Li Yun Zhao, Wu Hui, Liu Di, Yang Jun, Yang Jian, Ding Jia Wang, Zhou Gang, Zhang Jing, Zhang Dong
Institute of Cardiovascular Disease, China Three Gorges University, Yichang, 443003, China.
Department of Cardiology, Yichang Central People's Hospital, Yichang, 443003, China.
Cardiovasc Drugs Ther. 2023 Apr;37(2):225-238. doi: 10.1007/s10557-021-07280-1. Epub 2021 Nov 12.
Endoplasmic reticulum stress (ERS) plays a crucial role in myocardial ischemia-reperfusion injury (MIRI). Cellular FLICE-inhibitory protein (cFLIP) is an essential regulator of apoptosis and plays a major role in regulating ERS. The present study aimed to investigate the effects of long isoform cFLIP (cFLIP) on endogenous apoptosis and the mechanism of ERS in MIRI.
The cFLIP recombinant adenovirus vector was used to infect H9c2 cells and Sprague-Dawley (SD) rats. After infection for 72 h, ischemia was induced for 30 min, and reperfusion was then performed for 2 h to establish the MIRI model in SD rats. H9c2 cells were hypoxic for 4 h and then reoxygenated for 12 h to simulate ischemia/reperfusion (I/R) injury. Model parameters were evaluated by assessing cardiomyocyte viability, cell death (apoptosis), and ERS-related protein expression. In addition, tunicamycin (TM), an ERS agonist, was also added to the medium for pretreatment. Coimmunoprecipitation (Co-IP) of cFLIP and p38 MAPK protein was performed.
cFLIP expression was decreased in I/R injury and hypoxia/reoxygenation (H/R) injury, and cFLIP overexpression reduced myocardial infarction in vivo and increased the viability of H9c2 cells in vitro. I/R and H/R upregulated the protein expression of GRP78, IRE-1, and PERK to induce ERS and apoptosis. Interestingly, overexpression of cFLIP significantly inhibited ERS and subsequent apoptosis, which was reversed by an agonist of ERS. Moreover, Co-IP showed that cFLIP attenuated ERS and was associated with inhibiting the activation of p38 protein.
The expression of cFLIP is significantly downregulated in MIRI, and it is accompanied by excessive ERS and apoptosis. Upregulated cFLIP suppresses ERS to reduce myocardial apoptosis, which is associated with inhibiting the activity of p38 MAPK. Therefore, cFLIP may be a potential intervention target for MIRI.
内质网应激（ERS）在心肌缺血再灌注损伤（MIRI）中起关键作用。细胞FLICE抑制蛋白（cFLIP）是细胞凋亡的重要调节因子，在调节ERS中起主要作用。本研究旨在探讨长型cFLIP（cFLIPL）对MIRI内源性细胞凋亡的影响及其ERS机制。
采用cFLIP重组腺病毒载体感染H9c2细胞和Sprague-Dawley（SD）大鼠。感染72小时后，诱导缺血30分钟，然后进行再灌注2小时，以建立SD大鼠MIRI模型。H9c2细胞缺氧4小时，然后复氧12小时以模拟缺血/再灌注（I/R）损伤。通过评估心肌细胞活力、细胞死亡（凋亡）和ERS相关蛋白表达来评估模型参数。此外，ERS激动剂衣霉素（TM）也添加到培养基中进行预处理。进行cFLIP和p38 MAPK蛋白的免疫共沉淀（Co-IP）。
I/R损伤和缺氧/复氧（H/R）损伤中cFLIP表达降低，cFLIP过表达可减少体内心肌梗死并增加体外H9c2细胞活力。I/R和H/R上调GRP78、IRE-1和PERK的蛋白表达以诱导ERS和细胞凋亡。有趣的是，cFLIP过表达显著抑制ERS和随后的细胞凋亡，而ERS激动剂可逆转这种作用。此外，Co-IP显示cFLIP减轻ERS并与抑制p38蛋白的激活有关。
MIRI中cFLIP的表达显著下调，并伴有过度的ERS和细胞凋亡。上调cFLIP可抑制ERS以减少心肌细胞凋亡，这与抑制p38 MAPK的活性有关。因此，cFLIP可能是MIRI的潜在干预靶点。
