芍药苷对高糖环境下雪旺细胞线粒体动力学的影响

Effect of paeoniflorin on mitochondrial dynamics of Schwann cells in high glucose environment

目的观察芍药苷(PF)对高糖环境下雪旺细胞线粒体动力学的影响。方法选取大鼠雪旺细胞(RSC96细胞株)培养,分为空白对照组(Con,25 mmol/L葡萄糖)、高糖组(HG,150 mmol/L葡萄糖)、PF 1μmol/L组(PF 1,150 mmol/L葡萄糖+1μmol/L PF)、PF 10μmol/L组(PF10,150 mmol/L葡萄糖+10μmol/L PF)、PF 100μmol/L组(PF 100,150 mmol/L葡萄糖+100μmol/L PF),各组药物干预48 h。透射电镜观察线粒体形态,包括融合、分裂、溶胀、破裂等。Mitotracker-green标记线粒体联合活细胞激光扫描共聚焦显微镜,定量分析线粒体动力学参数,包括线粒体网状结构的平均分支长度和平均分支数。Western blot测定线粒体融合蛋白1(Mfn1)、Mfn2、视网膜萎缩蛋白1(Opa1)及线粒体分裂的动力相关蛋白1(Drp1)蛋白表达。结果与Con组比较,HG组线粒体多呈球状,线粒体网状结构的平均分支长度和数量减少(P<0.01),Mfn1、Mfn2和Opa1蛋白表达降低(P<0.01),Drp1蛋白表达升高(P<0.01);与HG组比较,PF 1、PF10、PF100组线粒体多呈棍状,线粒体网状结构平均分支长度和数量增加(P<0.01),Mfn1,Mfn2和Opa1蛋白表达升高(P<0.05或P<0.01),Drp1蛋白表达降低(P<0.05或P<0.01)。结论 PF能促进高糖环境下雪旺细胞线粒体融合,降低分裂,维持线粒体动力学平衡,改善线粒体形态与功能,降低雪旺细胞凋亡。

Objective To investigate the effect of paeoniflorin(PF)on mitochondrial dynamics of Schwann cell in high glucose environment.Methods Rat Schwann cells(RSC96 cell line)were cultured and divided into five groups:control group(Con,25 mmol/L glucose),high glucose control group(HG,150 mmol/L glucose),low,middle and high dose PF groups(150 mmol/L glucose+1 μmol/L,10 μmol/L and 100 μmol/L,PF 1,PF 10,PF 100). Each group was treated with drugs for 48 h. The morphology of mitochondria was observed by transmission electron microscope,including fusion,fission,swelling,and rupture. The parameters of mitochondrial dyanmics,including the average branch length and average branch number of mitochondrial reticular structure,were quantitatively analyzed by Mitotracker-green labeled mitochondria combined with laser scanning confocal microscope. The expressions of mitochondrial fusion protein mitofusin(Mfn)1,2,retinal atrophy protein 1(Opa1)and mitochondrial dynamic associated protein dynamin-relatedprotein 1(Drp1)were measured by Western blot.Results Compared with Con group,most of the mitochondria were sphericalin HG group,the average branch length and number of mitochondrial reticular structure decreased(P<0. 01),the expression of mitochondrial fusion proteins Mfn1,Mfn2 and Opa1 decreased(P<0. 01),while the expression of mitoprotein Drp1 increased in HG group (P<0. 01). Compared with HG group,most of the mitochondria are in the shape of a stick in PF treated groups,and the average branch length and number of mitochondrial reticular structure increased(P<0. 01),the expression of mitochondrial fusion protein Mfn1,Mfn2 and Opa1 increased(P<0. 01 or P<0. 05),while the expression of mitochondrial mitoprotein Drp1 decreased(P<0. 01 or P<0. 05)in PF 1,PF 10 and PF 100 groups.Conclusion PF can promote the mitochondrial fusion in SCs in high glucose environment,reduce division to maintain the balance of mitochondrial dyanmics,improve mitochondrial morphology,thus improve mitochondrial function and reduce apoptosis of SCs cells.