pathway in rats.

    Science.gov (United States)

    Chen, Lin; Wei, Xinbing; Hou, Yunfeng; Liu, Xiaoqian; Li, Senpeng; Sun, Baozhu; Liu, Xinyong; Liu, Huiqing

    2014-01-01

    CXC195 showed strongest protective effects among the ligustrazine derivatives in cells and prevented apoptosis induced by H2O2 injury. We recently demonstrated that CXC195 protected against cerebral ischemia/reperfusion (I/R) injury by its antioxidant activity. However, whether the anti-apoptotic action of CXC195 is involved in cerebral I/R injury is unknown. Here, we investigated the role of CXC195 in apoptotic processes induced by cerebral I/R and the possible signaling pathways. Male Wistar rats were submitted to transient middle cerebral artery occlusion for 2h, followed by 24h reperfusion. CXC195 was injected intraperitoneally at 2h and 12h after the onset of ischemia. The number of apoptotic cells was measured by TUNEL assay, apoptosis-related protein cleaved caspase-3, Bcl-2, Bax and the phosphorylation levels of Akt and GSK3β in ischemic penumbra were assayed by western blot. The results showed that administration of CXC195 at the doses of 3mg/kg and 10mg/kg significantly inhibited the apoptosis by decreasing the number of apoptotic cells, decreasing the level of cleaved caspase-3 and Bax, and increasing the level of Bcl-2 in rats subjected to I/R injury. Simultaneously, CXC195 treatment markedly increased the phosphorylation of Akt and GSK3β. Blockade of PI3K activity by wortmannin, dramatically abolished its anti-apoptotic effect and lowered both Akt and GSK3β phosphorylation levels. Our study firstly demonstrated that CXC195 protected against cerebral I/R injury by reducing apoptosis in vivo and PI3K/Akt/GSK3β pathway involved in the anti-apoptotic effect. Crown Copyright © 2014. Published by Elsevier Ltd. All rights reserved.

  18. Carbon monoxide protects against hepatic steatosis in mice by inducing sestrin-2 via the PERK-eIF2α-ATF4 pathway.

    Science.gov (United States)

    Kim, Hyo Jeong; Joe, Yeonsoo; Kim, Seul-Ki; Park, Se-Ung; Park, Jeongmin; Chen, Yingqing; Kim, Jin; Ryu, Jinhyun; Cho, Gyeong Jae; Surh, Young-Joon; Ryter, Stefan W; Kim, Uh-Hyun; Chung, Hun-Taeg

    2017-09-01

    Nonalcoholic fatty liver disease (NAFLD), the hepatic manifestation of the metabolic syndrome, has emerged as one of the most common causes of chronic liver disease in developed countries over the last decade. NAFLD comprises a spectrum of pathological hepatic changes, including steatosis, steatohepatitis, advanced fibrosis, and cirrhosis. Autophagy, a homeostatic process for protein and organelle turnover, is decreased in the liver during the development of NAFLD. Previously, we have shown that carbon monoxide (CO), a reaction product of heme oxygenase (HO) activity, can confer protection in NAFLD, though the molecular mechanisms remain unclear. We therefore investigated the mechanisms underlying the protective effect of CO on methionine/choline-deficient (MCD) diet-induced hepatic steatosis. We found that CO induced sestrin-2 (SESN2) expression through enhanced mitochondrial ROS production and protected against MCD-induced NAFLD progression through activation of autophagy. SESN2 expression was increased by CO or CO-releasing molecule (CORM2), in a manner dependent on signaling through the protein kinase R-like endoplasmic reticulum kinase (PERK), eukaryotic initiation factor-2 alpha (eIF2α)/ activating transcription factor-4 (ATF4)-dependent pathway. CO-induced SESN2 upregulation in hepatocytes contributed to autophagy induction through activation of 5'-AMP-activated protein kinase (AMPK) and inhibition of mechanistic target of rapamycin (mTOR) complex I (mTORC1). Furthermore, we demonstrate that CO significantly induced the expression of SESN2 and enhanced autophagy in the livers of MCD-fed mice or in MCD-media treated hepatocytes. Conversely, knockdown of SESN2 abrogated autophagy activation and mTOR inhibition in response to CO. We conclude that CO ameliorates hepatic steatosis through the autophagy pathway induced by SESN2 upregulation. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

  19. ZL006 protects spinal cord neurons against ischemia-induced oxidative stress through AMPK-PGC-1α-Sirt3 pathway.

    Science.gov (United States)

    Liu, Shu-Guang; Wang, Yun-Mei; Zhang, Yan-Jun; He, Xi-Jing; Ma, Tao; Song, Wei; Zhang, Yu-Min

    2017-09-01

    Spinal cord ischemia (SCI) induces a range of cellular and molecular cascades, including activation of glutamate receptors and downstream signaling. Post-synaptic density protein 95 (PSD-95) links neuronal nitric oxide synthase (nNOS) with the N-methyl-d-aspartic acid (NMDA) receptors to form a ternary complex in the CNS. This molecular complex-mediated cytotoxicity has been implicated in brain ischemia, but its role in SCI has not been determined. The goal of the study was to investigate the potential protective effects of ZL006, a small-molecule inhibitor of the PSD-95/nNOS interaction, in an in vitro SCI model induced by oxygen and glucose deprivation (OGD) in cultured spinal cord neurons. We found that ZL006 reduced OGD-induced lactate dehydrogenase (LDH) release, neuronal apoptosis and loss of cell viability. This protection was accompanied by the preservation of mitochondrial function, as evidenced by reduced mitochondrial oxidative stress, attenuated mitochondrial membrane potential (MMP) loss, and enhanced ATP generation. In addition, ZL006 stimulated mitochondrial enzyme activities and SOD2 deacetylation in a Sirt3-dependent manner. The results of western blot analysis showed that ZL006 increased the activation of AMPK-PGC-1α-Sirt3 pathway, and the beneficial effects of ZL006 was partially abolished by AMPK inhibitor and PGC-1α knockdown. Therefore, our present data showed that, by the AMPK-PGC-1α-Sirt3 pathway, ZL006 protects spinal cord neurons against ischemia through reducing mitochondrial oxidative stress to prevent apoptosis. Copyright © 2017 Elsevier Ltd. All rights reserved.

  20. The Extracts and Major Compounds Derived from Astragali Radix Alter Mitochondrial Bioenergetics in Cultured Cardiomyocytes: Comparison of Various Polar Solvents and Compounds

    Directory of Open Access Journals (Sweden)

    Yun Huang

    2018-05-01

    Full Text Available Astragali Radix (AR is a widely used “Qi-invigorating” herb in China for its tonic effects in strengthening biological tissues. The extract of AR contains abundant antioxidants, including astragalosides and isoflavonoids. However, very few reports have systematically measured the effects of the major components of AR on cell mitochondrial bioenergetics. Here, a systemic approach employing an extracellular flux analyzer was developed to evaluate mitochondrial respiration in cultured cardiomyocyte cells H9C2. The effects of different polar extractives, as well as of the major compounds of AR, were compared. The contents of astragaloside IV, calycosin, formononetin, and genistein in the AR extracts obtained by using water, 50% ethanol, and 90% ethanol were measured by liquid chromatograph-mass spectrometer (LC–MS. The antioxidant activities of the AR extracts, as well as of their major compounds, were determined by measuring the free radical scavenging activity and protective effects in tert-butyl hydroperoxide (tBHP-treated H9C2 cells. By monitoring the real-time oxygen consumption rate (OCR in tBHP-treated cardiomyocytes with a Seahorse extracellular flux analyzer, the tonic effects of the AR extracts and of their main compounds on mitochondrial bioenergetics were evaluated. AR water extracts possessed the strongest antioxidant activity and protective effects in cardiomyocytes exposed to oxidative stress. The protection was proposed to be mediated via increasing the spare respiratory capacity and mitochondrial ATP production in the stressed cells. The major compounds of AR, astragaloside IV and genistein, showed opposite effects in regulating mitochondrial bioenergetics. These results demonstrate that highly polar extracts of AR, especially astragaloside-enriched extracts, possess better tonic effects on mitochondrial bioenergetics of cultured cardiomyocytes than extracts with a lower polarity.

  1. Central estrogenic pathways protect against the depressant action of acute nicotine on reflex tachycardia in female rats

    International Nuclear Information System (INIS)

    El-Mas, Mahmoud M.; Fouda, Mohamed A.; El-gowilly, Sahar M.; Saad, Evan I.

    2012-01-01

    We have previously shown that acute exposure of male rats to nicotine preferentially attenuates baroreceptor-mediated control of reflex tachycardia in contrast to no effect on reflex bradycardia. Here, we investigated whether female rats are as sensitive as their male counterparts to the baroreflex depressant effect of nicotine and whether this interaction is modulated by estrogen. Baroreflex curves relating reflex chronotropic responses evoked by i.v. doses (1–16 μg/kg) of phenylephrine (PE) or sodium nitroprusside (SNP), were constructed in conscious freely moving proestrus, ovariectomized (OVX), and estrogen (50 μg/kg/day s.c., 5 days)-replaced OVX (OVXE 2 ) rats. Slopes of the curves were taken as a measure of baroreflex sensitivity (BRS PE and BRS SNP ). Nicotine (100 μg/kg i.v.) reduced BRS SNP in OVX rats but not in proestrus or OVXE 2 rats. The attenuation of reflex tachycardia by nicotine was also evident in diestrus rats, which exhibited plasma estrogen levels similar to those of OVX rats. BRS PE was not affected by nicotine in all rat preparations. Experiments were then extended to determine whether central estrogenic receptors modulate the nicotine–BRS SNP interaction. Intracisteral (i.c.) treatment of OVX rats with estrogen sulfate (0.2 μg/rat) abolished the BRS SNP attenuating effect of i.v. nicotine. This protective effect of estrogen disappeared when OVX rats were pretreated with i.c. ICI 182,780 (50 μg/rat, selective estrogen receptor antagonist). Together, these findings suggest that central neural pools of estrogen receptors underlie the protection offered by E 2 against nicotine-induced baroreceptor dysfunction in female rats. -- Highlights: ► Estrogen protects against the depressant effect of nicotine on reflex tachycardia. ► The baroreflex response and estrogen status affect the nicotine–BRS interaction. ► The protection offered by estrogen is mediated via central estrogen receptors.

  2. Protective Efficacy of Vitamins C and E on p,p′-DDT-Induced Cytotoxicity via the ROS-Mediated Mitochondrial Pathway and NF-κB/FasL Pathway

    Science.gov (United States)

    Jin, Xiaoting; Song, Li; Liu, Xiangyuan; Chen, Meilan; Li, Zhuoyu; Cheng, Long; Ren, Hua

    2014-01-01

    Dichlorodiphenoxytrichloroethane (DDT) is a known persistent organic pollutant and liver damage toxicant. However, there has been little emphasis on the mechanism underlying liver damage toxicity of DDT and the relevant effective inhibitors. Hence, the present study was conducted to explore the protective effects of vitamin C (VC) and vitamin E (VE) on the cytotoxicity of DDT in HL-7702 cells and elaborate the specific molecular mechanisms. The results demonstrated that p,p′-DDT exposure at over 10 µM depleted cell viability of HL-7702 cells and led to cell apoptotic. p,p′-DDT treatment elevated the level of reactive oxygen species (ROS) generation, induced mitochondrial membrane potential, and released cytochrome c into the cytosol, with subsequent elevations of Bax and p53, along with suppression of Bcl-2. In addition, the activations of caspase-3 and -8 were triggered. Furthermore, p,p′-DDT promoted the expressions of NF-κB and FasL. When the cells were exposed to the NF-κB inhibitor (PDTC), the up-regulated expression of FasL was attenuated. Strikingly, these alterations caused by DDT treatment were prevented or reversed by the addition of VC or VE, and the protective effects of co-treatment with VC and VE were higher than the single supplement with p,p′-DDT. Taken together, these findings provide novel experimental evidences supporting that VC or/and VE could reduce p,p′-DDT-induced cytotoxicity of HL-7702 cells via the ROS-mediated mitochondrial pathway and NF-κB/FasL pathway. PMID:25464339

  3. Overexpression of Cardiac-Specific Kinase TNNI3K Promotes Mouse Embryonic Stem Cells Differentiation into Cardiomyocytes.

    Science.gov (United States)

    Wang, Yin; Wang, Shi-Qiang; Wang, Li-Peng; Yao, Yu-Hong; Ma, Chun-Yan; Ding, Jin-Feng; Ye, Jue; Meng, Xian-Min; Li, Jian-Jun; Xu, Rui-Xia

    2017-01-01

    Backgroud/Aims: The biological function of cardiac troponin I-interacting kinase (TNNI3K), a cardiac-specific functional kinase, is largely unknown. We investigated the effect of human TNNI3K (hTNNI3K) on the differentiation of mouse embryonic stem cells (mESCs) into cardiomyocytes. First, the time-space expression of endogenous Tnni3k was detected by real-time polymerase chain reaction (PCR) and western blotting at 16 different time-points over a period of 28 days. Further, action potentials and calcium current with/without 5 µM nifedipine were measured by patch clamp for mESC-derived cardiomyocytes. HTNNI3K and mouse-derived siRNA were transfected into mESC using lentivirus vector to induce hTNNI3K overexpression and knock-down, respectively. The number of troponin-T (cTnT) positive cells was greater in the group with TNNI3K overexpression as compared to that in control group, while less such cells were detected in the mTnni3k knock-down group as evaluated on flow cytometry (FCM) and ImageXpress Micro system. After upregulation of connexin43, cardiac troponin-I (Ctni), Ctni, Gata4 were detected in mESCs with TNNI3K overexpression; however, overexpression of α-Actinin and Mlc2v was not detected. Interestingly, Ctnt, connexin40 and connexin45, the markers of ventricular, atrial, and pacemaker cells, respectively, were detected in by real-time PCR in TNNI3K overexpression group. our study indicated that TNNI3K overexpression promoted mESC differentiating into beating cardiomyocytes and induced up-regulating expression of cTnT by PKCε signal pathway, which suggested a modulation of TNNI3K activity as a potential therapeutic approach for ischemic cardiac disease. © 2017 The Author(s) Published by S. Karger AG, Basel.

  4. Pentazocine Protects SN4741 Cells Against MPP+-Induced Cell Damage via Up-Regulation of the Canonical Wnt/β-Catenin Signaling Pathway

    Directory of Open Access Journals (Sweden)

    Jiancai Wang

    2017-06-01

    Full Text Available The Wnt/β-catenin signaling pathway has been linked to many neurodegenerative diseases including Parkinson’s disease (PD. A glycoprotein named Dickkopf-1 (Dkk1 can combine with the receptor complex on cell membrane to inhibit Wnt/β-catenin signaling. Opioids, a series of compounds including morphine, fentanyl and pentazocine, have been reported to contribute to the up-regulation of Wnt/β-catenin signaling. Naloxone is an antagonist that has been used as an antidote to opioids through mu-opioid receptor. 1-methyl-4-phenylpyridinium (MPP+, which serves as a selective toxin for dopaminergic neurons, has been used to create experimental models of PD. In our study, we examined the protective effects of pentazocine against MPP+-induced cell death in the nigral dopaminergic cell line, SN4741 and tried to elucidate the molecular mechanisms underlying such protective effects. The data showed that pretreatment with pentazocine significantly rescued the SN4741 cell against MPP+. Moreover, the MPP+-exposed SN4741 cells exhibited a down-regulation of β-catenin, which could be restored by treatment with pentazocine. However, Dkk1 but not naloxonewas associated with the abrogation of protective effect of pentazocine. These results suggest that pentazocine alleviates MPP+-induced SN4741 cells apoptosis via the up-regulation of canonical Wnt/β-catenin signaling.

  5. Protective Effects of Green Tea Polyphenol Against Renal Injury Through ROS-Mediated JNK-MAPK Pathway in Lead Exposed Rats.

    Science.gov (United States)

    Wang, Haidong; Li, Deyuan; Hu, Zhongze; Zhao, Siming; Zheng, Zhejun; Li, Wei

    2016-06-30

    To investigate the potential therapeutic effects of polyphenols in treating Pb induced renal dysfunction and intoxication and to explore the detailed underlying mechanisms. Wistar rats were divided into four groups: control groups (CT), Pb exposure groups (Pb), Pb plus Polyphenols groups (Pb+PP) and Polyphenols groups (PP). Animals were kept for 60 days and sacrificed for tests of urea, serum blood urea nitrogen (BUN) and creatinine. Histological evaluations were then performed. In vitro studies were performed using primary kidney mesangial cells to reveal detailed mechanisms. Cell counting kit-8 (CCK-8) was used to evaluate cell viability. Pb induced cell apoptosis was measured by flow cytometry. Reactive oxygen species (ROS) generation and scavenging were tested by DCFH-DA. Expression level of tumor necrosis factor-α (TNF-α), interleukin-1-β (IL-1-β) and IL-6 were assayed by ELISA. Western blot and qPCR were used to measure the expression of ERK1/2, JNK1/2 and p38. Polyphenols have obvious protective effects on Pb induced renal dysfunction and intoxication both in vivo and in vitro. Polyphenols reduced Pb concentration and accumulation in kidney. Polyphenols also protected kidney mesangial cells from Pb induced apoptosis. Polyphenols scavenged Pb induced ROS generation and suppressed ROS-mediated ERK/JNK/p38 pathway. Downstream pro-inflammatory cytokines were inhibited in consistency. Polyphenol is protective in Pb induced renal intoxication and inflammatory responses. The underlying mechanisms lie on the antioxidant activity and ROS scavenging activity of polyphenols.

  6. Hsp20 Protects against Oxygen-Glucose Deprivation/Reperfusion-Induced Golgi Fragmentation and Apoptosis through Fas/FasL Pathway

    Directory of Open Access Journals (Sweden)

    Bingwu Zhong

    2015-01-01

    Full Text Available Cerebral ischemia-reperfusion injury plays an important role in the development of tissue injury after acute ischemic stroke. Finding effective neuroprotective agents has become a priority in the treatment of ischemic stroke. The Golgi apparatus (GA is a pivotal organelle and its protection is an attractive target in the treatment of cerebral ischemia-reperfusion injury. Protective effects of Hsp20, a potential cytoprotective agent due to its chaperone-like activity and involvement in regulation of many vital processes, on GA were assessed in an ischemia-reperfusion injury model. Mouse neuroblastoma Neuro2a (N2a cells were subjected to oxygen-glucose deprivation/reperfusion (OGDR insult. OGDR induces Golgi fragmentation, apoptosis, and p115 cleavage in N2a cells. However, transfection with Hsp20 significantly attenuates OGDR-induced Golgi fragmentation and apoptosis. Hsp20 interacts with Bax, decreases FasL and Bax expression, and inhibits caspases 3 and p115 cleavage in N2a cells exposed to OGDR. Our data demonstrate that increased Hsp20 expression protects against OGDR-induced Golgi fragmentation and apoptosis, likely through interaction with Bax and subsequent amelioration of the OGDR-induced elevation in p115 cleavage via the Fas/FasL signaling pathway. This neuroprotective potential of Hsp20 against OGDR insult and the underlying mechanism will pave the way for its potential clinical application for cerebral ischemia-reperfusion related disorders.

  7. Blockade of Inflammation and Apoptosis Pathways by siRNA Prolongs Cold Preservation Time and Protects Donor Hearts in a Porcine Model

    Directory of Open Access Journals (Sweden)

    Jia Wei

    2017-12-01

    Full Text Available In donor hearts from mini pigs, overtime cold preservation and ischemia-reperfusion injury cause poor graft quality and impaired heart function. Blockage of complement, apoptosis, and inflammation is considered a strategy for attenuating ischemia-reperfusion injury and protecting cardiac function. Minipig donor hearts were perfused and preserved in Celsior solution or transfection reagent containing Celsior solution with scramble siRNA or siRNAs targeting complement 3, caspase-8, caspase-3, and nuclear factor κB-p65 genes at 4°C and subsequently hemo-reperfused ex vivo (38°C or transplanted into recipients. The protective effect of the siRNA solution was evaluated by measuring cell apoptosis, structural alteration, protein markers for tissue damage and oxidative stress, and cardiac function. We found a reduction in cell apoptosis, myocardial damage, and tissue inflammation by reduced biochemistry and markers and protein expression of proinflammatory cytokines and improvement in cardiac function, as shown by the improved hemodynamic indices in 12-hr-preserved siRNA-treated hearts of both ex vivo and orthotopic transplantation models. These findings demonstrate that blockade of inflammation and apoptosis pathways using siRNA can prolong cold preservation time and better protect donor heart function in cardiac transplantation of large animals, which may be beneficial for human heart preservation.

