Bs 6050r

Immunofluorescence was performed as previously described. FSTL1 (1:250 dilution, bs-6050R, Bioss), VEGF (1:250 dilution, bs-4572R, Bioss), and vimentin (1:800 dilution, ab128507, Abcam) were used. Fluorescence intensity was analyzed using image software. Quantitative analysis was performed using ImageJ.

HRCECs and HUVECs were fixed in 4% paraformaldehyde in PBS for 10 min at room temperature and then permeabilized with PBS containing 0.1% Triton X-100 for another 10 min. Cells were blocked with bovine serum albumin (BSA) in PBS for 30 min, then incubated with FSTL1 (1:250 dilution, bs-6050R, Bioss), TGFβ1 (1:250 dilution, BA0290, Boster), CTGF (1:250 dilution, BA0752, Boster), VEGF (1:250 dilution, bs-4572R, Bioss), α-SMA (1:250 dilution, bs-10196R, Bioss), COL1A1 (1:250 dilution, BA0325, Boster), and FN (1:250 dilution, BA1771, Boster) antibodies at 4° C overnight. Cells were then washed with PBST and incubated with anti-rabbit secondary antibodies for 1 h at room temperature. The cells were then stained with DAPI for nuclear visualization, and fluorescence intensity was assessed under a microscope by digital analysis. Quantitative analysis was performed using ImageJ.