胰岛素样生长因子1基因转染对大鼠肌腱细胞生长的促进作用

Insulin-like growth factor 1 mediated gene transfer to rat tendon cells to promote the growth of the tendon cells

目的 探讨外源性胰岛素样生长因子 1(IGF 1)基因转染对离体培养的肌腱细胞分裂增殖的促进作用。方法 用逆转录多聚酶链反应 (RT PCR)和酶联免疫吸附试验 (ELISA )检测转染 48h后细胞IGF 1mRNA和活性蛋白的表达 ;3 H TdR掺入法检测转染后 2 4、48、72h肌腱细胞DNA的合成情况 ;并用激光共聚焦显微镜对转染 48h后细胞Ⅰ型胶原的免疫荧光染色行定量分析。结果 IGF 1mRNA和活性蛋白在转染细胞内正确表达 ;实验组每分钟闪烁值 (CPM )为2 3 5 5 .75± 5 41.98,而对照组CPM值为 114 9.0 0± 485 .3 0 ,两者差异有显著性 (P <0 .0 5 ) ;转染组Ⅰ型胶原荧光强度较对照组明显增强 (转染组 76.2 0± 3 2 .2 3 ,对照组 3 8.84± 11.10 ,P <0 .0 1)。结论 外源性IGF 1基因转染能够促进肌腱细胞的分裂增殖和Ⅰ型胶原的分泌。

Objective To explore the influence of extrinsic insulin-like growth factor 1 gene on the growth of tendon cells by introducing it into primary rat tendon cells.Methods Eukaryotic expression vector of pcDNA3.1(+)-IGF-1 was transfered into tendon cells mediated by lipopolyamine DOGS.48 h after transfection of IGF-1,the mRNA expression of IGF-1 was identified by RT-PCR and the active protein was determined by ELISA.Collagen type Ⅰsecretion of the tendon cells was detected by immunofluorescence assay.24 h,48 h,72 h after transfection respectively, 3H-TdR method was used to identify the DNA synthesis of the tendon cells.Results IGF-1mRNA and active protein expression were detected by RT-PCR and ELISA respectively.The CPM of the experimental group was 2?355.75± 541.98, significantly higher than that of the control group (1?149.00±485.30, P <0.05).The fluorescence intensity of collagen typeⅠof transfected cells was stronger than that of the controls (transfection group:76.20±32.23;control group:38.84±11.10, P <0.01).Conclusion IGF-1cDNA could be successfully transferred into primary rat tendon cells and stimulate DNA synthesis and collagen type Ⅰsecretion.