Citrus aurantiumL. Dry Extracts Ameliorate Adipocyte Differentiation of 3T3-L1 Cells Exposed to TNF alpha by Down-RegulatingmiR-155Expression

Citrus aurantiumL. dry extracts (CAde) improve adipogenesis in vitro. These effects are dependent from an early modulation of CCAAT/enhancer-binding protein beta (C/Ebp beta) expression and cyclic Adenosine Monophosphate (cAMP) response element-binding protein (CREB) activation.C/Ebp beta andCrebare also targets ofmiR-155. This study investigated whetherCAde regulatesmiR-155expression in the early stages of adipogenesis and whether it ameliorates adipocyte differentiation of cells exposed to tumor necrosis factor-alpha (TNF alpha). Adipogenic stimuli (AS) were performed in 3T3-L1 pre-adipocytes treated withCAde, TNF alpha, or both. Gene and miRNA expression were determined by quantitative real-time PCR. Adipogenesis was evaluated by Oil-Red O staining.CAde treatment enhanced AS effects during the early adipogenesis phases by further down-regulatingmiR-155expression and increasing bothC/Ebp beta andCrebmRNA and protein levels. At variance, TNF alpha inhibited 3T3-L1 adipogenesis and abolished AS effects onmiR-155,C/Ebp beta, andCrebexpression. However, in cells exposed to TNF alpha,CAde improved adipocyte differentiation and restored the AS effects on miRNA and gene expression at early time points. In conclusion, this study identifiedmiR-155down-regulation as part of the mechanism through whichCAde enhances adipogenesis of pre-adipocytes in vitro. Furthermore, it provides evidence ofCAde efficacy against TNF alpha negative effects on adipogenesis.