XCT-790导致成骨细胞活性下降和骨形成不足的机制  

Mechanism of XCT-790 reducing activity and osteogenesis of osteoblasts

作　　者：刘少津 万雷 乔荣勤 黄宏兴 王吉利[1] 李钊政 Liu Shaojin;Wan Lei;Qiao Rongqin;Huang Hongxing;Wang Jili;Li Zhaozheng(Guangzhou University of Chinese Medicine,Guangzhou 510405,Guangdong Province,China;the Third Affiliated Hospital of Guangzhou University of Chinese Medicine,Guangzhou 510240,Guangdong Province,China)

机构地区：[1]广州中医药大学,广东省广州市510405 [2]广州中医药大学第三附属医院,广东省广州市510240

出　　处：《中国组织工程研究》2019年第3期329-334,共6页Chinese Journal of Tissue Engineering Research

基　　金：国家自然科学基金项目(81302991;81673786)项目负责人:万雷;广东省自然科学基金项目(2014A030310127);广东省科技计划项目(2016A020216024);2014广东省"优青计划"项目(yq2014041);项目负责人:万雷~~

摘　　要：背景:Wnt信号通路在骨细胞的形成、分化和成熟过程中起着重要作用,DKK1、Sost在调节骨量和成骨细胞分化中起负调节作用,是Wnt信号通路的负调节因子。XCT-790是雌激素相关受体α的特异性抑制剂,可调节雌激素信号通路对成骨细胞的功能活动。目的:观察XCT-790对Wnt信号通路抑制因子DKK1、Sost过表达腺病毒载体转染的人成骨肉瘤细胞MG63细胞及其骨形成相关蛋白LRP5、骨形态发生蛋白2、骨桥蛋白、骨保护蛋白、结缔组织生长因子的作用。方法:将MG63细胞分成8组:空白对照组、过表达DKK1组、过表达Sost组、过表达DKK1+Sost组、XCT-790干预空载腺病毒组、XCT-790干预过表达DKK1组、XCT-790干预过表达Sost组、XCT-790干预过表达DKK1+Sost组,根据组别不同分别用上述重组腺病毒转染MG63细胞。应用MTT法检测MG63细胞的活性,碱性磷酸酶试剂盒检测碱性磷酸酶活性,流式分析钙离子浓度;Western blot检测低密度脂蛋白相关蛋白5、骨形态发生蛋白2、骨桥蛋白、骨保护蛋白、结缔组织生长因子的表达。结果与结论:①与空白对照组对比,过表达DKK1、Sost、DKK1+Sost组及XCT-790干预空载腺病毒组均可降低细胞活性、碱性磷酸酶活性,提高钙离子浓度;降低低密度脂蛋白相关蛋白5、骨形态发生蛋白2、骨桥蛋白、骨保护蛋白、结缔组织生长因子的表达量,组间比较差异均有显著性意义(P <0.05);②而当XCT-790干预过表达DKK1、Sost、DKK1+Sost组的MG63细胞时,可进一步降低细胞活性、碱性磷酸酶活性,提高钙离子浓度;降低低密度脂蛋白相关蛋白5、骨形态发生蛋白2、骨桥蛋白、骨保护蛋白、结缔组织生长因子的表达量,组间比较差异均有显著性意义(P <0.05);③结果说明,XCT-790直接调节Wnt信号通路的传导,同时也可协同DKK1、Sost共同抑制Wnt信号通路的开放,从而降低成骨细胞活性和骨形成相关蛋白LRP5、骨形态发生BACKGROUND:Wnt signaling pathway plays an important role in formation,differentiation and mature of osteocytes.As negative regulators of Wnt signaling pathway,DKK1 and Sost play negative roles in regulating bone mass and osteoblast differentiation.XCT-790,an inhibitor of estrogen receptor-related receptorα,can regulate the effect of estrogen signaling pathway on osteoclast function.OBJECTIVE:To observe the effects of XCT-790 on the proliferation of MG63 cells transfected with DKK 1 and Sost overexpression adenovirus vector and the expression of low-density lipoprotein receptor-related protein(LRP)5,bone morphologic protein 2,osteopontin,osteoprotegerin,and connective tissue growth factor(CTGF)protein.METHODS:MG63 cells were divided into eight groups:blank control,overexpressed DKK1(ad-DKK1),overexpressed Sost(ad-Sost),overexpressed DKK1+Sost(ad-DKK1+Sost),XCT-790-intervened blank adenovirus,XCT-790-intervened ad-Sost,XCT-790-intervened ad-DKK1-Sost groups.The cell viability was detected by MTT assay.The alkaline phosphatase activity was detected by alkaline phosphatase kit.Ca2+concentration was detected by flow cytometer.Expression of LRP5,bone morphologic protein 2,osteopontin,osteoprotegerin and CTGF was detected by western blot assay.RESULTS AND CONCLUSION:Compared with the blank control group,ad-DKK1,ad-Sost,ad-DKK1+Sost and XCT-790-intervened blank adenovirus groups could significantly decrease the cell activity,alkaline phosphatase activity,increase calcium concentration and down-regulate the expression of LRP5,bone morphologic protein 2,osteopontin,osteoprotegerin and CTGF(P<0.05).In the XCT-790-intervented ad-DKK1,ad-Sost,ad-DKK1+Sost groups,the cell activity,alkaline phosphatase activity,and the expression of LRP5,bone morphologic protein 2,osteopontin,osteoprotegerin and CTGF were significantly reduced,and the Ca2+concentration was significantly increased(P<0.05).In summary,XCT-790 can directly regulate Wnt signaling pathway,and can cooperate with DKK1 and Sost to inhibit Wnt signaling pathway,thereby

关 键 词：XCT-790 DKK1 SOST MG63细胞 雌激素相关受体α 组织构建 雌激素受体Α 腺病毒科 转染 组织工程 

分 类 号：R446[医药卫生—诊断学]