姜黄素通过上调LncRNA NORAD/miR-543-3p对MPP+诱导的帕金森病细胞模型的影响

The effects of curcumin on MPP~+-induced Parkinson’s cell model by upregulating LncRNA NORAD/mir-543-3p

目的探讨姜黄素对1-甲基-4-苯基吡啶离子(MPP+)诱导的帕金森病细胞模型的影响及分子机制。方法使用MPP+(5 mmol/L)处理MN9D细胞构建帕金森病细胞模型,记为MPP+组;正常培养的细胞作为对照组;细胞给予40μmol/L姜黄素处理后再用MPP+处理记为MPP++姜黄素组;将NORAD siRNA转染至细胞后分别用40μmol/L姜黄素和MPP+处理记为MPP++姜黄素+si-NORAD组;将NORAD siRNA和miR-543-3p inhibitor共转染细胞后分别用40μmol/L姜黄素和MPP+处理记为MPP++姜黄素+si-NORAD+anti-miR-543-3p组,实时荧光定量PCR(qRT-PCR)检测lncRNA NORAD和miR-543-3p表达水平,流式细胞术检测细胞凋亡,双荧光素酶实验检测lncRNA NORAD和miR-543-3p的靶向关系。结果与对照组比较,MPP+组MN9D细胞中lncRNA NORAD表达下调,miR-543-3p表达上调,细胞凋亡率升高, MDA水平明显升高,SOD水平明显降低;与MPP+组比较,MPP++姜黄素组lncRNA NORAD表达上调,miR-543-3p表达下调,细胞凋亡率降低, MDA水平明显降低,SOD水平明显升高;lncRNA NORAD靶向负调控miR-543-3p的表达;抑制lncRNA NORAD逆转了姜黄素对MN9D细胞凋亡和氧化应激的抑制作用;抑制miR-543-3p逆转了lncRNA NORAD的作用。结论姜黄素可抑制MPP+诱导的MN9D细胞凋亡和氧化应激,其机制可能与调控lncRNA NORAD/miR-543-3p通路有关。

Objective To investigate the effects of curcumin on 1-methyl-4-phenylpyridine(MPP~+) induced Parkinson’s cell model and its molecular mechanism. Methods MN9 D cells were treated with MPP+(5 mmol/L) to construct the Parkinson’s cell model, which was recorded as MPP+ group, and the normal cultured cells were used as control group. The cells were treated with 40 mol/L curcumin and then treated with MPP~+, which was denoted as MPP~++ curcumin group. NORAD siRNA was transfected into the cells and treated with 40 mol/L curcumin and MPP+, respectively, which was denoted as MPP~++ curcumin + si-NORAD group. After co-transfection with NORAD siRNA and miR-543-3 p inhibitor, the cells were treated with 40 mol/L curcumin and MPP~+, which was denoted as MPP~++ curcumin + si-NORAD + anti-miR-543-3 p group. The lncRNA NORAD and miR-543-3 p expressive levels were detected by real-time fluorescent quantitative PCR(qRT-PCR), apoptosis was detected by flow cytometry, and the targeting relationship between lncRNA NORAD and miR-543-3 p was detected by double luciferase assay. Results Compared with the control group, the lncRNA NORAD expressive level in MN9 D cells in MPP~+ group was down-regulated, the miR-543-3 p expressive level was up-regulated, the apoptosis rate was increased, and the MDA level was significantly increased, SOD level was significantly reduced. Compared with the MPP~+ group, the lncRNA NORAD expressive level in the MPP~++ curcumin group was up-regulated, the miR-543-3 p expressive level was down-regulated, the apoptosis rate was decreased, and the MDA level was significantly reduced, SOD level was increased significantly. LncRNA NORAD negatively regulated the miR-543-3 p expression. Inhibition of lncRNA NORAD reversed the inhibitory effect of curcumin on apoptosis and oxidative stress of MN9 D cells. Inhibition of miR-543-3 p reversed the role of lncRNA NORAD. Conclusion Curcumin could inhibit MPP~+-induced apoptosis and oxidative stress of MN9 D cells, and the mechanism might be related to the regulation of the lncRNA NORAD/miR-543-3 p pathway.