Ren Hou-Wei, Gu Bin, Zhang Yue-Zhan, Guo Ting, Wang Qian, Shen Yue-Qin, Wang Jun
Department of Emergency, Taizhou People's Hospital Affiliated to Nantong University, Taizhou, Jiangsu 225300, P.R. China.
Department of Emergency, The Fourth Affiliated Hospital of Zhejiang University, School of Medicine, Yiwu, Zhejiang 322000, P.R. China.
Exp Ther Med. 2021 Dec;22(6):1453. doi: 10.3892/etm.2021.10888. Epub 2021 Oct 15.
Cerebral ischemic stroke is the primary cause of stroke-associated mortality and disability, and current therapeutic options are limited and ineffective. The present study aimed to investigate the potential of apoptotic therapy and the role of microRNA (miR)-424 in cerebral ischemic stroke. PC12 cells, a cloned cell line from rat adrenal pheochromocytoma, were treated with CoCl to construct a cellular ischemia model. mRNA and protein levels of programmed cell death protein 4 (PDCD4), Bcl-2, Bax, caspase-3, PI3K and AKT were evaluated using reverse transcription-quantitative PCR and western blot analyses, respectively. Cell Counting Kit-8 assays were performed to examine cell viability in the ischemia model. Flow cytometry was conducted to evaluate the apoptosis of ischemic cells. Furthermore, a luciferase assay was performed to verify the target gene of miR-424. It was revealed that the expression level of miR-424 was downregulated in the ischemia model, while the expression of PDCD4 was upregulated. Moreover, the expression of miR-424 was increased after treatment with miR-424 mimics. The mRNA and protein expression of PDCD4 was upregulated after transfection with pcDNA3.1-PDCD4. PDCD4 was predicted and demonstrated to be a target of miR-424. Notably, overexpression of miR-424 increased cell viability and inhibited apoptosis in the ischemia model, which was reversed by co-treatment with pcDNA3.1-PDCD4. Furthermore, overexpression of miR-424 regulated the expression of PDCD4, Bax, Bcl-2, phosphorylated-PI3K/AKT and caspase-3, which was restored after co-transfection with pcDNA3.1-PDCD4. Collectively, the results indicated that miR-424 regulated the progression of cerebral ischemic stroke in a cellular model by targeting PDCD4.
脑缺血性中风是中风相关死亡率和残疾的主要原因，目前的治疗选择有限且效果不佳。本研究旨在探讨凋亡治疗的潜力以及微小RNA（miR）-424在脑缺血性中风中的作用。PC12细胞是一种源自大鼠肾上腺嗜铬细胞瘤的克隆细胞系，用氯化钴处理以构建细胞缺血模型。分别使用逆转录定量PCR和蛋白质印迹分析评估程序性细胞死亡蛋白4（PDCD4）、Bcl-2、Bax、半胱天冬酶-3、磷脂酰肌醇-3激酶（PI3K）和蛋白激酶B（AKT）的mRNA和蛋白质水平。进行细胞计数试剂盒-8检测以检查缺血模型中的细胞活力。进行流式细胞术以评估缺血细胞的凋亡。此外，进行荧光素酶测定以验证miR-424的靶基因。结果显示，在缺血模型中miR-424的表达水平下调，而PDCD4的表达上调。此外，用miR-424模拟物处理后miR-424的表达增加。用pcDNA3.1-PDCD4转染后，PDCD4的mRNA和蛋白质表达上调。预测并证明PDCD4是miR-424的靶标。值得注意的是，miR-424的过表达增加了缺血模型中的细胞活力并抑制了细胞凋亡，而与pcDNA3.1-PDCD4共同处理可逆转这种情况。此外，miR-424的过表达调节了PDCD4、Bax、Bcl-2、磷酸化-PI3K/AKT和半胱天冬酶-3的表达，与pcDNA3.1-PDCD4共转染后这些表达得以恢复。总体而言，结果表明miR-424在细胞模型中通过靶向PDCD4调节脑缺血性中风的进展。
