Profiling Synovial Cell Transcriptomes

We assayed 384 fibroblasts from four donors, two with OA and two with RA. For each donor, we collected fresh synovial tissue, isolated synovial cells by enzymatic digestion, and stained with antibodies against CD45, CD235a, CD31, CD146, PDPN, CD34, THY1, and CDH11. We sorted 96 single CD45^–CD235a^–CD31^–PDPN⁺ cells by FACSAria Fusion (BD), and assayed mRNA expression with the Smart-Seq2 protocol^{37 (link)}. Single-cell libraries were also prepared with the same protocol, and we aimed to sequence to a depth of 200K–12M reads per library^{38 (link)}. On average, we sequenced 5.4M fragments and detected 8,153 genes per cell with at least 1 TPM. We discarded 47 cells (12%) with fewer than 5,000 genes detected from further analysis.