生理性低氧及缺氧再复氧对小鼠骨髓间充质干细胞增殖及成骨分化的影响  

Effects of physiological hypoxia and anoxia-reoxygenation on proliferation and osteogenic differentiation of mouse bone marrow mesenchymal stem cells

作　　者：郭婷婷[1] 刘怡[2] 周建 GUO Ting-ting;LIU Yi;ZHOU Jian(Department of General Dentistry and Emergency Dental Care,Capital Medical University School of Stomatology,Beijing 100050,China)

机构地区：[1]首都医科大学口腔医学院急诊综合诊疗中心,北京100050 [2]首都医科大学口腔医学院牙周科,北京100050 [3]首都医科大学口腔医学院特诊特需科,北京100050

出　　处：《北京口腔医学》2023年第2期82-87,共6页Beijing Journal of Stomatology

基　　金：首都医科大学附属北京口腔医院学科建设基金(18-09-06)。

摘　　要：目的 探索生理性低氧及缺氧再复氧对于骨髓间充质干细胞(bone marrow mesenchymal stem cells,BMSCs)增殖、成骨分化能力的影响。方法 体外分离培养C57BL/6J小鼠的BMSCs,置于生理性低氧(3%O_(2))、常氧(21%O_(2))、缺氧再复氧环境,通过CCK8检测BMSCs增殖情况,同时成骨诱导,通过茜素红染色检测体外矿化能力,RT-PCR检测成骨向分化相关基因(Alp、Runx2、Opn)mRNA水平;Western blot检测RUNX2、OPN、LC3-Ⅱ/LC3-I、P62、cleaved-caspase3蛋白表达量。结果 生理性低氧较常氧CCK8表达显著升高,增加成骨相关基因(Runx2、Alp、Opn)mRNA水平并促进RUNX2、OPN蛋白表达;缺氧复氧后,常氧组LC3-Ⅱ/LC3-I表达减弱,cleaved-caspase3表达增强,Opn、OPN较低氧组降低。结论 生理性低氧较常氧能促进BMSCs的增殖、成骨分化能力。缺氧复氧过程中,生理性低氧较常氧可减缓无氧干预对细胞增殖、成骨分化的抑制作用。Objective To investigate the effects of physiological hypoxia and anoxia-reoxygenation on the proliferation and osteogenic differentiation of bone marrow mesenchymal stem cells(BMSCs)in vitro.Methods BMSCs of C57BL/6J mice were isolated and cultured in vitro and placed in physiological hypoxia(3%O_(2)),normoxia(21%O_(2)),and anoxia-reoxygenation environment.BMSCs proliferation was detected by CCK8,and osteogenic induction was conducted at the same time.In vitro mineralization ability was detected by alizarine red staining and calcium ion relative concentration.RT-PCR was used to detect mRNA levels of osteogenic differentiation-related genes(Alp,Runx2,Opn).Western blot was used to detect the protein expression levels of RUNX2,OPN,LC3-II/LC3-I,P62,and Cleaved-Caspase3.Results The expression of CCK8 was significantly increased in physiological hypoxia compared with normoxia,and the mRNA levels of osteogenic genes(Runx2,Alp,Opn)and the protein expressions of RUNX2 and OPN were increased.After anoxiareoxygenation,proliferation and osteogenesis were inhibited in both the hypoxia and normoxia groups compared with the control group.Autophagy was inhibited in the normoxia group,the expression of Cleaved-Caspase3 protein was enhanced,and Opn and OPN were further decreased compared with the hypoxia group.Conclusions 3%O_(2) physiological hypoxia can promote the proliferation and osteogenic differentiation of BMSCs compared with 21%O_(2) normoxia.In anoxia-reoxygenation,3%O2 hypoxia compared with 21%O_(2) normoxia could alleviate the inhibition of cell proliferation and osteogenic differentiation caused by the anaerobic intervention.

关 键 词：骨髓间充质干细胞 生理性低氧 缺氧复氧 细胞增殖 成骨分化 

分 类 号：R681.1[医药卫生—骨科学]