CircOGDH通过调控miR-195-5p/PGAM5轴对氧糖剥夺/复糖复氧诱导的小胶质细胞损伤的影响

doi:10.3969/j.issn.1006-6233.2024.09.05

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摘要 目的: 探讨环状RNA OGDH(CircOGDH)通过调控miR-195-5p/磷酸甘油酸变位酶5(PGAM5)轴对氧糖剥夺/复糖复氧(OGD/R)诱导的小胶质(MG)细胞损伤的影响。方法: 将体外培养的HMC3细胞分为:NC组(正常培养)、OGD/R、si-NC组、si-CircOGDH组、mimic NC组、miR-195-5p mimic组、si-CircOGDH+inhibitor NC组、si-CircOGDH+miR-195-5p inhibitor组。除NC组外,剩余组在转染对应物质前构建OGD/R模型。qRT-PCR法测定CircOGDH、miR-195-5p、PGAM5 mRNA表达水平;测定各组细胞增殖、凋亡、活性氧(ROS)及炎性因子水平;Western blot检测Cleaved caspase-3、Bax表达;双荧光素酶基因实验检测CircOGDH与miR-195-5p、PGAM5与miR-195-5p之间的靶向关系。结果: 与NC组对比,OGD/R组miR-195-5p表达、OD450值均降低,CircOGDH、PGAM5 mRNA表达、凋亡率、ROS及炎性因子水平、Cleaved caspase-3、Bax蛋白表达均升高(P<0.05);对比OGD/R组和si-NC组,si-CircOGDH组miR-195-5p表达、OD450值均升高,CircOGDH、PGAM5 mRNA表达、凋亡率、ROS及炎性因子水平、Cleaved caspase-3、Bax蛋白表达均降低(P<0.05);miR-195-5p mimic组分别与OGD/R组、mimic NC组对比,CircOGDH无显著差异(P>0.05),miR-195-5p表达、OD450值均升高,PGAM5 mRNA表达、凋亡率、ROS及炎性因子水平、Cleaved caspase-3、Bax蛋白表达均降低(P<0.05);与si-CircOGDH+inhibitor NC组对比,si-CircOGDH+miR-195-5p inhibitor组CircOGDH差异不显著(P>0.05),miR-195-5p表达、OD450值均降低,PGAM5 mRNA表达、凋亡率、ROS及炎性因子水平、Cleaved caspase-3、Bax蛋白表达均升高(P<0.05)。双荧光素酶基因实验显示miR-195-5p和CircOGDH、PGAM5均存在靶向关系。结论: 沉默CircOGDH可促进OGD/R诱导的HMC3细胞增殖,抑制其炎症反应和凋亡,可能与调控miR-195-5p/PGAM5轴有关。

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关键词 ：环状RNA OGDH, miR-195-5p, 磷酸甘油酸变位酶5, 氧糖剥夺/复糖复氧, 小胶质细胞

Abstract：Objective: To investigate the effect of circular RNA OGDH (CircOGDH) on microglia (MG) cell damage induced by oxygen-glucose deprivation/reoxygenation (OGD/R) by modulating the miR-195-5p/phosphoglycerate mutase 5 (PGAM5) axis.Methods: HMC3 cells cultured in vitro were separated into:NC group (normal culture), OGD/R, si-NC group, si-CircOGDH group, mimic NC group, miR-195-5p mimic group, si-CircOGDH+inhibitor NC group, and si-CircOGDH+miR-195-5p inhibitor group. Except for the NC group, the remaining groups constructed OGD/R models before transfecting the corresponding substances. QRT-PCR method was applied to determine the expression levels of CircOGDH, miR-195-5p, and PGAM5 mRNA. The levels of cell proliferation, apoptosis, reactive oxygen species (ROS), and inflammatory factors were measured in each group. Western blot was applied to detect the expression of Cleaved caspase-3 and Bax. Dual luciferase gene experiment was used to detect the targeting relationship between CircOGDH and miR-195-5p, and between PGAM5 and miR-195-5p.Results: Compared with the NC group, the expression of miR-195-5p and OD450 value were reduced in the OGD/R group, the mRNA expression of CircOGDH and PGAM5, apoptosis rate, ROS, and inflammatory cytokine levels, and the expression of Cleaved caspase-3 and Bax proteins were all elevated (P<0.05). Compared with the OGD/R group and the si-NC group, the expression of miR-195-5p and OD450 value were elevated in the si-CircOGDH group, the mRNA expression of CircOGDH and PGAM5, apoptosis rate, ROS and inflammatory cytokine levels, and the expression of Cleaved caspase-3 and Bax proteins were all reduced (P<0.05). Compared with the OGD/R group and the mimic NC group, the miR-195-5p mimic group showed no significant difference in CircOGDH (P>0.05), while the expression of miR-195-5p and OD450 values were increased, the expression of PGAM5 mRNA, apoptosis rate, ROS and inflammatory cytokine levels, and the expression of Cleaved caspase-3 and Bax protein were all reduced (P<0.05). Compared with the si-CircOGDH+inhibitor NC group, the CircOGDH of the si-CircOGDH+miR-195-5p inhibitor group was not significantly different (P>0.05), the expression of miR-195-5p and OD450 values were decreased, the expression of PGAM5 mRNA, apoptosis rate, ROS and inflammatory cytokine levels, and the expression of Cleaved caspase-3 and Bax protein were all elevated (P<0.05). A dual luciferase gene experiment showed that miR-195-5p had a targeting relationship with CircOGDH and PGAM5.Conclusion: Silencing CircOGDH can promote the proliferation of HMC3 cells induced by OGD/R, inhibit their inflammatory response and apoptosis, and may be related to the regulation of miR-195-5p/PGAM5 axis.

Key words： Circular RNA OGDH MiR-195-5p Phosphoglycerate mutase 5 Oxygen glucose deprivation/reoxygenation Microglia

基金资助:2024年度河北医学科学研究计划,(编号:20240045)

通讯作者: 张云鹤

引用本文:

李向男, 杨松涛, 李林, 张云鹤, 张志勇, 李建民, 付爱军. CircOGDH通过调控miR-195-5p/PGAM5轴对氧糖剥夺/复糖复氧诱导的小胶质细胞损伤的影响[J]. 河北医学, 2024, 30(9): 1440-1445.
LI Xiangnan, YANG Songtao, LI Lin, et al. Effect of CircOGDH on Oxygen-Glucose Deprivation/Reoxygenation-Induced Microglial Cell Injury through Modulation of the miR-195-5p/PGAM5 Axis. HeBei Med, 2024, 30(9): 1440-1445.

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http://www.hbyxzzs.cn/CN/10.3969/j.issn.1006-6233.2024.09.05 或 http://www.hbyxzzs.cn/CN/Y2024/V30/I9/1440

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