四甲基吡嗪通过调节LepR+BMSCs成骨分化治疗失重性骨丢失的初步研究  

A Preliminary Study of Tetramethylpyrazine in the Treatment of Mechanical Unloading-induced Bone Loss by Modulating the Osteogenic Differentiation of LepR+BMSCs

作　　者：宋世菊 范静 殷金花 郑超 杨柳 Song Shiju;Fan Jing;Yin Jinhua;Zheng Chao;Yang Liu(Department of Orthopaedic,Xijing Hospital,Air Force Medical University,Xi'an 710032,China)

机构地区：[1]空军军医大学附属西京医院骨科,西安710032

出　　处：《骨科》2024年第6期541-548,共8页ORTHOPAEDICS

基　　金：国家自然科学基金重点项目(82130070);国家自然科学基金面上项目(82372361)。

摘　　要：目的明确LepR+骨髓间充质干细胞(bone marrow mesenchymal stem cells,BMSCs)在失重性骨丢失发生中的作用并探究四甲基吡嗪(tetramethylpyrazine,TMP)对失重性骨丢失的治疗效果。方法本研究构建特异性示踪LepR+BMSCs的小鼠(LepR-Cre;R26tdTomato),取P7小鼠鼠尾进行基因鉴定,将3月龄LepR-Cre;R26tdTomato雄鼠分为四组:对照组、后肢去负荷(hindlimb unloading,HU)组、HU腹腔注射玉米油组、HU腹腔注射TMP药物组。显微CT分析对照组、HU组小鼠股骨骨小梁,以评估失重对骨量的影响;取两组股骨组织进行冰冻切片,行免疫荧光染色检测成骨细胞分子标志物Osterix(Osx)表达情况,统计骨软骨交界及骨小梁处LepR+BMSCs来源细胞中成骨细胞占比,分析失重对LepR+BMSCs成骨分化的影响。另外,取HU腹腔注射玉米油组、HU腹腔注射TMP药物组小鼠股骨组织行冰冻切片,通过HE染色检测TMP药物对骨量影响;在Osx染色后统计骨软骨交界及骨小梁处LepR+BMSCs来源细胞中成骨细胞占比,对比分析TMP对LepR+BMSCs成骨分化的影响。结果HU小鼠出现骨质疏松表型并且LepR+BMSCs来源的成骨细胞数目显著减少,给予HU小鼠TMP药物处理可以增加骨量,同时增加LepR+BMSCs来源的成骨细胞数目。结论LepR+BMSCs参与失重性骨丢失的发生,TMP通过促进LepR+BMSCs向成骨分化有助于治疗失重性骨丢失。Objective To determine the role of bone marrow mesenchymal stem cells(BMSCs)expressing leptin receptor(LepR)in bone loss under unloading conditions,and to explore the therapeutic effect of Tetramethylpyrazine(TMP)on unloading induced bone loss.Methods In this study,we constructed LepRCre;R26tdTomato mice to specifically trace LepR+BMSCs.DNA was extracted from mice's tails one week after birth and genotyping was performed based on the extracted DNA.The three-month-old LepR-Cre;R26tdTomato mice were selected and divided into four groups for experiments,including control group,hindlimb unloading(HU)group,HU with intraperitoneal injection of corn oil group,and HU with intraperitoneal injection of TMP drug group.After the experiment,mice's femurs of the control group and HU group were harvested to analyze the trabecular bone of the distal femur to evaluate the effect of HU on the bone mass through micro-CT analysis.To investigate the effect of unloading on the osteogenic differentiation ability of LepR+BMSCs,the femurs in the control group and HU group were cryo-sectioned to detect the expression of Osterix(Osx)by immunofluorescent staining.The proportion of osteoblasts in LepR+BMSCs-derived cells at the osteochondral junction and trabecular bone was calculated.In addition,the cryosections of mice femoral tissue in HU intraperitoneally injected with corn oil and TMP drug groups were used to detect the influence of TMP on bone mass by HE staining,and the percentage of osteoblasts in LepR+BMSCs-derived cells at the osteochondral junction and trabecular bone was analyzed after Osx staining.Thus,the effect of TMP on the osteogenic differentiation ability of LepR+BMSCs was detected by comparative analysis.Results HU mice showed an osteoporotic phenotype and the number of osteoblasts derived from LepR+BMSCs was significantly reduced.TMP treatment with HU mice could increase bone mass and the number of osteoblasts derived from LepR+BMSCs.Conclusion LepR+BMSCs were involved in the occurrence of unloading-induced bone loss,and

关 键 词：四甲基吡嗪 LepR+BMSCs 失重性骨丢失 后肢去负荷 谱系示踪 

分 类 号：R68[医药卫生—骨科学]