Sirt3在BMSCs成骨成脂双向命运分化调控的作用  

The regulatory role of Sirt3 in bidirectional fate determination of BMSCs during osteogenesis and adipogenesis

作　　者：彭力 辜诗淇 梁木春 高倩 张德茂 赵云[2] 杨静[1] PENG Li;GU Shi-Qi;LIANG Mu-Chun;GAO Qian;ZHANG De-Mao;ZHAO Yun;YANG Jing(State Key Laboratory of Oral Diseases&National Center for Stomatology&National Clinical Research Center for Oral Diseases&Dept.Of Cariology and Endontics West China Hospital of Stomatology,Sichuan University,Chengdu 610041,China;Key Laboratory of Bio-Resource and Eco-Environment of Ministry of Education,College of Life Sciences,Sichuan University,Chengdu 610065,China)

机构地区：[1]四川大学华西口腔医院口腔疾病国家重点实验室/国家口腔医学中心/国家口腔疾病临床研究中心/牙体牙髓科,成都610041 [2]四川大学生命科学学院生物资源与生态环境教育部重点实验室,成都610065

出　　处：《四川大学学报（自然科学版）》2024年第6期199-208,共10页Journal of Sichuan University(Natural Science Edition)

基　　金：国家自然科学基金(82470969,82001062);中国博士后科学基金(2022M722249)。

摘　　要：为探究Sirt3(Sirtuin 3)对骨形成过程中BMSCs成骨成脂双向分化的调控作用以及影响因素,本文通过组织学方式以及体外细胞培养的方式两种方式进行检测和验证.利用6个月的野生型小鼠和Sirt3基因敲除小鼠的长骨骨髓进行组织学切片染色,对骨髓中的成骨标志物Osterix进行免疫荧光染色,以及对成脂标志物Cebpα和Pparγ进行免疫荧光染色.同时观察Sirt3敲除对骨髓中炎症标志物IL-1β、IL-6、TNF-α的影响.同时对从小鼠骨髓提取出的BMSCs原代培养后,通过ELISA检测炎症因子IL-1β、IL-6、TNF-α的含量.成骨诱导后,通过碱性磷酸酶活性检测和茜素红染色,以及Western blot检测ALP、RUNX2、OCN的蛋白表达情况,探究Sirt3敲除对成骨分化的影响.而在成脂诱导BMSCs后,通过油红O染色和Western blot检测Cebpα和Pparγ的蛋白表达情况,研究Sirt3敲除对成脂分化的影响.以上结果表明敲除Sirt3基因会导致成年小鼠成骨降低成脂增加的不同方向分化的趋势,同时发现Sirt3敲除后炎症因子分泌显著上调,因此炎症可能作为Sirt3调控BMSCs成骨成脂双向命运分化的机制之一.To investigate the regulatory role of Sirt3 in the bidirectional differentiation of marrow mesenchy⁃mal stem cells(BMSCs)into osteogenic and adipogenic lineages during bone formation,we conducted histo⁃logical analyses and in vitro cell culture experiments.Histology sections of bone marrow from 6-month-old wild-type mice and Sirt3 knockout type mice were stained to assess the presence of osteogenic markers such as Osterix and adipogenic markers including CEBP-αand Pparγusing immunofluorescence.Additionally,the effects of Sirt3 knockout on inflammatory markers such as IL-1β,IL-6,and TNF-αin the bone marrow was observed.In parallel,primary BMSCs isolated from mouse bone marrow were cultured,and the levels of inflammatory cytokines IL-1β,IL-6,and TNF-αwere measured using ELISA.After osteogenic induc⁃tion,we evaluated the protein expressions of ALP,RUNX2,and OCN through alkaline phosphatase activity detection,alizarin red staining,and Western blot analysis to assess the impact of Sirt3 knockout on osteo⁃genic differentiation.Similarly,after inducing lipogenesis in BMSCs,we examined the protein expressions of CEBP-αand Pparγusing oil red O staining and Western blot analysis to study the effect of Sirt3 knockout on adipogenic differentiation.Our results indicate that the knockout of Sirt3 gene in adult mice leads to de⁃creased osteogenesis and increased adipogenesis in adult mice,and significantly upregulated the secretion of inflammatory factors.Therefore,inflammation may be one of the mechanisms through which Sirt3 regulates bidirectional differentiation of BMSCs during bone and fat formation.In summary,our findings shed light on the regulatory role of Sirt3 in the bidirectional differentiation of BMSCs and provide insights into its impact on bone formation.

关 键 词：Sirt3 成骨分化 成脂分化 炎症 骨髓间充质干细胞 

分 类 号：Q74[生物学—分子生物学]