Anti eif3b

Cytolysis buffer (RIPA buffer, Sigma-Aldrich, St. Louis, MO, USA) was used to extract proteins from HSCs. Proteins were quantified by Pierce BCA Protein Assay Kit (Thermo Scientific, Waltham, Massachusetts, USA), denatured in SDS loading buffer (Solarbio, Beijing, China) at 95°C, separated by SDS-PAGE, and then transferred to polyvinylidene difluoride (PVDF) membranes (Thermo Fisher Scientific, Waltham, MA, USA). The membranes were blocked with 5% skim milk, incubated with primary antibodies against α-SMA (diluted 1: 1000, ab32575, Abcam, Cambridge, UK), Collagen 1a1 (diluted 1: 500, AF7001, Affinity Biosciences, Cincinnati, OH, USA), EIF3B (diluted 1: 1000, ab133601, Abcam, Cambridge, UK), and GAPDH (diluted 1: 10000, AB0037, Shanghai Abways Biotechnology Co., Shanghai, China) overnight at 4°C. The secondary antibody incubations (diluted 1: 20 000, IRDye, 800CW 926-32211, LI-COR Biosciences, Lincoln, NE, USA) were for 1 h at room temperature. The final results were observed by Odyssey infrared imaging system (LI-COR Biosciences, Lincoln, NE, USA), and the protein densities were analyzed with ImageJ software (National Institutes of Health, Bethesda, MD, USA). The primary antibody of anti-α-SMA and anti-EIF3B were from Abcam, Cambridge, UK, anti-Collagen 1a1 was from Affinity Biosciences, Shanghai, China, and anti-GAPDH was from Cell Signaling Technology, Danvers, MA, USA.