Liver Protein Expression Analysis

Lysates were prepared by homogenizing liver pieces in 3 volumes of 0.25 M sucrose–10 mM phosphate buffer (pH 7.4), followed by centrifugation at 700 ×g for 10 min and collection of the supernatant. Western blotting was performed as described previously [8 (link)], using antibodies against alpha-smooth muscle actin (α-SMA) (ab5694, Abcam plc, Cambridge, UK) and platelet-derived growth factor receptor beta (PDGFR-β) (#3169, Cell Signaling Technology, Beverly, MA). Anti-glyceraldehyde-3-phosphate dehydrogenase (GAPDH) (sc-25778, Santa Cruz Biotechnology, Santa Cruz, CA, USA) was used for a loading control. Pierce 1-StepTM Ultra TMB-Blotting Solution (Pierce Biotechnology, Rockford, IL, USA) was used for signal development. Gray values for the target protein bands were measured with Image J (NIH, Maryland, USA) and normalized to the control group band value (defined as 1.0).