Anti fap

The antibodies used in this study were anti-GAPDH (60004-1-Ig, Proteintech, China), anti-α-SMA (14395-1-AP, Proteintech, China), anti-FAP (A11572, Abclonal, China), anti-VIM (60330-1-Ig, Proteintech, China), and anti-KI67 (27309-1-AP, Proteintech, China). flow cytometry Kit (Multi Sciences, China), CCK8 reagent (Dojindo, Japan), MTT Reagent (Sigma, Germany), BCA1-1KT kit (Sigma, Germany), RIPA lysate (Beyotime, China), and inhibitor of exosome GW4869 (MCE, Shanghai) were purchased from Baitengruida Biotechnology. Optical microscope (Nikon, Japan), HT7800 Transmission Electron microscope (Hitachi, Japan), microplate reader (Bio-RAD, USA), and flow cytometry (BIO-RAD, USA) were provided by the central lab of Renmin Hospital of Wuhan University. The Elekta Infinity™ linear accelerator (Elekta, Sweden) was provided by Renmin Hospital of Wuhan University. Four-week-old female BALB/c nude mice were purchased from Weitonglihua Experimental Animal Technology.

To assess RV histological changes under pressure overload, ventricle tissue was collected from PAB or sham rats at day 3, week 1, week 2, week 4, week 8 post-surgery, respectively. After collection, the heart from each animal were flushed with saline, and then fixed in 10% formaldehyde for 24 h. After paraffin embedding, sections were cut at 5 μm thickness for hematoxylin and eosin (HE) or Masson’s trichrome staining. Cardiomyocyte hypertrophy was assessed by measuring the cross-sectional area of cardiomyocytes, twenty cardiomyocytes were randomly selected from each RV to calculate the mean cross-sectional area of cardiomyocytes. And RV fibrosis was quantitated via calculating the ratios of the fibrosis area to the total area of RV free wall based on Masson’s trichrome staining. The immunofluorescence (IF) staining was performed as previously described [15 (link)]. Briefly, after deparaffinization and antigen retrieval, heart sections were blocked with 5% BSA for 1 h at room temperature and then incubated with primary antibody at 4 °C overnight, followed with species-specific Alexa Fluor-coupled secondary antibodies at room temperature for 2 h. The information of primary antibodies is displayed as follows: anti-CD31 (1: 500, GB12063, Servicebio, China), anti-FAP (1: 100, A6349, ABclonal, China). Image analysis was performed using Image J software (NIH, Bethesda, MD, USA).