高糖环境下大鼠骨髓间充质干细胞的成骨分化和线粒体自噬相关性的研究  

The correlation between osteogenic differentiation and mitochondrial autophagy of rat bone marrow mesenchymal stem cells under high glucose environment

作　　者：关淼升 王潇宇 刘琳 傅博[2] 王田田 沈婉君 李鸿波[4] GUAN Miao-sheng;WANG Xiao-yu;LIU Lin;FU Bo;WANG Tian-tian;SHEN Wan-jun;LI Hong-bo(Medical School of Chinese PLA,Beijing,100853,China;Department of Stomatology,the First Medical Centre,Chinese PLA General Hospital,Beijing,100853,China)

机构地区：[1]解放军医学院,解放军总医院第一医学中心口腔科,北京100853 [2]解放军总医院第一医学中心肾脏病科,北京100853 [3]解放军医学院,解放军总医院第一医学中心肾脏病科,北京100853 [4]解放军总医院第一医学中心口腔科,北京100853

出　　处：《口腔颌面修复学杂志》2020年第2期65-71,共7页Chinese Journal of Prosthodontics

基　　金：北京市科新星计划交叉学科合作课题(项目编号:Z181100006218128)。

摘　　要：目的:探讨在高糖环境下大鼠骨髓间充质干细胞线粒体自噬和成骨分化的相关性。方法:将rBMSCs在高糖环境下(葡萄糖浓度分为5.5mM低糖组、30mM高糖组和甘露醇等渗组)成骨诱导7天,通过Western Blot实验检测LC3 I、LC3II、ALP、OPN蛋白的表达情况;通过RT-PCR实验检测LC3 I、LC3II、ALP、OPN基因的表达情况;激光共聚焦共定位实验比较线粒体标记物Mito-tracker和自噬溶酶体标记物LC3的共定位情况,分析高糖环境下rBMSCs成骨分化和线粒体自噬的相关性。结果:30mM高糖组能显著抑制rBMSCs的增殖,与5.5mM组相比,30mM高糖组内自噬标记蛋白LC3 II/LC3 I(P<0.01)、LC3 II(P<0.05)以及成骨标记蛋白ALP(P<0.01)和OPN(P<0.05)的表达下降;自噬基因LC3 II/LC3 I(P<0.05)和成骨基因ALP、OPN(P<0.001)表达水平降低;Mito-tracker和LC3在5.5mM组定位情况(PCC=0.52)优于30mM组(PCC=0.26)。结论:高糖环境能够显著抑制rBMSCs的成骨分化和线粒体自噬能力,两者的变化可能存在相关性。Objective:To investigate the correlation between osteogenic differentiation and mitophagy of rat bone marrow derived mesenchymal stem cells(rBMSCs)under high glucose environment.Methods:rBMSCs were cultured in the osteogenesis induction medium under high glucose environment,which was divided into 5.5 mM low glucose group,30 mM high glucose group and mannitol isotonic group.Western blot assay was conducted to detect the protein expression of LC3 I,LC3II,ALP,OPN,RT-PCR was performed to detect the gene expression of LC3 I,LC3II,ALP,OPN.Laser confocal experiments was performed to compare the colocalization of mitochondrial markers mito-tracker and autolysosome markers LC3.The correlation between osteogenic differentiation and mitochondrial autophagy of rBMSCs in a high glucose environment were analyzed.Results:In the 30 mM high glucose group the proliferation of rBMSCs was significantly inhibited.Compared with the 5.5 mM group,the autophagy marker proteins LC3 II/LC3 I(P<0.01)and LC3 II(P<0.05)and the expression of osteogenic marker proteins ALP(P<0.01)and OPN(P<0.05)decreased significantly;The expression levels of the autophagy genes LC3 II/LC3 I(P<0.05)and the osteogenic genes ALP and OPN(P<0.001)were significantly reduced;the colocalization of mito-tracker and LC3 in the 5.5mM group was better(PCC=0.52)than that under the 30mM group(PCC=0.26).Conclusion:Mitochondrial autophagy may play an important role in the decreased osteogenic differentiation of rBMSCs under high glucose environment.

关 键 词：骨髓间充质干细胞 成骨分化 线粒体自噬 高糖环境 

分 类 号：R780.2[医药卫生—口腔医学]