figs1.

Figure S1. RGS14 protein expression in various metabolically tissues of control and Rgs14-HKO mice. Western bolt analysis of Rgs14 protein expression of Control and Rgs14-HKO mice in liver, heart, lung, kidney and adipose, β-Actin was served as internal control (n = 3 mice per group). A two-tailed Student t test analysis was used. ∗P < 0.05, ∗∗P < 0.01, n.s. indicates no significance between the two indicated groups. All data are shown as the mean ± s.d.; Figure S2. Overexpression of GNAI1/3 promotes hepatocyte steatosis and inflammation. (A) Western blot analysis of Giα1/3 expression in primary mouse hepatocytes transfected with indicated adenovirus (n = 3 independent experiments). (B) cAMP levels in Giα1 or Giα3 overexpressed primary mouse hepatocytes following PO exposure for 12 h (n = 3 independent experiments). (C) Representative images (n = 3 independent experiments) and quantification (n = 7 high power fields per group) of Nile Red staining in Giα1 or Giα3 overexpressed primary mouse hepatocytes under PO treatment. Scale bar, 10 um. (D) Intracellular TG content in in Giα1 or Giα3 overexpressed primary mouse hepatocytes under PO treatment (n = 3 independent experiments). (E–G) qPCR analysis of mRNA levels of fatty acid uptake and synthesis genes (E), fatty acid oxidation genes (F) and inflammation genes (G) in infected primary mouse hepatocytes treated with PO (n = 3 independent experiments). One-way ANOVA analysis was used. ∗P < 0.05, ∗∗P < 0.01, n.s. indicates no significance between the two indicated groups. All data are shown as the mean ± s.d. Abbreviations: Ad, Adenovirus; cAMP, cyclic adenosine monophosphate; TG, Triglyceride; PO, 0.5mM Palmitic Acid and 1mM Oleic Acid; Figure S3. Inhibition of GNAI1/3 improves hepatocyte steatosis and inflammation. (A) Western blot analysis of Giα1or Giα3 expression in primary mouse hepatocytes transfected with indicated adenovirus (n = 3 independent experiments). (B) cAMP levels in Giα1-knockdown, Giα3-knockdown or Giα1/3-knockdown primary mouse hepatocytes following PO exposure for 12 h (n = 3 independent experiments). (C) Representative images (n = 3 independent experiments) and quantification (n = 7 high power fields per group) of Nile Red staining in Giα1-knockdown, Giα3-knockdown or Giα1/3-knockdown primary mouse hepatocytes under PO treatment. Scale bar, 10 um. (D) Intracellular TG content in Giα1-knockdown, Giα3-knockdown or Giα1/3-knockdown primary mouse hepatocytes under PO treatment (n = 3 independent experiments). (E–G) qPCR analysis of the mRNA level of fatty acid uptake and synthesis genes (E), fatty acid oxidation genes (F) and inflammation genes (G) in Giα1-knockdown, Giα3-knockdown or Giα1/3-knockdown primary mouse hepatocytes under treated with PO 12 h (n = 3 independent experiments). A two-tailed Student t test (B) and one-way ANOVA (A and C–G) analysis was used. ∗P < 0.05, ∗∗P < 0.01, n.s. indicates no significance between the two indicated groups. All data are shown as the mean ± s.d. Abbreviations: Ad, Adenovirus; cAMP, cyclic adenosine monophosphate; TG, Triglyceride; PO, 0.5 mM Palmitic Acid and 1 mM Oleic Acid.