摘要
目的探讨骨疏方调节FoxO3a/Wnt信号通路对过氧化氢(H_(2)O_(2))诱导成骨细胞损伤的影响。方法CCK-8法检测不同浓度(0.01、0.1、1、10、100μmol/L)的骨疏方对H_(2)O_(2)诱导的MC3T3-E1细胞活力。随后设置对照组、H_(2)O_(2)(150μmol/L H_(2)O_(2))组、骨疏方组、骨疏方+空载体组、骨疏方+FoxO3a过表达组。CCK-8法、ELISA分别检测细胞活力、超氧化物歧化酶(SOD)、丙二醛(MDA);成骨诱导分化后,碱性磷酸酶(ALP)试剂盒及茜素红分析ALP活性、细胞骨矿化结节;qRT-PCR检测成骨相关基因OPN mRNA、Runx2 mRNA以及FoxO3a mRNA表达水平;Western blot检测通路相关蛋白FoxO3a、Wnt2、β-连环蛋白(β-catenin)表达水平。结果与对照组比较,H_(2)O_(2)组MDA、FoxO3a表达显著增加,细胞活力、SOD、ALP活性、骨矿化结节数量、OPN mRNA、Runx2 mRNA、Wnt2、β-catenin表达显著降低(P<0.05);与H_(2)O_(2)组比较,骨疏方组MDA、FoxO3a表达显著降低,细胞活力、SOD、ALP活性、骨矿化结节数量、OPN mRNA、Runx2 mRNA、Wnt2、β-catenin表达显著增加(P<0.05);与骨疏方+空载体组比较,骨疏方+FoxO3a过表达组MDA、FoxO3a表达显著增加,细胞活力、SOD、ALP活性、骨矿化结节数量、OPN mRNA、Runx2 mRNA、Wnt2、β-catenin表达显著降低(P<0.05)。结论骨疏方可改善H_(2)O_(2)诱导成骨细胞损伤,可能与抑制FoxO3a/Wnt信号通路有关。
Objective To investigate the impact of Gushu formula on hydrogen peroxide(H_(2)O_(2))induced osteoblast injury by regulating the FoxO3a/Wnt signaling pathway.Methods CCK-8 method was applied to detect the effects of different concentrations(0.01,0.1,1,10,100μmol/L)of Gushu formula on H_(2)O_(2)induced MC3T3-E1 cell viability.Subsequently,a control group,H_(2)O_(2)(150μmol/L H_(2)O_(2))group,Gushu formula group,Gushuformula+empty body group,and Gushu formula+FoxO3a overexpression group were set up.CCK-8 method and ELISA were applied to detect cell viability,superoxide dismutase(SOD),and malondialdehyde(MDA),respectively;after osteogenic induction and differentiation,alkaline phosphatase(ALP)assay kit and alizarin red were applied to analyze ALP activity and cellular bone mineralization nodules;qRT-PCR was applied to detect the expression levels of osteogenic related genes OPN mRNA;Runx2 mRNA,and FoxO3a mRNA;Western blot was applied to detect the expression levels of pathway related proteins FoxO3a;Wnt2;andβ-catenin.Results Compared with the control group,the expression of MDA and FoxO3a in the H_(2)O_(2)group was greatly increased,the cell viability,SOD and ALP activities,number of bone mineralization nodules,the expression of OPN mRNA,Runx2 mRNA,Wnt2,β-catenin were greatly reduced(P<0.05);compared with the H_(2)O_(2)group,the expression of MDA and FoxO3a in the Gushu formula group was obviously reduced,the cell viability,SOD and ALP activities,number of bone mineralization nodules,the expression of OPN mRNA,Runx2 mRNA,Wnt2,β-catenin were obviously increased(P<0.05);compared with the Gushuformula+empty-load group,the expression of MDA and FoxO3a in the Gushu formula+FoxO3a overexpression group was obviously increased,the cell viability,SOD and ALP activities,number of bone mineralization nodules,the expression of OPN mRNA,Runx2 mRNA,Wnt2,β-catenin were greatly reduced(P<0.05).Conclusion Gushu formula can improve H_(2)O_(2)induced osteoblast damage,which may be related to the inhibition of FoxO3a/Wnt signaling pathway.
作者
段波
陈丽川
马志毅
喻昭
刘静
王进军
DUAN Bo;CHEN Lichuan;MA Zhiyi;YU Zhao;LIU Jing;WANG Jinjun(Department of Rheumatology,Wuhan Hospital of Traditional Chinese Medicine,Wuhan 430050;Department of Acupuncture,Wuhan Hospital of Traditional Chinese Medicine,Wuhan430050,China)
出处
《中国骨质疏松杂志》
CAS
CSCD
北大核心
2024年第10期1426-1431,共6页
Chinese Journal of Osteoporosis
基金
武汉市医学科研项目(WZ19C17)。