  8. ACE variants interact with the RAS pathway to confer risk and protection against type 2 diabetic nephropathy

    DEFF Research Database (Denmark)

    Ahluwalia, Tarun Veer Singh; Ahuja, Monica; Rai, Taranjit Singh

    2009-01-01

    Genetic predisposition has been proposed to be a major determinant in the development of renal complications of diabetes. Among candidate genes examined for susceptibility to diabetic nephropathy, angiotensin-converting enzyme (ACE) gene has been found to be associated with pathogenesis......, in the present study, we evaluated the association of ACE haplotypes and the interactions of ACE, angiotensinogen (AGT), and angiotensin II receptor type I (AGTR1) gene polymorphisms with DNP in Asian Indians. We genotyped seven variants of the RAS pathway genes (ACE, AGT, and AGTR1) in type 2 diabetic cohorts...... and progression of diabetic nephropathy. However, the role of other renin-angiotensin system (RAS) polymorphisms and their possible interactions with different ACE I/D genotypes are less clearly defined. Recent studies also show that ACE haplotypes may be better predictors to disease susceptibility. Thus...

  9. Long-pulse gastric electrical stimulation protects interstitial cells of Cajal in diabetic rats via IGF-1 signaling pathway.

    Science.gov (United States)

    Li, Hai; Chen, Yan; Liu, Shi; Hou, Xiao-Hua

    2016-06-21

    To investigate the effects of different parameters of gastric electrical stimulation (GES) on interstitial cells of Cajal (ICCs) and changes in the insulin-like growth factor 1 (IGF-1) signal pathway in streptozotocin-induced diabetic rats. Male rats were randomized into control, diabetic (DM), diabetic with sham GES (DM + SGES), diabetic with GES1 (5.5 cpm, 100 ms, 4 mA) (DM + GES1), diabetic with GES2 (5.5 cpm, 300 ms, 4 mA) (DM + GES2) and diabetic with GES3 (5.5 cpm, 550 ms, 2 mA) (DM + GES3) groups. The expression levels of c-kit, M-SCF and IGF-1 receptors were evaluated in the gastric antrum using Western blot analysis. The distribution of ICCs was observed using immunolabeling for c-kit, while smooth muscle cells and IGF-1 receptors were identified using α-SMA and IGF-1R antibodies. Serum level of IGF-1 was tested using enzyme-linked immunosorbent assay. Gastric emptying was delayed in the DM group but improved in all GES groups, especially in the GES2 group. The expression levels of c-kit, M-SCF and IGF-1R were decreased in the DM group but increased in all GES groups. More ICCs (c-kit(+)) and smooth muscle cells (α-SMA(+)/IGF-1R(+)) were observed in all GES groups than in the DM group. The average level of IGF-1 in the DM group was markedly decreased, but it was up-regulated in all GES groups, especially in the GES2 group. The results suggest that long-pulse GES promotes the regeneration of ICCs. The IGF-1 signaling pathway might be involved in the mechanism underlying this process, which results in improved gastric emptying.

  10. Central estrogenic pathways protect against the depressant action of acute nicotine on reflex tachycardia in female rats

    Energy Technology Data Exchange (ETDEWEB)

    El-Mas, Mahmoud M., E-mail: mahelm@hotmail.com; Fouda, Mohamed A.; El-gowilly, Sahar M.; Saad, Evan I.

    2012-02-01

    We have previously shown that acute exposure of male rats to nicotine preferentially attenuates baroreceptor-mediated control of reflex tachycardia in contrast to no effect on reflex bradycardia. Here, we investigated whether female rats are as sensitive as their male counterparts to the baroreflex depressant effect of nicotine and whether this interaction is modulated by estrogen. Baroreflex curves relating reflex chronotropic responses evoked by i.v. doses (1–16 μg/kg) of phenylephrine (PE) or sodium nitroprusside (SNP), were constructed in conscious freely moving proestrus, ovariectomized (OVX), and estrogen (50 μg/kg/day s.c., 5 days)-replaced OVX (OVXE{sub 2}) rats. Slopes of the curves were taken as a measure of baroreflex sensitivity (BRS{sub PE} and BRS{sub SNP}). Nicotine (100 μg/kg i.v.) reduced BRS{sub SNP} in OVX rats but not in proestrus or OVXE{sub 2} rats. The attenuation of reflex tachycardia by nicotine was also evident in diestrus rats, which exhibited plasma estrogen levels similar to those of OVX rats. BRS{sub PE} was not affected by nicotine in all rat preparations. Experiments were then extended to determine whether central estrogenic receptors modulate the nicotine–BRS{sub SNP} interaction. Intracisteral (i.c.) treatment of OVX rats with estrogen sulfate (0.2 μg/rat) abolished the BRS{sub SNP} attenuating effect of i.v. nicotine. This protective effect of estrogen disappeared when OVX rats were pretreated with i.c. ICI 182,780 (50 μg/rat, selective estrogen receptor antagonist). Together, these findings suggest that central neural pools of estrogen receptors underlie the protection offered by E{sub 2} against nicotine-induced baroreceptor dysfunction in female rats. -- Highlights: ► Estrogen protects against the depressant effect of nicotine on reflex tachycardia. ► The baroreflex response and estrogen status affect the nicotine–BRS interaction. ► The protection offered by estrogen is mediated via central estrogen receptors.

  11. Salidroside Protection Against Oxidative Stress Injury Through the Wnt/β-Catenin Signaling Pathway in Rats with Parkinson’s Disease

    Directory of Open Access Journals (Sweden)

    Dong-Mei Wu

    2018-04-01

    Full Text Available Background/Aims: Parkinson’s disease (PD is the second most common neurodegenerative disease after Alzheimer’s disease, and recent studies suggested that oxidative stress (OS contributes to the cascade that leads to dopamine cell degeneration in PD. In this study, we hypothesized that salidroside (SDS offers protection against OS injury in 6-hydroxydopamine (6-OHDA unilaterally lesioned rats as well as the underlying mechanism. Methods: SDS and LiCl (activators of the Wnt/β-catenin signaling pathway administration alone and in combination with 6-OHDA injection in rats was performed 3 days before modeling for 17 consecutive days to verify the regulatory mechanism by which SDS affects the Wnt/β-catenin signaling pathway as well as to evaluate the protective effect of SDS on PD in relation to OS in vivo. In addition, pheochromocytoma 12 (PC12 cells were incubated with 10 µmol/L SDS or LiCl alone or with both in combination for 1 h followed by a 24-h incubation with 100 µmol/L 6-OHDA to obtain in vitro data. Results: In vivo the administration of LiCl was found to ameliorate behavioral deficits and dopaminergic neuron loss; increase superoxide dismutase (SOA activity, glutathione peroxidase (GSH-Px levels, and glycogen synthase kinase 3β phosphorylation (GSK-3β-Ser9; reduce malondialdehyde (MDA accumulation in the striatum and the GSK-3β mRNA level; as well as elevate β-catenin and cyclinD1 mRNA and protein levels in 6-OHDA-injected rats. This SDS treatment regimen was found to strengthen the beneficial effect of LiCl on 6-OHDA-injected rats. In vitro LiCl treatment decreased the toxicity of 6-OHDA on PC12 cells and prevented apoptosis. Additionally, LiCl treatment increased SOA activity, GSH-Px levels, and GSK-3β-Ser9 phosphorylation; decreased MDA accumulation in the striatum and GSK-3β mRNA levels; as well as increased β-catenin and cyclinD1 mRNA and protein levels in 6-OHDA-treated PC12 cells. Additionally, SDS treatment increased

  12. Predictive model identifies key network regulators of cardiomyocyte mechano-signaling.

    Directory of Open Access Journals (Sweden)

    Philip M Tan

    2017-11-01

    Full Text Available Mechanical strain is a potent stimulus for growth and remodeling in cells. Although many pathways have been implicated in stretch-induced remodeling, the control structures by which signals from distinct mechano-sensors are integrated to modulate hypertrophy and gene expression in cardiomyocytes remain unclear. Here, we constructed and validated a predictive computational model of the cardiac mechano-signaling network in order to elucidate the mechanisms underlying signal integration. The model identifies calcium, actin, Ras, Raf1, PI3K, and JAK as key regulators of cardiac mechano-signaling and characterizes crosstalk logic imparting differential control of transcription by AT1R, integrins, and calcium channels. We find that while these regulators maintain mostly independent control over distinct groups of transcription factors, synergy between multiple pathways is necessary to activate all the transcription factors necessary for gene transcription and hypertrophy. We also identify a PKG-dependent mechanism by which valsartan/sacubitril, a combination drug recently approved for treating heart failure, inhibits stretch-induced hypertrophy, and predict further efficacious pairs of drug targets in the network through a network-wide combinatorial search.

  13. Preclinical renal chemo-protective potential of Prunus amygdalus Batsch seed coat via alteration of multiple molecular pathways.

    Science.gov (United States)

    Pandey, Preeti; Bhatt, Prakash Chandra; Rahman, Mahfoozur; Patel, Dinesh Kumar; Anwar, Firoz; Al-Abbasi, Fahad; Verma, Amita; Kumar, Vikas

    2018-02-01

    Prunus amygdalus Batsch (almond) is a classical nutritive traditional Indian medicine. Along with nutritive with anti-oxidant properties, it is, clinically, used in the treatment of various diseases with underlying anti-oxidant mechanism. This study is an effort to scrutinise the renal protective effect of P. amygdalus Batsch or green almond (GA) seed coat extract and its underlying mechanism in animal model of Ferric nitrilotriacetate (Fe-NTA) induced renal cell carcinoma (RCC). RCC was induced in Swiss Albino Wistar rats by intraperitoneal injection of Fe-NTA. The rats were then treated with ethanolic extract of GA (25, 50 and 100 mg/kg per oral) for 22 weeks. Efficacy of GA administration was evaluated by change in biochemical, renal, macroscopical and histopathological parameters and alterations. Additionally, interleukin-6 (IL-6), tumour necrosis factor-α (TNF-α), interleukin-1β (IL-1β) and inflammatory mediator including prostaglandin E2 (PGE 2 ), nuclear factor-kappa B (NF-κB) were also observed to explore the possible mechanisms. The oral administration of GA significantly (p Bowman capsules and inflammatory cells. Hence, it can be concluded that GA possesses observable chemo-protective action and effect on Fe-NTA induced RCC via dual inhibition mechanism one by inhibiting free radical generation and second by inhibiting inflammation.

  14. Shikonin protects dopaminergic cell line PC12 against 6-hydroxydopamine-mediated neurotoxicity via both glutathione-dependent and independent pathways and by inhibiting apoptosis.

    Science.gov (United States)

    Esmaeilzadeh, Emran; Gardaneh, Mossa; Gharib, Ehsan; Sabouni, Farzaneh

    2013-08-01

    We have investigated the mechanism of shikonin function on protection of dopaminergic neurons against 6-OHDA-induced neurotoxicity. Treatment of rat pheochromocytoma cell line PC12 by serial dilutions of shikonin determined 10 μM of the compound as its optimum concentration for protection saving nearly 70 % of the cells against toxicity. Reverse transcription-PCR analysis of shikonin-treated cells showed threefold increase in mRNA levels of glutathione peroxidase-1 (GPX-1) as a representative component of the intracellular anti-oxidant defense system. To elucidate shikonin-GPX1 relationships and maximize protection, we transduced PC12 cells using recombinant lentivirus vectors that harbored GPX-1 coding sequence. This change upregulated GPX-1 expression, increased peroxidase activity and made neuronal cells resistant to 6-OHDA-mediated toxicity. More importantly, addition of shikonin to GPX1-overexpressing PC12 cells augmented GPX-1 protein content by eightfold leading to fivefold increase of enzymatic activity, 91 % cell survival against neurotoxicity and concomitant increases in intracellular glutathione (GSH) levels. Depletion of intracellular GSH rendered all cell groups highly susceptible to toxicity; however, shikonin was capable of partially saving them. Subsequently, GSH-independent superoxide dismutase mRNA was found upregulated by shikonin. As signs of apoptosis inhibition, the compound upregulated Bcl-2, downregulated Bax, and prevented cell nuclei from undergoing morphological changes typical of apoptosis. Also, a co-staining method demonstrated GPX-1 overexpression significantly increases the percent of live cells that is maximized by shikonin treatment. Our data indicate that shikonin as an antioxidant compound protects dopaminergic neurons against 6-OHDA toxicity and enhances their survival via both glutathione-dependent and direct anti-apoptotic pathways.

  15. Protective effect of lycopene on fluoride-induced ameloblasts apoptosis and dental fluorosis through oxidative stress-mediated Caspase pathways.

    Science.gov (United States)

    Li, Weishan; Jiang, Binghua; Cao, Xianglin; Xie, Yongjiang; Huang, Ting

    2017-01-05

    Fluoride is an environmental toxicant and induces dental fluorosis and oxidative stress. Lycopene (LYC) is an effective antioxidant that is reported to attenuate fluoride toxicity. To determine the effects of LYC on sodium fluoride (NaF) -induced teeth and ameloblasts toxicity, rats were treated with NaF (10 mg/kg) and/or LYC (10 mg/kg) by orally administration for 5 weeks; ameloblasts were treated with NaF (5 mM) and/or LYC (2 μM) for 6 h. We found that the concentrations of fluoride, malondialdehyde (MDA) and reactive oxygen species (ROS), gene expressions and activities of Caspase-9 and Caspase-3, and the gene expressions of Bax were significantly decreased, while the activities of superoxide dismutase (SOD) and glutathione peroxidase (GPX), the gene expression of Bcl-2 were significantly increased in the LYC + NaF-treated rats group; concentrations of MDA and ROS, gene expressions and activities of Caspase-9 and Caspase-3, and the gene expression of Bax, and ameloblasts apoptosis rate were significantly decreased, while the activities of SOD and GPX, the gene expression of Bcl-2 were significantly increased in the LYC + NaF-treated ameloblasts group. These results suggest that LYC significantly combated NaF-induced ameloblasts apoptosis and dental fluorosis by attenuation oxidative stress and down-regulation Caspase pathway. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  16. Protection against 2-chloroethyl ethyl sulfide (CEES) - induced cytotoxicity in human keratinocytes by an inducer of the glutathione detoxification pathway

    International Nuclear Information System (INIS)

    Abel, Erika L.; Bubel, Jennifer D.; Simper, Melissa S.; Powell, Leslie; McClellan, S. Alex; Andreeff, Michael; MacLeod, Michael C.; DiGiovanni, John

    2011-01-01

    Sulfur mustard (SM or mustard gas) was first used as a chemical warfare agent almost 100 years ago. Due to its toxic effects on the eyes, lungs, and skin, and the relative ease with which it may be synthesized, mustard gas remains a potential chemical threat to the present day. SM exposed skin develops fluid filled bullae resulting from potent cytotoxicity of cells lining the basement membrane of the epidermis. Currently, there are no antidotes for SM exposure; therefore, chemopreventive measures for first responders following an SM attack are needed. Glutathione (GSH) is known to have a protective effect against SM toxicity, and detoxification of SM is believed to occur, in part, via GSH conjugation. Therefore, we screened 6 potential chemopreventive agents for ability to induce GSH synthesis and protect cultured human keratinocytes against the SM analog, 2-chloroethyl ethyl sulfide (CEES). Using NCTC2544 human keratinocytes, we found that both sulforaphane and methyl-2-cyano-3,12-dioxooleana-1,9-dien-28-oate (CDDO-Me) stimulated nuclear localization of Nrf2 and induced expression of the GSH synthesis gene, GCLM. Additionally, we found that treatment with CDDO-Me elevated reduced GSH content of NCTC2544 cells and preserved their viability by ∼ 3-fold following exposure to CEES. Our data also suggested that CDDO-Me may act additively with 2,6-dithiopurine (DTP), a nucleophilic scavenging agent, to increase the viability of keratinocytes exposed to CEES. These results suggest that CDDO-Me is a promising chemopreventive agent for SM toxicity in the skin. - Highlights: → CDDO-Me treatment increased intracellular GSH in human keratinocytes. → CDDO-Me increased cell viability following exposure to the half-mustard, CEES. → The cytoprotective effect of CDDO-Me was likely due to scavenging with endogenous GSH.

  17. Amentoflavone protects dopaminergic neurons in MPTP-induced Parkinson's disease model mice through PI3K/Akt and ERK signaling pathways

    Energy Technology Data Exchange (ETDEWEB)

    Cao, Qin; Qin, Liyue; Huang, Fei, E-mail: Fei_H@hotmail.com; Wang, Xiaoshuang; Yang, Liu; Shi, Hailian; Wu, Hui; Zhang, Beibei; Chen, Ziyu; Wu, Xiaojun, E-mail: xiaojunwu320@126.com

    2017-03-15

    Parkinson's disease (PD) is characterized by the progressive degeneration of dopaminergic neurons in substantia nigra pars compacta (SNpc). Mitochondrial dysfunction and cell apoptosis are suggested to be actively involved in the pathogenesis of PD. In the present study, the neuroprotective effect of amentoflavone (AF), a naturally occurring biflavonoid from Selaginella tamariscina, was examined in PD models both in vitro and in vivo. On SH-SY5Y cells, AF treatment dose-dependently reduced 1-methyl-4-phenylpyridinium (MPP{sup +})-induced nuclear condensation and loss of cell viability without obvious cytotoxicity. It inhibited the activation of caspase-3 and p21 but increased the Bcl-2/Bax ratio. Further study disclosed that AF enhanced the phosphorylation of PI3K, Akt and ERK1/2 down-regulated by MPP{sup +} in SH-SY5Y cells, the effect of which could be blocked by LY294002, the inhibitor of PI3K. Consistently, AF alleviated the behavioral deterioration in pole and traction tests and rescued the loss of dopaminergic neurons in SNpc and fibers in striatum in methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) induced mice. It also could enhance the activation of PI3K and Akt as well as Bcl-2/Bax ratio in SN. Moreover, AF alleviated gliosis as well as the gene expression levels of IL-1β and iNOS in SN. Collectively, these results suggested that AF protected dopaminergic neurons against MPTP/MPP{sup +}-induced neurotoxicity, which might be mediated through activation of PI3K/Akt and ERK signaling pathways in dopaminergic neurons and attenuation of neuroinflammation. - Highlights: • AF protected dopaminergic neurons against MPTP/MPP{sup +}-induced neurotoxicity. • AF modulated PI3K/Akt and ERK signaling pathways. • AF could alleviate neuroinflammation in SN.

  18. Protective properties of sesamin against fluoride-induced oxidative stress and apoptosis in kidney of carp (Cyprinus carpio) via JNK signaling pathway.

    Science.gov (United States)

    Cao, Jinling; Chen, Jianjie; Xie, Lingtian; Wang, Jundong; Feng, Cuiping; Song, Jing

    2015-10-01

    Sesamin, a major lignan derived from sesame seeds, has been reported to have many benefits and medicinal properties. However, its protective effects against fluoride-induced injury in kidney of fish have not been clarified. Previously we found that fluoride exposure caused damage and apoptosis in the kidneys of the common carp, Cyprinus carpio. In this study, the effects of sesamin on renal oxidative stress and apoptosis in fluoride-exposed fish were determined. The results showed that sesamin alleviated significantly fluoride-induced renal damage and apoptosis of carp in a dose-dependent manner, indicated by the histopathological examination and ultrastructural observation. Moreover, treatment with sesamin also inhibited significantly fluoride-induced remarkable enhancement of reactive oxygen species (ROS) production and oxidative stress, such as the increase of lipid peroxidation level and the depletion of intracellular reduced glutathione (GSH) level in kidney. To explore the underlying mechanisms of sesamin action, we found that activities of caspase-3 were notably inhibited by treatment with sesamin in the kidney of fluoride-exposed fish. Sesamin decreased the levels of p-JNK protein in kidney, which in turn inactivated pro-apoptotic signaling events by restoring the balance between mitochondrial pro- and anti-apoptotic Bcl-2 and Bax proteins and by decreasing the release of mitochondrial cytochrome c in kidney of fluoride-exposed fish. JNK was also involved in the mitochondrial extrinsic apoptotic pathways of sesamin effects against fluoride-induced renal injury by regulating the levels of p-c-Jun, necrosis factor-alpha (TNF-α) and Bak proteins. These findings indicated that sesamin could protect kidney against fluoride-induced apoptosis by the oxidative stress downstream-mediated change in the inactivation of JNK signaling pathway. Taken together, sesamin plays an important role in maintaining renal health and preventing kidney from toxic damage induced by

  19. Standardized extracts of Bacopa monniera protect against MPP+- and paraquat-induced toxicity by modulating mitochondrial activities, proteasomal functions, and redox pathways.

