基因转染3T3细胞的hTGFβ_1表达

The Expression of Eukaryotic Expression Vector for Human Transforming Growth Factor β_1 in NIH3T3

目的鉴定重组pcDNA3.1-TGFβ1 载体在真核细胞中的表达 ,为其在软骨组织工程中的基因转染创造条件。 方法利用脂质体Fugene 6将TGFβ1 真核表达载体转染至NIH3T3细胞 ,RT -PCR、ELISA检测hTGFβ1 基因的表达 ,并利用CCL 6 4细胞株鉴定重组hTGFβ1 的生物学活性。 结果质粒转染 3T3细胞后 ,hTGFβ1 mRNA及其蛋白的表达均显著增强 ,且分泌表达的hTGFβ1 能明显抑制CCL 6 4细胞的生长。 结论重组载体在 3T3细胞中能表达具有一定生物学活性的hTGFβ1 。

Objective To determine the expression of transforming growth factor β 1 in eukaryotic 3T3 cells so as to analyze its biologic function and to study the possibility of gene transfection into chondrocytes. Methods The eukaryotic expression vector for TGFβ 1 (pcDNA3.1-TGFβ 1) was transfected into 3T3 cells by using Fugene6. The mRNA expression and protein production of transduced TGFβ 1 gene were determined by RT-PCR and ELISA methods. The inhibitory effect of TGFβ 1 on the growth of CCL/64 cells was also determined. Results The hTGFβ 1 gene transduced 3T3 cells showed prominently elevated mRNA and proterin products expressions of hTGFβ 1 and the secretive hTGFβ 1 expression could inhibit the growth of the cultured CCL/64 cells. Conclusion The results indicate that human TGFβ 1 is prominently expressed in 3T3 cells, and recombinogenic hTGFβ 1 secreted from gene transduced 3T3 cells possesses definite biologic activity. This establishes the base for hTGFβ 1 genetically modified tissue engineering of the cartilage.