Role of nuclear factor of activated T cells on impact of DEHP on IL-4 expression

ObjectiveTo investigate the role of nuclear factor of activated T cells(NFAT)on the impact of phthalate(2-ethylhexyl)ester(DEHP)on interleukin-4(IL-4)expression and to explore the immunotoxicity mechanism of DEHP.MethodsTwo activating agents of 10 ng/mL phorbol-12-myristate-13-acetate(PMA)+0.5 mg/mL ionomycin(Ion)and 25 ng/mL PMA+1 mg/mL Ion were used to activate spleen lymphocytes.The lymphocytes were exposed to 10μmol/L and 50μmol/L DEHP for 4 hours with or without 0.5μmol/L tacrolimus(FK506).IL-4,NFATp and NFATc mRNA were determined with real time-PCR test.NFATp and NFATc protein were measured with western blotting.ResultsUnder the activating of 10 ng/mL PMA+0.5 mg/mL Ion,the relative value of the expression of IL-4 mRNA was 2.53±0.42 in activating group and the value was 18.21±1.6 in 50μmol/L DEHP group,with significant difference between the two groups(P<0.01).The expression of IL-4 mRNA was 9.85±1.24 in FK506 group and the expression was obviously reduced compared with 50μmol/L DEHP group(P<0.01).The relative expression of NFATc mRNA was 2.00±0.32 in 50μmol/L DEHP group and the expression was 1.37±0.10 in activating group,with a significant difference between DEHP group and activating group(P<0.01).On the other hand,DEHP significantly enhanced NFATc protein expression in both cytoplasm and nucleus.The relative expression of NFATp mRNA was 2.51±0.45 in activating group and the expressions were 5.14±0.28 and 5.38±0.60 in 10μmol/L and 50μmol/L DEHP group,with statistically significant differences.But DEHP markedly decreased the expression of NFATp protein in both cytoplasm and nucleus after 4 hours'treatment.ConclusionNFAT plays an important role in the effect of DEHP on IL4 expression.