    Science.gov (United States)

    Singh, Manjeet; Murthy, Ven; Ramassamy, Charles

    2012-01-01

    Parkinson's disease (PD) is one of the most common age-related neurodegenerative diseases and affects millions of people worldwide. Strong evidence supports the role of free radicals, oxidative stress, mitochondrial, and proteasomal dysfunctions underlying neuronal death in PD. Environmental factors, especially pesticides, represent one of the primary classes of neurotoxic agents associated with PD, and several epidemiological studies have identified the exposure of the herbicide paraquat (PQ) as a potential risk factor for the onset of PD. The objective of our study was to investigate the neuroprotective effects of the standardized extracts of Bacopa monniera (BM) against PQ-induced and 1-methyl-4-phenyl-pyridinium iodide (MPP(+))-induced toxicities and to elucidate the mechanisms underlying this protection. Our results show that a pretreatment with the BM extract from 50 μg/ml protected the dopaminergic SK-N-SH cell line against MPP(+)- and PQ-induced toxicities in various cell survival assays. We demonstrate that BM pretreatment prevented the depletion of glutathione (GSH) besides preserving the mitochondrial membrane potential and maintaining the mitochondrial complex I activity. BM pretreatment from 10.0 μg/ml also prevented the generation of intracellular reactive oxygen species and decreased the mitochondrial superoxide level. BM treatment activated the nuclear factor erythroid 2-related factor 2 pathway by modulating the expression of Keap1, thereby upregulating the endogenous GSH synthesis. The effect of BM on the phosphorylation of Akt further strengthens its role in the promotion of cell survival. By preserving the cellular redox homeostasis and mitochondrial activities and by promoting cell survival pathways, BM extract may have therapeutic uses in various age-related neurodegenerative diseases such as PD.

  20. Amentoflavone protects dopaminergic neurons in MPTP-induced Parkinson's disease model mice through PI3K/Akt and ERK signaling pathways

    International Nuclear Information System (INIS)

    Cao, Qin; Qin, Liyue; Huang, Fei; Wang, Xiaoshuang; Yang, Liu; Shi, Hailian; Wu, Hui; Zhang, Beibei; Chen, Ziyu; Wu, Xiaojun

    2017-01-01

    Parkinson's disease (PD) is characterized by the progressive degeneration of dopaminergic neurons in substantia nigra pars compacta (SNpc). Mitochondrial dysfunction and cell apoptosis are suggested to be actively involved in the pathogenesis of PD. In the present study, the neuroprotective effect of amentoflavone (AF), a naturally occurring biflavonoid from Selaginella tamariscina, was examined in PD models both in vitro and in vivo. On SH-SY5Y cells, AF treatment dose-dependently reduced 1-methyl-4-phenylpyridinium (MPP + )-induced nuclear condensation and loss of cell viability without obvious cytotoxicity. It inhibited the activation of caspase-3 and p21 but increased the Bcl-2/Bax ratio. Further study disclosed that AF enhanced the phosphorylation of PI3K, Akt and ERK1/2 down-regulated by MPP + in SH-SY5Y cells, the effect of which could be blocked by LY294002, the inhibitor of PI3K. Consistently, AF alleviated the behavioral deterioration in pole and traction tests and rescued the loss of dopaminergic neurons in SNpc and fibers in striatum in methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) induced mice. It also could enhance the activation of PI3K and Akt as well as Bcl-2/Bax ratio in SN. Moreover, AF alleviated gliosis as well as the gene expression levels of IL-1β and iNOS in SN. Collectively, these results suggested that AF protected dopaminergic neurons against MPTP/MPP + -induced neurotoxicity, which might be mediated through activation of PI3K/Akt and ERK signaling pathways in dopaminergic neurons and attenuation of neuroinflammation. - Highlights: • AF protected dopaminergic neurons against MPTP/MPP + -induced neurotoxicity. • AF modulated PI3K/Akt and ERK signaling pathways. • AF could alleviate neuroinflammation in SN.

  1. Immaturity of human stem-cell-derived cardiomyocytes in culture: fatal flaw or soluble problem?

    NARCIS (Netherlands)

    Veerman, Christiaan C.; Kosmidis, Georgios; Mummery, Christine L.; Casini, Simona; Verkerk, Arie O.; Bellin, Milena

    2015-01-01

    Cardiomyocytes from human pluripotent stem cells (hPSC-CMs) are increasingly used to model cardiac disease, test drug efficacy and for safety pharmacology. Nevertheless, a major hurdle to more extensive use is their immaturity and similarity to fetal rather than adult cardiomyocytes. Here, we

  2. Microscale Generation of Cardiospheres Promotes Robust Enrichment of Cardiomyocytes Derived from Human Pluripotent Stem Cells

    Directory of Open Access Journals (Sweden)

    Doan C. Nguyen

    2014-08-01

    Full Text Available Cardiomyocytes derived from human pluripotent stem cells (hPSCs are a promising cell source for regenerative medicine, disease modeling, and drug discovery, all of which require enriched cardiomyocytes, ideally ones with mature phenotypes. However, current methods are typically performed in 2D environments that produce immature cardiomyocytes within heterogeneous populations. Here, we generated 3D aggregates of cardiomyocytes (cardiospheres from 2D differentiation cultures of hPSCs using microscale technology and rotary orbital suspension culture. Nearly 100% of the cardiospheres showed spontaneous contractility and synchronous intracellular calcium transients. Strikingly, from starting heterogeneous populations containing ∼10%–40% cardiomyocytes, the cell population within the generated cardiospheres featured ∼80%–100% cardiomyocytes, corresponding to an enrichment factor of up to 7-fold. Furthermore, cardiomyocytes from cardiospheres exhibited enhanced structural maturation in comparison with those from a parallel 2D culture. Thus, generation of cardiospheres represents a simple and robust method for enrichment of cardiomyocytes in microtissues that have the potential use in regenerative medicine as well as other applications.

  3. Dystrophin-deficient cardiomyocytes derived from human urine: New biologic reagents for drug discovery

    Directory of Open Access Journals (Sweden)

    Xuan Guan

    2014-03-01

    Full Text Available The ability to extract somatic cells from a patient and reprogram them to pluripotency opens up new possibilities for personalized medicine. Induced pluripotent stem cells (iPSCs have been employed to generate beating cardiomyocytes from a patient's skin or blood cells. Here, iPSC methods were used to generate cardiomyocytes starting from the urine of a patient with Duchenne muscular dystrophy (DMD. Urine was chosen as a starting material because it contains adult stem cells called urine-derived stem cells (USCs. USCs express the canonical reprogramming factors c-myc and klf4, and possess high telomerase activity. Pluripotency of urine-derived iPSC clones was confirmed by immunocytochemistry, RT-PCR and teratoma formation. Urine-derived iPSC clones generated from healthy volunteers and a DMD patient were differentiated into beating cardiomyocytes using a series of small molecules in monolayer culture. Results indicate that cardiomyocytes retain the DMD patient's dystrophin mutation. Physiological assays suggest that dystrophin-deficient cardiomyocytes possess phenotypic differences from normal cardiomyocytes. These results demonstrate the feasibility of generating cardiomyocytes from a urine sample and that urine-derived cardiomyocytes retain characteristic features that might be further exploited for mechanistic studies and drug discovery.

  4. Sirtuin7 is involved in protecting neurons against oxygen-glucose deprivation and reoxygenation-induced injury through regulation of the p53 signaling pathway.

    Science.gov (United States)

    Lv, Jianrui; Tian, Junbin; Zheng, Guoxi; Zhao, Jing

    2017-10-01

    Sirtuin7 (SIRT7) is known to regulate apoptosis and stress responses. So far, very little is known about the role of SIRT7 in cerebral ischemia/reperfusion injury. In this study, we aimed to investigate the potential role of SIRT7 in regulating oxygen-glucose deprivation and reoxygenation (OGD/R)-induced injury in neurons. We found a significant increase of SIRT7 expression in neurons in response to OGD/R treatment. Knockdown of SIRT7 aggravated OGD/R-induced injury. Knockdown of SIRT7 augmented the levels of total and acetylated p53 protein. Moreover, knockdown of SIRT7 markedly increased the transcriptional activity of p53 toward apoptosis and activated the p53-mediated proapoptotic signaling pathway. By contrast, overexpression of SIRT7 showed the opposite effects. Taken together, the results of our study suggest that SIRT7 is involved in protecting neurons against OGD/R-induced injury, possibly through regulation of the p53-mediated proapoptotic signaling pathway, indicating a potential therapeutic target for cerebral ischemia/reperfusion injury. © 2017 Wiley Periodicals, Inc.

  5. Altered calcium handling and increased contraction force in human embryonic stem cell derived cardiomyocytes following short term dexamethasone exposure

    NARCIS (Netherlands)

    Kosmidis, Georgios; Bellin, Milena; Ribeiro, Marcelo C.; van Meer, Berend; Ward-van Oostwaard, Dorien; Passier, Robert; Tertoolen, Leon G. J.; Mummery, Christine L.; Casini, Simona

    2015-01-01

    One limitation in using human pluripotent stem cell derived cardiomyocytes (hPSC-CMs) for disease modeling and cardiac safety pharmacology is their immature functional phenotype compared with adult cardiomyocytes. Here, we report that treatment of human embryonic stem cell derived cardiomyocytes

  6. Autonomous and Non-autonomous Defects Underlie Hypertrophic Cardiomyopathy in BRAF-Mutant hiPSC-Derived Cardiomyocytes

    Directory of Open Access Journals (Sweden)

    Rebecca Josowitz

    2016-09-01

    Full Text Available Germline mutations in BRAF cause cardio-facio-cutaneous syndrome (CFCS, whereby 40% of patients develop hypertrophic cardiomyopathy (HCM. As the role of the RAS/MAPK pathway in HCM pathogenesis is unclear, we generated a human induced pluripotent stem cell (hiPSC model for CFCS from three patients with activating BRAF mutations. By cell sorting for SIRPα and CD90, we generated a method to examine hiPSC-derived cell type-specific phenotypes and cellular interactions underpinning HCM. BRAF-mutant SIRPα+/CD90− cardiomyocytes displayed cellular hypertrophy, pro-hypertrophic gene expression, and intrinsic calcium-handling defects. BRAF-mutant SIRPα−/CD90+ cells, which were fibroblast-like, exhibited a pro-fibrotic phenotype and partially modulated cardiomyocyte hypertrophy through transforming growth factor β (TGFβ paracrine signaling. Inhibition of TGFβ or RAS/MAPK signaling rescued the hypertrophic phenotype. Thus, cell autonomous and non-autonomous defects underlie HCM due to BRAF mutations. TGFβ inhibition may be a useful therapeutic option for patients with HCM due to RASopathies or other etiologies.

  7. Testosterone-mediated upregulation of delayed rectifier potassium channel in cardiomyocytes causes abbreviation of QT intervals in rats.

    Science.gov (United States)

    Masuda, Kimiko; Takanari, Hiroki; Morishima, Masaki; Ma, FangFang; Wang, Yan; Takahashi, Naohiko; Ono, Katsushige

    2018-01-13

    Men have shorter rate-corrected QT intervals (QTc) than women, especially at the period of adolescence or later. The aim of this study was to elucidate the long-term effects of testosterone on cardiac excitability parameters including electrocardiogram (ECG) and potassium channel current. Testosterone shortened QT intervals in ECG in castrated male rats, not immediately after, but on day 2 or later. Expression of Kv7.1 (KCNQ1) mRNA was significantly upregulated by testosterone in cardiomyocytes of male and female rats. Short-term application of testosterone was without effect on delayed rectifier potassium channel current (I Ks ), whereas I Ks was significantly increased in cardiomyocytes treated with dihydrotestosterone for 24 h, which was mimicked by isoproterenol (24 h). Gene-selective inhibitors of a transcription factor SP1, mithramycin, abolished the effects of testosterone on Kv7.1. Testosterone increases Kv7.1-I Ks possibly through a pathway related to a transcription factor SP1, suggesting a genomic effect of testosterone as an active factor for cardiac excitability.

  8. Activation of β-Adrenoceptors by Dobutamine May Induce a Higher Expression of Peroxisome Proliferator-Activated Receptors δ (PPARδ in Neonatal Rat Cardiomyocytes

    Directory of Open Access Journals (Sweden)

    Ming-Ting Chou

    2012-01-01

    Full Text Available Recent evidence showed the role of peroxisome proliferator-activated receptors (PPARs in cardiac function. Cardiac contraction induced by various agents is critical in restoring the activity of peroxisome proliferator-activated receptors δ (PPARδ in cardiac myopathy. Because dobutamine is an agent widely used to treat heart failure in emergency setting, this study is aimed to investigate the change of PPARδ in response to dobutamine. Neonatal rat cardiomyocytes were used to examine the effects of dobutamine on PPARδ expression levels and cardiac troponin I (cTnI phosphorylation via Western blotting analysis. We show that treatment with dobutamine increased PPARδ expression and cTnI phosphorylation in a time- and dose-dependent manner in neonatal rat cardiomyocytes. These increases were blocked by the antagonist of β1-adrenoceptors. Also, the action of dobutamine was related to the increase of calcium ions and diminished by chelating intracellular calcium. Additionally, dobutamine-induced action was reduced by the inhibition of downstream messengers involved in this calcium-related pathway. Moreover, deletion of PPARδ using siRNA generated the reduction of cTnI phosphorylation in cardiomyocytes treated with dobutamine. Thus, we concluded that PPARδ is increased by dobutamine in cardiac cells.

  9. The Role of Sulfur Dioxide in the Regulation of Mitochondrion-Related Cardiomyocyte Apoptosis in Rats with Isopropylarterenol-Induced Myocardial Injury

    Directory of Open Access Journals (Sweden)

    Junbao Du

    2013-05-01

    Full Text Available The authors investigated the regulatory effects of sulfur dioxide (SO2 on myocardial injury induced by isopropylarterenol (ISO hydrochloride and its mechanisms. Wistar rats were divided into four groups: control group, ISO group, ISO plus SO2 group, and SO2 only group. Cardiac function was measured and cardiomyocyte apoptosis was detected. Bcl-2, bax and cytochrome c (cytc expressions, and caspase-9 and caspase-3 activities in the left ventricular tissues were examined in the rats. The opening status of myocardial mitochondrial permeability transition pore (MPTP and membrane potential were analyzed. The results showed that ISO-treated rats developed heart dysfunction and cardiac injury. Furthermore, cardiomyocyte apoptosis in the left ventricular tissues was augmented, left ventricular tissue bcl-2 expression was down-regulated, bax expression was up-regulated, mitochondrial membrane potential was significantly reduced, MPTP opened, cytc release from mitochondrion into cytoplasm was significantly increased, and both caspase-9 and caspase-3 activities were increased. Administration of an SO2 donor, however, markedly improved heart function and relieved myocardial injury of the ISO-treated rats; it lessened cardiomyocyte apoptosis, up-regulated myocardial bcl-2, down-regulated bax expression, stimulated mitochondrial membrane potential, closed MPTP, and reduced cytc release as well as caspase-9 and caspase-3 activities in the left ventricular tissue. Hence, SO2 attenuated myocardial injury in association with the inhibition of apoptosis in myocardial tissues, and the bcl-2/cytc/caspase-9/caspase-3 pathway was possibly involved in this process.

  10. Protective Effect of Edaravone Against Cyclosporine-Induced Chronic Nephropathy Through Antioxidant and Nitric Oxide Modulating Pathways in Rats

    Directory of Open Access Journals (Sweden)

    Elahe Sattarinezhad

    2017-03-01

    Full Text Available Background: Cyclosporine A (CsA is an immunosuppressant with therapeutic indications in various immunological diseases; however, its use is associated with chronic nephropathy. Oxidative stress has a crucial role in CsA-induced nephrotoxicity. The present study evaluates the protective effect of edaravone on CsA-induced chronic nephropathy and investigates its antioxidant and nitric oxide modulating property. Methods: Male Sprague-Dawley rats (n=66 were distributed into nine groups, including a control (group 1 (n=7. Eight groups received CsA (15 mg/kg for 28 days while being treated. The groups were categorized as: •\tGroup 2: Vehicle (n=10 •\tGroups 3, 4, and 5: Edaravone (1, 5, and 10 mg/kg (n=7 each •\tGroup 6: Diphenyliodonium chloride, a specific endothelial nitric oxide synthase (eNOS inhibitor (n=7 •\tGroup 7: Aminoguanidine, a specific inducible nitric oxide synthase (iNOS inhibitor (n=7 •\tGroup 8: Edaravone (10 mg/kg plus diphenyliodonium chloride (n=7 •\tGroup 9: Edaravone (10 mg/kg plus aminoguanidine (n=7 Blood urea nitrogen and serum creatinine levels, malondialdehyde, superoxide dismutase, and glutathione reductase enzyme activities were measured using standard kits. Renal histopathological evaluations and measurements of eNOS and iNOS gene expressions by RT-PCR were also performed. Data were analyzed using one-way analysis of variance (ANOVA followed by Tukey’s test (SPSS software version 18.0. Results: Edaravone (10 mg/kg significantly attenuated CsA-induced oxidative stress, renal dysfunction, and kidney tissue injury. Aminoguanidine improved the renoprotective effect of edaravone. Edaravone reduced the elevated mRNA level of iNOS, but could not alter the level of eNOS mRNA significantly. Conclusion: Edaravone protects against CsA-induced chronic nephropathy using antioxidant property and probably through inhibiting iNOS gene expression.

  11. Imaging alterations of cardiomyocyte cAMP microdomains in disease

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    Alexander eFroese

    2015-08-01

    Full Text Available 3’,5’-cyclic adenosine monophosphate (cAMP is an important second messenger which regulates heart function by acting in distinct subcellular microdomains. Recent years have provided deeper mechanistic insights into compartmentalized cAMP signaling and its link to cardiac disease. In this mini review, we summarize newest developments in this field achieved by cutting-edge biochemical and biophysical techniques. We further compile the data from different studies into a bigger picture of so far uncovered alterations in cardiomyocyte cAMP microdomains which occur in compensated cardiac hypertrophy and chronic heart failure. Finally, future research directions and translational perspectives are briefly discussed.

  12. Modulation of post-stroke degenerative and regenerative processes and subacute protection by site-targeted inhibition of the alternative pathway of complement.

    Science.gov (United States)

    Alawieh, Ali; Elvington, Andrew; Zhu, Hong; Yu, Jin; Kindy, Mark S; Atkinson, Carl; Tomlinson, Stephen

    2015-12-30

    Complement promotes neuroinflammation and injury in models of stroke. However, complement is also being increasingly implicated in repair and regeneration after central nervous system (CNS) injury, and some complement deficiencies have been shown to provide acute, but not subacute, protection after murine stroke. Here, we investigate the dual role of complement in injury and repair after cerebral ischemia and reperfusion. We used complement-deficient mice and different complement inhibitors in a model of transient middle cerebral artery occlusion to investigate complement-dependent cellular and molecular changes that occur through the subacute phase after stroke. C3 deficiency and site-targeted complement inhibition with either CR2-Crry (inhibits all pathways) or CR2-fH (inhibits alternative pathway) significantly reduced infarct size, reduced apoptotic cell death, and improved neurological deficit score in the acute phase after stroke. However, only in CR2-fH-treated mice was there sustained protection with no evolution of injury in the subacute phase. Whereas both inhibitors significantly reduced microglia/macrophage activation and astrogliosis in the subacute phase, only CR2-fH improved neurological deficit and locomotor function, maintained neurogenesis markers, enhanced neuronal migration, and increased VEGF expression. These findings in CR2-fH-treated mice correlated with improved performance in spatial learning and passive avoidance tasks. The complement anaphylatoxins have been implicated in repair and regenerative mechanisms after CNS injury, and in this context CR2-fH significantly reduced, but did not eliminate the generation of C5a within the brain, unlike CR2-Crry that completely blocked C5a generation. Gene expression profiling revealed that CR2-fH treatment downregulated genes associated with apoptosis, TGFβ signaling, and neutrophil activation, and decreased neutrophil infiltration was confirmed by immunohistochemistry. CR2-fH upregulated genes for

  13. A copper-induced quinone degradation pathway provides protection against combined copper/quinone stress in Lactococcus lactis IL1403.

    Science.gov (United States)

    Mancini, Stefano; Abicht, Helge K; Gonskikh, Yulia; Solioz, Marc

    2015-02-01

    Quinones are ubiquitous in the environment. They occur naturally but are also in widespread use in human and industrial activities. Quinones alone are relatively benign to bacteria, but in combination with copper, they become toxic by a mechanism that leads to intracellular thiol depletion. Here, it was shown that the yahCD-yaiAB operon of Lactococcus lactis IL1403 provides resistance to combined copper/quinone stress. The operon is under the control of CopR, which also regulates expression of the copRZA copper resistance operon as well as other L. lactis genes. Expression of the yahCD-yaiAB operon is induced by copper but not by quinones. Two of the proteins encoded by the operon appear to play key roles in alleviating quinone/copper stress: YaiB is a flavoprotein that converts p-benzoquinones to less toxic hydroquinones, using reduced nicotinamide adenine dinucleotide phosphate (NADPH) as reductant; YaiA is a hydroquinone dioxygenase that converts hydroquinone putatively to 4-hydroxymuconic semialdehyde in an oxygen-consuming reaction. Hydroquinone and methylhydroquinone are both substrates of YaiA. Deletion of yaiB causes increased sensitivity of L. lactis to quinones and complete growth arrest under combined quinone and copper stress. Copper induction of the yahCD-yaiAB operon offers protection to copper/quinone toxicity and could provide a growth advantage to L. lactis in some environments. © 2014 John Wiley & Sons Ltd.

