Study On The Effects Of TCDD/MEHP Co-exposure On MCF10A Cells

Objective: This study aimed to investigate the effects of 2,3,7,8-tetrachlorodibenzo-p-dioxin(TCDD)and mono-2-ethylhexyl phthalate MEHP)alone and co-exposed to normal human mammary gland cells MCF10 A,focusing on TCDD and MEHP on MCF10 A in AhR signaling pathway related genes and protein expression in order to initially explore TCDD and MEHP co-exposure to MCF10 A cell toxicity Mechanism.Method: 1.MTS method was used to detect TCDD / MEHP on normal human breast epithelial cells treated with MCF-10 A cell activity.MTS assay was used to detect the effects of 1pM,10 pM,100 pM,1n M,10 n M TCDD treatment on the cell viability of normal human mammary epithelial cells MCF10 A at 24 h and 48h;or 1?M,5?M,10?M,50?M and 100?M MEHP.The effects of 1pM,10 pM,100 pM,1n M TCDD and 100?M MEHP co-exposure treatment on the cell viability of normal human mammary epithelial cells MCF10 A after 24 h and 48 h were observed.2.The effect of TCDD / MEHP on the migration ability of normal human breast epithelial cells MCF 10 A was detected by scratch test.3.RT-PCR method was used to detect 100?M MEHP alone exposure,1n M TCDD alone exposure,100?M MEHP and 1n M TCDD co-exposure treatment for 48 h,The level of AhR,CYP1A1 and migration-invasion related genes MMP2,MMP9,VIM,E-calcium gene expression.4.RT-PCR and Western Blot methods were used to detect the expression level of AhR,CYP1A1,MMP2,MMP9,VIM,E-Ca gene and aroma after 100?M MEHP exposure alone,1n M TCDD alone exposure,100?M MEHP and 1n M TCDD exposure after si RNA interfered with AhR the effect of CYP1A1 protein expression.Result: 1.TCDD / MEHP on MCF10 A cell viability,TCDD can reduce cell viability,TCDD and MEHP co-exposure can also reduce cell viability.2.TCDD / MEHP on MCF10 A scratch test,TCDD treatment 48 h can make the cell healing ability to accelerate the trend than other groups,but no significant difference.3.Effect of TCDD/MEHP on MCF10 A migration related mRNA and AhR signaling pathway related gene expression.The TCDD treatment group could increase the expression level of CYP1A1 gene downstream of AhR.The expression level of CYP1A1 in MEHP and TCDD treatment group was lower than that in TCDD treatment group.The TCDD treatment group was able to inhibit E-Ca gene expression.4.Effect of TCDD/MEHP on AhR,CYP1A1 gene and protein expression levels in MCF10 A cells after AhR interference.AhR gene in si RNA-AhR group was significantly lower than si RNA-NC group,TCDD treatment group,In the MEHP /TCDD treatment group,after the si RNA interference,the expression level of CYP1A1 in the si RNA-AhR group was lower than that in the si RNA-NC group and there was a statistical difference.The AhR protein in the si RNA-AhR group was significantly lower than the si RNA-NC group,TCDD In treatment group,TCDD/MEHP treatment group,after interference with si RNA,the expression level of CYP1A1 protein in the si RNA-AhR group was lower than that in the si RNA-NC group.In conclusion: 1.TCDD alone exposure,as well as MEHP / TCDD co-exposure will affect cell viability and cell migration in varying degrees.2.TCDD may reduce the activity of MCF10 A cells and increase their migration ability by promoting the expression of MMP2,MMP9,AhR and CYP1A1 genes and decreasing the expression of E-Ca.3.MEHP may partially inhibit the toxic effects of TCDD exposure by affecting the AhR signaling pathway.