外泌体衍生的 exendin-4 通过 circRNA CDR1as/miR-671/GSK3β 信号通路逆转 PC12 细胞损伤。
Exendin-4 Reversed the PC12 Cell Damage Induced by circRNA CDR1as/miR-671/GSK3β Signaling Pathway.
机构信息
Department of Neurology, Shanghai Tenth People's Hospital, Tongji University School of Medicine, Shanghai, 200072, China.
Department of Central Laboratory, Clinical Laboratory, Jing'an District Center Hospital of Shanghai, Fudan University, Shanghai, 200040, China.
出版信息
J Mol Neurosci. 2021 Apr;71(4):778-789. doi: 10.1007/s12031-020-01698-2. Epub 2020 Sep 5.
The purpose of this paper is to study the effect of circRNA cerebellar degeneration-related protein 1 antisense RNA(CDR1as)/miR-671/GSK3β signaling pathway on PC12 cell injury and the mechanism of Exendin-4 (Ex-4) in PC12 cell injury protection. Quantitative reverse-transcription polymerase chain reaction (qRT-PCR) was used to detect the expression levels of circular RNA CDR1as and miR-671 in PC12 cells. By overexpressing or knocking out CDR1as, miR-671, and GSK3β, the role of CDR1as, miR-671, and GSK3β in PC12 cell injury was analyzed. The binding of CDR1as to miR-671 and GSK3β to miR-671 was verified by dual luciferase reporter assay. PC12 cells were treated with 1-methyl-4 phenyl-pyridine ion (MPP) to construct a PC12 cell damage model. PC12 cell transfection experiments were used to confirm the role of CDR1as/miR-671/GSK3β signal axis in PC12 cell damage, and the role of Ex-4 in the association of circRNA CDR1as/miR-671/GSK3β signaling axis and PC12 cell damage. PC12 cell damage was detected by 3-(4,5-dimethyl thiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) and cellular lactate dehydrogenase (LDH) release. Ex-4 reversed the phosphorylation levels of PI3K, AKT, and GSK-3β in MPP-treated PC12 cells, and reduced MPP-induced PC12 cell damage. CircRNA CDR1as upregulated the expression of GSK3β by sponge miR-671. Ex-4 downregulated CDR1as expression and upregulated miR-671 expression in MPP-induced PC12 cell. Silencing of CDR1as reduced MPP-induced PC12 cell damage. CDR1as transfection downregulated the expression of miR-671 in PC12 cells, promoted the expression and phosphorylated of GSK3β, and induced PC12 cell damage. GSK3β silencing reversed CDR1as-induced PC12 cell damage. CDR1as promoted the phosphorylation level of GSK3β in PC12 cells to cause cell damage; Ex-4 reversed the phosphorylation of GSK3β caused by CDR1as in PC12 cells and reduced the PC12 cell damage caused by CDR1as. Ex-4 reverses the damage of PC12 cells induced by CDR1as/miR-671/GSK3β signaling pathway.
本文旨在研究环状 RNA 小脑退化相关蛋白 1 反义 RNA（CDR1as）/miR-671/GSK3β 信号通路对 PC12 细胞损伤的影响及 Exendin-4（Ex-4）对 PC12 细胞损伤保护的作用机制。采用实时荧光定量聚合酶链反应（qRT-PCR）检测 PC12 细胞中环状 RNA CDR1as 和 miR-671 的表达水平。通过过表达或敲除 CDR1as、miR-671 和 GSK3β，分析 CDR1as、miR-671 和 GSK3β 在 PC12 细胞损伤中的作用。通过双荧光素酶报告基因实验验证 CDR1as 与 miR-671、GSK3β 与 miR-671 的结合。用 1-甲基-4-苯基-1,2,3,6-四氢吡啶离子（MPP）处理 PC12 细胞构建 PC12 细胞损伤模型。通过 PC12 细胞转染实验证实 CDR1as/miR-671/GSK3β 信号轴在 PC12 细胞损伤中的作用，以及 Ex-4 与 circRNA CDR1as/miR-671/GSK3β 信号轴和 PC12 细胞损伤的关联。通过 3-（4,5-二甲基噻唑-2-基）-2,5-二苯基四氮唑溴盐（MTT）和细胞乳酸脱氢酶（LDH）释放检测 PC12 细胞损伤。Ex-4 逆转了 MPP 处理的 PC12 细胞中 PI3K、AKT 和 GSK-3β 的磷酸化水平，并减轻了 MPP 诱导的 PC12 细胞损伤。环状 RNA CDR1as 通过海绵 miR-671 上调 GSK3β 的表达。Ex-4 下调了 MPP 诱导的 PC12 细胞中 CDR1as 的表达，上调了 miR-671 的表达。沉默 CDR1as 可减轻 MPP 诱导的 PC12 细胞损伤。CDR1as 转染可下调 PC12 细胞中 miR-671 的表达，促进 GSK3β 的表达和磷酸化，并诱导 PC12 细胞损伤。沉默 GSK3β 可逆转 CDR1as 诱导的 PC12 细胞损伤。CDR1as 促进 PC12 细胞中 GSK3β 的磷酸化水平导致细胞损伤；Ex-4 逆转了 CDR1as 在 PC12 细胞中引起的 GSK3β 磷酸化，并减轻了 CDR1as 引起的 PC12 细胞损伤。Ex-4 逆转了 CDR1as/miR-671/GSK3β 信号通路诱导的 PC12 细胞损伤。
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