  14. Vanillin Protects Dopaminergic Neurons against Inflammation-Mediated Cell Death by Inhibiting ERK1/2, P38 and the NF-κB Signaling Pathway.

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    Yan, Xuan; Liu, Dian-Feng; Zhang, Xiang-Yang; Liu, Dong; Xu, Shi-Yao; Chen, Guang-Xin; Huang, Bing-Xu; Ren, Wen-Zhi; Wang, Wei; Fu, Shou-Peng; Liu, Ju-Xiong

    2017-02-12

    Neuroinflammation plays a very important role in the pathogenesis of Parkinson's disease (PD). After activation, microglia produce pro-inflammatory mediators that damage surrounding neurons. Consequently, the inhibition of microglial activation might represent a new therapeutic approach of PD. Vanillin has been shown to protect dopaminergic neurons, but the mechanism is still unclear. Herein, we further study the underlying mechanisms in lipopolysaccharide (LPS)-induced PD models. In vivo, we firstly established rat models of PD by unilateral injection of LPS into substantia nigra (SN), and then examined the role of vanillin in motor dysfunction, microglial activation and degeneration of dopaminergic neurons. In vitro, murine microglial BV-2 cells were treated with vanillin prior to the incubation of LPS, and then the inflammatory responses and the related signaling pathways were analyzed. The in vivo results showed that vanillin markedly improved the motor dysfunction, suppressed degeneration of dopaminergic neurons and inhibited microglial over-activation induced by LPS intranigral injection. The in vitro studies demonstrated that vanillin reduces LPS-induced expression of inducible nitric oxide (iNOS), cyclooxygenase-2 (COX-2), IL-1β, and IL-6 through regulating ERK1/2, p38 and NF-κB signaling. Collectively, these data indicated that vanillin has a role in protecting dopaminergic neurons via inhibiting inflammatory activation.

  15. Vanillin Protects Dopaminergic Neurons against Inflammation-Mediated Cell Death by Inhibiting ERK1/2, P38 and the NF-κB Signaling Pathway

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    Xuan Yan

    2017-02-01

    Full Text Available Neuroinflammation plays a very important role in the pathogenesis of Parkinson’s disease (PD. After activation, microglia produce pro-inflammatory mediators that damage surrounding neurons. Consequently, the inhibition of microglial activation might represent a new therapeutic approach of PD. Vanillin has been shown to protect dopaminergic neurons, but the mechanism is still unclear. Herein, we further study the underlying mechanisms in lipopolysaccharide (LPS-induced PD models. In vivo, we firstly established rat models of PD by unilateral injection of LPS into substantia nigra (SN, and then examined the role of vanillin in motor dysfunction, microglial activation and degeneration of dopaminergic neurons. In vitro, murine microglial BV-2 cells were treated with vanillin prior to the incubation of LPS, and then the inflammatory responses and the related signaling pathways were analyzed. The in vivo results showed that vanillin markedly improved the motor dysfunction, suppressed degeneration of dopaminergic neurons and inhibited microglial over-activation induced by LPS intranigral injection. The in vitro studies demonstrated that vanillin reduces LPS-induced expression of inducible nitric oxide (iNOS, cyclooxygenase-2 (COX-2, IL-1β, and IL-6 through regulating ERK1/2, p38 and NF-κB signaling. Collectively, these data indicated that vanillin has a role in protecting dopaminergic neurons via inhibiting inflammatory activation.

  16. Atorvastatin Protects Vascular Smooth Muscle Cells From TGF-β1-Stimulated Calcification by Inducing Autophagy via Suppression of the β-Catenin Pathway

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    Demin Liu

    2014-01-01

    Full Text Available Background: Arterial calcification is a major event in the progression of atherosclerosis. It is reported that statins exhibit various protective effects against vascular smooth muscle cell (VSMC inflammation and proliferation in cardiovascular remodeling. Although statins counteract atherosclerosis, the molecular mechanisms of statins on the calcium release from VSMCs have not been clearly elucidated. Methods: Calcium content of VSMCs was measured using enzyme-linked immunosorbent assay (ELISA. The expression of proteins involved in cellular transdifferentiation was analyzed by western blot. Cell autophagy was measured by fluorescence microscopic analysis for acridine orange staining and transmission electron microscopy analysis. The autophagic inhibitors (3-MA, chloroquine, NH4Cl and bafilomycin A1 and β-catenin inhibitor JW74 were used to assess the effects of atorvastatin on autophagy and the involvement of β-catenin on cell calcification respectively. Furthermore, cell transfection was performed to overexpress β-catenin. Results: In VSMCs, atorvastatin significantly suppressed transforming growth factor-β1 (TGF-β1-stimulated calcification, accompanied by the induction of autophagy. Downregulation of autophagy with autophagic inhibitors significantly suppressed the inhibitory effect of atorvastatin on cell calcification. Moreover, the beneficial effect of atorvastatin on calcification and autophagy was reversed by β-catenin overexpression. Conversely, JW74 supplement enhanced this effect. Conclusion: These data demonstrated that atorvastatin protect VSMC from TGF-β1-stimulated calcification by inducing autophagy through suppression of the β-catenin pathway, identifying autophagy induction might be a therapeutic strategy for use in vascular calcification.

  17. Protective effect of thymoquinone improves cardiovascular function, and attenuates oxidative stress, inflammation and apoptosis by mediating the PI3K/Akt pathway in diabetic rats.

    Science.gov (United States)

    Liu, Hui; Liu, Hong-Yang; Jiang, Yi-Nong; Li, Nan

    2016-03-01

    Thymoquinone is the main active monomer extracted from black cumin and has anti‑inflammatory, antioxidant and anti‑apoptotic functions. However, the protective effects of thymoquinone on cardiovascular function in diabetes remain to be fully elucidated. The present study aimed to investigate the molecular mechanisms underling the beneficial effects of thymoquinone on the cardiovascular function in streptozotocin‑induced diabetes mellitus (DM) rats. Supplement thymoquinone may recover the insulin levels and body weight, inhibit blood glucose levels and reduce the heart rate in DM‑induced rats. The results indicated that the heart, liver and lung to body weight ratios, in addition to the blood pressure levels, were similar for each experimental group. Treatment with thymoquinone significantly reduced oxidative stress damage, inhibited the increased endothelial nitric oxide synthase protein expression and suppressed the elevation of cyclooxygenase‑2 levels in DM‑induced rats. In addition, thymoquinone significantly suppressed the promotion of tumor necrosis factor‑α and interleukin‑6 levels in the DM‑induced rats. Furthermore, administration of thymoquinone significantly reduced caspase‑3 activity and the promotion of phosphorylated‑protein kinase B (Akt) protein expression levels in DM‑induced rats. These results suggest that the protective effect of thymoquinone improves cardiovascular function and attenuates oxidative stress, inflammation and apoptosis by mediating the phosphatidylinositol 3‑kinase/Akt pathway in DM‑induced rats.

  18. Neuron-derived IgG protects dopaminergic neurons from insult by 6-OHDA and activates microglia through the FcγR I and TLR4 pathways.

    Science.gov (United States)

    Zhang, Jie; Niu, Na; Wang, Mingyu; McNutt, Michael A; Zhang, Donghong; Zhang, Baogang; Lu, Shijun; Liu, Yuqing; Liu, Zhihui

    2013-08-01

    Oxidative and immune attacks from the environment or microglia have been implicated in the loss of dopaminergic neurons of Parkinson's disease. The role of IgG which is an important immunologic molecule in the process of Parkinson's disease has been unclear. Evidence suggests that IgG can be produced by neurons in addition to its traditionally recognized source B lymphocytes, but its function in neurons is poorly understood. In this study, extensive expression of neuron-derived IgG was demonstrated in dopaminergic neurons of human and rat mesencephalon. With an in vitro Parkinson's disease model, we found that neuron-derived IgG can improve the survival and reduce apoptosis of dopaminergic neurons induced by 6-hydroxydopamine toxicity, and also depress the release of NO from microglia triggered by 6-hydroxydopamine. Expression of TNF-α and IL-10 in microglia was elevated to protective levels by neuron-derived IgG at a physiologic level via the FcγR I and TLR4 pathways and microglial activation could be attenuated by IgG blocking. All these data suggested that neuron-derived IgG may exert a self-protective function by activating microglia properly, and IgG may be involved in maintaining immunity homeostasis in the central nervous system and serve as an active factor under pathological conditions such as Parkinson's disease. Crown Copyright © 2013. Published by Elsevier Ltd. All rights reserved.

  19. Mucuna pruriens Protects against MPTP Intoxicated Neuroinflammation in Parkinson's Disease through NF-κB/pAKT Signaling Pathways.

    Science.gov (United States)

    Rai, Sachchida N; Birla, Hareram; Singh, Saumitra S; Zahra, Walia; Patil, Ravishankar R; Jadhav, Jyoti P; Gedda, Mallikarjuna R; Singh, Surya P

    2017-01-01

    analysis of the Mp seed extract have shown L-DOPA, gallic acid, phytic acid, quercetin, and catechin equivalents as the major components which might cause neuroprotection in PD mice. Our result suggested that Mp extract treatment containing L-DOPA and a mixture of rich novel phytochemicals significantly alleviates the MPTP induced neurotoxicity by NF-κB and pAkt pathway. The findings observed thereby indicate that Mp extract have suggestively ameliorated MPTP induced neuroinflammation, restored the biochemical and behavioral abnormalities in PD mouse and thus provided a scientific basis for its traditional claim.

  20. The IRE1α/XBP1s Pathway Is Essential for the Glucose Response and Protection of β Cells.

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    Justin R Hassler

    2015-10-01

    Full Text Available Although glucose uniquely stimulates proinsulin biosynthesis in β cells, surprisingly little is known of the underlying mechanism(s. Here, we demonstrate that glucose activates the unfolded protein response transducer inositol-requiring enzyme 1 alpha (IRE1α to initiate X-box-binding protein 1 (Xbp1 mRNA splicing in adult primary β cells. Using mRNA sequencing (mRNA-Seq, we show that unconventional Xbp1 mRNA splicing is required to increase and decrease the expression of several hundred mRNAs encoding functions that expand the protein secretory capacity for increased insulin production and protect from oxidative damage, respectively. At 2 wk after tamoxifen-mediated Ire1α deletion, mice develop hyperglycemia and hypoinsulinemia, due to defective β cell function that was exacerbated upon feeding and glucose stimulation. Although previous reports suggest IRE1α degrades insulin mRNAs, Ire1α deletion did not alter insulin mRNA expression either in the presence or absence of glucose stimulation. Instead, β cell failure upon Ire1α deletion was primarily due to reduced proinsulin mRNA translation primarily because of defective glucose-stimulated induction of a dozen genes required for the signal recognition particle (SRP, SRP receptors, the translocon, the signal peptidase complex, and over 100 other genes with many other intracellular functions. In contrast, Ire1α deletion in β cells increased the expression of over 300 mRNAs encoding functions that cause inflammation and oxidative stress, yet only a few of these accumulated during high glucose. Antioxidant treatment significantly reduced glucose intolerance and markers of inflammation and oxidative stress in mice with β cell-specific Ire1α deletion. The results demonstrate that glucose activates IRE1α-mediated Xbp1 splicing to expand the secretory capacity of the β cell for increased proinsulin synthesis and to limit oxidative stress that leads to β cell failure.

  1. Quercetin protects against aluminium induced oxidative stress and promotes mitochondrial biogenesis via activation of the PGC-1α signaling pathway.

    Science.gov (United States)

    Sharma, Deep Raj; Sunkaria, Aditya; Wani, Willayat Yousuf; Sharma, Reeta Kumari; Verma, Deepika; Priyanka, Kumari; Bal, Amanjit; Gill, Kiran Dip

    2015-12-01

    The present investigation was carried out to elucidate a possible molecular mechanism related to the protective effect of quercetin administration against aluminium-induced oxidative stress on various mitochondrial respiratory complex subunits with special emphasis on the role of PGC-1α and its downstream targets, i.e. NRF-1, NRF-2 and Tfam in mitochondrial biogenesis. Aluminium lactate (10mg/kg b.wt./day) was administered intragastrically to rats, which were pre-treated with quercetin 6h before aluminium (10mg/kg b.wt./day, intragastrically) for 12 weeks. We found a decrease in ROS levels, mitochondrial DNA oxidation and citrate synthase activity in the hippocampus (HC) and corpus striatum (CS) regions of rat brain treated with quercetin. Besides this an increase in the mRNA levels of the mitochondrial encoded subunits - ND1, ND2, ND3, Cyt b, COX1, COX3 and ATPase6 along with increased expression of nuclear encoded subunits COX4, COX5A and COX5B of electron transport chain (ETC). In quercetin treated group an increase in the mitochondrial DNA copy number and mitochondrial content in both the regions of rat brain was observed. The PGC-1α was up regulated in quercetin treated rats along with NRF-1, NRF-2 and Tfam, which act downstream from PGC-1α. Electron microscopy results revealed a significant decrease in the mitochondrial cross-section area, mitochondrial perimeter length and increase in mitochondrial number in case of quercetin treated rats as compared to aluminium treated ones. Therefore it seems quercetin increases mitochondrial biogenesis and makes it an almost ideal flavanoid to control or limit the damage that has been associated with the defective mitochondrial function seen in many neurodegenerative diseases. Copyright © 2015 Elsevier Inc. All rights reserved.

  2. Edaravone protects the retina against ischemia/reperfusion‑induced oxidative injury through the PI3K/Akt/Nrf2 pathway.

    Science.gov (United States)

    Xu, Yi-Pin; Han, Fang; Tan, Jian

    2017-12-01

    Retinal ischemia/reperfusion (I/R) injury can occur as a result of a number of ocular diseases or ischemic events in the brain, leading to possible vision loss if not treated properly. The overproduction of reactive oxygen species is important in the process of I/R injury. Edaravone, a free radical scavenger, has been demonstrated to have a neuroprotective effect in cerebral ischemia; however, its effect against retinal I/R injury remains to be fully elucidated. Therefore, the present study investigated the effects of edaravone on the oxidative parameters, retinal inflammation and apoptosis induced by I/R injury, and treated photoreceptor‑derived 661W cells with hydrogen peroxide (H2O2) and edaravone to examine the underlying mechanism. For the in vivo study, oxidative parameters (malondialdehyde, DNA fragmentation, total antioxidant status, superoxide dismutase and glutathione) in the retina, retinal thickness, and apoptotic index in the ganglionic cell layer and inner nuclear layer were measured. For the in vitro study, the effects of edaravone or nuclear factor erythroid‑2‑related factor 2 (Nrf2) small interfering RNA or phosphatidylinositol 3‑kinase (PI3K)/Akt inhibitors on cell viability, membrane integrity, levels of phosphorylated‑Akt, Akt and nuclear Nrf2 of H2O2‑treated 661W cells were examined. The results demonstrated that edaravone inhibited the oxidative injury in the retina induced by the retinal I/R procedure and increased retinal inflammation, and apoptosis. The results of the in vitro experiments demonstrated that edaravone effectively protected the viability and the membrane integrity of the H2O2‑treated 661W cells via the phosphatidylinositol 3‑kinase (PI3K)/Akt/Nrf2pathway. These results indicated the potential protective effect of edaravone against retinal I/R injury and provided a novel explanation for the protective effects of edaravone.

  3. Hydrogen protects against hyperoxia-induced apoptosis in type II alveolar epithelial cells via activation of PI3K/Akt/Foxo3a signaling pathway.

    Science.gov (United States)

    Wu, Dan; Liang, Mulin; Dang, Hongxing; Fang, Fang; Xu, Feng; Liu, Chengjun

    2018-01-08

    Oxidative stress is regarded as a key regulator in the pathogenesis of prolonged hyperoxia-induced lung injury, which causes injury to alveolar epithelial cells and eventually leads to development of bronchopulmonary dysplasia (BPD). Many studies have shown that hydrogen has a protective effect in a variety of cells. However, the mechanisms by which hydrogen rescues cells from damage due to oxidative stress in BPD remains to be fully elucidated. This study sought to evaluate the effects of hydrogen on hyperoxia-induced lung injury and to investigate the underlying mechanism. Primary type II alveolar epithelial cells (AECIIs) were divided into four groups: control (21% oxygen), hyperoxia (95% oxygen), hyperoxia + hydrogen, and hyperoxia + hydrogen + LY294002 (a PI3K/Akt inhibitor). Proliferation and apoptosis of AECIIs were assessed using MTS assay and flow cytometry (FCM), respectively. Gene and protein expression were detected by quantitative polymerase chain reaction (q-PCR) and western blot analysis. Stimulation with hyperoxia decreased the expression of P-Akt, P- FoxO3a, cyclinD1 and Bcl-2. Hyperoxic conditions increased levels of Bim, Bax, and Foxo3a, which induced proliferation restriction and apoptosis of AECIIs. These effects of hyperoxia were reversed with hydrogen pretreatment. Furthermore, the protective effects of hydrogen were abrogated by PI3K/Akt inhibitor LY294002. The results indicate that hydrogen protects AECIIs from hyperoxia-induced apoptosis by inhibiting apoptosis factors and promoting the expression of anti-apoptosis factors. These effects were associated with activation of the PI3K/Akt/FoxO3a pathway. Copyright © 2017 Elsevier Inc. All rights reserved.

  4. Liver-Enriched Gene 1, a Glycosylated Secretory Protein, Binds to FGFR and Mediates an Anti-stress Pathway to Protect Liver Development in Zebrafish.

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    Minjie Hu

    2016-02-01

    Full Text Available Unlike mammals and birds, teleost fish undergo external embryogenesis, and therefore their embryos are constantly challenged by stresses from their living environment. These stresses, when becoming too harsh, will cause arrest of cell proliferation, abnormal cell death or senescence. Such organisms have to evolve a sophisticated anti-stress mechanism to protect the process of embryogenesis/organogenesis. However, very few signaling molecule(s mediating such activity have been identified. liver-enriched gene 1 (leg1 is an uncharacterized gene that encodes a novel secretory protein containing a single domain DUF781 (domain of unknown function 781 that is well conserved in vertebrates. In the zebrafish genome, there are two copies of leg1, namely leg1a and leg1b. leg1a and leg1b are closely linked on chromosome 20 and share high homology, but are differentially expressed. In this report, we generated two leg1a mutant alleles using the TALEN technique, then characterized liver development in the mutants. We show that a leg1a mutant exhibits a stress-dependent small liver phenotype that can be prevented by chemicals blocking the production of reactive oxygen species. Further studies reveal that Leg1a binds to FGFR3 and mediates a novel anti-stress pathway to protect liver development through enhancing Erk activity. More importantly, we show that the binding of Leg1a to FGFR relies on the glycosylation at the 70th asparagine (Asn(70 or N(70, and mutating the Asn(70 to Ala(70 compromised Leg1's function in liver development. Therefore, Leg1 plays a unique role in protecting liver development under different stress conditions by serving as a secreted signaling molecule/modulator.

  5. Differentiation of mouse embryonic stem cells into cardiomyocytes via the hanging-drop and mass culture methods.

    Science.gov (United States)

    Fuegemann, Christopher J; Samraj, Ajoy K; Walsh, Stuart; Fleischmann, Bernd K; Jovinge, Stefan; Breitbach, Martin

    2010-12-01

    Herein, we describe two protocols for the in vitro differentiation of mouse embryonic stem cells (mESCs) into cardiomyocytes. mESCs are pluripotent and can be differentiated into cells of all three germ layers, including cardiomyocytes. The methods described here facilitate the differentiation of mESCs into the different cardiac subtypes (atrial-, ventricular-, nodal-like cells). The duration of cell culture determines whether preferentially early- or late-developmental stage cardiomyocytes can be obtained preferentially. This approach allows the investigation of cardiomyocyte development and differentiation in vitro, and also allows for the enrichment and isolation of physiologically intact cardiomyocytes for transplantation purposes. © 2010 by John Wiley & Sons, Inc.

  6. Light Chain Amyloid Fibrils Cause Metabolic Dysfunction in Human Cardiomyocytes.

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    Helen P McWilliams-Koeppen

    Full Text Available Light chain (AL amyloidosis is the most common form of systemic amyloid disease, and cardiomyopathy is a dire consequence, resulting in an extremely poor prognosis. AL is characterized by the production of monoclonal free light chains that deposit as amyloid fibrils principally in the heart, liver, and kidneys causing organ dysfunction. We have studied the effects of amyloid fibrils, produced from recombinant λ6 light chain variable domains, on metabolic activity of human cardiomyocytes. The data indicate that fibrils at 0.1 μM, but not monomer, significantly decrease the enzymatic activity of cellular NAD(PH-dependent oxidoreductase, without causing significant cell death. The presence of amyloid fibrils did not affect ATP levels; however, oxygen consumption was increased and reactive oxygen species were detected. Confocal fluorescence microscopy showed that fibrils bound to and remained at the cell surface with little fibril internalization. These data indicate that AL amyloid fibrils severely impair cardiomyocyte metabolism in a dose dependent manner. These data suggest that effective therapeutic intervention for these patients should include methods for removing potentially toxic amyloid fibrils.

