Hum icell s010

Human Primary Synovial Fibroblasts Cells (HUM-iCell-s010) and Human Primary Rheumatoid Arthritis Synovial Fibroblasts Cells (HUM-iCell-s010RA) were obtained from iCell Bioscience (Shanghai, China). The two cell lines were grown in Dulbecco′s Modified Eagle′s Medium (DMEM) (Thermo Fisher Scientific Inc., Waltham, MA, USA) supplemented with 20% fetal bovine serum (FBS) and 1% Penicillin-Streptomycin solution (Thermo) at 37 °C with 5% CO₂. Cells were expanded up to 3 passages for further use.

Synovial tissue samples were obtained from RA patients who had undergone joint replacement surgery. Briefly, the samples were cut into small fragments and cultured in Dulbecco's modified Eagle medium (DMEM) medium containing penicillin (100 U/ml) and streptomycin (0.1 mg/ml). The immortalized cell line of RA‐FLS was obtained by using lentivirus vector‐mediated SV40+ T antigen transfection.¹⁷ The second generation of the cells was used for identification, and the level of Vimentin protein was identified by immunofluorescence (IF) cytochemistry. RA‐FLS were thereafter fixed by paraformaldehyde and incubated with Triton X‐100 for 0.5 h. The primary anti‐vimentin (Bs‐8533R; Bioss) was then added and incubated at 4°C for 8 h. The secondary antibody (1:400, goat anti‐rabbit) was then added to all the sections and incubated for 1 h. After DAPI staining, the tablets were sealed and observed under a fluorescence microscope (BA410E; Motic). Based on this preliminary study, 3–5 generations of cells were selected for the subsequent experiments. FLS for NC (NC‐FLS, HUM‐iCell‐s010; iCell Bioscience Inc.) were cultured in DMEM medium containing 10% FBS (sh30256.01; Hyclone).