The Potential Role Of Anion Channel CFTR In Bisphenol A-induced Malignant Transformation Of Prostate Cells

Objectives:Bisphenol A（BPA）is a synthetic monomer of polycarbonate and epoxy resin,which is widely used in food packaging materials,water pipes,fireproof materials and other products.Our previous studies showed that BPA exposure levels were detectable in people of all age groups in China;moreover,other studies found that very low levels of BPA may lead to a variety of biological changes in prostate cells;in case-control studies of primary prostate cancer,BPA was found to be a risk factor.Cystic fibrosis transmembrane conduction regulator（CFTR）is an ATP gated and c AMP dependent anion channel.CFTR is associated with the occurrence and development of a variety of cancers,such as breast cancer,prostate cancer and so on.Our paper aims to explore the potential mechanism of CFTR in the effect of BPA on human normal prostate cells.This study is helpful to reveal the mechanism of reproductive system related tumors of environmental endocrine disruptors and to promote the development of anti-cancer drugs or techniques.Methods:First,we determined the concentration and time of BPA exposure to human normal prostatic epithelial cells by CCK-8 assay.Then,under the effect of appropriate concentration and time of BPA,the proliferation,migration and invasion of human normal prostate cells were determined by soft agar colony formation assay,wound healing assay and Transwell invasion assay.ELISA and Western blot were used to detect the contents of ATP,CFTR,Bcl-2and Bax in BPA treated prostate cells.Finally,the apoptosis level of BPA treated prostate cells was measured by flow cytometry.Using SPSS analysis data by single factor variance and factorial design,the data are expressed as mean value±standard deviation,P<0.05 is statistically significant.Results:The CCK-8 results showed that there was a nonlinear relationship between RWPE-1 cell viability and BPA concentration.In the concentration range of 0μM-10μM,there was no significant change in cell activity after BPA treatment for 24 hours.When the concentration was higher than 10μM,the cell viability decreased significantly.After making the growth curve,we calculated that IC₅₀ was 160μM.In toxicology,one eighth of IC₅₀ is usually taken as the exposure concentration,so the optimal concentration of BPA in this study is 20μM.We then explored the relationship between RWPE-1 cell activity and time at the optimal BPA concentration.The results showed that the cell activity was independent of time at 20μM BPA concentration.We also detected the apoptosis level of the cells after exposure to 20μM BPA for 48 h by flow cytometry.Compared with the control group,there was no difference in the apoptosis level of the cells in the experimental group According to the above experiments this concentration has no acute damage to RWPE-1 cells and does not change the apoptosis level.We determined that the exposure time of RWPE-1 cells was 4weeks.After that,the clone ability,migration ability and invasion ability of the transformed cells were evaluated by soft agar colony formation assay,wound healing assay and Transwell invasion assay.Compared with the control group,the abilities of the transformed cells were enhanced.After that,the level of ATP in the transformed cells was detected by ELISA.It was found that the content of ATP in the experimental group was significantly lower than that in the control group.The levels of CFTR,Bax and Bcl-2 were detected by Western blot.Compared with the control group,the levels of CFTR and Bax in transformed human normal prostate cells were significantly decreased,while the expression level of Bcl-2 protein was significantly increased.Conclusions:（1）BPA may induce the malignant transformation of RWPE-1 cells;（2）BPA may induce malignant transformation of human normal prostate epithelial cells by inhibiting anion channel CFTR expression,causing mitochondrial dysfunction and inhibiting apoptosis.