  7. The Correlation of PPARα Activity and Cardiomyocyte Metabolism and Structure in Idiopathic Dilated Cardiomyopathy during Heart Failure Progression

    Directory of Open Access Journals (Sweden)

    E. Czarnowska

    2016-01-01

    Full Text Available This study aimed to define relationship between PPARα expression and metabolic-structural characteristics during HF progression in hearts with DCM phenotype. Tissue endomyocardial biopsy samples divided into three groups according to LVEF ((I 45–50%, n=10; (II 30–40%, n=15; (III 60%, n=6 were investigated. The PPARα mRNA expression in the failing hearts was low in Group (I, high in Group (II, and comparable to that of the control in Group (III. There were analogous changes in the expression of FAT/CD36 and CPT-1 mRNA in contrast to continuous overexpression of GLUT-4 mRNA and significant increase of PDK-4 mRNA in Group (II. In addition, significant structural changes of cardiomyocytes with glycogen accumulation were accompanied by increased expression of PPARα. For the entire study population with HF levels of FAT/CD36 mRNA showed a strong tendency of negative correlation with LVEF. In conclusion, PPARα elevated levels may be a direct cause of adverse remodeling, both metabolic and structural. Thus, there is limited time window for therapy modulating cardiac metabolism and protecting cardiomyocyte structure in failing heart.

  8. Leptin suppresses non-apoptotic cell death in ischemic rat cardiomyocytes by reduction of iPLA{sub 2} activity

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    Takatani-Nakase, Tomoka, E-mail: nakase@mukogawa-u.ac.jp; Takahashi, Koichi, E-mail: koichi@mukogawa-u.ac.jp

    2015-07-17

    Caspase-independent, non-apoptotic cell death is an important therapeutic target in myocardial ischemia. Leptin, an adipose-derived hormone, is known to exhibit cytoprotective effects on the ischemic heart, but the mechanisms are poorly understood. In this research, we found that pretreatment of leptin strongly suppressed ischemic-augmented nuclear shrinkage and non-apoptotic cell death on cardiomyocytes. Leptin was also shown to significantly inhibit the activity of iPLA{sub 2}, which is considered to play crucial roles in non-apoptotic cell death, resulting in effective prevention of ischemia-induced myocyte death. These findings provide the first evidence of a protective mechanism of leptin against ischemia-induced non-apoptotic cardiomyocyte death. - Highlights: • Myocardial ischemia-model induces in caspase-independent, non-apoptotic cell death. • Leptin strongly inhibits ischemic-augmented non-apoptotic cell death. • Leptin reduces iPLA{sub 2} activity, leading to avoidance of non-apoptotic cell death.

  9. Leptin suppresses non-apoptotic cell death in ischemic rat cardiomyocytes by reduction of iPLA2 activity

    International Nuclear Information System (INIS)

    Takatani-Nakase, Tomoka; Takahashi, Koichi

    2015-01-01

    Caspase-independent, non-apoptotic cell death is an important therapeutic target in myocardial ischemia. Leptin, an adipose-derived hormone, is known to exhibit cytoprotective effects on the ischemic heart, but the mechanisms are poorly understood. In this research, we found that pretreatment of leptin strongly suppressed ischemic-augmented nuclear shrinkage and non-apoptotic cell death on cardiomyocytes. Leptin was also shown to significantly inhibit the activity of iPLA 2 , which is considered to play crucial roles in non-apoptotic cell death, resulting in effective prevention of ischemia-induced myocyte death. These findings provide the first evidence of a protective mechanism of leptin against ischemia-induced non-apoptotic cardiomyocyte death. - Highlights: • Myocardial ischemia-model induces in caspase-independent, non-apoptotic cell death. • Leptin strongly inhibits ischemic-augmented non-apoptotic cell death. • Leptin reduces iPLA 2 activity, leading to avoidance of non-apoptotic cell death

  10. Hippo pathway deficiency reverses systolic heart failure after infarction.

    Science.gov (United States)

    Leach, John P; Heallen, Todd; Zhang, Min; Rahmani, Mahdis; Morikawa, Yuka; Hill, Matthew C; Segura, Ana; Willerson, James T; Martin, James F

    2017-10-12

    Mammalian organs vary widely in regenerative capacity. Poorly regenerative organs, such as the heart are particularly vulnerable to organ failure. Once established, heart failure commonly results in mortality. The Hippo pathway, a kinase cascade that prevents adult cardiomyocyte proliferation and regeneration, is upregulated in human heart failure. Here we show that deletion of the Hippo pathway component Salvador (Salv) in mouse hearts with established ischaemic heart failure after myocardial infarction induces a reparative genetic program with increased scar border vascularity, reduced fibrosis, and recovery of pumping function compared with controls. Using translating ribosomal affinity purification, we isolate cardiomyocyte-specific translating messenger RNA. Hippo-deficient cardiomyocytes have increased expression of proliferative genes and stress response genes, such as the mitochondrial quality control gene, Park2. Genetic studies indicate that Park2 is essential for heart repair, suggesting a requirement for mitochondrial quality control in regenerating myocardium. Gene therapy with a virus encoding Salv short hairpin RNA improves heart function when delivered at the time of infarct or after ischaemic heart failure following myocardial infarction was established. Our findings indicate that the failing heart has a previously unrecognized reparative capacity involving more than cardiomyocyte renewal.

  11. The hippo pathway in heart development, regeneration, and diseases.

    Science.gov (United States)

    Zhou, Qi; Li, Li; Zhao, Bin; Guan, Kun-Liang

    2015-04-10

    The heart is the first organ formed during mammalian development. A properly sized and functional heart is vital throughout the entire lifespan. Loss of cardiomyocytes because of injury or diseases leads to heart failure, which is a major cause of human morbidity and mortality. Unfortunately, regenerative potential of the adult heart is limited. The Hippo pathway is a recently identified signaling cascade that plays an evolutionarily conserved role in organ size control by inhibiting cell proliferation, promoting apoptosis, regulating fates of stem/progenitor cells, and in some circumstances, limiting cell size. Interestingly, research indicates a key role of this pathway in regulation of cardiomyocyte proliferation and heart size. Inactivation of the Hippo pathway or activation of its downstream effector, the Yes-associated protein transcription coactivator, improves cardiac regeneration. Several known upstream signals of the Hippo pathway such as mechanical stress, G-protein-coupled receptor signaling, and oxidative stress are known to play critical roles in cardiac physiology. In addition, Yes-associated protein has been shown to regulate cardiomyocyte fate through multiple transcriptional mechanisms. In this review, we summarize and discuss current findings on the roles and mechanisms of the Hippo pathway in heart development, injury, and regeneration. © 2015 American Heart Association, Inc.

  12. Remote ischemic preconditioning protects liver ischemia-reperfusion injury by regulating eNOS-NO pathway and liver microRNA expressions in fatty liver rats.

    Science.gov (United States)

    Duan, Yun-Fei; An, Yong; Zhu, Feng; Jiang, Yong

    2017-08-15

    Ischemic preconditioning (IPC) is a strategy to reduce ischemia-reperfusion (I/R) injury. The protective effect of remote ischemic preconditioning (RIPC) on liver I/R injury is not clear. This study aimed to investigate the roles of RIPC in liver I/R in fatty liver rats and the involvement of endothelial nitric oxide synthase-nitric oxide (eNOS-NO) pathway and microRNA expressions in this process. A total of 32 fatty rats were randomly divided into the sham group, I/R group, RIPC group and RIPC+I/R group. Serum alanine aminotransferase (ALT), aspartate aminotransferase (AST) and nitric oxide (NO) were measured. Hematoxylin-eosin staining was used to observe histological changes of liver tissues, TUNEL to detect hepatocyte apoptosis, and immunohistochemistry assay to detect heat shock protein 70 (HSP70) expression. Western blotting was used to detect liver inducible NOS (iNOS) and eNOS protein levels and real-time quantitative polymerase chain reaction to detect miR-34a, miR-122 and miR-27b expressions. Compared with the sham and RIPC groups, serum ALT, AST and iNOS in liver tissue were significantly higher in other two groups, while serum NO and eNOS in liver tissue were lower, and varying degrees of edema, degeneration and inflammatory cell infiltration were found. Cell apoptosis number was slightly lower in the RIPC+I/R group than that in I/R group. Compared with the sham group, HSP70 expressions were significantly increased in other three groups (all Pfatty liver I/R injury by affecting the eNOS-NO pathway and liver microRNA expressions. Copyright © 2017 The Editorial Board of Hepatobiliary & Pancreatic Diseases International. Published by Elsevier B.V. All rights reserved.

  13. Protective effects of curcumin against mercury-induced hepatic injuries in rats, involvement of oxidative stress antagonism, and Nrf2-ARE pathway activation.

    Science.gov (United States)

    Liu, W; Xu, Z; Li, H; Guo, M; Yang, T; Feng, S; Xu, B; Deng, Yu

    2017-09-01

    Mercury (Hg) represents a ubiquitous environmental heavy metal that could lead to severe toxic effects in a variety of organs usually at a low level. The present study focused on the liver oxidative stress, one of the most important roles playing in Hg hepatotoxicity, by evaluation of different concentrations of mercuric chloride (HgCl 2 ) administration. Moreover, the protective potential of curcumin against Hg hepatotoxic effects was also investigated. Eighty-four rats were randomly divided into six groups for a three-days experiment: control, dimethyl sulfoxide control, HgCl 2 treatment (0.6, 1.2, and 2.4 mg kg -1 day -1 ), and curcumin pretreatment (100 mg kg -1 day -1 ) groups. Exposure of HgCl 2 resulted in acute dose-dependent hepatotoxic effects. Administration of 2.4 mg kg -1 HgCl 2 significantly elevated total Hg, nonprotein sulfhydryl, reactive oxygen species formation, malondialdehyde, apoptosis levels, serum lactate dehydrogenase, and alanine transaminase activities, with an impairment of superoxide dismutase and glutathione peroxidase in the liver. Moreover, HgCl 2 treatment activated nuclear factor-E2-related factor 2-antioxidant response element (Nrf2-ARE) signaling pathway in further investigation, with a significant upregulation of Nrf2, heme oxygenase-1, and γ-glutamylcysteine synthetase heavy subunit expression, relative to control. Pretreatment with curcumin obviously prevented HgCl 2 -induced liver oxidative stress, which may be due to its free radical scavenging or Nrf2-ARE pathway-inducing properties. Taking together these data suggest that curcumin counteracts HgCl 2 hepatotoxicity through antagonizing liver oxidative stress.

  14. Benfotiamine improves functional recovery of the infarcted heart via activation of pro-survival G6PD/Akt signaling pathway and modulation of neurohormonal response.

    Science.gov (United States)

    Katare, Rajesh; Caporali, Andrea; Emanueli, Costanza; Madeddu, Paolo

    2010-10-01

    Benfotiamine (BFT) is a transketolase activator that directs glucose to the pentose phosphate pathway. The present study investigated whether BFT improves the recovery after myocardial infarction (MI) and explored underlying mechanisms of protection. Non-diabetic and streptozotocin-induced type 1 diabetic mice were supplemented with BFT (70 mg/kg/day in drinking water) for 4 weeks and then subjected to MI or sham operation. Cardiac function was monitored by echocardiography. At two weeks post-MI, intra-ventricular pressure was measured by Millar tip-catheter and hearts were collected for biochemical, immunohistochemical and expressional analyses. No treatment effect was observed in sham-operated mice. Post-MI mortality was higher in diabetic mice and hemodynamic studies confirmed the worsening effect of diabetes on functional recovery. Furthermore, diabetic mice demonstrated increased cardiomyocyte apoptosis, reduced reparative angiogenesis, larger scars, enhanced oxidative stress, and blunted activation of the pro-survival VEGF receptor-2/Akt/Pim-1 signaling pathway. BFT improved post-MI survival, functional recovery and neovascularization and reduced cardiomyocyte apoptosis and neurohormonal activation in diabetic as well as in non-diabetic mice. In addition, BFT stimulated the activity of pentose phosphate pathway enzymes, leading to reduction of oxidative stress, phosphorylation/activation of VEGF receptor-2 and Akt and increased Pim-1, pBad and Bcl-2 levels. These effects were contrasted on silencing glucose-6-phosphate dehydrogenase, the key enzyme in pentose phosphate pathway, or inhibiting Akt. BFT benefits post-MI recovery through stimulation of pro-survival mechanisms and containment of neurohormonal response. These results may have implications for the treatment of myocardial ischemia. Copyright © 2010 Elsevier Ltd. All rights reserved.

  15. Taurine prevents arsenic-induced cardiac oxidative stress and apoptotic damage: Role of NF-κB, p38 and JNK MAPK pathway

    International Nuclear Information System (INIS)

    Ghosh, Jyotirmoy; Das, Joydeep; Manna, Prasenjit; Sil, Parames C.

    2009-01-01

    Cardiac dysfunction is a major cause of morbidity and mortality worldwide due to its complex pathogenesis. However, little is known about the mechanism of arsenic-induced cardiac abnormalities and the use of antioxidants as the possible protective agents in this pathophysiology. Conditionally essential amino acid, taurine, accounts for 25% to 50% of the amino acid pool in myocardium and possesses antioxidant properties. The present study has, therefore, been carried out to investigate the underlying mechanism of the beneficial role of taurine in arsenic-induced cardiac oxidative damage and cell death. Arsenic reduced cardiomyocyte viability, increased reactive oxygen species (ROS) production and intracellular calcium overload, and induced apoptotic cell death by mitochondrial dependent caspase-3 activation and poly-ADP ribose polymerase (PARP) cleavage. These changes due to arsenic exposure were found to be associated with increased IKK and NF-κB (p65) phosphorylation. Pre-exposure of myocytes to an IKK inhibitor (PS-1145) prevented As-induced caspase-3 and PARP cleavage. Arsenic also markedly increased the activity of p38 and JNK MAPKs, but not ERK to that extent. Pre-treatment with SP600125 (JNK inhibitor) and SB203580 (p38 MAPK inhibitor) attenuated NF-κB and IKK phosphorylation indicating that p38 and JNK MAPKs are mainly involved in arsenic-induced NF-κB activation. Taurine treatment suppressed these apoptotic actions, suggesting that its protective role in arsenic-induced cardiomyocyte apoptosis is mediated by attenuation of p38 and JNK MAPK signaling pathways. Similarly, arsenic intoxication altered a number of biomarkers related to cardiac oxidative stress and other apoptotic indices in vivo and taurine supplementation could reduce it. Results suggest that taurine prevented arsenic-induced myocardial pathophysiology, attenuated NF-κB activation via IKK, p38 and JNK MAPK signaling pathways and could possibly provide a protection against As

  16. Innervating sympathetic neurons regulate heart size and the timing of cardiomyocyte cell cycle withdrawal.

    Science.gov (United States)

    Kreipke, R E; Birren, S J

    2015-12-01

    Sympathetic drive to the heart is a key modulator of cardiac function and interactions between heart tissue and innervating sympathetic fibres are established early in development. Significant innervation takes place during postnatal heart development, a period when cardiomyocytes undergo a rapid transition from proliferative to hypertrophic growth. The question of whether these innervating sympathetic fibres play a role in regulating the modes of cardiomyocyte growth was investigated using 6-hydroxydopamine (6-OHDA) to abolish early sympathetic innervation of the heart. Postnatal chemical sympathectomy resulted in rats with smaller hearts, indicating that heart growth is regulated by innervating sympathetic fibres during the postnatal period. In vitro experiments showed that sympathetic interactions resulted in delays in markers of cardiomyocyte maturation, suggesting that changes in the timing of the transition from hyperplastic to hypertrophic growth of cardiomyocytes could underlie changes in heart size in the sympathectomized animals. There was also an increase in the expression of Meis1, which has been linked to cardiomyocyte cell cycle withdrawal, suggesting that sympathetic signalling suppresses cell cycle withdrawal. This signalling involves β-adrenergic activation, which was necessary for sympathetic regulation of cardiomyocyte proliferation and hypertrophy. The effect of β-adrenergic signalling on cardiomyocyte hypertrophy underwent a developmental transition. While young postnatal cardiomyocytes responded to isoproterenol (isoprenaline) with a decrease in cell size, mature cardiomyocytes showed an increase in cell size in response to the drug. Together, these results suggest that early sympathetic effects on proliferation modulate a key transition between proliferative and hypertrophic growth of the heart and contribute to the sympathetic regulation of adult heart size. © 2015 The Authors. The Journal of Physiology © 2015 The Physiological Society.

  17. Effects of gamma-ray radiation on activity and apoptosis of rat cardiomyocytes in vitro

    International Nuclear Information System (INIS)

    Hu Shunying; Jiang Changsheng; Chen Guowei; Duan Haifeng; Wang Rongliang; Wu Bin; Guo Zikuan; Wang Lisheng

    2007-01-01

    Objective: It is reported that radiation-induced myocardial degeneration in the rat is preceded by changes in capillary structure and function. The aim of the present study is to investigate direct effect of gamma ray radiation on activity and apoptosis of cultured rat cardiomyocytes in vitro. Methods: The study was performed using primary cell cultures of cardiomyocytes isolated from hearts of now-born rats. After being cultured for 72h in vitro, cardiomyocytes were irradiated with single dose of 5 Gy, 10 Gy, 20 Gy of gamma ray respectively. At 48h post-irradiation, the concentration of LDH in the supernatant of cell culture was tested using methods introduced by International Federation of clinical chemistry (IFCC), and apoptosis was determined with flow cytometry. The viability of myocytes was determined with crystal violet test and MTT test at 48h and 120h post-irradiation respectively. Results: LDH concentration in the supernatant of cell culture of cardiomyocytes were increased significantly with the irradiation dose augment. Flow cytometry confirmed the induction of apoptosis in response to different gamma ray doses irradiation at 48h after irradiation. The viable cardiomyocytes irradiated by gamma ray were significantly declined at 120h after irradiation compared to un-irradiated cells, however there were no significant difference between two groups at 48h post-irradiation. Dose-effect relationship was demonstrated between cardiomyocyte apoptosis, viability and irradiation dose in the study. Conclusion: The study demonstrates gamma ray radiation can cause direct damage to cultured cardiomyocytes, including inhibiting activity and inducing apoptosis of cardiomyocytes in vitro, which shows dose effect relationship. The mechanism of gamma ray irradiation induced injury to cardiomyocytes should be investigated further. (authors)

  18. Cardiomyocyte architectural plasticity in fetal, neonatal, and adult pig hearts delineated with diffusion tensor MRI.

    Science.gov (United States)

    Zhang, Lei; Allen, John; Hu, Lingzhi; Caruthers, Shelton D; Wickline, Samuel A; Chen, Junjie

    2013-01-15

    Cardiomyocyte organization is a critical determinant of coordinated cardiac contractile function. Because of the acute opening of the pulmonary circulation, the relative workload of the left ventricle (LV) and right ventricle (RV) changes substantially immediately after birth. We hypothesized that three-dimensional cardiomyocyte architecture might be required to adapt rapidly to accommodate programmed perinatal changes of cardiac function. Isolated fixed hearts from pig fetuses or pigs at midgestation, preborn, postnatal day 1 (P1), postnatal day 5, postnatal day 14 (P14), and adulthood (n = 5 for each group) were acquired for diffusion-weighted magnetic resonance imaging. Cardiomyocyte architecture was visualized by three-dimensional fiber tracking and was quantitatively evaluated by the measured helix angle (α(h)). Upon the completion of MRI, hearts were sectioned and stained with hematoxylin/eosin (H&E) to evaluate cardiomyocyte alignment, with picrosirius red to evaluate collagen content, and with anti-Ki67 to evaluate postnatal cell proliferation. The helical architecture of cardiomyocyte was observed as early as the midgestational period. Postnatal changes of cardiomyocyte architecture were observed from P1 to P14, which primary occurred in the septum and RV free wall (RVFW). In the septum, the volume ratio of LV- vs. RV-associated cardiomyocytes rapidly changed from RV-LV balanced pattern at birth to LV dominant pattern by P14. In the RVFW, subendocardial α(h) decreased by ~30° from P1 to P14. These findings indicate that the helical architecture of cardiomyocyte is developed as early as the midgestation period. Substantial and rapid adaptive changes in cardiac microarchitecture suggested considerable developmental plasticity of cardiomyocyte form and function in the postnatal period in response to altered cardiac mechanical function.

