Combination of endostatin gene transfer and radiation therapy against lung adenocarcinoma model

摘要

To investigate the combined effects of endostatin gene therapy and radiation in lung adenaocarcinoma model. Human lung adenocarcinoma A549 cell line was transfected with retroviral vector pLESSN which encoded endostatin gene to obtain A549/Endo cells. PCR amplification was used to verify the endostatin gene in the DNA of A549/Endo cells. The transfectant A549/Endo cell's growth in vitro was estimated by cell counting assay. And the inhibitive effect of expressed endostatin on the proliferation of endothelial cell EA.hy926 in vitro was assayed by MTT method. In vivo study, the nude mouse subcutaneous tumor growths of 4 groups (A549, A549/Endo, A549+irradiation, A549/Endo+irradiation) were observed and compared. And the microvessel density (MVD) of each of 4 group tumors was assayed with Immunohistochemical analysis. PCR confirmed that there was a 600-bp endostatin gene fragment in the DNA of A549/Endo cells, but not in the DNA of A549 cells. Cell counting assay showed that there was no cell-proliferation difference between A549/Endo cells and A549 cells in vitro. MTT assay revealed that the expressed product of endostatin gene could significantly inhibit the proliferation of endothelial cells EA.hy926 in vitro by 47.3%. Tumor growth of A549/Endo cells was delayed significantly compared with A549 cells (p<0.01). Although radiation had a definite effect on A549 tumors, the tumor growth inhibitive effect of endostatin gene transfer plus radiation was greater than any their alone therapy (p<0.01,respectively). The results of immunohistochemical detection showed that the MVD of A549/Endo and A549/Endo+Irradiation tumor were both significantly decreased compared with A549 tumor(p<0.01, respectively). But further compared between A549/Endo and A549/Endo+Irradiation tumor, the MVD of A549/Endo+Irradiation tumor was further significant declined (p<0.01). Retrovirus vector can effectively transfer endostatin gene into A549 cells. Endostatin gene transfer combined with radiation has a synergistical inhibition effect on the growth of lung adenocarcinoma tumor model.