Hdac1

Reduced samples were loaded onto NuPAGE Bis-Tris (Thermo Fisher Scientific) gels in MOPS buffer and separated proteins were transferred onto a PVDF membrane for immunodetection with anti-PPARγ (C26H12, 1:2000: Cell Signaling Technologies, Danvers, MA, USA), HSP90α (GTX109753: Gene Tex, Irvine, CA, USA, 1:2000), HDAC1 (GTX100513: Gene Tex, 1:2000), GAPDH (GTX100118: Gene Tex, 1:1000), H3K27ac (39134: Active Motif, Carlsbad, CA, USA, 1:2000), H3K9ac (39918: Active Motif, 1:2000), H3k27me3 (C36B11: Cell Signaling Technologies, 1:2000), and Histone H3 (D2B12: Cell Signaling Technologies, 1:2000) antibodies as primary antibodies, and the HRP-conjugated goat anti-rabbit antibody (#7074: Cell Signaling Technologies, 1:2000) as the secondary antibody.

Magnolol was purchased from Shanghai BS Bio-Tech Co., Ltd. (Shanghai, China). MM1 was prepared as described by Lin et al. (43 (link)). The purity of magnolol and MM1 was <99%, as determined by high-precision liquid chromatography (HPLC) analysis. Magnolol and MM1 were each dissolved in dimethyl sulfoxide (DMSO) to obtain a stock concentration of 100 mM, which was then stored at −20°C before use. DMSO 0.1% v/v was used as the vehicle control. Sorafenib was purchased from Sigma-Aldrich (St. Louis, MO, USA). Antibodies against human class I HDACs (HDAC1, HDAC2, HDAC3, and HDAC8), acetyl-histone H3, acetyl-p53, p53, p21, Ki-67, E-cadherin, N-cadherin, vimentin, Snail, Slug, and β-actin were purchased from GeneTex (Irvine, CA, USA) and Cell Signaling Technology (Beverly, MA, USA). The antibody to cyclin D1 was purchased from ABclonal Technology (Woburn, MA, USA), and the antibodies against CDK4 were purchased from Proteintech (Rosemont, IL, USA). Secondary antibodies were purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA).

The primary antibodies against the following proteins were used: E-cadherin (BD Biosciences 610182, 1:5000), N-cadherin (BD Biosciences 610920, 1:5000), Vimentin (BD Biosciences 550513, 1:8000), Fibronectin (BD Biosciences 610077, 1:2000); β-actin (Sigma-Aldrich A1978, 1:10000); Snail (Abcam ab63371, 1:1000), Slug (Abcam ab27568, 1:1000); Zeb1(Santa Cruz sc-25388, 1:500), HDAC3 (Santa Cruz sc-17795, 1:1000); HDAC1(GeneTex, GTX27028, 1:2000); HDAC2(GeneTex, GTX12169, 1:2000); HDAC4(GeneTex, GTX50484, 1:2000); Smad2/3(Cell Signaling Technology 5678S, 1:1000), p-Smad2/3 (Cell Signaling Technology 8828S, 1:1000), RUNX2(Cell Signaling Technology 12556S, 1:1000); H3-Ac (Millipore #06-599, 1:1000), H4-Ac (Millipore #06-598, 1:1000). The secondary antibodies used were: goat anti-mouse and goat anti-rabbit (ZSGB-BIO, Beijing, China, 1:2000).
Recombinant Human TGF-β1 was purchased from R&D Systems.