Monoclonal anti yap

BMP2 (R&D, Minnesota, USA, 355-BM-100), DKK1 (BioLegend, California, USA, 759602), 4′,6-diamidino-2-phenylindole (DAPI, Solarbio, Beijing, PRC, C0060), carboxyfluorescein succinimidyl ester (CFSE, eBioscience, California, USA, 65-0850-84) and gallocyanine (Selleck, Texas, USA, S5602) were used. The antibodies used in this study were monoclonal anti-YAP (Cell Signaling Technology, Boston, USA, 14074), anti-pYAP S127 (Cell Signaling Technology, Boston, USA, 13008), anti-actin alpha2, smooth muscle (anti-α-Sma, Servicebio, Wuhan, PRC, GB111364), anti-osteocalcin (anti-OCN, BioByt, Cambridge, UK, orb259644), tetramethylrhodamine isothiocyanate (TRITC)-conjugated anti-rabbit antibody (ABclonal, Wuhan, PRC, AS040), fluorescein 5-isothiocyanate (FITC)-conjugated anti-mouse antibody (ABclonal, Wuhan, PRC, AS001) and anti-β-actin (Sigma, St. Louis, USA, A1978). Tris-HCl, NaCl and other chemicals were purchased from Sigma.

Monoclonal anti-YAP, monoclonal anti-cyclin D1, polyclonal anti-cleaved caspase 3 and polyclonal anti-pYAP antibodies were obtained from Cell Signalling Technology (Danvers, MA). Polyclonal anti-CCN1 (Cyr61), anti-CCN2 (CTGF) and monoclonal anti-GAPDH antibodies were obtained from Santa Cruz Biotechnology (Santa Cruz, CA). Horseradish peroxidase-conjugated goat anti-mouse IgG and goat anti-rabbit IgG were obtained from Bio-Rad Laboratories (Hercules, CA). S1P was obtained from Avanti Polar Lipids (Alabaster, AL). Verteporfin (VP) was obtained from VWR International (Randor, PA). For the VP experiments, cells were treated with 10 μm of VP for 10 min, washed and then treated with 300 nM of S1P for additional 2 h. Cycloheximide (#1041) was obtained from BioVision (Mountain View, CA).