Background MicroRNAs (miRNAs) play as regulators of rheumatoid arthritis (RA) and other diseases. MicroRNA is edited by the adenosine deaminase double-stranded RNA (ADAR) as well as mRNA and other noncoding RNA after transcription. A to I RNA editing is occurred limited variety of miRNAs. MicroRNA-381 (MiR-381) is one of limited miRNAs and was down regulated in synovium derived from patients with RA. However, the function of miR-381 and A to I RNA editing against RA is not clear.
Objectives To clarify a role of miR-381 and A to I RNA editing in synovium derived from RA or OA.
Methods 1) RNA was purified from the synovium derive from patients with RA (n=6) or OA (n=6) and analyzed with next generation sequencing (NGS). MiRNA editing was annotated containing 2 nucleotide mismatch. 2) The expression of wild type (WT) and edited miR-381 of synovium derived from RA (n=10) was compared to OA (n=10) with real time PCR. 3) Fibroblast-like synoviocytes (FLS) purified from RA synovium (n=3) were treated TNFα and IL-1β to clear miR-381 expression mechanism. 4) FLS were transfected with WT or edited miRNA expression vectors. These migration, proliferation and MMP production were analyzed. The target genes was predicted and determined by luciferase assay. 5) Male DBA/1 mice were intraperitoneally injected anti-collagen antibodies (collagen antibody-induced arthritis, CAIA) on day0 and LPS on day3. MiR-381 mimic was injected intra articular space on day0 and 3 after antibody injection. Arthritis was evaluated by arthritis score and histological score.
Results 1) Two types of A to I RNA editing were detected in miR-381 seed sequence using NGS (edit1/edit2). 2) These expression of both WT, edit1 and edit2 miR-381 in RA synovium were lower than OA (p<0.05). 3) The expression of series of miR-381s were decreased by TNFα and IL-1β stimulation. 4) Overexpression of WT and edit1 miR-381 reduced the migration of FLS (p<0.01). Overexpression of edit1 and 2 miR-381 repressed proliferation of FLS (p<0.01), which may be caused by reducing CUL3 mRNA expression. CUL3 was regulator of cell cycle and was a direct target gene of edited miR-381s by luciferase assay. All types of miR-381 reduced MMP13 expression (p<0.05). 5) Treatment with miR-381 mimic increased expression of miR-381 in synovium but not in other tissues and significantly reduced arthritis score (scramble vs. miR-381 mimic: 11.5±0.76 vs. 8.4±2.06) and cartilage destruction.
Conclusions These findings suggested that miR-381 was added functions by A to I RNA editing and controls FLS and arthritis.
Disclosure of Interest Y. Tanaka Grant/research support from: Resesarch Fellow of Japan Society for the Promotion of Science, S. Takada Grant/research support from: CREST, Japan Science and Technology Agency, H. Iizasa: None declared, A. Hatzigeorgiou: None declared, S.-I. Miyazawa: None declared, T. Furumatsu: None declared, K. Nishida: None declared, H. Asahara Grant/research support from: CREST, Japan Science and Technology Agency