Macrophage polarization controlled by fibroblast crosstalk and targeted by medications in rheumatoid arthritis

In this study, we performed RNAseq on human macropahges following a tumor necrosis factor response with and without human fibroblasts, the representative stromal cell type in the synovial lining of joints that become activated during inflammatory arthritis. Our RNAseq analysis reveals a crosstalk pathway between macrophages and fibroblasts that emerges as a result of co-culture that we call the MTF polarization phenotype. We test multiple drugs with RA indications and candidate therapies and report that the genes that shaped by the MTF phenotype are opposed in their expression by known therapies. Our RA joint inflammation model data provides insights into the cellular crosstalk mechanisms between macrophages and fibroblasts as well additional knowledge into RA drug mechanisms of action. Human CD14+ MCSF-differentiated macrophages were cultured with or without synovial fibroblasts in transwell chambers. Samples include an unstimulated condition, TNF condition (added at Day 0 and harvested at Day 1), and drug treatments including Auronfin (Aur), Dexamethasone (Dex), Tofacatinib (Tofa), Hydroxychloroquine (Hydrox), Methotrexate (Meth), Diclofenac (Diclo), Sulfasalazine (Sulf), AG ( AG 1478), Naproxen (Naprox), TNF was added at Day 0, macrophages were harvested at Day 1. Macrophage RNA was purified using RNeasy mini kit (Qiagen). Tru-seq sample preparation kits (Illumina) were used to purify poly-A transcripts and generate libraries with multiplexed barcode adaptors. All samples passed quality control on a Bioanalyzer 2100 (Agilent). Single-end reads (50 x 2 cycles per sample) were obtained on an Illumina HiSeq 4000. The STAR program was used to align the reads to the UCSC Hg19 human reference genome and reads were counted by the HTSeq program. Additional coculture samples [GSM2527917-GSM2527920] include replicates of the previous study [GSE57723] following the treatment protocols described in GSE57723 (PMID: 25057003). The major distinction of these samples and the drug samples is the TNF condition is over 2 days and the RNA-seq libraries were paired end. The 'coculture_htseq_counts.txt' contains processed data for the additional samples.