氯化锂对OVX小鼠骨丢失和JAK/STAT3信号通路的影响

Effect of lithium chloride on bone loss and JAK/STAT3 signaling pathway in OVX mice

目的研究氯化锂(lithium chloride,LiCl)干预对双侧卵巢切除(ovariectomized,OVX)小鼠骨丢失、Janus激酶/信号转导与转录激活子(The janus kinase/signal transducer and activator of tran-ions,JAK/STAT)信号通路及成骨细胞(osteoblast,OB)的影响。方法体内实验:选取8周龄雌性C57BL6/J小鼠24只,分为假手术组(Sham组),OVX组和卵巢切除+氯化锂干预(OVX+LiCl)组。收集动物左右侧股骨及右侧胫骨标本,进行Micro-CT、免疫组织化学和骨生物力学测试。体外实验:处死3日龄C57BL/6小鼠,分离颅骨培养原代OB,分为空白对照组(Control组)和LiCl组进行细胞活性检测、碱性磷酸酶染色、茜素红染色、实时荧光定量PCR检测。结果Micro-CT分析示OVX组骨量减少,骨小梁厚度变薄,骨小梁数量减少,骨小梁分离度增加,LiCl干预显著改善骨丢失。生物力学示OVX组的最大载荷、最大应力均显著低于Sham组(P<0.05),OVX+LiCl组的最大载荷、最大应力较OVX组显著升高(P<0.05)。免疫组化结果示,OVX+LiCl组JAK2和STAT3表达显著高于Sham组(P<0.05);OVX组P-STAT3表达明显低于Sham组(P<0.05),而OVX+LiCl组表达显著升高(P<0.05)。LiCl组的OB增殖活性显著高于对照组(P<0.05)。LiCl组的碱性磷酸酶活性和钙结节数量显著高于OVX组(P<0.05)。LiCl组STAT3 mRNA和OCN mRNA表达显著高于对照组(P<0.05)。结论LiCl通过调控OB改善OVX小鼠骨量和骨微结构,其机制可能与调节JAK/STAT3信号通路有关。

Objective To study the effects of lithium chloride(LiCl)preconditioning on bone loss,JAK/STAT signaling pathway,and osteoblast(OB)in bilaterally ovariectomized mice.Methods In vivo experimental:24 female 8-week-old C57BL6/J mice were randomly divided into sham operation group(Sham),ovariectomized operation group(OVX),and ovariectomized operation+Lithium chloride intervention(OVX+LiCl)group.The left and right femurs and the right tibia of the animals were collected and tested using micro-CT,immunohistochemistry,and bone biomechanics.In vitro experiment:3-day-old C57BL/6 mice were sacrificed.The calvarial bone was isolated to culture primary osteoblasts and randomly divided into blank group(Control)and LiCl group(LiCl).Alkaline phosphatase staining,Alizarin red staining,cell activity detection,and real-time quantitative PCR analysis were used.Results Micro-CT analysis result showed that the bone microstructure in the Sham group was generally normal.However,the microstructure in the OVX group changed abnormally,mainly including decreased bone mass,thinning of trabecular thickness,and decreased number of trabecular bone.The degree of separation of trabecular bone was increased.The intervention with LiCl significantly improved bone loss.Biomechanics result showed that the maximum load and maximum stress in mice of the OVX group were significantly lower than those of the Sham group(P<0.05),and the maximum load and maximum stress of the OVX+LiCl group were significantly higher than those of the OVX group(P<0.05).Immunohistochemical staining showed that the expressions of JAK2 and STAT3 in the OVX+LiCl group were significantly higher than those in the OVX group(P<0.05).The expression of P-STAT3 in the OVX group was significantly lower than that in the Sham group(P<0.05 vs.Sham),while the expression in OVX+LiCl group increased significantly(P<0.05 vs.OVX).The proliferation activity of osteoblasts in the LiCl group was significantly higher than that in the control group(P<0.05).The alkaline phosphatase activity and the number of calcium nodules in the LiCl group were significantly higher than those in the control group(P<0.05).The expressions of STAT3 mRNA and OCN mRNA in LiCl group were significantly higher than those in control group(P<0.05).Conclusion LiCl improves bone mass and bone microstructure of OVX mice by regulating OB.The mechanism may be related to the regulation of the JAK/STAT3 signaling pathway.