Figure 3
Overexpression of miR-494-3p downregulates the browning and thermogenic gene program in beige adipocytes. (A,C) Western blotting analysis of 3T3-L1 beige adipocytes transfected with miR-494-3p precursor (A), inhibitor (C) or controls. Immunoblot shown is representative of three independent experiments. Cells were harvested at day 8 after iso (10 µM) treatment for 8 h. n = 6. (B,D) Densitometric analysis of proteins from western blot in (A) and (C). Levels were normalized to HSP70 as an internal control. n = 6 each group. (E) mtDNA copy number was analysed in cells harvested at day 8 after iso (10 µM) treatment for 8 h as described in Method (supplementary file 1) (F,G) RT-qPCR analysis of mitochondrial biogenesis markers. Cells were harvested at day 8 with iso (10 µM) treatment for 8 h. mRNA levels were normalized using 36B4. n = 6. (H,I) Oxygen consumption rate (OCR) under miR-494-3p overexpression or inhibition in 3T3-L1 beige adipocytes transfected with miR-494-3p precursor, inhibitor or control with 50 µM isoproterenol treatment at 24 min. n = 5. *p < 0.05, **p < 0.01, ***p < 0.001. Full-length blots are presented in Supplementary Fig. 5.
