Mir-34a In Rheumatoid Arthritis: Characterisation Of Elevated Synovial Expression and Association With Treatment Resistance

Background/Purpose: Cells of the monocyte/macrophage lineage are critical to RA pathogenesis: unravelling mechanisms underlying macrophage inflammatory gene expression should elucidate novel disease-associated pathways and thereby biomarkers and therapeutic targets. MicroRNA (miR) comprise small, non-coding RNA species that are key regulators of mammalian gene expression. We sought to define the expression and regulation of novel miR species that regulate RA macrophage biology. Methods: Blood (PB; healthy controls and RA patients with defined treatment characteristics) or synovial fluid (SF; RA) CD14+ cells were purified using histopaque centrifugation and autoMACS bead separation. CD14+ cells were matured with M-CSF or GM-CSF for up to 7 days, or stimulated with various TLR ligands. For T cell -macrophage interaction, CD4+ T cells were cultured with TNF, IL-2, IL-6, prior to 24hr co-culture with M-CSF-matured CD14+ cells. miR expression and copy number was variously quantified in cells or tissues via TLDA, validatory qPCR and in situ hybridisation with specific primers and DIG or FITC labelled probes, respectively. Results: Prior TLDA elicited several dysregulated miRs in RA SF compared to PB CD14+ cells, including miR-34a. Fold change and copy number PCR assays confirmed elevated miR-34a in RA SF cells (n=10; p Conclusion: miR-34a expression is elevated in RA SF macrophages and in PB monocytes of treatment resistant RA patients where it correlates with clinical disease activity measures. miR-34a is upregulated by the maturation rich synovial microenvironment and by interactions with a activated T cells. Putative miR-34a molecular targets include NOTCH1 rendering it a plausible novel immune regulator in synovitis.