Silencing long non-coding RNA NEAT1 attenuates rheumatoid arthritis via the MAPK/ERK signalling pathway by downregulating microRNA-129 and microRNA-204

Abstract

ABSTRACT The participation of long noncoding RNAs (lncRNAs) and microRNAs (miRs) in the progression of rheumatoid arthritis (RA) is a key area of investigation. The current study aimed to investigate the action of lncRNA nuclear paraspeckle assembly transcript 1 (NEAT1) in fibroblast-like synoviocyte (FLS) proliferation and synovitis in RA. A rat model of RA was established. LncRNA NEAT1 expression in the synovial tissues of patients with RA and FLSs from the RA rat model was determined using RT-qPCR. Next, dual luciferase reporter gene assay was applied to investigate the relationship between miR-129/204 and mitogen-activated protein kinase (MAPK)/extracellular regulated protein kinase (ERK). A putative binding relationship between miR-204 and lncRNA NEAT1 was evaluated by RIP assay, and miR-129 promoter methylation was determined using MSP. After the expression of lncRNA NEAT1, miR-129 or miR-204 was altered in FLSs, the extent of ERK1/2 phosphorylation was assessed. In addition, FLS synovitis and proliferation were determined by ELISA and EdU assay, respectively. In RA rats, lncRNA NEAT1 was silenced and miR-129/miR-204 was overexpressed to explore their roles in vivo. LncRNA NEAT1 was upregulated, while miR-129 and miR-204 were downregulated in RA synovial tissues and FLSs. MAPK1 was target gene of both miR-129 and miR-204. LncRNA NEAT1 bound to miR-204 and promoted miR-129 promoter methylation. Silencing lncRNA NEAT1 or overexpressing miR-129/miR-204 enhanced miR-129/miR-204 expression, but reduced the extent of ERK1/2 phosphorylation, proliferation of FLSs, and synovitis in RA. Collectively, silencing lncRNA NEAT1 promoted miR-129 and miR-204 to inhibit the MAPK/ERK signalling pathway, reducing FLS synovitis in RA. Abbreviations: ACR: American College of Rheumatology; ELISA: Enzyme-linked immunosorbent assay; ERK: extracellular signal-regulated kinase; FLS: fibroblast-like synoviocyte; GADPH: glyceraldehyde-3-phosphate dehydrogenase; HRP: horseradish peroxidase; IFA: Incomplete Freund’s Adjuvant; lncRNAs: long noncoding RNAs; MSP: Methylation-specific PCR; NC: negative control; NEAT1: nuclear paraspeckle assembly transcript 1; OD: optical density; RA: rheumatoid arthritis; RIPA: Radio Immunoprecipitation Assay; RLU: relative light units; RT-qPCR: reverse transcription quantitative polymerase chain reaction; UTR: untranslated region.