Butylated hydroxyanisole isomers induce distinct adipogenesis in 3T3-L1 cells

Butylated hydroxyanisole (BHA) isomers, as the widely used anthropogenic antioxidants in food, have been revealed to induce endocrine disrupting effects, while the mechanism how BHA isomers regulate the lipogenic differentiation remains to be elucidated. Using 3T3-L1 differentiation model, the effects of BHA isomers, including 2-tert-butyl-4-hydroxyanisole (2-BHA), 3-tert-butyl-4-hydroxyanisole (3-BHA) and their mixture (BHA), on adipogenesis were tested. The results showed that 3-BHA and BHA promoted adipocyte differentiation and enhanced the cellular lipid accumulation through the regulation of the transcriptional and protein levels of the adipogenetic biomarkers, while 2-BHA had no effect. The effective window for 3-BHA induced lipogenesis was the first four days during 3T3-L1 differentiation. BHA isomers showed no binding affinities for peroxisome proliferator activated receptor gamma (PPAR gamma). Instead, the upstream of PPAR gamma signaling pathway, i.e. the phosphorylation of cAMP-response element binding protein (CREB), upregulation of CAAT/enhancer-binding proteins beta (C/EBP beta) and elevated cell proliferation during postconfluent mitosis stage were induced by 3-BHA exposure. Altogether, this study revealed the adipogenic effect of 3-BHA through interference with the upstream events of the PPAR gamma signaling pathway. The authorized usage of BHA as food additives and its occurrence in human sera can potentially contribute to the incidence of obesity, which is of high concern.