  19. Signaling Pathways in Cardiac Myocyte Apoptosis

    Science.gov (United States)

    Xia, Peng; Liu, Yuening

    2016-01-01

    Cardiovascular diseases, the number 1 cause of death worldwide, are frequently associated with apoptotic death of cardiac myocytes. Since cardiomyocyte apoptosis is a highly regulated process, pharmacological intervention of apoptosis pathways may represent a promising therapeutic strategy for a number of cardiovascular diseases and disorders including myocardial infarction, ischemia/reperfusion injury, chemotherapy cardiotoxicity, and end-stage heart failure. Despite rapid growth of our knowledge in apoptosis signaling pathways, a clinically applicable treatment targeting this cellular process is currently unavailable. To help identify potential innovative directions for future research, it is necessary to have a full understanding of the apoptotic pathways currently known to be functional in cardiac myocytes. Here, we summarize recent progress in the regulation of cardiomyocyte apoptosis by multiple signaling molecules and pathways, with a focus on the involvement of these pathways in the pathogenesis of heart disease. In addition, we provide an update regarding bench to bedside translation of this knowledge and discuss unanswered questions that need further investigation. PMID:28101515

  20. Sodium Ferulate Prevents Daunorubicin - Induced Apoptosis in H9c2 Cells via Inhibition of the ERKs Pathway

    Directory of Open Access Journals (Sweden)

    Zhi-Juan Wu

    2015-07-01

    Full Text Available Background: Daunorubicin (DNR-induced cardiotoxicity, which is closely associated with cardiomyocyte apoptosis, limits the drug's clinical application. The activation of the extracellular regulated protein kinases (ERKs pathway is responsible for the pro-apoptosis effect of DNR Sodium ferulate (SF has recently been found to attenuate both DNR-induced cardiotoxicity and mitochondrial apoptosis in juvenile rats. Nonetheless, the precise mechanism underlying SF-induced cardio-protection remains unclear. Methods: The DNR-injured H9c2 cell model was prepared by incubating the cells in 1 µM DNR for 24 h. Amounts of 15.6, 31.3 or 62.5 µM SF were simultaneously added to the cells. The effect of SF on the cytotoxic and apoptotic parameters of the cells was studied by monitoring apoptosis regulation via the ERKs pathway. Results: SF attenuated DNR-induced cell death (particularly apoptotic death, cTnI and β-tubulin degradation, and cellular morphological changes. SF reduced mitochondrial membrane potential depolarization, cytochrome c leakage, and caspase-9 and caspase-3 activation. SF also decreased ERK1/2, phospho-ERK1/2, p53 and Bax expression and increased Bcl-2 expression. These effects were similar to the results observed when using the pharmacological ERKs phosphorylation inhibitor, AZD6244. Conclusion: We determined that SF protects H9c2 cells from DNR-induced apoptosis through a mechanism that involves the interruption of the ERKs signaling pathway.

  1. Fluoxetine protects against methamphetamine‑induced lung inflammation by suppressing oxidative stress through the SERT/p38 MAPK/Nrf2 pathway in rats.

    Science.gov (United States)

    Wang, Yun; Gu, Yu-Han; Liu, Ming; Bai, Yang; Wang, Huai-Liang

    2017-02-01

    Methamphetamine (MA) abuse is a major public health and safety concern throughout the world and a growing burden on healthcare costs. The purpose of the present study was to investigate the protective effect of fluoxetine against MA‑induced chronic pulmonary inflammation and to evaluate the potential role of nuclear factor erythroid 2-related factor 2 (Nrf2)-mediated antioxidative stress. Wistar rats were divided into control, MA and two fluoxetine‑treated groups. Rats in the MA and the two fluoxetine‑treated groups were treated daily with intraperitoneal injection of 10 mg/kg MA twice daily. Rats in the two fluoxetine‑treated groups were injected intragastrically with fluoxetine (2 and 10 mg/kg) once daily, respectively. After 5 weeks, the rats were euthanized and hematoxylin and eosin staining, immunohistochemistry, western blot analysis and redox assay were performed. It was demonstrated that chronic exposure to MA can induce pulmonary inflammation in rats, with the symptoms of inflammatory cell infiltration, crowded lung parenchyma, thickened septum and a reduced number of alveolar sacs. Fluoxetine attenuated pulmonary inflammation and the expression of interleukin‑6 and tumor necrosis factor‑α in rat lungs. Fluoxetine inhibited MA‑induced increases in the expression levels of serotonin transporter (SERT) and p‑p38 mitogen‑activated protein kinase (MAPK), and reversed the MA‑induced decrease in nuclear Nrf2 and human heme oxygenase‑1 in lungs. Fluoxetine at 10 mg/kg significantly reversed the reduced glutathione (GSH) level, the ratio of GSH/oxidized glutathione, and the reactive oxygen species level in rat lungs from the MA group. These findings suggested that fluoxetine, a SERT inhibitor, has a protective effect against MA‑induced lung inflammation by suppressing oxidative stress through the SERT/p38 MAPK/Nrf2 pathway in rats.

  2. Regulatory T cells protect mice against coxsackievirus-induced myocarditis through the transforming growth factor beta-coxsackie-adenovirus receptor pathway.

    Science.gov (United States)

    Shi, Yu; Fukuoka, Masahiro; Li, Guohua; Liu, Youan; Chen, Manyin; Konviser, Michael; Chen, Xin; Opavsky, Mary Anne; Liu, Peter P

    2010-06-22

    Coxsackievirus B3 infection is an excellent model of human myocarditis and dilated cardiomyopathy. Cardiac injury is caused either by a direct cytopathic effect of the virus or through immune-mediated mechanisms. Regulatory T cells (Tregs) play an important role in the negative modulation of host immune responses and set the threshold of autoimmune activation. This study was designed to test the protective effects of Tregs and to determine the underlying mechanisms. Carboxyfluorescein diacetate succinimidyl ester-labeled Tregs or naïve CD4(+) T cells were injected intravenously once every 2 weeks 3 times into mice. The mice were then challenged with intraperitoneal coxsackievirus B3 immediately after the last cell transfer. Transfer of Tregs showed higher survival rates than transfer of CD4(+) T cells (P=0.0136) but not compared with the PBS injection group (P=0.0589). Interestingly, Tregs also significantly decreased virus titers and inflammatory scores in the heart. Transforming growth factor-beta and phosphorylated AKT were upregulated in Tregs-transferred mice and coxsackie-adenovirus receptor expression was decreased in the heart compared with control groups. Transforming growth factor-beta decreased coxsackie-adenovirus receptor expression and inhibited coxsackievirus B3 infection in HL-1 cells and neonatal cardiac myocytes. Splenocytes collected from Treg-, CD4(+) T-cell-, and PBS-treated mice proliferated equally when stimulated with heat-inactivated virus, whereas in the Treg group, the proliferation rate was reduced significantly when stimulated with noninfected heart tissue homogenate. Adoptive transfer of Tregs protected mice from coxsackievirus B3-induced myocarditis through the transforming growth factor beta-coxsackie-adenovirus receptor pathway and thus suppresses the immune response to cardiac tissue, maintaining the antiviral immune response.

  3. Taurine protects HK-2 cells from oxidized LDL-induced cytotoxicity via the ROS-mediated mitochondrial and p53-related apoptotic pathways

    Energy Technology Data Exchange (ETDEWEB)

    Chang, Chun-Yu [Graduate Institute of Biomedical Sciences, National Chung Hsing University, Taichung, Taiwan (China); Shen, Chao-Yu [School of Medical Imaging and Radiological Sciences, Chung Shan Medical University, Taichung, Taiwan (China); Department of Medical Imaging, Chung Shan Medical University Hospital, Taichung, Taiwan (China); School of Medicine, Chung Shan Medical University, Taichung, Taiwan (China); Kang, Chao-Kai [Department of Life Sciences, National Chung Hsing University, Taichung, Taiwan, (China); Sher, Yuh-Pyng [Graduate Institute of Clinical Medical Science, China Medical University, Taichung, Taiwan (China); Center for Molecular Medicine, China Medical University Hospital, Taichung 404, Taiwan (China); Sheu, Wayne H.-H. [Graduate Institute of Biomedical Sciences, National Chung Hsing University, Taichung, Taiwan (China); Division of Endocrinology and Metabolism, Department of Internal Medicine, Taichung Veterans General Hospital, Taichung, Taiwan (China); School of Medicine, National Yang Ming University, Taipei, Taiwan (China); School of Medicine, National Defense Medical Center, Taipei, Taiwan (China); Chang, Chia-Che, E-mail: chia_che@dragon.nchu.edu.tw [Graduate Institute of Biomedical Sciences, National Chung Hsing University, Taichung, Taiwan (China); Agricultural Biotechnology Center, National Chung Hsing University, Taichung, Taiwan (China); Lee, Tsung-Han, E-mail: thlee@email.nchu.edu.tw [Department of Life Sciences, National Chung Hsing University, Taichung, Taiwan, (China); Graduate Institute of Clinical Medical Science, China Medical University, Taichung, Taiwan (China); Agricultural Biotechnology Center, National Chung Hsing University, Taichung, Taiwan (China); Department of Biological Science and Technology, China Medical University, Taichung, Taiwan (China)

    2014-09-15

    Oxidized LDL (oxLDL) induces a pro-oxidative environment and promotes apoptosis, causing the progression of renal diseases in humans. Taurine is a semi-essential amino acid in mammals and has been shown to be a potent endogenous antioxidant. The kidney plays a pivotal role in maintaining the balance of taurine. However, the mechanisms underlying the protective effects of taurine against oxLDL-induced injury in renal epithelial cells have not been clarified. In the present study, we investigated the anti-apoptotic effects of taurine on human proximal tubular epithelial (HK-2) cells exposed to oxLDL and explored the related mechanisms. We observed that oxLDL increased the contents of ROS and of malondialdehyde (MDA), which is a lipid peroxidation by-product that acts as an indicator of the cellular oxidation status. In addition, oxLDL induced cell death and apoptosis in HK-2 cells. Pretreatment with taurine at 100 μM significantly attenuated the oxLDL-induced cytotoxicity. We determined that oxLDL triggered the phosphorylation of ERK and, in turn, the activation of p53 and other apoptosis-related events, including calcium accumulation, destabilization of the mitochondrial permeability and disruption of the balance between pro-apoptotic Bax and anti-apoptotic Bcl-2 proteins. The malfunctions induced by oxLDL were effectively blocked by taurine. Thus, our results suggested that taurine exhibits potential therapeutic activity by preventing oxLDL-induced nephrotoxicity. The inhibition of oxLDL-induced epithelial apoptosis by taurine was at least partially due to its anti-oxidant activity and its ability to modulate the ERK and p53 apoptotic pathways. - Highlights: • Oxidized LDL induced cytotoxicity and apoptosis in HK-2 cells. • Pretreatment with taurine attenuated oxLDL-induced nephrotoxicity. • Taurine protected against renal damages through inhibition of ROS generation. • Taurine prevented apoptosis through modulation of the p53 phosphorylation.

  4. Interferon-β induced in female genital epithelium by HIV-1 glycoprotein 120 via Toll-like-receptor 2 pathway acts to protect the mucosal barrier.

    Science.gov (United States)

    Nazli, Aisha; Dizzell, Sara; Zahoor, Muhammad Atif; Ferreira, Victor H; Kafka, Jessica; Woods, Matthew William; Ouellet, Michel; Ashkar, Ali A; Tremblay, Michel J; Bowdish, Dawn Me; Kaushic, Charu

    2018-03-19

    More than 40% of HIV infections occur via female reproductive tract (FRT) through heterosexual transmission. Epithelial cells that line the female genital mucosa are the first line of defense against HIV-1 and other sexually transmitted pathogens. These sentient cells recognize and respond to external stimuli by induction of a range of carefully balanced innate immune responses. Previously, we have shown that in response to HIV-1 gp120, the genital epithelial cells (GECs) from upper reproductive tract induce an inflammatory response that may facilitate HIV-1 translocation and infection. In this study, we report that the endometrial and endocervical GECs simultaneously induce biologically active interferon-β (IFNβ) antiviral responses following exposure to HIV-1 that act to protect the epithelial tight junction barrier. The innate antiviral response was directly induced by HIV-1 envelope glycoprotein gp120 and addition of gp120 neutralizing antibody inhibited IFNβ production. Interferon-β was induced by gp120 in upper GECs through Toll-like receptor 2 signaling and required presence of heparan sulfate on epithelial cell surface. The induction of IFNβ was dependent upon activation of transcription factor IRF3 (interferon regulatory factor 3). The IFNβ was biologically active, had a protective effect on epithelial tight junction barrier and was able to inhibit HIV-1 infection in TZM-bl indicator cells and HIV-1 replication in T cells. This is the first report that recognition of HIV-1 by upper GECs leads to induction of innate antiviral pathways. This could explain the overall low infectivity of HIV-1 in the FRT and could be exploited for HIV-1 prophylaxis.Cellular and Molecular Immunology advance online publication, 19 March 2018; doi:10.1038/cmi.2017.168.

  5. Taurine protects HK-2 cells from oxidized LDL-induced cytotoxicity via the ROS-mediated mitochondrial and p53-related apoptotic pathways

    International Nuclear Information System (INIS)

    Chang, Chun-Yu; Shen, Chao-Yu; Kang, Chao-Kai; Sher, Yuh-Pyng; Sheu, Wayne H.-H.; Chang, Chia-Che; Lee, Tsung-Han

    2014-01-01

    Oxidized LDL (oxLDL) induces a pro-oxidative environment and promotes apoptosis, causing the progression of renal diseases in humans. Taurine is a semi-essential amino acid in mammals and has been shown to be a potent endogenous antioxidant. The kidney plays a pivotal role in maintaining the balance of taurine. However, the mechanisms underlying the protective effects of taurine against oxLDL-induced injury in renal epithelial cells have not been clarified. In the present study, we investigated the anti-apoptotic effects of taurine on human proximal tubular epithelial (HK-2) cells exposed to oxLDL and explored the related mechanisms. We observed that oxLDL increased the contents of ROS and of malondialdehyde (MDA), which is a lipid peroxidation by-product that acts as an indicator of the cellular oxidation status. In addition, oxLDL induced cell death and apoptosis in HK-2 cells. Pretreatment with taurine at 100 μM significantly attenuated the oxLDL-induced cytotoxicity. We determined that oxLDL triggered the phosphorylation of ERK and, in turn, the activation of p53 and other apoptosis-related events, including calcium accumulation, destabilization of the mitochondrial permeability and disruption of the balance between pro-apoptotic Bax and anti-apoptotic Bcl-2 proteins. The malfunctions induced by oxLDL were effectively blocked by taurine. Thus, our results suggested that taurine exhibits potential therapeutic activity by preventing oxLDL-induced nephrotoxicity. The inhibition of oxLDL-induced epithelial apoptosis by taurine was at least partially due to its anti-oxidant activity and its ability to modulate the ERK and p53 apoptotic pathways. - Highlights: • Oxidized LDL induced cytotoxicity and apoptosis in HK-2 cells. • Pretreatment with taurine attenuated oxLDL-induced nephrotoxicity. • Taurine protected against renal damages through inhibition of ROS generation. • Taurine prevented apoptosis through modulation of the p53 phosphorylation

  6. Emodin Protects Against Concanavalin A-Induced Hepatitis in Mice Through Inhibiting Activation of the p38 MAPK-NF-κB Signaling Pathway

    Directory of Open Access Journals (Sweden)

    Jihua Xue

    2015-03-01

    Full Text Available Background/Aims: To investigate the effects of emodin on concanavalin A (Con A-induced hepatitis in mice and to elucidate its underlying molecular mechanisms. Methods: A fulminant hepatitis model was established successfully by the intravenous administration of Con A (20 mg/kg to male Balb/c mice. Emodin was administered to the mice by gavage before and after Con A injection. The levels of pro-inflammatory cytokines and chemokines, numbers of CD4+ and F4/80+ cells infiltrated into the liver, and amounts of phosphorylated p38 MAPK and NF-γB in mouse livers and RAW264.7 and EL4 cells were measured. Results: Pretreatment with emodin significantly protected the animals from T cell-mediated hepatitis, as shown by the decreased elevations of serum alanine aminotransferase (ALT and aspartate aminotransferase (AST, as well as reduced hepatic necrosis. In addition, emodin pretreatment markedly reduced the intrahepatic expression of pro-inflammatory cytokines and chemokines, including tumor necrosis factor (TNF-a, interferon (IFN-γ, interleukin (IL-1ß, IL-6, IL-12, inducible nitric oxide synthase (iNOS, integrin alpha M (ITGAM, chemokine (C-C motif ligand 2 (CCL2, macrophage inflammatory protein 2 (MIP-2 and chemokine (CXC motif receptor 2 (CXCR2. Furthermore, emodin pretreatment dramatically suppressed the numbers of CD4+ and F4/80+ cells infiltrating into the liver as well as the activation of p38 MAPK and NF-γB in Con A-treated mouse livers and RAW264.7 and EL4 cells. Conclusion: The results indicate that emodin pretreatment protects against Con A-induced liver injury in mice; these beneficial effects may occur partially through inhibition of both the infiltration of CD4+ and F4/80+ cells and the activation of the p38 MAPK-NF-γB pathway in CD4+ T cells and macrophages.

  7. Inhibition of Autophagy via Activation of PI3K/Akt Pathway Contributes to the Protection of Ginsenoside Rb1 against Neuronal Death Caused by Ischemic Insults

    Directory of Open Access Journals (Sweden)

    Tianfei Luo

    2014-09-01

    Full Text Available Lethal autophagy is a pathway leading to neuronal death caused by transient global ischemia. In this study, we examined the effect of Ginsenoside Rb1 (GRb1 on ischemia/reperfusion-induced autophagic neuronal death and investigated the role of PI3K/Akt. Ischemic neuronal death in vitro was induced by using oxygen glucose deprivation (OGD in SH-SY5Y cells, and transient global ischemia was produced by using two vessels occlusion in rats. Cellular viability of SH-SY5Y cells was assessed by MTT assay, and CA1 neuronal death was evaluated by Hematoxylin-eosin staining. Autophagic vacuoles were detected by using both fluorescent microscopy in combination with acridine orange (AO and Monodansylcadaverine (MDC staining and transmission electronic microscopy. Protein levels of LC3II, Beclin1, total Akt and phosphor-Akt at Ser473 were examined by western blotting analysis. GRb1 inhibited both OGD and transient ischemia-induced neuronal death and mitigated OGD-induced autophagic vacuoles in SH-SY5Y cells. By contrast, PI3K inhibitor LY294002 counteracted the protection of GRb1 against neuronal death caused by either OGD or transient ischemia. LY294002 not only mitigated the up-regulated protein level of phosphor Akt at Ser473 caused by GRb1, but also reversed the inhibitory effect of GRb1 on OGD and transient ischemia-induced elevation in protein levels of LC3II and Beclin1.

  8. L-3-n-Butylphthalide Protects HSPB8 K141N Mutation-Induced Oxidative Stress by Modulating the Mitochondrial Apoptotic and Nrf2 Pathways

    Directory of Open Access Journals (Sweden)

    Xiao-Dong Yang

    2017-07-01

    Full Text Available Charcot–Marie–Tooth disease (CMT, also known as hereditary motor and sensory neuropathy, is the most common inherited peripheral nerve disorder. Missense mutations, such as K141N, in the small heat shock protein HSPB8 are known to cause distal hereditary motor neuropathy 2A (dHMN2A or Charcot-Marie-Tooth neuropathy type 2L (CMT2L. However, of critical clinical significance, very few specific therapies for this disease exist. In the present study, we investigated the impact of mutant K141N HSPB8 on mitochondrial distribution and function in a cellular model of CMT2L. Our results indicate that K141N HSPB8 induced mitochondrial aggregation and caused increased oxidative stress injury. As an extraction from Chinese celery Apium graveolens Linn seeds, L-3-n-Butylphthalide (NBP, has been reported to exert many neuroprotective effects, we interrogated whether NBP could elicit a protective effect on the cell injury typically caused by HSPB8 K141N mutations. We found NBP could reverse the pathological processes induced by HSPB8 K141N mutation via an antioxidant effect, modulation of the Bax/Bcl-2 mitochondrial apoptotic and Nrf2 pathways. We propose a novel function of HSPB8, highlighting the consequence of the K141N pathogenic mutation. Furthermore, we suggest NBP may have promising therapeutic potential in the treatment of CMT2L.

  9. L-cysteine protects intestinal integrity, attenuates intestinal inflammation and oxidant stress, and modulates NF-κB and Nrf2 pathways in weaned piglets after LPS challenge.

    Science.gov (United States)

    Song, Ze he; Tong, Guo; Xiao, Kan; Jiao, Le fei; Ke, Ya lu; Hu, Cai hong

    2016-04-01

    In this study we investigated whetherL-cysteine (L-cys) could alleviate LPS-induced intestinal disruption and its underlying mechanism. Piglets fed with anL-cys-supplemented diet had higher average daily gain.L-cys alleviated LPS-induced structural and functional disruption of intestine in weanling piglets, as demonstrated by higher villus height, villus height (VH) to crypt depth (CD) ratio, and transepithelial electrical resistance (TER) and lower FITC-dextran 4 (FD4) kDa flux in jejunum and ileum. Supplementation withL-cys up-regulated occludin and claudin-1 expression, reduced caspase-3 activity and enhanced proliferating cell nuclear antigen expression of jejunum and ileum relative to LPS group. Additionally,L-cys suppressed the LPS-induced intestinal inflammation and oxidative stress, as demonstrated by down-regulated TNF-α, IL-6 and IL-8 mRNA levels, increased catalase, superoxide dismutase, glutathione peroxidase activity, glutathione (GSH) contents and the ratio of GSH and oxidized glutathione in jejunum and ileum. Finally, a diet supplemented withL-cys inhibited NF-κB(p65) nuclear translocation and elevated NF erythroid 2-related factor 2 (Nrf2) translocation compared with the LPS group. Collectively, our results indicated the protective function ofL-cys on intestinal mucosa barrier may closely associated with its anti-inflammation, antioxidant and regulating effect on the NF-κB and Nrf2 signaling pathways. © The Author(s) 2016.

