去势大鼠骨量丢失过程中骨髓间充质干细胞的增殖分化功能  

Proliferation and differentiation of bone marrow mesenchymal stem cells in process of bone loss in ovariectomized rats

作　　者：梁世桢[1] 王国轩[2] 范龙坤[2] 吴高义[3] 金岩[4] 朱国雄[3] 

机构地区：[1]辽宁医学院济南军区总医院研究生培养基地口腔科,山东济南250031 [2]辽宁医学院研究生学院,辽宁锦州121001 [3]济南军区总医院口腔科,山东济南250031 [4]第四军医大学口腔医学院,陕西西安710032

出　　处：《中国病理生理杂志》2012年第3期398-403,共6页Chinese Journal of Pathophysiology

基　　金：国家自然科学基金资助项目(No.2011CB964700)

摘　　要：目的:研究骨髓间充质干细胞(BMSCs)增殖分化功能在去势大鼠骨质疏松发病过程中对骨量丢失的作用。方法:选用10周龄健康雌性SD大鼠行双侧卵巢切除术(OVX),建立骨质疏松症(OP)的动物模型;选用同一批次、周龄相同、体重相近的健康雌性SD大鼠行双侧卵巢附近脂肪组织部分切除术,建立假手术(sham)组,假手术大鼠组(sham group)。采用离心法、贴壁法和有限稀释法分离、培养、纯化大鼠BMSCs,体外培养传至3～4代后用于实验:流式细胞术进行BMSCs表型鉴定;克隆形成实验检测BMSCs增殖状况;MTT法测定BMSCs生长曲线;成脂诱导后脂滴油红O染色法检测比较2组大鼠BMSCs成脂能力;成骨诱导后钙化结节茜素红染色法检测比较2组大鼠BMSCs成骨能力;RT-PCR法检测大鼠BMSCs成骨相关蛋白Runx2、骨钙素(OCN)和骨桥蛋白(OPN)mRNA的表达。结果:与sham组大鼠BMSCs相比,OVX组大鼠BMSCs克隆形成能力减弱,增殖能力降低,成脂向分化增强,成骨向分化减弱(P<0.05)。结论:去卵巢骨质疏松大鼠BMSCs增殖及成骨分化减弱,成脂分化增强;这导致去势大鼠快速的骨量丢失,在去势大鼠OP发病过程中发挥着重要作用。AIM：To study the function of proliferation and differentiation of bone marrow mesenchymal stem cells（BMSCs） for bone loss in the pathogenesis of osteoporosis（OP） in ovariectomized rats.METHODS：Animal model of OP was established by ovariectomy（OVX,bilateral ovarian resection） in 10-week-old healthy female Sprague-Dawley （SD） rats.BMSCs were isolated,cultured and purified by the combination of density gradient centrifugation,adhesion separation and limited dilution method,and cultured in vitro to the 3rd～4th passage in all experiments.The BMSCs phenotype appraisal was studied by flow cytometry.Colony-forming assay was applied to detect the BMSCs proliferation ability. The MTT method was used to analyze the growth curves of BMSCs.After adipogenic induction（ADI）,lipid drops were observed by oil red 0 staining to compare the adipogenic potential between the 2 kinds of BMSCs.After osteogenic induction （OSI）,calcium nodules were observed by alizarin red staining（ARS）.The mRNA expression levels of BMSCs osteogenesis-related proteins,for instance,Runx2,osteocalcin（OCN） and osteopontin（OPN ） were measured by RT-PCR. RESULTS：Compared with sham group,the colony-forming ability of BMSCs in OVX group became decreased,the proliferation capacity was declined,the osteogenic potential was decreased,and the adipogenic potential was increased （P 0.05）.CONCLUSION：In ovariectomized OP rats,the proliferation and osteogenesis of BMSCs decrease,and the adipogenesis of BMSCs increases,which may cause rapid bone loss and play an important role in the pathogenesis of OP.

关 键 词：骨髓间充质干细胞 卵巢切除术 大鼠 骨生成 脂肪形成 

分 类 号：R361.3[医药卫生—病理学]