lncRNA CRNDE靶向miR-212-5p调控氧糖剥夺诱导的大鼠皮质神经元损伤

吴明锋; 韩晓艳; 郭建磊; 张士忠

doi:10.13898/j.cnki.issn.1000-2200.2023.06.003

lncRNA CRNDE靶向miR-212-5p调控氧糖剥夺诱导的大鼠皮质神经元损伤

LncRNA CRNDE regulates oxygen and glucose deprivation-induced rat cortical neuron injury by targeting miR-212-5p

摘要

摘要:
目的探讨lncRNA CRNDE对氧糖剥夺(OGD)诱导的大鼠皮质神经元损伤的影响及作用机制。
方法培养大鼠皮质神经元，分为对照组(Con组)、OGD组、OGD+si-NC组、OGD+si-CRNDE组、OGD+miR-NC组、OGD+miR-212-5p组、OGD+si-CRNDE+anti-miR-NC组和OGD+si-CRNDE+anti-miR-212-5p组，RT-qPCR法检测细胞中CRNDE和miR-212-5p表达水平，CCK-8法检测细胞存活率，试剂盒法检测细胞培养上清液中乳酸脱氢酶(LDH)漏出率，流式细胞术检测细胞凋亡率，Western blotting检测细胞中Bcl-2、Bax蛋白表达。双荧光素酶报告基因实验验证CRNDE和miR-212-5p的调控关系。
结果与Con组比较，OGD组CRNDE水平、LDH漏出率、细胞凋亡率、Bax蛋白水平升高(P < 0.01)，miR-212-5p水平、细胞存活率、Bcl-2蛋白水平降低(P < 0.01)。与OGD+si-NC组比较，OGD+si-CRNDE组细胞存活率、Bcl-2蛋白水平升高(P < 0.01)，LDH漏出率、细胞凋亡率和Bax蛋白水平降低(P < 0.01)。与OGD+miR-NC组比较，OGD+miR-212-5p组细胞存活率、Bcl-2蛋白水平升高(P < 0.01)，LDH漏出率、细胞凋亡率和Bax蛋白水平降低(P < 0.01)。与OGD+si-CRNDE+anti-miR-NC组比较，OGD+si-CRNDE+anti-miR-212-5p组细胞存活率、Bcl-2蛋白水平降低(P < 0.01)，LDH漏出率、细胞凋亡率和Bax蛋白水平升高(P < 0.01)。CRNDE靶向负调控miR-212-5p表达。
结论干扰CRNDE表达可靶向上调miR-212-5p促进OGD诱导的大鼠皮质神经元增殖，并抑制神经元凋亡及LDH漏出率，减轻了神经元损伤。

Abstract:
ObjectiveTo investigate the effect and mechanism of lncRNA CRNDE on oxygen and glucose deprivation (OGD)-induced rat cortical neurons injury.
MethodsRat cortical neurons were cultured and divided into Con group, OGD group, OGD+si-NC group, OGD+si-CRNDE group, OGD+miR-NC group, OGD+miR-212-5p group, OGD+si-CRNDE+anti-miR-NC group and OGD+si-CRNDE+anti-miR-212-5p group. The expression levels of CRNDE and miR-212-5p in the cells were detected by RT-qPCR, the cell survival rate was detected by CCK-8 method, lactic dehydrogenase(LDH) leakage rate in the cell culture supernatant was detected by the related kit, the apoptosis rate was detected by flow cytometry, and the expression of Bcl-2 and Bax proteins was detected by Western blotting. The dual luciferase reporter gene experiment was used to verify the regulatory relationship between CRNDE and miR-212-5p.
ResultsCompared with the Con group, the CRNDE level, LDH leakage rate, apoptosis rate, and Bax protein level in the OGD group were increased (P < 0.01), but miR-212-5p level, cell survival rate, and Bcl-2 protein level were decreased (P < 0.01). Compared with the OGD+si-NC group, the cell survival rate and Bcl-2 protein level in the OGD+si-CRNDE group were increased (P < 0.01), while LDH leakage rate, apoptosis rate and Bax protein level were decreased (P < 0.01). Compared with the OGD+miR-NC group, the cell survival rate and Bcl-2 protein level in the OGD+miR-212-5p group were increased (P < 0.01), while LDH leakage rate, apoptosis rate and Bax protein level were decreased (P < 0.01). Compared with the OGD+si-CRNDE+anti-miR-NC group, the cell survival rate and Bcl-2 protein level in the OGD+si-CRNDE+anti-miR-212-5p group were decreased (P < 0.01), meanwhile, LDH leakage rate, apoptosis rate and Bax protein level were increased (P < 0.01). CRNDE negatively regulated the expression of miR-212-5p.
ConclusionsInterfering CRNDE expression can promote the proliferation of OGD-induced rat cortical neurons and inhibit apoptosis and LDH leakage rate by up-regulating miR-212-5p, which reduces neuron injury.

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