| Objectives To investigate the effect of VEGFR signal pathway inhibitor apatinib on tamoxifen resistant breast cancer cell line LCC2 and its possible mechanism.Methods 1 MCF-7 and LCC2 cell lines were selected as study cells,and the morphological differences between parent cell line MCF-7 and tamoxifen resistant cell line LCC2 were observed when cultured in logarithmic phase.2 CCK-8 assay was used to determine the proliferation inhibition rate of MCF-7 and LCC2 cells treated with different concentrations of 4-hydroxy-tamoxifen for 48 hours,and to detect the change of cell proliferation inhibition rate after LCC2 was treated with different concentrations of apatinib for 48 hours,and to detect the proliferation inhibition rate of LCC2 cells treated with different concentrations of 4-hydroxy-tamoxifen combined with 10 μM apatinib for 48 hours.3 After LCC2 cells were treated with 4-hydroxy-tamoxifen,apatinib and their combination for 48 hours,the apoptosis rate was detected by flow cytometry(FCM).4 Western blotting was used to detect the expression of ERα,PI3 K,Akt and p-Akt pathway proteins in MCF-7 cells and LCC2 cells,and the changes of Bcl-2,Bax apoptosis protein and ERα,PI3 K,Akt and p-Akt protein expression in LCC2 cells treated with 4-hydroxy-tamoxifen,apatinib and their combination for 48 hours.Every experiment was repeated more than 3 times in each group.All data were analyzed by SPSS21.0 statistical software.The measurement data were expressed by (?)±s.One-way ANOVA was used to compare multiple groups.P<0.05 was considered statistically significant.Results 1 The morphology of MCF-7 cells was slightly different from that of LCC2 cells.The MCF-7 of sensitive cells was fusiform or polygonal with a short long axis,while the LCC2 of drug-resistant cells was round or oval.2 The results of CCK-8 assay showed that the inhibition rate of LCC2 cells was lower than that of MCF-7 cells treated with tamoxifen at the same time and the same concentration,and the results of CCK-8 showed that apatinib could inhibit the proliferation of LCC2 cells in different degrees after 48 hours,and the inhibition rate of cell proliferation increased with the increase of apatinib concentration(P<0.05).CCK-8 detection of different concentrations of tamoxifen and 10 μM apatinib compared with single drug tamoxifen,the inhibition rate of cell proliferation was significantly increased(P<0.05).3 FCM showed that the apoptosis rate in the experimental group was higher than that in the control group(P<0.05),and the apoptosis rate in the combination group was higher than that in the tamoxifen group(P<0.01).4 Western blotting showed that the expression of ERα,PI3 K,Akt and p-Akt protein in the resistant cell line was higher than that in the sensitive cell line(P<0.05).After 48 hours of treatment with 4-hydroxy-tamoxifen,apatinib and the combination of apatinib and apatinib,the expression of apoptotic protein Bax protein increased and the expression of Bcl-2 protein and Bcl-2/Bax ratio decreased in the combination group compared with the single drug group(P<0.01).The expression of ERα,p-Akt protein and the ratio of p-Akt/Akt in the combination group were lower than those in the tamoxifen group(P<0.01).Conclusions The difference of drug resistance was determined by the action of 4-hydroxy-tamoxifen on drug-resistant strains and sensitive strains respectively,it was found that LCC2 showed high resistance to 4-hydroxy-tamoxifen,apatinib could inhibit the growth of breast cancer drug-resistant cells in a dose-dependent manner,and the combination of apatinib and tamoxifen had synergistic enhancement.Apatinib can inhibit the proliferation of drug-resistant cells and enhance the sensitivity of tamoxifen,which may be related to the up-regulation of Bax protein and down-regulation of Bcl-2 protein and the expression of estrogen receptor pathway proteins ERα,PI3 K,Akt and p-Akt.Figure[9];Table[2];Reference[136]... |
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