Effects Of Recombinant Human Interleukin-1β On Functions Of Peripheral Blood Lymphocytes From Human And The Effects Of Paeoniflorin

OBJECTIVETo clarify the role of interleukin-1 (IL-1) in the pathogenesis of rheumatoid arthritis (RA) and whether paeoniflorin (Pae) exerts its therapeutical effects on RA by modulating the abnormal activation, proliferation, differentiation and immunotolerance of peripheral blood lymphocyte (PBLC) from human. In this article, we examined the effects of recombinant human IL-1β(rhIL-1β) on the functions of human PBLC, and the effects of Pae.METHODSAfter informed consent, the peripheral venous blood were drew from healthy volunteers, and the PBLC was seperated by density gradient centrifugation and further purified by adherence. The effects on human PBLC proliferation, the production of IL-2 and IL-10, the expression of CD25 and the proportion of CD4⁺CD25^high Treg of rhIL-1βin different concentration and time, and the effects of Pae were observed in vitro.The proliferative response of human PBLC was measured by MTT assay. The level of IL-2 and IL-10 in human PBLC culture supernatants were detected by radioimmunoassay (RIA) and enzyme-linked immunsorbent assay (ELISA) respectively. The expression of CD25 and the proportion of CD4⁺CD25^high regulatory T cell (CD4⁺CD25^high Treg) were analysed by flow cytometry (FCM).RESULTS1. The inhibition of Pae on the proliferation of human PBLC promoted by rhIL-1β.The proliferation of human PBLC was promoted by rhIL-1β(0.1,1.0,10.0,100.0μg·L^-1). In vitro Pae (10-8,10^-7,10^-6,10^-5 mol·L^-1) inhibited the human PBLC proliferation treated by rhIL-1β. It demonstrated that Pae could recover the abnormal PBLC proliferation treated by rhIL-1βto normal level.2. The effects of Pae on the level of IL-2 and IL-10 produced by human PBLC treated by rhIL-1β.RhIL-1β(0.1,1.0,10.0,100.0μg·L^-1) increased the level of IL-2 produced by human PBLC, and significantly increased the level of IL-2 after 24 h . In vitro Pae (10-8,10^-7,10^-6,10^-5 mol·L^-1) decreased the level of IL-2 produced by human PBLC treated by rhIL-1β. RhIL-1β(0.1,1.0,10.0,100.0μg·L^-1) decreased the level of IL-10 produced by human PBLC. In vitro Pae (10-8,10^-7,10^-6,10^-5 mol·L^-1) significantly elevated the level of IL-10. It suggested that Pae induce Th0 cell to differentiate into Th2 cell and recover the balance of Th1/Th2 in human PBLC treated by rhIL-1β.3. The effects of Pae on the expression of CD25 on human PBLC and the proportion of CD4⁺CD25^high Treg in human PBLC treated by rhIL-1β.The expression of CD25 on the surface of human PBLC was upregulated by rhIL-1β(0.1,1.0,10.0,100.0μg·L^-1), and it was significantly upregulated after 24 h. In vitro Pae (10^-8,10^-7,10^-6,10^-5 mol·L^-1) significantly downregulated the expression of CD25 of human PBLC treated by rhIL-1β. RhIL-1βdidn't influence the proportion of CD4⁺CD25^high Treg in human PBLC. In vitro Pae (10^-7,10^-6,10^-5 mol·L^-1) significantly elevated the proportion of CD4⁺CD25^high Treg in human PBLC treated by rhIL-1β. It demonstrated that Pae reversed the activation of CD4+CD25- T cell induced by IL-1, single IL-1 could not influence the proportion of CD4⁺CD25^high Treg, and Pae induced immunotolerance to play its role in immunoregulation by enhancing the proportion of CD4⁺CD25^high Treg in human PBLC treated by IL-1.CONCLUSIONS1. Pae reversed the proliferation of human PBLC promoted by rhIL-1β.2. Pae inhibited IL-2 production and enhanced IL-10 production of human PBLC treated by rhIL-1β.3. Pae inhibited the activation of human PBLC, downregulated the expression of CD25 and enhanced the proportion of CD4⁺CD25^high Treg in human PBLC treated by rhIL-1β.