Wang Yibiao, Xu Min
Department of Neurosurgery, Hainan Affiliated Hospital of Hainan Medical University, Haikou City, Hainan Province, 570311, China.
Department of Neurosurgery, Kunshan Hospital of Traditional Chinese Medicine, Kunshan Affiliated Hospital of Nanjing University of Chinese Medicine, No. 189 Chaoyang Road, Kunshan City, Jiangsu Province, 215300, China.
Transl Neurosci. 2021 May 18;12(1):210-217. doi: 10.1515/tnsci-2020-0172. eCollection 2021 Jan 1.
This study aimed to explore the role of miR-380-5p in cerebral ischemia/reperfusion (CIR) injury-induced neuronal cell death and the potential signaling pathway involved.
Human neuroblastoma cell line SH-SY5Y cells were used in this study. Oxygen and glucose deprivation/reperfusion (OGD/R) model was used to mimic ischemia/reperfusion injury. CCK-8 assay and flow cytometry were used to examine cell survival. Quantitative real time PCR (RT-qPCR) assay and Western blotting were used to measure the change of RNA and protein expression, respectively. TargetScan and Luciferase assay was used to confirm the target of miR-380-5p. Malondialdehyde (MDA) superoxide dismutase (SOD) and glutathione peroxidase (GSHPx) were measured using commercial kits.
miR-380-5p was downregulated in SH-SY5Y cells after OGD/R. Cell viability was increased by miR-380-5p, while cell apoptosis was reduced by miR-380-5p mimics. MDA was reduced by miR-380-5p mimics, while SOD and GSHPx were increased by miR-380-5p. Results of TargetScan and luciferase assay have showed that BACH1 is the direct target of miR-380-5p. Expression of NRF2 was upregulated after OGD/R, but was not affected by miR-380-5p. mRNA expression of HO-1 and NQO1 and ARE activity were increased by miR-380-5p. Overexpression of BACH1 reversed the antioxidant and neuroprotective effects of miR-380-5p.
miR-380-5p inhibited cell death induced by CIR injury through target BACH1 which also facilitated the activation of NRF2, indicating the antioxidant and neuroprotective effects of miR-380-5p.
本研究旨在探讨miR - 380 - 5p在脑缺血/再灌注（CIR）损伤诱导的神经元细胞死亡中的作用以及潜在的信号通路。
本研究使用人神经母细胞瘤细胞系SH - SY5Y细胞。采用氧糖剥夺/再灌注（OGD/R）模型模拟缺血/再灌注损伤。使用CCK - 8法和流式细胞术检测细胞存活率。分别采用定量实时PCR（RT - qPCR）法和蛋白质印迹法检测RNA和蛋白质表达的变化。使用TargetScan和荧光素酶测定法确认miR - 380 - 5p的靶标。使用商业试剂盒测定丙二醛（MDA）、超氧化物歧化酶（SOD）和谷胱甘肽过氧化物酶（GSHPx）。
OGD/R后SH - SY5Y细胞中miR - 380 - 5p表达下调。miR - 380 - 5p提高细胞活力，同时miR - 380 - 5p模拟物降低细胞凋亡。miR - 380 - 5p模拟物降低MDA水平，同时miR - 380 - 5p提高SOD和GSHPx水平。TargetScan和荧光素酶测定结果表明BACH1是miR - 380 - 5p的直接靶标。OGD/R后NRF2表达上调，但不受miR - 380 - 5p影响。miR - 380 - 5p提高HO - 1和NQO1的mRNA表达以及ARE活性。BACH1的过表达逆转了miR - 380 - 5p的抗氧化和神经保护作用。
miR - 380 - 5p通过靶向BACH1抑制CIR损伤诱导的细胞死亡，这也促进了NRF2的激活，表明miR - 380 - 5p具有抗氧化和神经保护作用。
