论文部分内容阅读
目的探讨携带人转化生长因子β1(hTGF-β1)的腺病毒载体(adeno-hTGF-β1)转染人脂肪干细胞(ADSCs)及其表达的可行性。方法重组adeno-hTGF-β1经293细胞包装、扩增后,转染人ADSCs。转染72h后,RT-PCR检测重组腺病毒转染后ADSCs的hTGF-β1 mRNA表达,免疫细胞化学染色定性和酶联免疫吸附(ELASA)定量测定hTGF-β1蛋白的表达。结果转染后72h,ADSCs能有效表达hTGF-β1 mRNA,免疫细胞化学证实重组腺病毒转染后ADSCs胞浆内有大量棕黄色的hTGF-β1蛋白表达,ELISA结果提示其hTGF-βl表达量是转染Adeno-lacZ的3.85倍(<0.05)。结论重组腺病毒adeno-hTGF-β1能够被导入ADSCs并表达hTGF-β1蛋白,为hTGF-β1基因转染的ADSCs在软骨组织工程应用中奠定了基础。
Objective To investigate the feasibility of transfection of human adipose stem cells (ADSCs) with adeno-hTGF-β1 carrying human transforming growth factor-β1 (hTGF-β1) and its expression. Methods Recombinant adeno-hTGF-β1 was packaged in 293 cells and amplified. The recombinant adeno-hTGF-β1 was transfected into human ADSCs. The expression of hTGF-β1 mRNA in ADSCs was detected by RT-PCR 72 h after transfection. The expression of hTGF-β1 protein was quantified by immunocytochemical staining and enzyme linked immunosorbent assay (ELISA). Results ADSCs could effectively express hTGF-β1 mRNA at 72h after transfection. Immunocytochemistry confirmed that a large amount of hTGF-β1 protein was expressed in cytoplasm of ADSCs after transfection. The results of ELISA showed that the expression of hTGF-βl was 3.85-fold (<0.05) transfection with Adeno-lacZ. Conclusion The recombinant adenovirus adeno-hTGF-β1 can be introduced into ADSCs and express hTGF-β1 protein. The ADSCs transfected with hTGF-β1 gene have laid the foundation for the application of cartilage tissue engineering.