Study Of Drug-Resistant Cells From Cycle-Induction Combined Chemotherapy And Its Relationship With CSC

Object:To investigate the possibility that induction of cell cycle entry enhences chemotherapy sensitivity and the mechanism.Materials and Methods:In vitro, cells as experimental subjects, cultured human colon cancer HCT116 cell lines and primary colon cancer cell respectively were given the epidermal growth factor (EGF).After a certain time, cell proliferation and changes in the proportion of CD133+ cells were detected by flow cytometry, and in CD133+ cells sorted by flow cytometry cell proliferation was detected after EGF stimulation; after tumor cells were given EGF combined with 5-Fu, apoptosis and proliferation of tumor cells were detected by flow cytometry. In vitro, Subcutaneous xenograft mod of HCT116 human colon cancer cell lines and primary colon cancer cells in nude mice were established. After EGF and 5-Fu treatment, by recording the indicators of tumor growth analysis was performed to evaluate the efficiency of cycle-induction combined chemotherapy.Result:In vitro:1) After the EGF stimulation, the cell proportion of S-G2/M phase was increased as well as Ki67 expression, indicating more cells entering the cell cycle to promote proliferation; simultaneously the CD133+ cell percentage also increased, and among them more cells entered the proliferative phase; 2) 5-Fu decreased the tumor cells in S-G2/M phase, while cell apoptosis increased; 3) After combined treatment of EGF and 5-Fu, the apoptosis of tumor cells is higher than 5-Fu alone Group. Animal experiments:1) given EGF, S-G2/M phase cells in xnografts were increased as well as Ki67 expression; CD133+ cells increased in proportion.2) After the interventions of different options, EGF +5-Fu group tumor weight were less than 5-Fu alone group. Conclusion:1)EGF can promote the proliferation of tumor cells by entering the cell cycle, including CD133+ cells.2)EGF can enhance tumor chemotherapy efficincy, which may resulted in the cancer stem cells entry into the cell cycle and proliferation, increasing their sensitivity to chemotherapeutic drugs. Object:To investigate the drug-resistant cells from cycle-induction combined chemotherapyMaterials and Methods:In vitro, drug-resistant cells obtained after administration were investigated,and the expression of the major cyclins as well as DNA content were detected by flow cytometry; CFSE fluorescent was analysised for cell division and proliferation status; Drug resistance of every group of cells was analysised. In animal experiments, the major cyclin expression of each xenograft and DNA distribution were detected; growth of every xenograft were recorded after transplantation; Resistance of each group was investigated.Result:1) In vitro:the major cyclins expression was significantly lower in EGF +5-Fu group than in the others, However, no significant cycle arrest observable in each group; the division and proliferation rate of cells in the EGF +5-Fu group was significantly lower than that of other groups; after the second 5-Fu administration, apoptosis rate of EGF +5-Fu was significantly lower than other groups.2) Animal experiments:the major cyclins expression was significantly lower in EGF +5-Fu group than in the others, However, no significant cycle arrest observable in each group; after passaged to the second recipient,,tumors of EGF +5-Fu group showed a significantly higher growth rate and more potential resistance than other groups; HE sections also showed a higher degree of malignancy in combined group.Conclusion:Residual tumor cells from cycle-induction combined chemotherapy is characterized as a distinct subpopulation of enhenced drug resistance with low cyclin expression and slow cell cycle, and also shows a higher degree of malignancy, most likely cause of resistance leading to tumor recurrence Object:To investigate whether the drug-resistant cells possess sthe property of cancer stem cellMaterials and Methods:In vitro, self-renewal capacity were analysised by series passage; sphere formation was performed in testing tumor initiation capability. In animal experiments, limite dilution assay was used to determine the tumor initiation ability; expression of CSCs associated gene and surface markers were quantitated by real-time PCR.Results:In vitro experiment:EGF +5-Fu group were of stronger self-renewal capacity and ability of initiating tumors than other groups. Animal experiments:EGF +5- Fu group of tumor cells in nude mice is stronger than the other cells, expression of sox2 and notch2 in EGF +5-Fu group was higher than other groups, as well as that of surface mark associated.Conclusion:resistant cells from cycle-induction combind chemotherapy have a potential capacity of self-renewal and tumor initiation, possessing the basic characteristics of CSCs...