摘要
目的观察促分裂原活化蛋白激酶2敲除(Mekk2-/-)小鼠的表型变化以及贞术调脂胶囊(FTZ)含药血清对MEKK2表达影响,探讨FTZ对MEKK2-β-catenin通路以及β-catenin去泛素化的作用机制。方法建立Mekk2-/-小鼠,通过MicroCT检测其与野生型的表型差别;通过FTZ干预Mekk2-/-小鼠和野生型的成骨细胞,运用Western blot观察对β-catenin磷酸化水平的作用;通过FTZ与FGF2干预野生型小鼠原代细胞,观察对MEKK2磷酸化水平的作用;通过C3H10T1/2细胞转染后进行FTZ和Wnt3a干预,观察β-catenin活性变化。结果Mekk2-/-小鼠与野生型小鼠相比,BV/TV、Tb.N、Tb.Th值均较后者降低(P<0.05),Tb.sp无差异变化(P>0.05)。FTZ含药血清增强了野生型小鼠细胞β-catenin的磷酸化水平(P<0.05),但是在Mekk2-/-小鼠细胞中则明显降低(P<0.05)。FTZ含药血清和FGF2干预后的原代细胞中,p-MEKK2/3表达水平均比不干预时细胞增强(P<0.05),且二者提高水平不存在明显差异(P>0.05)。FTZ含药血清和Wnt3a均促进β-catenin活化,而FTZ含药血清和Wnt3a共同干预细胞后,可见β-catenin活化程度更高,效果呈累加效应(P<0.05)。结论FTZ通过激活MEKK2通路,独立于Wnt经典通路促进β-catenin的去泛素化进程。
Objective To observe the phenotypic changes of mitogen-activated protein kinase 2 knockout(Mekk2-/-)mice and the effect of FTZ containing serum on the expression of MEKK2;To investigate the mechanism of FTZ on the MEKK2-β-catenin pathway and the deubiquitination ofβ-catenin.Methods Mekk2-/-mice were established by CRISPR/Cas9.The phenotypic differences between Mekk2-/-mouse and wildtype mouse were detected by MicroCT.The osteoblasts of Mekk2-/-mouse and wild-type mouseare treated by FTZ and the phosphorylation ofβ-catenin was detected by Western blot.The phosphorylation level of MEKK2 was observed in osteoblasts wildtype mousetreated by FTZ and FGF2;β-catenin activity was evaluated in transfection of C3H10T1/2 cells treated by FTZ and Wnt3a.Results The BV/TV,Tb.N,and Tb.Th values of Mekk2-/-mice were lower than those of the wildtype mouse(P<0.05),and there was no difference in Tb.sp(P>0.05).FTZ containing serum enhanced the phosphorylation ofβ-catenin in osteoblasts of wild type mice(P<0.05),but significantly decreased in Mekk2-/-mouse(P<0.05).In the primary osteoblasts after FTZ containing serum and FGF2 treatments,the expression of p-MEKK2/3 was higher than that of non-treatment group(P<0.05),and there was no significant difference between the two levels(P>0.05).Both FTZcontaining serum and Wnt3a promoted the activation ofβ-catenin activation.After FTZ-containing serum and Wnt3a treatment in the cells,the activation ofβ-catenin was higher and the co-treatment effect was additive(P<0.05).Conclusion FTZ promotes the deubiquitination ofβ-catenin by activating the MEKK2 pathway,independent of the Wnt classical pathway.
作者
孙平
李风英
杨国柱
韩晓蕊
陈镇秋
何伟
洪郭驹
郭姣
SUN Ping;LI Fengying;YANG Guozhu;HAN Xiaorui;CHEN Zhenqiu;HE Wei;HONG Guoju;GUO Jiao(Department of Endocrinology, the First Affiliated Hospital of Guangdong Pharmaceutical University, Guangzhou 510080, China;Department of Endocrinology, Qingdao Jimo People’s Hospital, Qingdao 266200, China;School of Life Sciences and Biopharmaceutics, Guangdong Pharmaceutical University, Guangzhou 510006, China;School of Medicine, South China University of Technology, Guangzhou 510006, China;Department of Orthopedics, the First Affiliated Hospital of Guangzhou University of Chinese Medicine, Guangzhou 510405, China;Lab of Orthopedics and Traumatology of Chinese Medicine of Linnan Medical Research Center of Guangzhou University of Chinese Medicine, Guangzhou 510405, China;Division of Orthopedics, Faculty of Medicine and Dentists, the University of Alberta, Edmonton T6G 2R3, Canada;Guangdong Pharmaceutical University, Guangzhou 510006, China)
出处
《中国骨质疏松杂志》
CAS
CSCD
北大核心
2020年第12期1726-1731,共6页
Chinese Journal of Osteoporosis
基金
国家自然科学基金项目青年科学基金(81603641)。
