Anti hsp90

Cell or tissue lysates and immunoblot analysis were performed as described previously [11 (link), 12 (link)]. Densitometric analysis was conducted using ImageJ software. The primary antibodies used include: anti-STIP1 (1:200, Boster Biotechnology, Wuhan, China), anti-ALK2 (1:400, Boster Biotechnology, Wuhan, China), anti-SMAD1/5 (1:200, Boster Biotechnology, Wuhan, China), anti-CTSK (1:500, Boster Biotechnology, Wuhan, China), anti-ERK1/2 (1:500, Boster Biotechnology, Wuhan, China), anti-Hsp90 (1:500, Boster Biotechnology, Wuhan, China), anti-Ki67 (1:500, Boster Biotechnology, Wuhan, China) and anti-GAPDH (1:500, Boster Biotechnology, Wuhan, China).

VSMCs were lysed on ice for 30 min with SDS Lysis Buffer. The concentration of acquired protein in cell lysis was measured by BCA protein quantitative kit (Thermo Scientific, Carlsbad, CA). Equal amounts of protein samples were added into the lane of SDS-PAGE gels. After electrophoresis and transmembrane, the membrane was incubated with primary antibodies and following secondary antibodies. Then, the immunoreactive bands were visualized by chemiluminescence (Thermo Scientific, Carlsbad, CA) and analysed by Image J software (Bethesda, MD). β-Actin was used as loading control. The primary antibodies included anti-OPN (1:1000 dilution, Abcam, Cambridge, UK), anti-Runx2 (1:1000 dilution, Abcam, Cambridge, UK), anti-HSF1 (1:500 dilution, Proteintech, Wuhan, China), anti-HSP70 (1:500 dilution, Boster, Wuhan, China), anti-HSP90 (1:500 dilution, Boster, Wuhan, China) and anti-β-actin (1:1000 dilution, Proteintech, Wuhan, China).