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目的:探讨脂肪间充质干细胞作为软骨组织工程的种子细胞和基因增强的组织工程的可行性。方法:取3周龄Lewis大鼠的腹股沟脂肪垫,消化法获得脂肪间充质干细胞,通过脂质体介导,将hTGFβ2基因转染到体外培养的脂肪间充质干细胞中,采用免疫组织化学染色、RT-PCR和Western blotting的方法检测目的基因与软骨特异性蛋白——Ⅱ型胶原和蛋白多糖表达的情况。结果:从成体大鼠脂肪组织中培养出脂肪间充质干细胞,体外培养呈成纤维细胞样,能大量稳定增殖传代。原代脂肪间充质干细胞能自发分化为脂肪细胞,传代细胞在胰岛素和地塞米松的作用下生成脂滴向脂肪细胞分化;hTGFβ2基因转入脂肪间充质干细胞能获得瞬时及稳定表达,并促使Ⅱ型胶原和蛋白多糖合成。结论:从脂肪组织中可获得具有多分化潜能的间充质干细胞,并能在体外稳定增殖传代,可自发或经诱导后分化为脂肪细胞;pcDNA3.1(+)/hTGFβ2真核表达载体成功转染脂肪间充质干细胞,诱导其向软骨细胞分化。
OBJECTIVE: To investigate the feasibility of adipose tissue-derived mesenchymal stem cells as tissue engineering engineered cartilage cells for enhanced tissue engineering. METHODS: Adipose-derived mesenchymal stem cells were obtained from the inguinal fat pads of 3-week-old Lewis rats. The hTGFβ2 gene was transfected into adipose-derived mesenchymal stem cells by liposome-mediated transfection in vitro. Immunohistochemistry Staining, RT-PCR and Western blotting were used to detect the expression of target gene and cartilage-specific type II collagen and proteoglycan. Results: Adipose-derived mesenchymal stem cells were cultured from adult rat adipose tissue and cultured fibroblast-like in vitro. Primary adipose-derived mesenchymal stem cells can differentiate into adipocytes spontaneously, and subcultured cells produce lipid droplets differentiated into adipocytes under the action of insulin and dexamethasone. Transient hTGFβ2 gene transfection into adipose-derived mesenchymal stem cells can achieve transient and stable expression Promote type II collagen and proteoglycan synthesis. CONCLUSION: Mesenchymal stem cells with multidifferentiation potential can be obtained from adipose tissue and can stably proliferate and passage in vitro, which can differentiate into adipocytes either spontaneously or after induction. The eukaryotic expression vector pcDNA3.1 (+) / hTGFβ2 is successful Transfection of adipose-derived mesenchymal stem cells induces their differentiation into chondrocytes.