低氧预处理改善去卵巢大鼠骨髓间充质干细胞的成骨分化潜能  被引量:5

Hypoxic precondition rescues osteogenic potential of bone marrow mesenchymal stem cells derived from ovariectomized rats

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作  者:黄晓雄 陈维凯 刘滔[1] 杨惠林[1,2] 何帆[1,2] Huang Xiaoxiong;Chen Weikai;Liu Tao;Yang Huilin;He Fan(Department of Orthopedics,the First Affiliated Hospital of Soochow University,Suzhou 215006,Jiangsu Province,China;Institute of Orthopaedics at Soochow University,Suzhou 215007,Jiangsu Province,China)

机构地区:[1]苏州大学附属第一医院骨科,江苏省苏州市215006 [2]苏州大学骨科研究所,江苏省苏州市215007

出  处:《中国组织工程研究》2022年第13期2034-2039,共6页Chinese Journal of Tissue Engineering Research

基  金:国家自然科学基金(31771063),项目负责人:何帆。

摘  要:背景:低氧是调节血管生成-成骨分化过程的主要驱动力之一,细胞在分化前进行低氧预处理能够保持干细胞干性和增强干细胞的成骨分化能力,但低氧预处理能否改善去卵巢大鼠骨髓间充质干细胞的成骨分化能力尚不清楚。目的:探讨低氧预处理对去卵巢大鼠骨髓间充质干细胞成骨分化能力的影响以及低氧诱导因子1α在其中的作用。方法:首先建立去卵巢SD大鼠模型,提取假手术大鼠和去卵巢大鼠骨髓间充质干细胞。将细胞分为3组,①正常组:假手术组大鼠骨髓间充质干细胞在常氧环境培养(体积分数21%氧气);②骨质疏松组:去卵巢大鼠骨髓间充质干细胞在常氧环境培养(体积分数21%氧气);③低氧组:去卵巢大鼠骨髓间充质干细胞在低氧环境培养(体积分数5%氧气)。采用CCK-8法检测第1,3,5,7天的吸光度值来反映各组细胞的增殖能力;在低氧环境培养72 h后,转移至正常培养箱进行成骨诱导分化14 d,采用茜素红染色和qRT-PCR检测成骨相关基因表达水平。最后将骨质疏松组骨髓间充质干细胞转染小干扰RNA沉默低氧诱导因子1α表达,在低氧环境培养72 h,转移至正常培养箱进行成骨诱导分化14 d,观察沉默低氧诱导因子1α后成骨能力变化。结果与结论:①去卵巢大鼠骨髓间充质干细胞增殖能力下降,同时成骨分化能力受损;②低氧环境可以促进去卵巢大鼠骨髓间充质干细胞增殖,提高钙结节的沉积以及成骨相关基因的表达;③沉默低氧诱导因子1α后,低氧预处理改善成骨能力的效应被消除;④结果表明,低氧预处理可以改善去卵巢大鼠骨髓间充质干细胞的增殖及成骨分化能力,低氧诱导因子1α在低氧预处理促进成骨分化过程中起到重要作用。BACKGROUND:Hypoxia is one of the main driving forces regulating the angiogenesis-osteogenesis differentiation process.Hypoxia treatment of cells before differentiation is considered to maintain the stemness and enhance the osteogenic potential of stem cells.However,it is not clear whether hypoxic precondition could improve the osteogenic potential of bone marrow mesenchymal stem cells derived from osteoporosis rat.OBJECTIVE:To investigate the effects of hypoxic precondition on the osteogenic potential of bone marrow mesenchymal stem cells derived from osteoporosis rat and the role of hypoxia inducible factor-1α.METHODS:Ovariectomized SD rat models were established.Bone marrow mesenchymal stem cells were isolated from femurs of female rats in ovariectomized group and sham operation group.The bone marrow mesenchymal stem cells were divided into three groups.In the normal group,the bone marrow mesenchymal stem cells extracted from the sham rats were cultured in the incubator with normal oxygen concentration(21%O_(2)).In the osteoporosis group,the bone marrow mesenchymal stem cells extracted from the ovariectomized rats were cultured in the incubator with normal oxygen concentration(21%O_(2)).In the hypoxia group,the bone marrow mesenchymal stem cells extracted from the ovariectomized rats were cultured in the hypoxic incubator with hypoxic oxygen concentration(5%O_(2)).CCK-8 assay was used to assess the proliferation capacity of bone marrow mesenchymal stem cells by measuring absorbance values at 1,3,5,and 7 days.After 72 hours of incubation in hypoxic environment,cells were moved to normal incubator for osteogenic differentiation with other two groups for 14 days.Alizarin red staining and qRT-PCR were utilized to detect osteogenic related gene expression levels.Finally,the bone marrow mesenchymal stem cells of the osteoporosis group were transfected with small interfering RNA to silence the expression of hypoxia inducible factor-1α,cultured in a hypoxic environment for 72 hours,and transferred to a normal incubat

关 键 词:干细胞 骨髓间充质干细胞 骨质疏松 低氧 细胞增殖 成骨分化 低氧诱导因子1Α 

分 类 号:R459.9[医药卫生—治疗学] R318[医药卫生—临床医学]

 

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