A preliminary study on the pathogenesis of neutrophil extracellular traps in rheumatoid arthritis

Abstract

Objective To explore the potential effects of neutrophil extracellular traps (NETs) on rheumatoid arthritis synovial fibroblasts (RA-FLSs). Methods The synovial tissues of RA patients were isolated and cultured in vitro. Peripheral blood neutrophils were extracted from healthy volunteers and used to stimulate NETs' formation, following with NETs' extraction. MTS proliferation assay was used to evaluate the effect of NETs on the proliferation of RA-FLSs. QRT-polymerase chain reaction (q-PCR) was used to determine the expression of connective tissue growth factor (CTGF) mRNA in cells treated with NETs-stimulated RA-FLSs for 60 h. The results were processed using paired sample t-test and one-way analysis of variance (ANOVA) . Results The isolated and purified neutrophils could form NETs by in vitro stimulation. The concentration of extracted NETs-DNA was 58.5 ng/μl (1×106 cells). Compared with the control group (0 μl NETs), NETs could promote the proliferation of RA-FLSs. With the increase of NETs' concentration, the proliferation of RA-FLSs was also enhanced (F=99.519, P<0.05). Compared with the control group (0 μl NETs), 10 μl NETs could signi-ficantly promote the proliferation of RA-FLSs (t=-12.226, P<0.01). Pretreatment of NETs with DNase Ⅰ inhi-bited its effect on promoting the proliferation of RA-FLSs (t=-2.376, P=0.049), NETs stimulated the up-re-gulation of CTGF mRNA expression in RA-FLSs [(30.7±0.5), t=12.13, P<0.01]. Conclusion NETs can pro-mote the proliferation of RA-FLSs and stimulate the up-regulation of CTGF mRNA in RA-FLSs in vitro. Key words: Neutrophil extracellular traps; Arthritis, rheumatoid; Rheumatoid arthritis fibroblast-like synoviocytes; Connective tissue growth factor