AIM: To construct interleukin-2 gene-modified humanhepatocyte line (L-02/IL-2) and investigate the changes ofthe function of liver cells and IL-2 secretion in culture withmicrocarrier, laying the foundation for furthe...AIM: To construct interleukin-2 gene-modified humanhepatocyte line (L-02/IL-2) and investigate the changes ofthe function of liver cells and IL-2 secretion in culture withmicrocarrier, laying the foundation for further experimentationon hepatocyte transplantation.METHODS: hIL-2 gene was transduced into L-02hepatocytes by recombinant retroviral vector pLNCIL-2, andthe changes of morphology and clonogeneicity rate of thetransduced cells were observed, the secretion levels of hIL-2 in cultural supernatant were detected by ELISA and NeoRgene was amplified by PCR. The growth of L-02/IL-2, thespecial biochemistry items and the levels of IL-2 weredetected after cultivation with microcarrier.RESULTS: The clonogeneicity rate of the L-02/IL-2 cellswas lower than that of L-02/Neo cells and L-02 cells. Thelevels of hIL-2 could reach 32 000 pg/106 cells per day andkept secreting for more than ten weeks. NeoR gene segmentwas respectively obtained by PCR from both L-02/IL-2 andL-02/Neo cell's genomic DNA. At the 6th day in culture withmicrocarrier, the matrix-induced liver cell aggregates wereformed, the number of alive L-02/IL-2 cell were 16.8±0.53x106/flask and the levels of ALB and UREA were 52.54±1.28mg/L and 5.29±0.17 mmol/L, respectively. These data hadnot significantly changed as compared with those of L-02cells (P＞0.05); However, the levels of IL-2 in IL-2/L-02 cellsremarkably exceeded that in L-02 cells in the whole cultureprocess (P＜0.001).CONCLUSION: The IL-2 gene-modified hepatooyte line hasbeen successfully constructed. The L-02/IL-2 cellularaggregates cultured with microcarrier have a high capacityof IL-2 production as well as protein synthesis and aminoacid metabolism.展开更多
基金Science and Technology Development Foundation of Fujian Province,No.98-Z-214
文摘AIM: To construct interleukin-2 gene-modified humanhepatocyte line (L-02/IL-2) and investigate the changes ofthe function of liver cells and IL-2 secretion in culture withmicrocarrier, laying the foundation for further experimentationon hepatocyte transplantation.METHODS: hIL-2 gene was transduced into L-02hepatocytes by recombinant retroviral vector pLNCIL-2, andthe changes of morphology and clonogeneicity rate of thetransduced cells were observed, the secretion levels of hIL-2 in cultural supernatant were detected by ELISA and NeoRgene was amplified by PCR. The growth of L-02/IL-2, thespecial biochemistry items and the levels of IL-2 weredetected after cultivation with microcarrier.RESULTS: The clonogeneicity rate of the L-02/IL-2 cellswas lower than that of L-02/Neo cells and L-02 cells. Thelevels of hIL-2 could reach 32 000 pg/106 cells per day andkept secreting for more than ten weeks. NeoR gene segmentwas respectively obtained by PCR from both L-02/IL-2 andL-02/Neo cell's genomic DNA. At the 6th day in culture withmicrocarrier, the matrix-induced liver cell aggregates wereformed, the number of alive L-02/IL-2 cell were 16.8±0.53x106/flask and the levels of ALB and UREA were 52.54±1.28mg/L and 5.29±0.17 mmol/L, respectively. These data hadnot significantly changed as compared with those of L-02cells (P＞0.05); However, the levels of IL-2 in IL-2/L-02 cellsremarkably exceeded that in L-02 cells in the whole cultureprocess (P＜0.001).CONCLUSION: The IL-2 gene-modified hepatooyte line hasbeen successfully constructed. The L-02/IL-2 cellularaggregates cultured with microcarrier have a high capacityof IL-2 production as well as protein synthesis and aminoacid metabolism.
