Paired Related Homoeobox1 Promotes The Docetaxel Resistance Of Breast Cancer Cell By Inducing Epithelial-mesenchymal Transition

【 Objective 】 To investigate the effect of Prrx1-induced epithelial-mesenchymal transition on Docetaxel resistance in breast cancer cell in vitro.【Methods】1.The lowest expression of Prrx1 of Breast cancer cell lines(including BT-549,MDA-MB-468 and MCF-7)were screened by real-time PCR.2.Then the cells were transfected with Prrx1 over-expression letiviral vectors.The morphology changes of the breast cancer cells were observed under the white light and fluorescence of fluorescence microscopy after the transfection was verified to be accomplished through Real-time PCR.3.The influence of Prrx1 on the migration ability of the breast cancer cell was detected by Transwell assay.4.CCK-8 was used to detect the IC50 of breast cancer MCF-7 cell,and the IC50 of the experimental group,the blank control group and the negative control group(NC)were measured after letiviral vectors infection in 24 hours.5.Effects of Prrx1 over-expression on apoptosis rate of breast cancer cells After the three groups cells were treated with docetaxel in 24 h by flow cytometry.6.Western blot was used to detect the expression of Twist1 and multidrug resistance-associated protein 1(MDR1/ABCB1 / p-gp)of the three groups cells.【Results】 Of these three cells,the lowest expression of Prrx1 was MCF-7 cells(p <0.001).Then the MCF-7 cells were transfected by letiviral vectors to be Prrx1 over-expressing.We observed that the morphology of these cells changed from "paving stone" to long spindle appearance and the fluorescence efficiency of 80% or more.The real-time PCR results showed that the expression of Prrx1,E-adherin and Vimentin was 252.09,0.64,and 2.65 times compared with groups without transfection respectively(t=44.30,p(27)0.001),(t=4.91,p(27)0.001),(t=12.05,p(27)0.001).Transwell assay showed that the number of the migratory cells in over-expression group(71.69±6.66)was significantly higher than NC group(40.87±4.16)(t=15.05,p(27)0.001)和 Blank control group(41.2±4.62)(15.76,p(27)0.001)while the difference between the two control groups were not statistically significant(t=0.21,p=0.84).CCK-8 assay showed that the IC50 values of docetaxel on MCF-7 cells in 24 h and 48 h were(10.94 ± 0.64),(10.66 ± 1.68)(t=0.27,p=0.80),After transfection,The IC50 of Prrx1 over-expressing MCF-7 cells was significantly higher than the blank control groups(13.30,p(27)0.001)and NC group(t=0.91,p(27)0.001)and there was no significant difference between the two control groups(t=0.22,p=0.83).Flow cytometry analysis showed that the apoptotic rate of Prrx1 over-expressing MCF-7 cells was significantly lower than blank(t=9.70,p(27)0.001)and negative control groups(t=9.32,p(27)0.001),and the difference was statistically significant,while two control groups were no statistically significant(t=0.48,p=0.65).Western blot results showed that the expression of Twist1 and ABCB1 in Prrx1 over-expressing MCF-7 cells was significantly higher than the two control group cells(p <0.001),while two control groups were no statistically significant(p(29)0.05).【Conclusion】 Prrx1 could effectively enhance the Docetaxel resistance of breast cancer cell by inducing epithelial-mesenchymal transition.