A–G: Reduced β3AR and Epac1 protein in HFD adipocytes as well as blunted epinephrine-stimulated adiponectin secretion upon β3AR and Epac1 siRNA knockdown.

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A–G: Reduced β3AR and Epac1 protein in HFD adipocytes as well as blunted epinephrine-stimulated adiponectin secretion upon β3AR and Epac1 siRNA knockdown. A–G: Reduced β3AR and Epac1 protein in HFD adipocytes as well as blunted epinephrine-stimulated adiponectin secretion upon β3AR and Epac1 siRNA knockdown. A: Representative bright-field and TIRF images of chow and HFD adipocytes immunostained with anti–β3-antibody followed by incubation with Alexa Fluor 488 secondary antibody. Negative control was incubated with secondary antibody only. B: Mean intensity/area in images of adipocytes from 20 chow-fed and 20 HFD-fed mice. C: Gaussian distribution of mean intensity for images of chow and HFD adipocytes. D: Epac1 content in chow and HFD adipocytes. E: Relative mRNA expression in 3T3-L1 adipocytes transfected with β3AR (Adrb3) or Epac1 (Rapgef3) siRNA. The siRNA expression level for each gene is normalized against its expression in concurrently scramble transfected cells. F and G: Epinephrine (EPI)-stimulated adiponectin secretion (30 min) in the β3AR or Epac1 silenced as well as scrambled siRNA-transfected adipocytes in E. Data in B and C are from cells isolated from three chow-fed and three HFD-fed animals. Data in D represent results from 20 animals (10 chow-fed and 10 HFD-fed). Results in E, F, and G are from five separate experiments. **P < 0.01; ***P < 0.001. Ali M. Komai et al. Diabetes 2016;65:3301-3313 ©2016 by American Diabetes Association