RETROVIRUS-MEDIATED TNF-α GENE TRANSFER INTO TCA8113 CELLS

摘要

Objective To investigate whether TNF-α gene-modified Tca8113 cells (Tca8113/TNF-α) can be used as vaccine for oral squamous cell carcinoma. Methods TNF-α gene was transduced into Tca8113 cells in vitro with retroviral vector earring genes for both TNF-α and Neo~R. After that, presence and expression of exogenous gene in the transgenic cells, expression of HLA antigen on the cells, expression of TNF-α and survival rate of the cells after irradiation and cryopreservation, and mutagenic activity of the cells were analyzed by PCR technique, ELISA technique, FACS technique, ~(60)Co irradiation inactivation test, cryopreservation test, and Ames test, respectively. Results The presence of both TNF-α and Neo~R gene and expression of TNF-α gene were demonstrated in transgenic cells. The levels of the HLA-A, B, C, DR expressed by Tca8113/TNF-α were higher than by the parental cells. Tca8113/TNF-α continued to secrete TNF-α for 14 d, there was a secretion peak time from d4 to d6;and, all the cells died by d14 after irradiation. The Level of TNF-α secreted by Tca8113/TNF-α cryopreserved for 48 h was no different from that cryopreserved for 1 week after irradiation , the level of TNF-α secreted by the cryopreserved cells was just a little lower than that secreted by the noncryopreserved cells. Both DNA and supernatant of the cells have no mutagenic activity. Conclusion TNF-α gene can be transduced into Tca8113 cells with retroviral vector, and the cells can express TNF-α. Expression of HLA Ⅰ, Ⅱ antigens on Tca8113 cells can be increased by TNF-α gene transduction. Irradiation is a reliable inactivation method, and cryopreservation is a feasible conservation method for Tca8113/TNF-α. Ames test result indicate that Tca8113/TNF-α has no mutagenic activity.