Study Of DEHP On Proliferation And Cell Cycle In MIN6 Cells

Objective Di(2-ethylhexyl)phthalate(DEHP)is a plasticizer widely distributed in our daily life and considered as an environmental endocrine disruptor that can cause toxic effect on pancreatic β-cell.The purpose of this study is to analyze the effect of DEHP on cell proliferation and cell cycle,which can further elucidate the toxic effect of DEHP on MIN6 cells.Methods The research object was MIN 6 cells,which were cultured in 25,50,100 μM DEHP or vehicle(DMSO,<0.1)for 48 h,respectively.The effect of DEHP on cell proliferation in MIN6 cells was detected by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide(MTT)assay.The effect of DEHP on glucose-stimulated insulin secretion(GSIS)in MIN6 cells was detected by the enzyme linked immunosorbent assay(ELISA).Flow cytometry was used to observe the effect of DEHP on cell cycle in MIN 6 cells.The effect of DEHP on the activity of SOD in MIN 6 cells was detected by Water-soluble tetrazolium salt(WST-1)assay.The effect of DEHP on the content of MDA in MIN 6 cells was detected by thiobarbituric acid(TBA)assay.The effect of DEHP on the expression of Cyclin D1 and Cyclin-dependent kinase 4(CDK4)in MIN 6 cells were detected by WB.Results1.MTT assay showed that 25μM,50μM and 100μM DEHP inhibited the cell viability of MIN6 cells(P<0.05).2.GSIS results showed that DEHP did not affect glucose-stimulated insulin secretion in MIN6 cells under basal glucose stimulation,while 50μM and 100μM DEHP inhibited glucose-stimulated insulin secretion in MIN6 cells under high glucose stimulation(P<0.01).3.The results of SOD activity showed that the activity of SOD in 100μM DEHP group decreased(P<0.01).4.The results of MDA content showed that the content of MDA in 50μM and 100μM DEHP group increased(P<0.001).5.The results of cell cycle showed that the percentage of cells in G0/G1 phase of MIN6 cells increased(P<0.01)while the percentage of cells in S phase and G2/M phase decreased in 25μM,50μM and 100μM DEHP group(P<0.05).6.The results of WB showed decreased expression of Cyclin D1 and CDK4 in 25μM,50μM and 100μM DEHP group compared to vehicle(P<0.05).Conclusions1.DEHP inhibits the viability and GSIS,also induces the cell cycle arrest and downregulates the expression of Cyclin D1 and CDK4 of MIN6 cells,which is related to oxidative stress.2.DEHP induces oxidative stress via inhibiting the activity of SOD and stimulating the production of MDA in MIN 6 cells.