ALKBH5 Regulates Apoptosis of Rheumatoid Arthritis Fibroblast-like Synoviocytes by Modulating miR-181b-5p Maturation via m6A Demethylation.

OBJECTIVE

To investigate the role of ALKBH5, a member of the AlkB family of Fe(II)/α-KG-dependent dioxygenases that functions as an RNA demethylase removing N6-methyladenosine (m6A), in regulating apoptosis of rheumatoid arthritis fibroblast-like synoviocytes (RA-FLS), with a focus on miR-181b-5p maturation.

METHODS

RA-FLS were isolated and analyzed for ALKBH5, pre-miR-181b-1, and miR-181b-5p expression using qRT-PCR. ALKBH5 protein levels were assessed by Western blotting. Cell proliferation and apoptosis were evaluated using MTT assay and flow cytometry, respectively. m6A modification on pre-miR-181b-1 was measured via MeRIP, and its binding to DGCR8 was assessed using co-immunoprecipitation.

RESULTS

ALKBH5 expression was significantly downregulated in RA-FLS. ALKBH5 overexpression inhibited proliferation and promoted apoptosis, while its knockdown had the opposite effect. ALKBH5 decreased m6A modification of pre-miR-181b-1, thereby increasing levels of pre-miR-181b-1 and mature miR-181b-5p. Inhibition of miR-181b-5p attenuated the effects of ALKBH5.

CONCLUSION

Overexpression of ALKBH5 promotes apoptosis and inhibits proliferation in RA-FLS by demethylating m6A on pre-miR-181b-1, thereby enhancing miR-181b-5p maturation. These findings suggest a novel therapeutic target for rheumatoid arthritis.