摘要
目的探讨重组大鼠转化生长因子β1基因(pcDNA3+TGFβ1)单独转染及其与重组大鼠胰岛素样生长因子1基因(pAT153+IGF1)共转染兔软骨细胞后细胞增殖及所分泌的TGFβ1因子、IGF1因子、Ⅱ型胶原的变化。方法兔软骨细胞体外分别用pcDNA3+TGFβ1单转染、pcDNA3+TGFβ1和pAT153+IGF1共转染,筛选阳性克隆后,进行原位杂交、免疫组织化学、免疫荧光检测、流式细胞仪检测、3HTdR(3H标记的胸腺嘧啶脱氧核苷)检测。结果基因转染组与空白组相比,TGFβ1、IGF1、Ⅱ型胶原的含量均明显提高,空白组、基因单转染组、基因双转染组软骨细胞位于S期的比例分别为5.6%、33.4%、40.1%,差异有统计学意义(P<0.05)。结论基因pcDNA3+TGFβ1、pAT153+IGF1转染软骨细胞后,细胞分泌的TGFβ1、IGF1及Ⅱ型胶原显著增多,细胞增殖明显增强,上述基因转染有助于软骨细胞活力的提高;pcDNA3+TGFβ1和pAT153+IGF1双基因共转染软骨细胞后,细胞分裂增生活跃程度及分泌的TGFβ1、IGF1和Ⅱ型胶原含量高于pcDNA3+TGFβ1单基因转染,多基因共转染作为将来骨性关节炎基因治疗的方法,其治疗效果可能会优于单基因转染。
Objective To investigate the effects of transfeetion of the pcDNA3 ＋ TGF-β1 with or without pAT153 ＋ IGF-1 on rabbit chondrocytes proliferating and synthesizing TGF-betal, IGF-1 and collagen Ⅱ . Methods Monolayer culture of rabbit articular chondrocytes was infected with recombinant rat gene pcDNA3 ＋ TGF-betal, and co-transfected by pcDNA3 ＋ TGF-betal, pAT153 ＋ IGF-1. The synthesis of TGF-betal, IGF-1, and type Ⅱ collagen was examined by in situ hybridization, immunohistochemistry, immunofluoroscopy, flow cytemeter and 3 H-TdR radiolabeling. Results The expression of TGF-beta 1, IGF-1 and type Ⅱ collagen was increased in transfection groups （P 〈 0.05 ）. The eo-transfection could elevate TGF-beta 1, IGF-1 and type Ⅱ collagen synthesis beyond the levels of recombinant pcDNA3 ＋ TGF-beta 1 （P〈0.05）. The chondrocytes ratio at stage S in different groups was respectively 5.6%, 33.4 % and 40.1% respectively. Conclusion Transfer of TGF-betal and IGF-1 genes to articular chondrocytes can greatly increases proliferation and synthesis of chondrocytes ex vivo. The co-transfeetion can provoke a more increase in the synthesis of TGF-beta 1, IGF-1 and type Ⅱ collagen than pcDNA3 ＋ TGF-beta 1 transfection, so it may be more useful to osteoarthritis therapy. The results encourage the further development of gene therapy for osteoarthritis.
出处
《中华实验外科杂志》
CAS
CSCD
北大核心
2005年第10期1243-1245,共3页
Chinese Journal of Experimental Surgery
基金
国家自然科学基金资助项目(30371440)
山西省自然科学基金资助项目(20041117)
