HGF基因转染关节软骨细胞及其生物学特性影响

The biological influences of HGF cDNA labeled with GFP to chondrocytes of the rabbit condylar transfected by HGF cDNA

目的观察携带有绿色荧光蛋白(gree fluorescent prote in,GFP)标记的肝细胞生长因子(hepatocyte growth factor,HGF)cDNA转染兔软骨细胞后的主要生物学特性。方法取兔膝关节软骨细胞体外培养至第二代,HGFcDNA转染软骨细胞,空白质粒组为对照组;噻唑蓝(MTT)检测软骨细胞的增殖能力,SABC免疫组化及原位杂交法测其II型胶原在转染前后的变化与mRNA的表达。结果软骨细胞在转染后可较长时间的保持其软骨细胞的生物学特性,并可促进软骨细胞的增殖,SABC免疫组化测第VI代软骨细胞的II型胶原表达仍呈强阳性,而对照组软骨细胞则明显减弱,说明HGF基因转染软骨细胞后可表达其生物学功能。基因瞬间转染率大约为31.23(。结论GFP标记的HGF基因以脂质体为载体转染兔膝关节软骨细胞后可发挥其生物学功能,加速细胞的生长,在荧光显微镜下可直观的计算软骨细胞的瞬间转染率。

Objective： To study the main biological chacteristics of chondroeytes after gene-transfected by HGF eDNA labled with GFP. Methods： The chondroeytes of the rabbit condylar were derived from rabbit, and were cultured to the second generation in vitro,then HGFcDNA transfected the cells. The capacity of cell proliferation was tested by MTI＇,The deferences of the type II collagen were tested by SABC immunohistoehemical of the type Ⅱ collagen after infected;the mRNA levels were tested nd so on the mRNA levels were tested by in situ hybridization of the type II collagen between the transfection group and comparison group . The chondrocytes transfected was counted ten times under fluoreseen inversion microscope, made the average. Results： The condylar chondroeytes transfected by HGF could keep the main biological chacteristics of chondroeytes when it was the sixth generation,then comparison group couldnt ; HGFcould accelerate the speed of cell proliferation, improved cell proliferation. The expression of collagen Ⅱ is stronger than the comparison group on the seventh generation. The percent of the chondrocytes transfected was 31.23%. Conclusion： The chondroeytes transfected by HGF eDNA Labeled with GFP through liposome as vector was successful,The HGF could express its biological chacteristics, then it could accelerate the cell＇s proliferation, and it could be indirectly accounted that how many chondroevtes were transfected.