| Biological drugs paclitaxel and adriamycin are clinically important drugs for cancer chemotherapy. But it is the important cause of treatment failure that chemotherapy drugs resistance produced in the process of tumor therapy. And deeply understanding about the mechanism of drug-resistance is the basic of solving the chemo-resistance. Human teratomas derived growth factor(TDGF1/Cripto1) is a biomarker of stem cells, and TDGF1 is not expression in the normal tissues. But recently researches, TDGF1 are re-expression in human malignant tumors. The highly expression in tumors is often involved in chemotherapy with poor prognosis. It is a new sight of understanding tumor development. Our study used cell line A2780 as study material, a cell line of ovarian carcinoma which is seriously affecting the health of female. Studying on the relationship between A2780 drug-resistance and TDGF1, and the activation of signal pathways related TDGF1 expected to deeply understanding the function of TDGF1 in the process of ovarian cancer drug resistance.Firstly, we used MTT to measure the drug sensitivity to paclitaxel of the A2780/WT and A2780/PTX cells. The IC50 of the two cell lines respectively: 23.88 μg/m L and 240.9 μg/m L. The IC50 of A2780/PTX cells were higher 10-folds than A2780/WT cells, indicated that A2780/PTX cells were resistant to paclitaxel. RT-PCR and fluorescence staining were carried out to test the expression of TDGF1 in A2780/WT and A2780/PTX cells. The results indicated that the expression of TDGF1 in A2780/PTX cells was significant higher than A2780/WT cells. The expression of TDGF1 usually resulted in cell proliferation. We draw the growth curve and measured the proliferation index(PI) of A2780/WT and A2780/PTX cells, and the results showed that the proliferative capacity of A2780/PTX cells was stronger than A2780/WT cells. This part confirmed that A2780 resistant to paclitaxel is related to high expression of TDGF1, and the high expression of TDGF1 enhanced the ability of cell proliferation.Secondly, to deeply confirm A2780 drug resistance was involved in high re-expression of TDGF1, we selected the first-line chemotherapy drugs paclitaxel and adriamycin to induce A2780/PTX and A2780/ADM cells by low concentration dose method. During this process, we acquired A2780/PTX series cells: A2780/PTX1 and A2780/PTX2 cells. MTT assay was carried out to test the IC50 of the induced A2780/PTX2 cells. The IC50 of A2780/PTX2 cells was 255.3 μg/m L, 10-folds than A2780/WT cells, indicated that the induced A2780/PTX2 cells were resistant to paclitaxel. RT-PCR and fluorescence staining results showed that the induce A2780/PTX series cells were expressed TDGF1 higher than A2780/WT cells, and the growth curve and PI of the induce A2780/PTX series cells showed that the proliferative capacity was stronger than A2780/WT cells. The same results derived in the induce A2780/ADM series cells. This part confirmed the ovarian tumor cell lines A2780 resistant to paclitaxel/adrimycin would resulted the high expression of TDGF1.Finally, the high expression of TDGF1 in A2780/PTX cells was activated the Nodal1-dependent TGFβ signal pathway. RT-PCR measured the genes related TDGF1 signal pathways, and the results showed that the expression of Nodal1 in A2780/PTX cells was higher than A2780/WT cells. It preliminarily indicated that the high expression of TDGF1 was actived the pathways about Nodal1. The results of western blot discovered that Smad3 was high expression and Smad3 phosphorylation in A2780/PTX cells. It finally confirmed that the high expression of TDGF1 in A2780/PTX cells actived the Nodal1-dependent TGFβ signal pathway.Our study researched the relationship between ovarian cancer cell lines A2780 drug-resistance and the high expression of TDGF1. Confirmed the TDGF1 was involved in the process of ovarian cancer cell drug-resistance, but deeply mechanism of drug-resistance of TDGF1 must be carried out in the future study. |
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