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

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 Ca 2+ ([Ca 2+ ] 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 [Ca 2+ ] i levels increased in a timedependent 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 [Ca 2+ ] 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 [Ca 2+ ] i level, and result in the apoptosis of H9c2 cells.

<p>Dexmedetomidine protects H9c2 cardiomyocytes against oxygen-glucose deprivation/reoxygenation-induced intracellular calcium overload and apoptosis through regulating FKBP12.6/RyR2 signaling</p>

“…[Ca 2+ ]i overload eventually initiates the apoptotic process where caspase-3 protein acts as a critical executioner. 12,24 A study found that DEX protected spinal cord against I/R-induced injury through inhibiting apoptosis and caspase-3 activation, 12 which was in line with the current results in H9c2 cells.…”

“…21,22 Studies also revealed that the regulation of Ca 2+ in SR and cytoplasm was associated with the apoptotic signaling pathway. 12,23,24 In the apoptotic process, caspase-3 is known as an executioner that functions in DNA fragmentation and degradation of cytoskeletal proteins. 25 In this study, the effect of DEX pretreatment on Ca 2+ signaling and apoptosis in H9c2 cardiomyocytes during oxygen-glucose deprivation/reoxygenation (OGD/R) was investigated.…”

TRPC Channels: Dysregulation and Ca2+ Mishandling in Ischemic Heart Disease

“…They also demonstrated significant improvement in the viability of cardiomyocytes after SKF96365 treatment. Moreover, Meng et al [77] observed that in vitro I/R increases TRPC6 protein expression, [Ca 2+ ] i levels, and cell apoptotic rate in a time-dependent manner in H9C2 cell line. In addition, they suggested TRPC6 as a possible target for cardioprotection in H9C2 cells since the administration of danshensu, an active component of Salvia miltiorrhiza, protects against I/R injury by reducing TRPC6 expression via the c-Jun N-terminal kinases (JNK) signaling pathway [77].…”

“…Moreover, Meng et al [77] observed that in vitro I/R increases TRPC6 protein expression, [Ca 2+ ] i levels, and cell apoptotic rate in a time-dependent manner in H9C2 cell line. In addition, they suggested TRPC6 as a possible target for cardioprotection in H9C2 cells since the administration of danshensu, an active component of Salvia miltiorrhiza, protects against I/R injury by reducing TRPC6 expression via the c-Jun N-terminal kinases (JNK) signaling pathway [77]. Hang et al [78] also demonstrated that brain-derived neurotrophic factor (BDNF) protects against MI and inhibits H/R-mediated cardiomyocyte apoptosis through TRPC3 and TRPC6 regulation.…”

Metabolite identification of tanshinol borneol ester in rats by high‐performance liquid chromatography combined with electrospray ionization quadrupole time‐of‐flight mass spectrometry

“…DSS is a predominant bioactive component in aqueous extract of Salvia miltiorrhiza , which is the Jun (emperor) medicinal of FFDS (Zhao et al, ). It can delay the senescence of rat aortic endothelial cells (Wang et al, ), inhibit platelet aggregation and thrombus formation (Yu et al, ), and protect against cardio‐cerebral ischaemia/reperfusion injury (Fan et al, ; Guo et al, ; Meng et al, ; Yin et al, ). In addition to the above‐mentioned effects, DSS has also been reported to inhibit β ‐adrenergic receptor‐mediated cardiac fibrosis and carbon tetrachloride‐induced hepatic fibrosis (Lu et al, ; Qu, Huang, Li, & Qin, ).…”