3-tert-Butyl-4-hydroxyanisole Perturbs Differentiation of C3H10T1/2 Mesenchymal Stem Cells into Brown Adipocytes through Regulating Smad Signaling

Exposure to 3-BHA induced a shift from a brown to white-likephenotype in C3H10T1/2 mesenchymal stem cells by promoting the phosphorylationof Smad1/5/8. 3-tert-Butyl-4-hydroxyanisole (3-BHA),one ofthe most commonly used antioxidants in foodstuffs, has been identifiedas an environmental endocrine disruptor (EED) with obesogenic activity.Given the increasing concern on EED-caused dysfunction in lipid metabolism,whether 3-BHA could influence the development of brown adipocytesis worthy of being explored. In this study, the effect of 3-BHA onthe differentiation of C3H10T1/2 mesenchymal stem cells (MSCs) intobrown adipocytes was investigated. Exposure to 3-BHA promoted lipogenesisof the differentiated cells, as evidenced by the increased intracellularlipid accumulation and elevated expressions of adipogenic biomarkers,including peroxisome proliferator-activated receptor & gamma; (PPAR & gamma;), Perilipin, Adiponectin, and fatty acid binding protein 4 (FABP4). Surprisingly,the thermogenic capacity of the differentiated cells was compromisedas a result of 3-BHA exposure, because neither intracellular mitochondrialcontents nor expressions of thermogenic biomarkers, including uncouplingprotein 1 (UCP1), peroxisome proliferator-activatedreceptor & gamma; coactivator 1 & alpha; (PGC1 & alpha;), cell-death-inducing DNA fragmentation factor & alpha; subunit-likeeffector A (CIDEA), and PR domain containing 16 (PRDM16), were increased by this chemical. The underlyingmolecular mechanism exploration revealed that, in contrast to p38MAPK, 3-BHA stimulation induced phosphorylation of Smad1/5/8 in anexposure time-dependent manner, suggesting that this chemical-triggeredSmad signaling was responsible for the shift of C3H10T1/2 MSC differentiationfrom a brown to white-like phenotype. The finding herein, for thefirst time, revealed the perturbation of 3-BHA in the developmentof brown adipocytes, uncovering new knowledge about the obesogenicpotential of this emerging chemical of concern.