title: Danshensu protects against ischemia/reperfusion injury and inhibits the apoptosis of H9c2 cells by reducing the calcium overload through the p-JNK-NF-κB-TRPC6 pathway.
abstract: Ischemia-reperfusion (I/R) plays an important role in myocardial injury. In the present study, we aimed to examine the protective effects of Danshensu (DSS) against I/R injury and to elucidate the underlying mechanisms. For this purpose, H9c2 cells were cultured in hypoxic solution in a hypoxic incubator for 2 h, and then cultured in a high oxygen incubator for various periods of time and pre-treated with or without DSS, ammonium pyrrolidine dithiocarbamate (PDTC) or SP600125 [a c-Jun N-terminal kinase (JNK) inhibitor]. Cell apoptosis and cytosolic free Ca2+ ([Ca2+]i) levels were analyzed by flow cytometry. The protein expression levels of JNK, phosphorylated (p-)JNK, nuclear factor-κB (NF-κB) and transient receptor potential cation channel, subfamily C, member 6 (TRPC6) were measured by western blot analysis. The mRNA expression levels of JNK were measured by RT-qPCR. The results revealed that TRPC6 protein expression, the cell apoptotic rate and the [Ca2+]i levels increased in a time-dependent manner in the H9c2 cells following the induction of I/R injury. The apoptotic rate and TRPC6 protein expression decreased when the cells were treated with DSS prior to the induction of I/R injury. The knockdown of JNK expression by siRNA decreased the p-JNK and TRPC6 protein expression levels in the H9c2 cells subjected to I/R injury. The protein expression levels of p-JNK and NF-κB in the nucleus increased significantly when the H9c2 cells were subjected to I/R injury, whereas NF-κB expression in the cytoplasm decreased in a time‑dependent manner. However, p-JNK, NF-κB and TRPC6 protein expression, the [Ca2+]i level and cell apoptosis decreased when the H9c2 cells were pre-treated with DSS or SP600125. Therefore, our data suggest that DSS prevents myocardial I/R injury by inhibiting p-JNK activation and NF-κB translocation, which potentially upregulate TRPC6 expression, increase the [Ca2+]i level, and result in the apoptosis of H9c2 cells.
authors.affiliation: Department of Pharmacology, Key Laboratory of Anti-inflammatory and Immunopharmacology, Ministry of Education, Anhui Medical University, Hefei, Anhui 230032, P.R. China.
authors.forename: Ying
authors.initials: Y
authors.lastname: Meng
authors.affiliation: Department of Pharmacology, Key Laboratory of Anti-inflammatory and Immunopharmacology, Ministry of Education, Anhui Medical University, Hefei, Anhui 230032, P.R. China.
authors.forename: Wei-Zhu
authors.initials: WZ
authors.lastname: Li
authors.affiliation: Department of Cardiovascular Medicine, Binhu Hospital of Hefei City, Hefei, Anhui 230011, P.R. China.
authors.forename: You-Wei
authors.initials: YW
authors.lastname: Shi
authors.affiliation: Department of Cardiovascular Medicine, Binhu Hospital of Hefei City, Hefei, Anhui 230011, P.R. China.
authors.forename: Bing-Feng
authors.initials: BF
authors.lastname: Zhou
authors.affiliation: Department of Integrative Physiology and Cardiovascular Research Institute, University of North Texas Health Science Center, Fort Worth, TX 76107, USA.
authors.forename: Rong
authors.initials: R
authors.lastname: Ma
authors.affiliation: Department of Pharmacology, Key Laboratory of Anti-inflammatory and Immunopharmacology, Ministry of Education, Anhui Medical University, Hefei, Anhui 230032, P.R. China.
authors.forename: Wei-Ping
authors.initials: WP
authors.lastname: Li
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D004365:Drugs, Chinese Herbal / Q000494:pharmacology*; D005786:Gene Expression Regulation / Q000187:drug effects; D007773:Lactates / Q000494:pharmacology*; D048670:MAP Kinase Kinase 4 / Q000235:genetics / Q000378:metabolism; D020935:MAP Kinase Signaling System / Q000187:drug effects; D015428:Myocardial Reperfusion Injury / Q000235:genetics / Q000378:metabolism / Q000517:prevention & control*; D016328:NF-kappa B / Q000235:genetics / Q000378:metabolism; D020011:Protective Agents / Q000494:pharmacology*; D051381:Rats; D015398:Signal Transduction / Q000187:drug effects*; D050052:TRPC Cation Channels / Q000235:genetics / Q000378:metabolism*
chemical_list: D004365:Drugs, Chinese Herbal; D007773:Lactates; D016328:NF-kappa B; D020011:Protective Agents; D050052:TRPC Cation Channels; C492145:Trpc6 protein, rat; C035055:3,4-dihydroxyphenyllactic acid; D048670:MAP Kinase Kinase 4; D002118:Calcium