Construction of mouse fibroblast NIH/3T3 strain stably expressing HAb18G/CD147 and its implication

    HUANG Yong, JIANG Jianª²Li, ZHOU Jun, YAO Xiª²Ying, DOU Keª²Feng, CHEN Zhiª²Nan

    1Department of Hepatobiliary Surgery, Xijing Hospital, 2Cell Engineering Research Center, School of Basic Medicine, Fourth Military Medical University, Xi¬ðan 710033, China

    ¡¾Abstract¡¿ AIM: To observe the possibility and implication of stable transfection of fullª²length HAb18G cDNA into NIH/3T3 cell line. METHODS: The eukaryotic expression vector pcDNA3.0 containing human fullª²length HAb18G cDNA and empty vector pcDNA3.0 were introduced respectively into NIH/3T3 fibroblast cells by liposomeª²mediated gene transfection method. NIH/3T3ª²HAb18G was selected by G418 and the positive clone strain t3T3 was gained. The expression of HAb18G in t3T3 was identified by flow cytometry and indirect immunoflourescence staining and the effects on 3T3 excreting MMPs after transfecting were studied by gelatin zymography and RTª²PCR. RESULTS: Flow cytometry and indirect immunoflourescence staining showed that HAb18G was stably transfected into NIH/3T3 cells and agarose gel electrophoresis showed that the RTª²PCR products of MMPª²2 and MMPª²9 mRNA increased after transfection. The gelatin zymography confirmed that the ability of fibroblast to excrete MMPs was enhanced after transfection. CONCLUSION: Successfully constructed fibroblast strain with stable HAb18G expression can supply a cell model in studying the mechanism of MMPs excretion stimulating HAb18G. ......