Abstract
BackgroundHydrogen sulfide (H2S), a third member of gasotransmitter family along with nitric oxide and carbon monoxide, generated from mainly catalyzed by cystathionine-lyase, possesses important functions in the cardiovascular system. Ischemic post-conditioning (PC) strongly protects against the hypoxia/reoxygenation (H/R)-induced injury and apoptosis of cardiomyocytes. However, PC protection is ineffective in the aging cardiomyocytes. Whether H2S restores PC-induced cardioprotection by decrease of reactive oxygen species (ROS) level in the aging cardiomyocytes is unknown.MethodsThe aging cardiomyocytes were induced by treatment of primary cultures of neonatal cardiomyocytes using d-galactose 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. ROS level was analyzed using spectrofluorimeter. Related protein expressions were detected through Western blot.ResultsTreatment of NaHS (a H2S donor) protected against H/R-induced apoptosis, cell damage, the expression of cleaved caspase-3 and cleaved caspase-9, the release of cytochrome c (Cyt c). The supplementation of NaHS also decreased the activity of LDH and CK, MDA contents, ROS levels and the phosphorylation of IκBα, NF-κB, JNK2 and STAT3, and increased cell viability, the expression of Bcl-2, the activity of SOD, CAT and GSH-PX. PC alone did not provide cardioprotection in H/R-treated aging cardiomyocytes, which was significantly restored by the addition of NaHS. The beneficial role of NaHS was similar to the supply of N-acetyl-cysteine (NAC, an inhibitor of ROS), Ammonium pyrrolidinedithiocarbamate (PDTC, an inhibitor of NF-κB) and AG 490 (an inhibitor of JNK2), respectively, during PC.ConclusionOur results suggest that exogenous H2S contributes to recovery of PC-induced cardioprotection by decrease of ROS level via down-regulation of NF-κB and JAK2/STAT3 pathways in the aging cardiomyocytes.
Highlights
Hydrogen sulfide (H2S), a third member of gasotransmitter family along with nitric oxide and carbon monoxide, generated from mainly catalyzed by cystathionine-lyase, possesses important functions in the cardiovascular system
We focused on the mechanism of reactive oxygen species (ROS) generation, including NF-κB and janus kinase-2 (JAK2)-signal transducer and activator of transcription 3 (STAT3) pathways, and the role that exogenous H2S played during this process
Assay kits for malondialdehyde (MDA), superoxide dismutase (SOD), Glutathione peroxidase (GSH-PX), catalase (CAT), lactate dehydrogenase (LDH), creatine kinase (CK) and ROS were purchased from Nanjing Jiancheng Bioengineering Institute (Nanjing, China)
Summary
Hydrogen sulfide (H2S), a third member of gasotransmitter family along with nitric oxide and carbon monoxide, generated from mainly catalyzed by cystathionine-lyase, possesses important functions in the cardiovascular system. Ischemic post-conditioning (PC) strongly protects against the hypoxia/reoxygenation (H/R)-induced injury and apoptosis of cardiomyocytes. Whether H2S restores PC-induced cardioprotection by decrease of reactive oxygen species (ROS) level in the aging cardiomyocytes is unknown. Myocardial ischemia (hypoxia)/reperfusion (reoxygenation) causes cardiomyocytes injury, including cardiomyocytes apoptosis and necrosis by increase of free radical generation, calcium overload and the adhesion of leukocytes [7, 8]. Reactive oxygen species (ROS) are increased during I/R lead to cell death [9]. PC can inhibit I/R-induced cardiomyocytes injury via decrease of ROS. The beneficial effect of PC has been documented in the young heart of every species tested [13, 14], but this response is attenuated or loss in the aged heart [8, 9, 15, 16]
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