华西口腔医学杂志

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Ⅰ型骨质疏松症大鼠骨髓基质细胞体外培养及分化研究

唐尤超,汤炜,田卫东,陈希哲,李声伟   

  1. 口腔生物医学工程教育部重点实验室,四川大学,四川 成都610041
  • 收稿日期:2005-04-25 修回日期:2005-04-25 出版日期:2005-04-20 发布日期:2005-04-20
  • 通讯作者: 汤 炜,Tel:028-85503406
  • 作者简介:唐尤超(1971-),男,江西人,主治医师,博士
  • 基金资助:

    四川省应用基础研究资助项目(02SY029-116)

Cultivation and Induced Differentiation of Bone Marrow Stromal Cells of SD Rats with TypeⅠOsteoporosisin vitro

TANG You-chao, TANG Wei, TIANWei-dong, CHENXi-zhe, LISheng-wei   

  1. KeyLab. ofOralBiomedicalEngineering ofMin- istry ofEducation, Sichuan University, Chengdu610041, China
  • Received:2005-04-25 Revised:2005-04-25 Online:2005-04-20 Published:2005-04-20

摘要: 目的 研究在构建的Ⅰ型骨质疏松症大鼠动物模型中,骨髓基质细胞( BMSCs)的生物学特性及其成骨和 成脂分化能力。方法 16只6月龄雌性SD大鼠随机分为实验组和对照组,实验组行双侧卵巢切除建立Ⅰ型骨质 疏松动物模型,对照组切除卵巢周围少量脂肪组织。术后3个月处死动物,非连续密度梯度离心分离骨质疏松大 鼠的BMSCs,传代后分别在成骨及成脂培养基中诱导培养14 d,观察细胞形态,通过组织特染对BMSCs的成骨、成脂 表型进行鉴定,计数阳性染色细胞百分比。结果 构建的Ⅰ型骨质疏松大鼠模型大鼠体重及整体股骨骨矿物质密 度(BMD)与对照组比较差异有显著性(P<0.01 );诱导培养后两组大鼠BMSCs碱性磷酸酶(ALP )、Von Kossa′s染色 和油红O染色实验阳性,且两组细胞阳性染色率无明显差别(P>0.05)。结论 本实验成功建立了大鼠骨质疏松 症模型,骨质疏松症大鼠BMSCs保持分化能力。

关键词: 骨质疏松, 骨髓基质细胞, 成体干细胞

Abstract:

Objective To study the biological features and osteoblast/adipocyte phenotypes of bone marrow stromal cells (BMSCs) of Sprague-Dawley (SD) rats withTypeⅠosteoporosis by induced culture.Methods Six-month-old SD rats were used in this study.16 female rats were randomly divided into two groups. eight rats were ovariectomied as experimental group to estab- lish the modle ofTypeⅠosteoporosis, while other rats received sham-operation. Three month later BMSCs of 16 rats were isolated by discontinueous gradient centrifugation and then plated inα-MEM medium as primary culture. Secondary harvested cells were cultured for 14 days inα-MEMmedium supplemented with dexamethasone, ascorbic acid, vitamin D3,β-glycerophosphate or dex- amethasone, 3-isobutyl-1-methylxanthine, insuline, and indomethine. The cells were screened by inverted microscope each day and cell growth was studied with cell counting. The osteoblast and adipocyte phenotypes were verified by cytochemistry staining, counted the percentage of positive stained cells.Results The weight and bone mineral density of rats were statistically different between experimental group and control group. Gomori and Von Kossa′s staining demonstrated positive osteoblast phenotypes of al- kaline phosphatase and mineralized nods by osteogenic inducer, while Oil Red O staining identified BMSCs treatedwith adipogenic medium resulted in adipocyte formation and there was no significant difference in the percentage of positive stained cells between two groups.Conclusion The model of TypeⅠosteoporosis has been established successfully. BMSCs from SD rats with osteo- porosis maintain their differentiation potential.

Key words: osteoporosis, marrow stromal cell, adult stem cell