RESUMO
The present study aimed to investigate the role of the soluble programmed death1 (sPD-1) protein, which is released by peripheral blood regulatory T cells (Treg) during the progression of rheumatoid arthritis (RA). From October 2012 to May 2014, 82 RA patients (RA group) and 90 healthy volunteers (healthy controls; HC) were recruited. Cluster of differentiation (CD)4, CD25 and forkhead/winged helix transcription factor p3 (Foxp3) and expression of cytotoxic T lymphocyte associated antigen 4 (CTLA-4) and Foxp3 were detected by flow cytometry. Expression of sPD1 in Treg was detected by western blot analysis. Immunosuppressive activity of CD4+CD25 Treg was measured via thiazolyl blue in an MTT assay. ELISA was used to detect interleukin10 (IL10), transforming growth factor beta (TGF-ß), interleukin-4 (IL-4), interferonγ (IFN-γ) and nuclear factor of activated T cells (NFAT). It was observed that in peripheral blood, CD4+CD25-FOXP3+/CD4+ levels were reduced in the RA group (P<0.001), and sPD1 levels were markedly higher (P<0.001), compared with the HC group. Additionally, it was observed that relative sPD1 protein expression in the small interfering RNA (siRNA)-sPD-1 treated group was reduced compared with the untreated and scrambled siRNA groups (all P<0.0001). The mean fluorescence intensity of CTLA-4 and Foxp3 decreased markedly upon transfection with siRNA-sPD-1 (P<0.001). Compared with the normal CD4+CD25 T group, optical density (OD)540 values, IFN-γ/IL-4 concentration ratio and NFAT activity in siRNA untreated and scramble groups reduced significantly (all P<0.001). OD540 value, IFN-γ/IL-4 concentration ratio and NFAT activity in the siRNAsPD1 group were significantly upregulated (all P<0.001). Therefore, sPD-1 may suppress the level of CD4+CD25 Tregs in the peripheral blood of RA patients, and may be involved in a variety of immune processes mediated by CD4+CD25 Tregs.
