Effects Of Sulforaphane On Transdifferentiation Of3T3-L1Adipocytes And The Molecular Mechanism

Objective To investigate the effect of sulforaphane on transdifferentiation of3T3-L1adipoc-ytes from white into brown and explore the underling mechanism.Methods In the study, murine3T3-L1pre-adipocytes were cultured and differentiated intomature adipocytes by classic cocktail method. To screen the optimal time of intervention,mitochondrial content was detected by the Mito-Tracker Green staining in the differentiating3T3-L1pre-adipocytes differentiation. Then,3T3-L1mature adipocytes were incubated with lowdose of SFN(0to10.0μmol/L) for48h and mitochondrial biosynthesis was measured byMito-Tracker Green staining method and Mito Biogenesis In-Cell ELISA Kit. Furthermore,mitochondrial ultrastructural was observed under transmission electron microscope of3T3-L1mature adipocytes treated with0and1.0μmol/L of SFN for48h. In addition, the activities ofmitochondrial Complex-I and citrate synthase were investigated by specific assay kit to evaluatethe mitochongdrial activity. At the same time, mitochondrial function was measured by ATPdetection kit. Finally, leves of Nrf2, Sirt1, PGC-1α, NRF1and UCP1proteins were detected bywestern blotting.Results In the present study, Mito-Tracker Green staining confirmed a gradual increase in thenumber of mitochondrion during3T3-L1pre-adipocytes differentiation and being stable at the day10. Thus, the mature adipocytes at day10of differentiation were chosed to be treated with SFN. Itwas demonstrated that SFN treatment evidently increased the fluorescence intensity of Mito-Tracker Green suggesting an increase in mitochondrial mass. Besides, mitochondrial biogenesiswas confirmed to be significantly multiplied by SFN. Meanwhile, transmission electronmicroscope showed that a large number of mitochondria, with larger size and smaller lipid dropletover populate in3T3-L1adipocytes treated by1.0μmol/L SFN. In addition, ATP levels weremarkedly up-regulated in mature adipocytes treated by SFN for48h. Moreover, complex-I andcitrate synthase also were significantly activated. Finally, western blotting results showed that SFNwas able to strongly up-regulate the expession of Nrf2and UCP1protein. Besides, theup-regulation of PGC-1α protein as well as Sirt1and NRF1was observed, especially at1.0 μmol/L of SFN for48h.Conclusion Low level of SFN(0-10.0μmol/L)could up-regulate expession of key factorsfor regulating mitochondrial biosynthesis, such as Nrf2, NRF1, Sirt1and PGC-1α. By activatingSirt1/PGC-1α and Nrf2/PGC-1α pathways, SFN induced3T3-L1white mature adipocytes totransdifferentiate into brown adipocytes and obtain strong respiratory and thermogenesis function,which was great beneficial for weight control. Above all, the study provided powerful evidence forthe prevention and therapy of obesity and other metabolism syndrome targeting at WAT.