  10. Lycopene protects human SH-SY5Y neuroblastoma cells against hydrogen peroxide-induced death via inhibition of oxidative stress and mitochondria-associated apoptotic pathways

    Science.gov (United States)

    FENG, CHUNSHENG; LUO, TIANFEI; ZHANG, SHUYAN; LIU, KAI; ZHANG, YANHONG; LUO, YINAN; GE, PENGFEI

    2016-01-01

    Oxidative stress, which is characterized by excessive production of reactive oxygen species (ROS), is a common pathway that results in neuronal injury or death due to various types of pathological stress. Although lycopene has been identified as a potent antioxidant, its effect on hydrogen peroxide (H2O2)-induced neuronal damage remains unclear. In the present study, pretreatment with lycopene was observed to protect SH-SY5Y neuroblastoma cells against H2O2-induced death via inhibition of apoptosis resulting from activation of caspase-3 and translocation of apoptosis inducing factor (AIF) to the nucleus. Furthermore, the over-produced ROS, as well as the reduced activities of anti-oxidative enzymes, superoxide dismutase and catalase, were demonstrated to be alleviated by lycopene. Additionally, lycopene counteracted H2O2-induced mitochondrial dysfunction, which was evidenced by suppression of mitochondrial permeability transition pore opening, attenuation of the decline of the mitochondrial membrane potential, and inhibition of the increase of Bax and decrease of Bcl-2 levels within the mitochondria. The release of cytochrome c and AIF from the mitochondria was also reduced. These results indicate that lycopene is a potent neuroprotectant against apoptosis, oxidative stress and mitochondrial dysfunction, and could be administered to prevent neuronal injury or death. PMID:27035331

  11. Pomegranate protects liver against cecal ligation and puncture-induced oxidative stress and inflammation in rats through TLR4/NF-κB pathway inhibition.

    Science.gov (United States)

    Makled, Mirhan N; El-Awady, Mohammed S; Abdelaziz, Rania R; Atwan, Nadia; Guns, Emma T; Gameil, Nariman M; Shehab El-Din, Ahmed B; Ammar, Elsayed M

    2016-04-01

    Acute liver injury secondary to sepsis is a major challenge in intensive care unit. This study was designed to investigate potential protective effects of pomegranate against sepsis-induced acute liver injury in rats and possible underlying mechanisms. Pomegranate was orally given (800mg/kg/day) for two weeks before sepsis induction by cecal ligation and puncture (CLP). Pomegranate improved survival and attenuated liver inflammatory response, likely related to downregulation of mRNA expression of toll like recptor-4, reduced nuclear translocation and DNA binding activity of proinflammatory transcription factor NF-κB subunit p65, decreased mRNA and protein expression of tumor necrosis factor-alpha and reduction in myeloperoxidase activity and mRNA expression. Pomegranate also decreased CLP-induced oxidative stress as reflected by decreased malondialdehyde content, and increased reduced glutathione level and superoxide dismutase activity. These results confirm the antiinflammatory and antioxidant effects of pomegranate in CLP-induced acute liver injury mediated through inhibiting TLR4/NF-κB pathway, lipid peroxidation and neutrophil infiltration. Copyright © 2016 Elsevier B.V. All rights reserved.

  12. Endothelial microparticles released by activated protein C protect beta cells through EPCR/PAR1 and annexin A1/FPR2 pathways in islets.

    Science.gov (United States)

    Kreutter, Guillaume; Kassem, Mohamad; El Habhab, Ali; Baltzinger, Philippe; Abbas, Malak; Boisrame-Helms, Julie; Amoura, Lamia; Peluso, Jean; Yver, Blandine; Fatiha, Zobairi; Ubeaud-Sequier, Geneviève; Kessler, Laurence; Toti, Florence

    2017-11-01

    Islet transplantation is associated with early ischaemia/reperfusion, localized coagulation and redox-sensitive endothelial dysfunction. In animal models, islet cytoprotection by activated protein C (aPC) restores islet vascularization and protects graft function, suggesting that aPC triggers various lineages. aPC also prompts the release of endothelial MP that bear EPCR, its specific receptor. Microparticles (MP) are plasma membrane procoagulant vesicles, surrogate markers of stress and cellular effectors. We measured the cytoprotective effects of aPC on endothelial and insulin-secreting Rin-m5f β-cells and its role in autocrine and paracrine MP-mediated cell crosstalk under conditions of oxidative stress. MP from aPC-treated primary endothelial (EC) or β-cells were applied to H 2 O 2 -treated Rin-m5f. aPC activity was measured by enzymatic assay and ROS species by dihydroethidium. The capture of PKH26-stained MP and the expression of EPCR were probed by fluorescence microscopy and apoptosis by flow cytometry. aPC treatment enhanced both annexin A1 (ANXA1) and PAR-1 expression in EC and to a lesser extent in β-cells. MP from aPC-treated EC (eM aPC ) exhibited high EPCR and annexin A1 content, protected β-cells, restored insulin secretion and were captured by 80% of β cells in a phosphatidylserine and ANXA1-dependent mechanism. eMP activated EPCR/PAR-1 and ANXA1/FPR2-dependent pathways and up-regulated the expression of EPCR, and of FPR2/ALX, the ANXA1 receptor. Cytoprotection was confirmed in H 2 O 2 -treated rat islets with increased viability (62% versus 48% H 2 O 2 ), reduced apoptosis and preserved insulin secretion in response to glucose elevation (16 versus 5 ng/ml insulin per 10 islets). MP may prove a promising therapeutic tool in the protection of transplanted islets. © 2017 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

  13. The Protective Effects of Κ-Opioid Receptor Stimulation in Hypoxic Pulmonary Hypertension Involve Inhibition of Autophagy Through the AMPK-MTOR Pathway

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    Yaguang Zhou

    2017-12-01

    stimulation protects against HPH by inhibiting PASMCs autophagy via the AMPK-mTOR pathway.

  14. Targeting Cardiomyocyte Ca2+ Homeostasis in Heart Failure

    Science.gov (United States)

    Røe, Åsmund T.; Frisk, Michael; Louch, William E.

    2015-01-01

    Improved treatments for heart failure patients will require the development of novel therapeutic strategies that target basal disease mechanisms. Disrupted cardiomyocyte Ca2+ homeostasis is recognized as a major contributor to the heart failure phenotype, as it plays a key role in systolic and diastolic dysfunction, arrhythmogenesis, and hypertrophy and apoptosis signaling. In this review, we outline existing knowledge of the involvement of Ca2+ homeostasis in these deficits, and identify four promising targets for therapeutic intervention: the sarcoplasmic reticulum Ca2+ ATPase, the Na+-Ca2+ exchanger, the ryanodine receptor, and t-tubule structure. We discuss experimental data indicating the applicability of these targets that has led to recent and ongoing clinical trials, and suggest future therapeutic approaches. PMID:25483944

  15. Defects of mtDNA Replication Impaired Mitochondrial Biogenesis During Trypanosoma cruzi Infection in Human Cardiomyocytes and Chagasic Patients: The Role of Nrf1/2 and Antioxidant Response

    Science.gov (United States)

    Wan, Xianxiu; Gupta, Shivali; Zago, Maria P.; Davidson, Mercy M.; Dousset, Pierre; Amoroso, Alejandro; Garg, Nisha Jain

    2012-01-01

    Background Mitochondrial dysfunction is a key determinant in chagasic cardiomyopathy development in mice; however, its relevance in human Chagas disease is not known. We determined if defects in mitochondrial biogenesis and dysregulation of peroxisome proliferator-activated receptor gamma (PPARγ) coactivator-1 (PGC-1)–regulated transcriptional pathways constitute a mechanism or mechanisms underlying mitochondrial oxidative-phosphorylation (OXPHOS) deficiency in human Chagas disease. Methods and Results We utilized human cardiomyocytes and left-ventricular tissue from chagasic and other cardiomyopathy patients and healthy donors (n>6/group). We noted no change in citrate synthase activity, yet mRNA and/or protein levels of subunits of the respiratory complexes were significantly decreased in Trypanosoma cruzi–infected cardiomyocytes (0 to 24 hours) and chagasic hearts. We observed increased mRNA and decreased nuclear localization of PGC-1-coactivated transcription factors, yet the expression of genes for PPARγ-regulated fatty acid oxidation and nuclear respiratory factor (NRF1/2)–regulated mtDNA replication and transcription machinery was enhanced in infected cardiomyocytes and chagasic hearts. The D-loop formation was normal or higher, but mtDNA replication and mtDNA content were decreased by 83% and 40% to 65%, respectively. Subsequently, we noted that reactive oxygen species (ROS), oxidative stress, and mtDNA oxidation were significantly increased, yet NRF1/2-regulated antioxidant gene expression remained compromised in infected cardiomyocytes and chagasic hearts. Conclusions The replication of mtDNA was severely compromised, resulting in a significant loss of mtDNA and expression of OXPHOS genes in T cruzi–infected cardiomyocytes and chagasic hearts. Our data suggest increased ROS generation and selective functional incapacity of NRF2-mediated antioxidant gene expression played a role in the defects in mtDNA replication and unfitness of mtDNA for

  16. The location of energetic compartments affects energetic communication in cardiomyocytes

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    Rikke eBirkedal

    2014-09-01

    Full Text Available The heart relies on accurate regulation of mitochondrial energy supply to match energy demand. The main regulators are Ca2+ and feedback of ADP and Pi. Regulation via feedback has intrigued for decades. First, the heart exhibits a remarkable metabolic stability. Second, diffusion of ADP and other molecules is restricted specifically in heart and red muscle, where a fast feedback is needed the most. To explain the regulation by feedback, compartmentalization must be taken into account. Experiments and theoretical approaches suggest that cardiomyocyte energetic compartmentalization is elaborate with barriers obstructing diffusion in the cytosol and at the level of the mitochondrial outer membrane (MOM. A recent study suggests the barriers are organized in a lattice with dimensions in agreement with those of intracellular structures. Here, we discuss the possible location of these barriers. The more plausible scenario includes a barrier at the level of MOM. Much research has focused on how the permeability of MOM itself is regulated, and the importance of the creatine kinase system to facilitate energetic communication. We hypothesize that at least part of the diffusion restriction at the MOM level is not by MOM itself, but due to the close physical association between the sarcoplasmic reticulum (SR and mitochondria. This will explain why animals with a disabled creatine kinase system exhibit rather mild phenotype modifications. Mitochondria are hubs of energetics, but also ROS production and signaling. The close association between SR and mitochondria may form a diffusion barrier to ADP added outside a permeabilised cardiomyocyte. But in vivo, it is the structural basis for the mitochondrial-SR coupling that is crucial for the regulation of mitochondrial Ca2+-transients to regulate energetics, and for avoiding Ca2+-overload and irreversible opening of the mitochondrial permeability transition pore.

  17. Importance of Thickness in Human Cardiomyocyte Network for Effective Electrophysiological Stimulation Using On-Chip Extracellular Microelectrodes

    Science.gov (United States)

    Hamada, Tomoyo; Nomura, Fumimasa; Kaneko, Tomoyuki; Yasuda, Kenji

    2012-06-01

    We have developed a three-dimensionally controlled in vitro human cardiomyocyte network assay for the measurements of drug-induced conductivity changes and the appearance of fatal arrhythmia such as ventricular tachycardia/fibrillation for more precise in vitro predictive cardiotoxicity. To construct an artificial conductance propagation model of a human cardiomyocyte network, first, we examined the cell concentration dependence of the cell network heights and found the existence of a height limit of cell networks, which was double-layer height, whereas the cardiomyocytes were effectively and homogeneously cultivated within the microchamber maintaining their spatial distribution constant and their electrophysiological conductance and propagation were successfully recorded using a microelectrode array set on the bottom of the microchamber. The pacing ability of a cardiomyocyte's electrophysiological response has been evaluated using microelectrode extracellular stimulation, and the stimulation for pacing also successfully regulated the beating frequencies of two-layered cardiomyocyte networks, whereas monolayered cardiomyocyte networks were hardly stimulated by the external electrodes using the two-layered cardiomyocyte stimulation condition. The stability of the lined-up shape of human cardiomyocytes within the rectangularly arranged agarose microchambers was limited for a two-layered cardiomyocyte network because their stronger force generation shrunk those cells after peeling off the substrate. The results indicate the importance of fabrication technology of thickness control of cellular networks for effective extracellular stimulation and the potential concerning thick cardiomyocyte networks for long-term cultivation.

  18. Overexpressed connective tissue growth factor in cardiomyocytes attenuates left ventricular remodeling induced by angiotensin II perfusion.

    Science.gov (United States)

    Zhang, Ying; Yan, Hua; Guang, Gong-Chang; Deng, Zheng-Rong

    2017-01-01

    To evaluate the improving effects of specifically overexpressed connective tissue growth factor (CTGF) in cardiomyocytes on mice with hypertension induced by angiotensin II (AngII) perfusion, 24 transgenic mice with cardiac-restricted overexpression of CTGF (Tg-CTGF) were divided into two equal groups that were perfused with acetic acid and AngII, respectively, for 7 days. Another 24 cage-control wild-type C57BL/6 mice (NLC) were divided and treated identically. Blood pressure was detected by caudal artery cannulation. Cardiac structural and functional changes were observed by echocardiography. Cardiac fibrosis was detected by Masson staining. After AngII perfusion, blood pressures of NLC and Tg-CTGF mice, especially those of the formers, significantly increased. Compared with NLC + AngII group, Tg-CTGF + AngII group had significantly lower left ventricular posterior wall thickness at end-diastole and left ventricular posterior wall thickness at end-systole as well as significantly higher left ventricular end-systolic diameter and left ventricular end-diastolic diameter (P tissues (P < 0.05). Tg-CTGF can protect AngII-induced cardiac remodeling of mice with hypertension by mitigating inflammatory response. CTGF may be a therapy target for hypertension-induced myocardial fibrosis, but the detailed mechanism still needs in-depth studies.

  19. Electrophysiological properties and calcium handling of embryonic stem cell-derived cardiomyocytes

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    Jae Boum Youm

    2016-03-01

    Full Text Available Embryonic stem cell-derived cardiomyocytes (ESC-CMs hold great interest in many fields of research including clinical applications such as stem cell and gene therapy for cardiac repair or regeneration. ESC-CMs are also used as a platform tool for pharmacological tests or for investigations of cardiac remodeling. ESC-CMs have many different aspects of morphology, electrophysiology, calcium handling, and bioenergetics compared with adult cardiomyocytes. They are immature in morphology, similar to sinus nodal-like in the electrophysiology, higher contribution of trans-sarcolemmal Ca2+ influx to Ca2+ handling, and higher dependence on anaerobic glycolysis. Here, I review a detailed electrophysiology and Ca2+ handling features of ESC-CMs during differentiation into adult cardiomyocytes to gain insights into how all the developmental changes are related to each other to display cardinal features of developing cardiomyocytes.

  20. A novel type of self-beating cardiomyocytes in adult mouse ventricles

    International Nuclear Information System (INIS)

    Omatsu-Kanbe, Mariko; Matsuura, Hiroshi

    2009-01-01

    This study was designed to investigate the presence of resident heart cells that are distinct from terminally-differentiated cardiomyocytes. Adult mouse heart was coronary perfused with collagenase, and ventricles were excised and further digested. After spinning cardiomyocyte-containing fractions down, the supernatant fraction was collected and cultured without adding any chemicals. Two to five days after plating, some of rounded cells adhered to the culture dish, gradually changed their shape and then started self-beating. These self-beating cells did not appreciably proliferate but underwent a further morphological maturation process to form highly branched shapes with many projections. These cells were mostly multinucleated, well sarcomeric-organized and expressed cardiac marker proteins, defined as atypically-shaped cardiomyocytes (ACMs). Patch-clamp experiments revealed that ACMs exhibited spontaneous action potentials arising from the preceding slow diastolic depolarization. We thus found a novel type of resident heart cells in adult cardiac ventricles that spontaneously develop into self-beating cardiomyocytes.

  1. CTCF counter-regulates cardiomyocyte development and maturation programs in the embryonic heart

    NARCIS (Netherlands)

    M. Gomez-Velazquez (Melisa); C. Badia-Careaga (Claudio); Lechuga-Vieco, A.V. (Ana Victoria); Nieto-Arellano, R. (Rocio); Tena, J.J. (Juan J.); Rollan, I. (Isabel); Alvarez, A. (Alba); Torroja, C. (Carlos); Caceres, E.F. (Eva F.); Roy, A. (Anna); N.J. Galjart (Niels); Delgado-Olguin, P. (Paul); F. Sánchez-Cabo (Fátima); Enriquez, J.A. (Jose Antonio); Gomez-Skarmeta, J.L. (Jose Luis); M. Manzanares (Miguel)

    2017-01-01

    textabstractCardiac progenitors are specified early in development and progressively differentiate and mature into fully functional cardiomyocytes. This process is controlled by an extensively studied transcriptional program. However, the regulatory events coordinating the progression of such

  2. Role of microRNA-195 in cardiomyocyte apoptosis induced by ...

    Indian Academy of Sciences (India)

    drinking water and sterilized standard diet. The mice were ... was performed with the in situ cell death detection kit ... facturer's protocol to detect apoptotic cardiomyocytes. The ..... ulate the leakage of Cyt-c and initiate apoptosis through the.

  3. ShenFu Preparation Protects AML12 Cells Against Palmitic Acid-Induced Injury Through Inhibition of Both JNK/Nox4 and JNK/NFκB Pathways

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    Jia-Fu Ji

    2018-02-01

    Full Text Available Background/Aims: Nonalcoholic steatohepatitis includes steatosis along with liver inflammation, hepatocyte injury and fibrosis. In this study, we investigated the protective role and the potential mechanisms of a traditional Chinese medicine ShenFu (SF preparation in an in vitro hepatic steatosis model. Methods: In palmitic acid (PA-induced murine hepatic AML12 cell injury, effects of SF preparation on cellular apoptosis and intracellular triglyceride (iTG level were assessed using TUNEL and TG Colorimetric Assay. Reactive oxygen species (ROS and mitochondrial membrane potential (MMP levels were measured using DCF and JC-1 assay. Cytokine levels were evaluated using ELISA assay. Immunoblot was used to compare the activation level of c-Jun N terminal kinase (JNK, NADPH oxidase (Nox4, and NFκB pathways. Results: Addition of SF preparation prevented PA-mediated increase of apoptosis and iTG as well as IL-8 and IL-6. In PA-treated cell, SF preparation reduced the level of Nox4 and ROS, while increasing the level of MMP and the expression of manganese superoxide dismutase (MnSOD and catalase, indicating emendation of mitochondrial dysfunction. Nox4 inhibitor GKT137381 prevented PA-induced increase of ROS and apoptosis, while decreasing iTG slightly and not influencing the level of IL-8 and IL-6. SF preparation prevented PA-induced upregulation of phospho-JNK. JNK inhibitor SP600125 prevented PA-mediated increase of Nox4, IL-8, IL-6 and iTG. Nuclear translocation of NFκB/p65 was detected in PA-treated cells, which was prevented by SF preparation. An IκB degradation inhibitor, BAY11-7082, prevented PA-induced increase of IL-8 and IL-6 as well as iTG, whereas it only decreased ROS levels slightly and showed no influence on cellular apoptosis. Conclusion: SF preparation shows a beneficial role in prevention of hepatocyte injury by attenuating oxidative stress and cytokines production at least partially through inhibition of JNK/Nox4 and JNK

  4. Role of Nrf2/ARE pathway in protective effect of electroacupuncture against endotoxic shock-induced acute lung injury in rabbits.

