MiR9 is increased in Rheumatoid Arthritis Monocytes and Regulates Monocyte Migration

Abstract

PurposeRheumatoid Arthritis (RA) is characterized by inflammation and joint degradation. Monocytes, a key leukocyte in inflammation, migrate from the blood to damaged tissues where they differentiate into macrophages that serve multiple immune functions. MicroRNA (miRNA), small noncoding regions of RNA, act as regulators of gene expression, bind to and degrade mRNA. We have previously shown that miR9 is up regulated in THP1 monocytes and U937 monocytes with TNFα stimulation. Here we hypothesize that miR9 is up regulated in the inflammatory disease RA, and that mir9 regulates monocyte migration.Methods and ResultsMonocytes were isolated from blood of patients with RA (N=3) and normal subjects (N=6). MiRNA was isolated, cDNA was prepared, and qPCR was performed using primers specific to miR9. miR9 was significantly upregulated in RA monocytes by 2.3 fold compared to normal monocytes (P < 0.05). We searched for miR9 gene targets using miRWALK, MiRanda, and NCBI databases and found 8,778 gene targets. Targets were narrowed to biological pathways specific to human monocytes and inflammation, leaving 39 gene targets. Of these 39 targets, suppressor of cytokine signaling 4 (SOCS 4) was chosen for further study because it showed the highest seed length for miRNA binding. An inhibitor and mimic for miR9 were used to assess its ability in modulating SOCS4 expression. Results showed a 0.2 fold reduction in expression with the miR9 mimic, and a negligible change in expression from the miR9 inhibitor. Chemotaxis assays were performed on THP‐1 monocytes transfected with miR9 mimic or inhibitor to measure the effect of miR9 on monocyte migration. THP‐1 cells treated with the miR9 mimic displayed an increase in migration by 40% (n=10 replicates) compared to control cells, whereas the inhibitor showed no change in migration (n=10 replicates).ConclusionOur results suggest miR9 is upregulated in RA monocytes and that miR9 plays a role in monocyte migration by regulating genes such as SOCS4.Support or Funding InformationHewlett‐Mellon Fund for Faculty Development at Albion College