1,25(OH)₂D₃通过稳定线粒体DNA对GCDC诱导的HiBECs凋亡的作用研究

The Role of 1,25(OH)₂D₃ on GCDC-Induced Apoptosis in HiBECs by Stabilising Mitochondrial DNA

目的:探讨骨化三醇(1,25(OH)2D3)对甘氨鹅脱氧胆酸盐(GCDC)诱导的人肝内胆管上皮细胞(HiBECs)凋亡的影响,并初步阐明其潜在的作用机制。方法:采用1 nM GCDC处理HiBECs建立原发性胆汁性胆管炎细胞模型,并采用不同浓度(0.1 nM、1 nM、10 nM、100 nM) 1,25(OH)2D3处理12 h。流式细胞术检测细胞凋亡水平,ELISA检测细胞培养液中炎症因子IL-6和IL-8水平,qPCR检测PDC-E2、PGC-1α、NrF-1和NrF-2的mRNA相对表达水平,Western blot检测细胞内Bcl-2、PDC-E2表达水平。结果:1 mM GCDC可以降低HiBECs细胞增殖活性,诱导HiBECs凋亡,提高细胞培养液中IL-6和IL-8水平,上调PDC-E2蛋白表达,下调Bcl-2蛋白表达,抑制COX-1、PGC-1α、NrF-1和NrF-2 mRNA相对表达水平。可以提高GCDC诱导的HiBECs细胞增殖活性,抑制GCDC诱导HiBECs细胞凋亡,降低细胞培养液中IL-6和IL-8水平,下调PDC-E2蛋白表达水平,提高COX-1、PGC-1α、NrF-1和NrF-2 mRNA相对表达水平。结论:1,25(OH)2D3可抑制诱导HiBECs细胞凋亡,其作用机制可能与线粒体DNA稳定有关。

Objective: To investigate the effects of osteotriol (1,25(OH)2D3) on Glycochenodeoxycholate (GCDC)-induced apoptosis of human intrahepatic biliary epithelial cells (HiBECs) and to preliminarily elucidate its potential mechanism of action. Methods: Primary biliary cholangitis cell model was established by treating HiBECs with 1 nM GCDC and treated with different concentrations (0.1 nM, 1 nM, 10 nM, 100 nM) of 1,25(OH)2D3 for 12 h. The apoptosis level was detected by flow cytometry, the levels of the inflammatory factors IL-6 and IL-8 were detected by ELISA in the cell culture fluid, and qPCR was performed to detect the relative mRNA expression levels of PDC-E2, PGC-1α, NrF-1 and NrF-2 were detected by qPCR, and the intracellular expression levels of Bcl-2 and PDC-E2 were detected by Western blot. Results: 1 mM GCDC could reduce the proliferative activity of HiBECs cells, induce apoptosis of HiBECs, increase the levels of IL-6 and IL-8 in the cell culture medium, up-regulate the expression of PDC-E2 protein, down-regulate the expression of Bcl-2 protein, and inhibit the relative expression levels of COX-1, PGC-1α, NrF-1, and NrF-2 mRNA. It can increase the proliferative activity of GCDC-induced HiBECs cells, inhibit GCDC-induced apoptosis of HiBECs cells, reduce the levels of IL-6 and IL-8 in the cell culture medium, down-regulate the level of PDC-E2 protein expression, and increase the level of COX-1, PGC-1α, NrF-1 and NrF-2 mRNA relative expression. Conclusion: 1,25(OH)2D3 can inhibit the induction of apoptosis in HiBECs, and its mechanism of action may be related to mitochondrial DNA stabilisation.