Abstract
BackgroundA gasotransmitter hydrogen sulfide (H2S) plays an important physiological and pathological role in cardiovascular system. Ischemic post-conditioning (PC) provides cardioprotection in the young hearts but not in the aged hearts. Exogenous H2S restores PC-induced cardioprotection by inhibition of mitochondrial permeability transition pore opening and oxidative stress and increase of autophagy in the aged hearts. However, whether H2S contributes to the recovery of PC-induced cardioprotection via down-regulation of endoplasmic reticulum stress (ERS) in the aged hearts is unclear.MethodsThe aged H9C2 cells (the cardiomyocytes line) were induced using H2O2 and were exposed to H/R and PC protocols. Cell viability was observed by CCK-8 kit. Apoptosis was detected by Hoechst 33342 staining and flow cytometry. Related protein expressions were detected through Western blot.ResultsIn the present study, we found that 30 μM H2O2 induced H9C2 cells senescence but not apoptosis. Supplementation of NaHS protected against H/R-induced apoptosis, the expression of cleaved caspase-3 and cleaved caspase-9 and the release of cytochrome c. The addition of NaHS also counteracted the reduction of cell viability caused by H/R and decreased the expression of GRP 78, CHOP, cleaved caspase-12, ATF 4, ATF 6 and XBP-1 and the phosphorylation of PERK, eIF 2α and IRE 1α. Additionally, NaHS increased Bcl-2 expression. PC alone did not provide cardioprotection in H/R-treated aged cardiomyocytes, which was significantly restored by the supplementation of NaHS. The beneficial role of NaHS was similar to the supply of 4-PBA (an inhibitor of ERS), GSK2656157 (an inhibitor of PERK), STF083010 (an inhibitor of IRE 1α), respectively, during PC.ConclusionOur results suggest that the recovery of myocardial protection from PC by exogenous H2S is associated with the inhibition of ERS via down-regulating PERK-eIF 2α-ATF 4, IRE 1α-XBP-1 and ATF 6 pathways in the aged cardiomyocytes.
Highlights
A gasotransmitter hydrogen sulfide (H2S) plays an important physiological and pathological role in cardiovascular system
The primary antibodies for anti-Cyclin D1, p21Cip/WAF−1, cleaved caspase-3 and -9, Bcl-2, cytochrome c (Cyt c), GRP 78, C/EBP homologous protein (CHOP), cleaved caspase-12, activating transcription factor 4 (ATF 4), ATF 6, X-box-binding protein-1 (XBP-1) and GAPDH were from Proteintech (Wuhan, China)
The data showed that we successfully induced the senescence of the H9C2 cells with 30 μM H 2O2 and the number of senescent cells was increased in a dose-dependent manner (Fig. 2a), it led to apoptosis when exposed to the concentrations of H 2O2 more than 30 μM and 30 μM H 2O2 had no apparent effect on caspase-3 activity (Fig. 2b)
Summary
A gasotransmitter hydrogen sulfide (H2S) plays an important physiological and pathological role in cardiovascular system. Exogenous H2S restores PC-induced cardioprotection by inhibition of mitochondrial permeability transition pore opening and oxidative stress and increase of autophagy in the aged hearts. Whether H 2S contributes to the recovery of PC-induced cardioprotection via down-regulation of endoplasmic reticulum stress (ERS) in the aged hearts is unclear. The abnormal metabolism and functions of the CSE/ H2S pathway have been linked to various cardiovascular diseases, including ischemia/reperfusion (I/R) injury, atherosclerosis, hypertension, heart failure, myocardial infarction etc. I/R (or hypoxia/reoxygenation, H/R) injury increases endoplasmic reticulum stress (ERS) and induces apoptosis [6]. ERS related proteins, glucose regulated protein 78 (GRP78), C/EBP homologous protein (CHOP) and cleaved caspase-12, and related signaling pathways, pancreatic endoplasmic reticulum kinase (PERK)-eukaryotic translation initiation factor 2α (eIF 2α)-activating transcription factor 4 (ATF 4), inositolrequiring enzyme1α (IRE 1α)- X-box-binding protein-1 (XBP-1) and ATF 6 are increased during I/R (or H/R) [7,8,9,10]. It was recently reported that PC loses its cardioprotection in aged hearts [3, 4, 10,11,12,13,14]
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