核受体NR6A1通过上调RIPK3基因表达诱导血管平滑肌细胞凋亡

目的:探讨核受体亚家族6A1(NR6A1)对血管平滑肌细胞凋亡的影响及可能的分子机制。方法:将腺病毒Ad-NR6A1感染大鼠血管平滑肌细胞,分别在感染后0 h、24 h和48 h时进行MTT实验,以时间为横坐标,A570为纵坐标绘制细胞生长曲线,观察NR6A1对细胞生长的影响;进行DAPI染色、TUNEL染色及caspase活性检测,观察细胞凋亡情况;进一步通过基因芯片技术,寻找NR6A1的靶基因;采用siRNA介导的基因沉默技术,观察受体相互作用丝氨酸/苏氨酸蛋白激酶3(RIPK3)基因沉默对NR6A1诱导的血管平滑肌细胞凋亡的影响。结果:腺病毒Ad-NR6A1感染细胞48 h时,NR6A1过表达组细胞数量较对照组(重组腺病毒载体Ad-Lac Z)明显减少;DAPI染色显示NR6A1过表达诱导血管平滑肌细胞出现核浓缩和核碎裂的凋亡表型,TUNEL染色显示NR6A1过表达引起细胞凋亡,caspase活性检测结果显示NR6A1过表达细胞内caspase-3、caspase-8和caspase-9活性均较对照组高;基因芯片技术检测发现,NR6A1过表达上调血管平滑肌细胞中RIPK3基因表达;RIPK3基因沉默可以显著抑制NR6A1诱导的平滑肌细胞凋亡。结论:NR6A1通过上调RIPK3基因表达诱导血管平滑肌细胞凋亡。

NR6A1 promotes vascular smooth muscle cell apoptosis by up-regulating RIPK3 gene expression

AIM: To determine the role of nuclear receptor subfamily 6, group A, member 1 (NR6A1) in vascular smooth muscle cell (VSMC) apoptosis.METHODS: NR6A1 protein was over-expressed in the VSMCs by infection of adenovirus. The effect of NR6A1 on the viability of VSMCs was measured by MTT assay. DAPI staining, TUNEL staining and caspase activity assay were conducted. DNA microarray was used to quickly screen the target genes of NR6A1. The effect of receptor-interacting serine/threonine-protein kinase 3 (RIPK3) silencing on NR6A1-induced apoptosis of the VSMCs was further analyzed.RESULTS: Adenovirus-mediated over-expression of NR6A1 induced the apoptosis of VSMCs. The RIPK3 gene expression was up-regulated by NR6A1 over-expression in the VSMCs. NR6A1-induced VSMC apoptosis was inhibited by RIPK3 silencing.CONCLUSION: NR6A1 promotes VSMC apoptosis by up-regulating the RIPK3 gene expression.