    Directory of Open Access Journals (Sweden)

    Jian-bo Yu

    Full Text Available NF-E2 related factor 2 (Nrf2 is a major transcription factor and acts as a key regulator of antioxidant genes to exogenous stimulations. The aim of current study was to determine whether Nrf2/ARE pathway is involved in the protective effect of electroacupuncture on the injured lung in a rabbit model of endotoxic shock. A dose of lipopolysaccharide (LPS 5 mg/kg was administered intravenously to replicate the model of acute lung injury induced by endotoxic shock. Electroacupuncture pretreatment was handled bilaterally at Zusanli and Feishu acupoints for five consecutive days while sham electroacupuncture punctured at non-acupoints. Fourty anesthetized New England male rabbits were randomized into normal control group (group C, LPS group (group L, electroacupuncture + LPS group (group EL and sham electroacupuncture + LPS (group SEL. At 6 h after LPS administration, the animals were sacrificed and the blood samples were collected for biochemical measurements. The lungs were removed for calculation of wet-to-dry weight ratios (W/D, histopathologic examination, determination of heme oxygenase (HO-1 protein and mRNA, Nrf2 total and nucleoprotein, as well as Nrf2 mRNA expression, and evaluation of the intracellular distribution of Nrf2 nucleoprotein. LPS caused extensive morphologic lung damage, which was lessened by electroacupuncture treatment. Besides, lung W/D ratios were significantly decreased, the level of malondialdehyde was inhibited, plasma levels of TNF-α and interleukin-6 were decreased, while the activities of superoxide dismutase, glutathione peroxidase and catalase were enhanced in the electroacupucnture treated animals. In addition, electroacupuncture stimulation distinctly increased the expressions of HO-1 and Nrf2 protein including Nrf2 total protein and nucleoprotein as well as mRNA in lung tissue, while these effects were blunted in the sham electroacupuncture group. We concluded that electroacupuncture treatment at ST36 and BL13

  5. Role of Nrf2/ARE pathway in protective effect of electroacupuncture against endotoxic shock-induced acute lung injury in rabbits.

    Science.gov (United States)

    Yu, Jian-bo; Shi, Jia; Gong, Li-rong; Dong, Shu-an; Xu, Yan; Zhang, Yuan; Cao, Xin-shun; Wu, Li-li

    2014-01-01

    NF-E2 related factor 2 (Nrf2) is a major transcription factor and acts as a key regulator of antioxidant genes to exogenous stimulations. The aim of current study was to determine whether Nrf2/ARE pathway is involved in the protective effect of electroacupuncture on the injured lung in a rabbit model of endotoxic shock. A dose of lipopolysaccharide (LPS) 5 mg/kg was administered intravenously to replicate the model of acute lung injury induced by endotoxic shock. Electroacupuncture pretreatment was handled bilaterally at Zusanli and Feishu acupoints for five consecutive days while sham electroacupuncture punctured at non-acupoints. Fourty anesthetized New England male rabbits were randomized into normal control group (group C), LPS group (group L), electroacupuncture + LPS group (group EL) and sham electroacupuncture + LPS (group SEL). At 6 h after LPS administration, the animals were sacrificed and the blood samples were collected for biochemical measurements. The lungs were removed for calculation of wet-to-dry weight ratios (W/D), histopathologic examination, determination of heme oxygenase (HO)-1 protein and mRNA, Nrf2 total and nucleoprotein, as well as Nrf2 mRNA expression, and evaluation of the intracellular distribution of Nrf2 nucleoprotein. LPS caused extensive morphologic lung damage, which was lessened by electroacupuncture treatment. Besides, lung W/D ratios were significantly decreased, the level of malondialdehyde was inhibited, plasma levels of TNF-α and interleukin-6 were decreased, while the activities of superoxide dismutase, glutathione peroxidase and catalase were enhanced in the electroacupucnture treated animals. In addition, electroacupuncture stimulation distinctly increased the expressions of HO-1 and Nrf2 protein including Nrf2 total protein and nucleoprotein as well as mRNA in lung tissue, while these effects were blunted in the sham electroacupuncture group. We concluded that electroacupuncture treatment at ST36 and BL13 effectively

  6. High Uric Acid Induces Insulin Resistance in Cardiomyocytes In Vitro and In Vivo.

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    Li Zhi

    Full Text Available Clinical studies have shown hyperuricemia strongly associated with insulin resistance as well as cardiovascular disease. Direct evidence of how high uric acid (HUA affects insulin resistance in cardiomyocytes, but the pathological mechanism of HUA associated with cardiovascular disease remains to be clarified. We aimed to examine the effect of HUA on insulin sensitivity in cardiomyocytes and on insulin resistance in hyperuricemic mouse model. We exposed primary cardiomyocytes and a rat cardiomyocyte cell line, H9c2 cardiomyocytes, to HUA, then quantified glucose uptake with a fluorescent glucose analog, 2-NBDG, after insulin challenge and detected reactive oxygen species (ROS production. Western blot analysis was used to examine the levels of insulin receptor (IR, phosphorylated insulin receptor substrate 1 (IRS1, Ser307 and phospho-Akt (Ser473. We monitored the impact of HUA on insulin resistance, insulin signaling and IR, phospho-IRS1 (Ser307 and phospho-Akt levels in myocardial tissue of an acute hyperuricemia mouse model established by potassium oxonate treatment. HUA inhibited insulin-induced glucose uptake in H9c2 and primary cardiomyocytes. It increased ROS production; pretreatment with N-acetyl-L-cysteine (NAC, a ROS scavenger, reversed HUA-inhibited glucose uptake induced by insulin. HUA exposure directly increased the phospho-IRS1 (Ser307 response to insulin and inhibited that of phospho-Akt in H9C2 cardiomyocytes, which was blocked by NAC. Furthermore, the acute hyperuricemic mice model showed impaired glucose tolerance and insulin tolerance accompanied by increased phospho-IRS1 (Ser307 and inhibited phospho-Akt response to insulin in myocardial tissues. HUA inhibited insulin signaling and induced insulin resistance in cardiomyocytes in vitro and in vivo, which is a novel potential mechanism of hyperuricemic-related cardiovascular disease.

  7. Hypertrophic stimulation increases beta-actin dynamics in adult feline cardiomyocytes.

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    Sundaravadivel Balasubramanian

    2010-07-01

    Full Text Available The myocardium responds to hemodynamic stress through cellular growth and organ hypertrophy. The impact of cytoskeletal elements on this process, however, is not fully understood. While alpha-actin in cardiomyocytes governs muscle contraction in combination with the myosin motor, the exact role of beta-actin has not been established. We hypothesized that in adult cardiomyocytes, as in non-myocytes, beta-actin can facilitate cytoskeletal rearrangement within cytoskeletal structures such as Z-discs. Using a feline right ventricular pressure overload (RVPO model, we measured the level and distribution of beta-actin in normal and pressure overloaded myocardium. Resulting data demonstrated enriched levels of beta-actin and enhanced translocation to the Triton-insoluble cytoskeletal and membrane skeletal complexes. In addition, RVPO in vivo and in vitro hypertrophic stimulation with endothelin (ET or insulin in isolated adult cardiomyocytes enhanced the content of polymerized fraction (F-actin of beta-actin. To determine the localization and dynamics of beta-actin, we adenovirally expressed GFP-tagged beta-actin in isolated adult cardiomyocytes. The ectopically expressed beta-actin-GFP localized to the Z-discs, costameres, and cell termini. Fluorescence recovery after photobleaching (FRAP measurements of beta-actin dynamics revealed that beta-actin at the Z-discs is constantly being exchanged with beta-actin from cytoplasmic pools and that this exchange is faster upon hypertrophic stimulation with ET or insulin. In addition, in electrically stimulated isolated adult cardiomyocytes, while beta-actin overexpression improved cardiomyocyte contractility, immunoneutralization of beta-actin resulted in a reduced contractility suggesting that beta-actin could be important for the contractile function of adult cardiomyocytes. These studies demonstrate the presence and dynamics of beta-actin in the adult cardiomyocyte and reinforce its usefulness in measuring

  8. Group B streptococcal beta-hemolysin/cytolysin directly impairs cardiomyocyte viability and function.

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    Mary E Hensler

    Full Text Available BACKGROUND: Group B Streptococcus (GBS is a leading cause of neonatal sepsis where myocardial dysfunction is an important contributor to poor outcome. Here we study the effects of the GBS pore-forming beta-hemolysin/cytolysin (Bh/c exotoxin on cardiomyocyte viability, contractility, and calcium transients. METHODOLOGY/PRINCIPAL FINDINGS: HL-1 cardiomyocytes exposed to intact wild-type (WT or isogenic Deltabeta h/c mutant GBS, or to cell-free extracts from either strain, were assessed for viability by trypan blue exclusion and for apoptosis by TUNEL staining. Functionality of exposed cardiomyocytes was analyzed by visual quantitation of the rate and extent of contractility. Mitochondrial membrane polarization was measured in TMRE-loaded cells exposed to GBS beta h/c. Effects of GBS beta h/c on calcium transients were studied in fura-2AM-loaded primary rat ventricular cardiomyocytes. Exposure of HL-1 cardiomyocytes to either WT GBS or beta h/c extracts significantly reduced both rate and extent of contractility and later induced necrotic and apoptotic cell death. No effects on cardiomyocyte viability or function were observed after treatment with Deltabeta h/c mutant bacteria or extracts. The beta h/c toxin was associated with complete and rapid loss of detectable calcium transients in primary neonatal rat ventricular cardiomyocytes and induced a loss of mitochondrial membrane polarization. These effects on viability and function were abrogated by the beta h/c inhibitor, dipalmitoyl phosphatidylcholine (DPPC. CONCLUSIONS/SIGNIFICANCE: Our data show a rapid loss of cardiomyocyte viability and function induced by GBS beta h/c, and these deleterious effects are inhibited by DPPC, a normal constituent of human pulmonary surfactant.. These findings have clinical implications for the cardiac dysfunction observed in neonatal GBS infections.

  9. Hydrogen sulfide protects against chemical hypoxia-induced injury by inhibiting ROS-activated ERK1/2 and p38MAPK signaling pathways in PC12 cells.

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    Aiping Lan

    Full Text Available Hydrogen sulfide (H(2S has been proposed as a novel neuromodulator and neuroprotective agent. Cobalt chloride (CoCl(2 is a well-known hypoxia mimetic agent. We have demonstrated that H(2S protects against CoCl(2-induced injuries in PC12 cells. However, whether the members of mitogen-activated protein kinases (MAPK, in particular, extracellular signal-regulated kinase1/2(ERK1/2 and p38MAPK are involved in the neuroprotection of H(2S against chemical hypoxia-induced injuries of PC12 cells is not understood. We observed that CoCl(2 induced expression of transcriptional factor hypoxia-inducible factor-1 alpha (HIF-1α, decreased cystathionine-β synthase (CBS, a synthase of H(2S expression, and increased generation of reactive oxygen species (ROS, leading to injuries of the cells, evidenced by decrease in cell viability, dissipation of mitochondrial membrane potential (MMP , caspase-3 activation and apoptosis, which were attenuated by pretreatment with NaHS (a donor of H(2S or N-acetyl-L cystein (NAC, a ROS scavenger. CoCl(2 rapidly activated ERK1/2, p38MAPK and C-Jun N-terminal kinase (JNK. Inhibition of ERK1/2 or p38MAPK or JNK with kinase inhibitors (U0126 or SB203580 or SP600125, respectively or genetic silencing of ERK1/2 or p38MAPK by RNAi (Si-ERK1/2 or Si-p38MAPK significantly prevented CoCl(2-induced injuries. Pretreatment with NaHS or NAC inhibited not only CoCl(2-induced ROS production, but also phosphorylation of ERK1/2 and p38MAPK. Thus, we demonstrated that a concurrent activation of ERK1/2, p38MAPK and JNK participates in CoCl(2-induced injuries and that H(2S protects PC12 cells against chemical hypoxia-induced injuries by inhibition of ROS-activated ERK1/2 and p38MAPK pathways. Our results suggest that inhibitors of ERK1/2, p38MAPK and JNK or antioxidants may be useful for preventing and treating hypoxia-induced neuronal injury.

  10. Inhibition of ErbB2 by receptor tyrosine kinase inhibitors causes myofibrillar structural damage without cell death in adult rat cardiomyocytes

    International Nuclear Information System (INIS)

    Pentassuglia, Laura; Graf, Michael; Lane, Heidi; Kuramochi, Yukio; Cote, Gregory; Timolati, Francesco; Sawyer, Douglas B.; Zuppinger, Christian; Suter, Thomas M.

    2009-01-01

    Inhibition of ErbB2 (HER2) with monoclonal antibodies, an effective therapy in some forms of breast cancer, is associated with cardiotoxicity, the pathophysiology of which is poorly understood. Recent data suggest, that dual inhibition of ErbB1 (EGFR) and ErbB2 signaling is more efficient in cancer therapy, however, cardiac safety of this therapeutic approach is unknown. We therefore tested an ErbB1-(CGP059326) and an ErbB1/ErbB2-(PKI166) tyrosine kinase inhibitor in an in-vitro system of adult rat ventricular cardiomyocytes and assessed their effects on 1. cell viability, 2. myofibrillar structure, 3. contractile function, and 4. MAPK- and Akt-signaling alone or in combination with Doxorubicin. Neither CGP nor PKI induced cardiomyocyte necrosis or apoptosis. PKI but not CGP caused myofibrillar structural damage that was additive to that induced by Doxorubicin at clinically relevant doses. These changes were associated with an inhibition of excitation-contraction coupling. PKI but not CGP decreased p-Erk1/2, suggesting a role for this MAP-kinase signaling pathway in the maintenance of myofibrils. These data indicate that the ErbB2 signaling pathway is critical for the maintenance of myofibrillar structure and function. Clinical studies using ErbB2-targeted inhibitors for the treatment of cancer should be designed to include careful monitoring for cardiac dysfunction.

  11. An essential role of Nrf2 in American ginseng-mediated anti-oxidative actions in cardiomyocytes.

    Science.gov (United States)

    Li, Jinqing; Ichikawa, Tomonaga; Jin, Yu; Hofseth, Lorne J; Nagarkatti, Prakash; Nagarkatti, Mitzi; Windust, Anthony; Cui, Taixing

    2010-07-20

    administration of American ginseng markedly increased Nrf2 activity in murine hearts. These results demonstrate that American ginseng suppresses oxidative stress and oxidative stress-induced cell death in cardiomyocytes through activating the Nrf2 pathway, thereby providing cardioprotection against pathological cardiac remodeling. Copyright 2010 Elsevier Ireland Ltd. All rights reserved.

  12. Protective effects of Cassia tora leaves in experimental cataract by modulating intracellular communication, membrane co-transporters, energy metabolism and the ubiquitin-proteasome pathway.

    Science.gov (United States)

    Sreelakshmi, V; Abraham, Annie

    2017-12-01

    Cataract is the clouding of eye lens which causes impairment in vision and accounts for the leading factor of global blindness. Functional food-based prevention of cataract finds application in vision research because of its availability and easy access to all classes of the society. Cassia tora Linn. (Caesalpinaceae) is an edible plant mentioned in the traditional systems of medicine for whole body health, especially to the eyes. The present study evaluates the potential of ethyl acetate fraction of Cassia tora leaves (ECT) on experimental cataract. Cataract was induced by a single subcutaneous injection of sodium selenite (4 μg/g body weight) on 10th day. ECT was supplemented orally from 8th day up to 12th day at a concentration of 5 μg/g body weight and marker parameters were evaluated after 30 days. The production of MPO and the activation of calpain were reduced 52.17% and 36.67% by ECT in lens tissue, respectively. It modulated the energy status by significantly increasing the activity of CCO 1 (55.56%) and ATP production (41.88%). ECT maintained the ionic balance in the lens by reducing the level of sodium (50%) and increasing the level of potassium (42.5%). It also reduced cell junction modifications and preserved a functional ubiquitin-proteasome pathway. The results reinforce the growing attention on wild plant food resources for preventive protection against cataract. The data suggest the value of Cassia tora leaves as a functional food for ameliorating cataract pathology.

  13. Arctigenin protects against steatosis in WRL68 hepatocytes through activation of phosphoinositide 3-kinase/protein kinase B and AMP-activated protein kinase pathways.

    Science.gov (United States)

    Chen, Kung-Yen; Lin, Jui-An; Yao, Han-Yun; Hsu, An-Chih; Tai, Yu-Ting; Chen, Jui-Tai; Hsieh, Mao-Chih; Shen, Tang-Long; Hsu, Ren-Yi; Wu, Hong-Tan; Wang, Guey Horng; Ho, Bing-Ying; Chen, Yu-Pei

    2018-04-01

    Arctigenin (ATG), a lignin extracted from Arctium lappa (L.), exerts antioxidant and anti-inflammatory effects. We hypothesized that ATG exerts a protective effect on hepatocytes by preventing nonalcoholic fatty liver disease (NAFLD) progression associated with lipid oxidation-associated lipotoxicity and inflammation. We established an in vitro NAFLD cell model by using normal WRL68 hepatocytes to investigate oleic acid (OA) accumulation and the potential bioactive role of ATG. The results revealed that ATG inhibited OA-induced lipid accumulation, lipid peroxidation, and inflammation in WRL68 hepatocytes, as determined using Oil Red O staining, thiobarbituric acid reactive substance assay, and inflammation antibody array assays. Quantitative RT-PCR analysis demonstrated that ATG significantly mitigated the expression of acetylcoenzyme A carboxylase 1 and sterol regulatory element-binding protein-1 and significantly increased the expression of carnitine palmitoyltransferase 1 and peroxisome proliferator-activated receptor alpha. The 40 targets of the Human Inflammation Antibody Array indicated that ATG significantly inhibited the elevation of the U937 lymphocyte chemoattractant, ICAM-1, IL-1β, IL-6, IL-6sR, IL-7, and IL-8. ATG could activate the phosphoinositide 3-kinase/protein kinase B (PI3K/AKT) and AMP-activated protein kinase (AMPK) pathways and could increase the phosphorylation levels of Akt and AMPK to mediate cell survival, lipid metabolism, oxidation stress, and inflammation. Thus, we demonstrated that ATG could inhibit NAFLD progression associated with lipid oxidation-associated lipotoxicity and inflammation, and we provided insights into the underlying mechanisms and revealed potential targets to enable a thorough understanding of NAFLD progression. Copyright © 2018 Elsevier Inc. All rights reserved.

  14. BF*F allotype of the alternative pathway of complement: A marker of protection against the development of antiphospholipid antibodies in patients with systemic lupus erythematosus.

    Science.gov (United States)

    Picceli, V F; Skare, T L; Nisihara, R M; Nass, F R; Messias-Reason, I T; Utiyama, S R R

    2016-04-01

    B factor (BF) from the alternative complement pathway seems to participate in the pathophysiology of systemic lupus erythematosus (SLE) and antiphospholipid syndrome (APS). To study the allotypic variability of BF in SLE and their associations with clinical and autoantibodies profile. BF allotypes were determined by high-voltage agarose gel electrophoresis, under constant cooling, followed by immunofixation with anti-human BF antibody, in 188 SLE patients and 103 controls. Clinical and serological data were obtained from medical examination and records. No significant differences of BF variants between patients and controls were found, neither in relation to epidemiologic or clinical manifestations. Associations of phenotype BF SS07 and allotype BF*S07 were found with anticardiolipin IgM (aCl-IgM) antibodies (p = 0.014 and p = 0.009 respectively), but not with aCl-IgG, lupus anticoagulant (LA), anti β2GPI or clinical APS. A significant decrease in BF*F allotype (p = 0.043) and BF SF phenotype (p = 0.018) was detected in patients with anti-phospholipid antibodies as a whole (aCl-IgG, aCl-IgM, LA and anti β2GPI). There is a link between phenotype BF SS07 and allotype BF*S07 with aCl-IgM in SLE patients; BF*F allotype could be considered a marker of protection against the development of antiphospholipid antibodies in these patients. © The Author(s) 2015.

  15. Puerarin protects differentiated PC12 cells from H₂O₂-induced apoptosis through the PI3K/Akt signalling pathway.

    Science.gov (United States)

    Zhang, Qin; Huang, Wei-Dong; Lv, Xue-Ying; Yang, Yun-Mei

    2012-05-01

    Oxidative stress has been implicated as a major mechanism underlying the pathogenesis of neurodegenerative disorders. ROS (reactive oxygen species) can cause cell death via apoptosis. NGF (nerve growth factor) differentiated rat PC12 cells have been extensively used to study the differentiation and apoptosis of neurons. This study has investigated the protective effects of puerarin in H2O2-induced apoptosis of differentiated PC12 cells, and the possible molecular mechanisms involved. Differentiated PC12 cells were incubated with 700 μM H2O2 in the absence or presence of different doses of puerarin (4, 8 and 16 μM). Apoptosis was assessed by MTS [3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium] assay, TUNEL (terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling) analysis and Annexin V-PI (propidium iodide) double staining flow cytometry. Protein levels of phospho-Akt and phospho-BAD (Bcl-2/Bcl-XL-antagonist, causing cell death) were assayed by Western blotting. After stimulation with H2O2 for 18 h, the viability of differentiated PC12 cells decreased significantly and a large number of cells underwent apoptosis. Differentiated PC12 cells were rescued from H2O2-induced apoptosis at different concentrations of puerarin in a dose-dependent manner. This was through increased production of phospho-Akt and phospho-BAD, an effect that could be reversed by wortmannin, an inhibitor of PI3K (phosphoinositide 3-kinase). The results suggest that puerarin may have neuroprotective effect through activation of the PI3K/Akt signalling pathway.

  16. Zurampic Protects Pancreatic β-Cells from High Uric Acid Induced-Damage by Inhibiting URAT1 and Inactivating the ROS/AMPK/ERK Pathways

    Directory of Open Access Journals (Sweden)

    Ying Xin

    2018-05-01

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