摘要
背景:肥胖是牙周炎及机体其他疾病的易感因素,因肥胖与瘦素密切相关,目前尚无通过瘦素基因突变小鼠骨髓间充质干细胞的分化情况探索相关机制的研究。目的:通过研究ob/ob遗传性肥胖小鼠骨髓间充质干细胞在脂多糖刺激下的成骨能力及成骨相关分子的表达,从分子及细胞水平探讨肥胖与成骨能力的相关机制。方法:选用8周龄雄性肥胖ob/ob小鼠、8周龄雄性C57小鼠(由中国医学科学院实验动物研究所提供)各8只,取两组小鼠双侧股骨,采用全骨髓法分离培养纯化得到ob/ob小鼠骨髓间充质干细胞及C57小鼠骨髓间充质干细胞,加入含有100μg/L脂多糖的成骨诱导液进行成骨诱导分化,通过碱性磷酸酶染色、茜素红染色和碱性磷酸酶活性检测观察比较两组小鼠骨髓间充质干细胞成骨能力。采用RT-PCR检测两组小鼠骨髓间充质干细胞中Alp、Runx2、Ocn、Nf-κb mRNA表达水平。结果与结论:(1)成骨诱导1周后,与C57小鼠骨髓间充质干细胞比较,ob/ob小鼠骨髓间充质干细胞碱性磷酸酶染色减弱,碱性磷酸酶活性显著降低(P <0.01);成骨诱导液中添加脂多糖后,碱性磷酸酶活性较未添加脂多糖组显著下降(P <0.01),ob/ob小鼠骨髓间充质干细胞碱性磷酸酶活性亦较C57小鼠骨髓间充质干细胞显著降低(P <0.05);(2)成骨诱导2周后,与C57小鼠骨髓间充质干细胞比较,ob/ob小鼠骨髓间充质干细胞茜素红染成橘红色的矿化结节减少,而成骨诱导液中添加脂多糖后,两组骨髓间充质干细胞均未见矿化结节形成;(3)成骨诱导后,与C57小鼠骨髓间充质干细胞比较,ob/ob小鼠骨髓间充质干细胞中Alp、Runx2、Ocn mR NA表达降低(P <0.05);诱导液中加入脂多糖后,Alp、Runx2、Ocn表达较未添加脂多糖组均显著减少(P <0.05),且在ob/ob小鼠骨髓间充质干细胞中Ocn、Runx2表达较C57小鼠骨髓间充质干细胞明显降低(P <0.05);(4)结果表明,瘦素缺乏的肥
BACKGROUND:Obesity is a predisposing factor for periodontitis and other diseases in the body.Increasing evidence has proved the close correlation between obesity and leptin.However,there is still no study on the differentiation of bone marrow mesenchymal stem cells from mice with a mutation in leptin gene.OBJECTIVE:To investigate the osteogenic differentiation and expression of osteogenic related molecules of bone marrow mesenchymal stem cells derived from ob/ob mice by stimulation of lipopolysaccharide,and to explore the correlation between obesity and osteogenic ability at molecular and cellular levels.METHODS:Eight8-week-old male ob/ob mice with hereditary obesity were used as experimental group,and eight8-week-old C57mice were used as control group.All the mice were provided by the Institute of Laboratory Animal Sciences,Chinese Academy of Medical Sciences&Peking Union Medical College in China.Bone marrow mesenchymal stem cells were isolated from bilateral femurs of all the mice using whole bone marrow culture method,cultured and purified.The cells were then cultured in the osteoinduction medium containing100μg/L lipopolysaccharide.Alkaline phosphatase staining,alizarin red staining and alkaline phosphatase activity detection were utilized to compare the osteogenic ability of bone marrow mesenchymal stem cells between the two groups.RT-PCR was used to detect the mRNA levels of Alp,Runx2,Ocn and Nf-κb.RESULTS AND CONCLUSION:(1)After1week of osteogenic induction,alkaline phosphatase staining and activity in the experimental group significantly weakened compared with the control group(P<0.01),and the addition of lipopolysaccharide further decreased the alkaline phosphatase activity(P<0.01),especially in the experimental group(P<0.01).(2)After2weeks of osteogenic induction,less mineralized nodes were observed in the experimental group than the control group.After the addition of lipopolysaccharide,no mineralized nodes formed in the two groups.(3)After osteogenic induction,the expression of Alp,Runx2,Ocn and Nf-
作者
刘大勇
周伟伟
肖蕊
王梅蕊
赵梦明
贾智
Liu Dayong;Zhou Weiwei;Xiao Rui;Wang Meirui;Zhao Mengming;Jia Zhi(Department of Dental Endodontics, Tianjin Medical University School & Hospital of Stomatology, Tianjin 300070, China;;Emergency Comprehensive Medical Center, Beijing Stomatological Hospital, Capital Medical University, Beijing 100069, China)
出处
《中国组织工程研究》
CAS
北大核心
2019年第5期703-709,共7页
Chinese Journal of Tissue Engineering Research
基金
国家自然科学基金(81670953)
项目负责人:刘大勇~~
