Generating SARS-CoV-2 Spike Protein in FLS

Human fibroblast-like synoviocytes (FLS) were grown in complete DMEM (12800017; Thermo Fisher Scientific, Waltham, MA, USA) supplemented with 10% (v/v) fetal bovine serum (FBS) (#16000044; Thermo Fisher Scientific) at 37 °C in an atmosphere of 5% CO₂. For generation of lentivirus carrying SARS-CoV-2 spike protein [23 ], HEK293T cells were transfected with a plasmid encoding SARS-CoV-2 spike protein (pBOB-CAG-SARS-CoV-2-Spike-HA,#141347; Addgene, Watertown, MA, USA) and packaging vectors (pMD2.G, #12259 and psPAX2, #12260; Addgene) using Lipofectamine 2000 (#11668030; Thermo Fisher Scientific). Culture supernatants were harvested at 48 h posttransfection and aliquots of 2 × 10⁵ human FLS were transduced with the generated recombinant lentivirus. At 48 h after lentiviral transduction, cells and supernatant were harvested for qPCR and ELISA, respectively.