Effects Of Prostaglandin E₂ Receptors And G Protein Coupled Signal Transduction On Fibroblast-like Synoviocytes Of Rats With Collagen-induced Arthritis By RIL-1α And The Effects Of Total Glucosides Of Paeony

Rheumatoid arthritis (RA) is a common chronic autoimmune and destructive arthropathy that cannot be cured. The chronic nature of this disease results in a progressive joint destruction, which lead to personal, social, and economic costs. RA is characterized by the persistent synovial inflammation, destructive of bone and cartilage, and numerous systemic manifestations. In particular, the higher secretion of synovial cells activated by proinflammatory factors such as interleukin-1(IL-1), tumor necrosis factor-α(TNF-α) and prostaglandin E2 (PGE2) are thought to be a crucial process in the destruction of cartilaginous and bony tissues in RA joints. Up to date, the etiology of RA in not fully understood. Although there are a few anti-rheumatic drugs of treating RA, their persistence and side effects call for new and nature drugs.Total glucosides of paeony (TGP) is an active compound, extracted from roots of paeonia lactiflora Pall, a Chinese traditional herbal medicine (CTM) , have been recognized as the valuable traditional herbs used in treatment for RA. Chemical constituents of TGP contains paeoniflorin (more than 90%, PF), albiflorin, hydroxy-paeoniflorin, paeonin and benzoylpaeoniflorin et al. However, it is unclear whether TGP exerts its effect on the chronic autoimmune diseases such as RA. In this article, we show that the effects of TGP on the secondary inflammatory and synoviocytes proliferation in rat CIA in vivo. In addition, we examine recombinant rat IL-1α(rIL-1α)-stimulated cytokines level and expression of EP4 receptor protein of fibroblast-like synoviocytes (FLS) in CIA rats in vitro, and TGP inhibit synoviocytes proliferation and cytokines level may be through modulating PGE2-EP-G protein-cAMP signal transduction pathway.OBJECTIVETo clarify the effects and mechanisms of total glucosides of paeony (TGP), an effective compound of Chinese traditional herbal medicine (CTM), on functions and activities of FLS. In this article, we examine the effects of cytokines level such as PGE2,TNF-αand cAMP by rIL-1αand explore the therapeutic mechanisms of TGP may be through modulating G-protein coupling signal transduction pathway.METHODSThe model of CIA rats was induced by injection of chicken typeⅡcollagen(CⅡ) in Freund's completed adjuvant. Rats were divided into six guoups and were administrated intragastrically TGP (25, 50, 100 mg·kg-1, d14～d28) and glucosides of triterygium wifordii (GTW) (40 mg·kg-1, d14～d28) as control groups. And for the groups of normal and CIA group, rats were given an equal volume of vehicle (CMC-Na) at the same time.Secondary paw swelling of CIA rats was measured with volume meter. The level of anti-CⅡIgG antibody in serum was examined by enzyme linked immunosorbent assay (ELISA). Synoviocytes proliferation was determined by MTT assay. PGE2,TNF-αand cAMP levle by synoviocytes were measured by radioimmunoassay (RIA). The expression of EP4 protein and Gαi2 protein were detected by Western blot analysis.RESULTS1.The effects of TGP on inhibiting secondary paw swelling,synoviocytes proliferation and IL-1,PGE2 and TNF-αlevel by synoviocytes of CIA ratsNoninjected hind paw volume was pre-measured before the first injection (d0), and then measured (Δml) at 4-day intervals. The noninjected paw swelling on d14 was significantly increased in CIA rats compared with normal rats. The peak incidence occured on d22 after immunization. Treatment with TGP (25, 50, 100 mg·kg-1, d14～d28) diminished dramatically the secondary paw swelling. In vivo TGP (25, 50, 100 mg·kg-1, d14～d28) significantly inhibited proliferative synoviocytes and reduced the level of IL-1,PGE2 and TNF-α.2. The effects of TGP on inhibiting PGE2 and TNF-αlevel of FLS stimulated by rIL-1αin vitroIn RA,activated synoviocytes excrete many proinflammatory cytokines such as IL-1, TNF-αand PGE2. So we examined the other cytokines level such as PGE2 and TNF-αof FLS stimulated by rIL-1αin vitro. TGP (2.5, 12.5, 62.5, 125, 250μg·ml-1) inhibited level of PGE2 and TNF-αin a dose-dependent manner. It suggested that TGP regulated FLS functions and activities by decreasing its secretion action.3. The effects of TGP on modulating PGE2-EP-G protein-cAMP signal transduction pathway, which may be the most mechanisms of TGP preventing RAMany studies showed that EP4 is involved in the pathophysiology of RA. In the present study, TGP elevating EP4 protein expression,decreasing Gαi2 protein expression,regulating cAMP level and then inhibiting synoviocytes proliferation and cytokines level. Combining with the above results, we demonstrated that TGP inhibited synoviocytes action and decreased inflammatory cytokines by modulating PGE2-EP- G-protein-cAMP signal transduction pathway.CONCLUSIONS1. TGP has an improvement effects on paw swelling in CIA rats;TGP decreases PGE2,TNF-αand IL-1 level and inhibites synoviocytes proliferation.2. TGP inhibits the level of PGE2 and TNF-αof FLS by rIL-1α. It is important characteristic effect of TGP inhibiting accentuate secretion of synoviocytes.3. TGP inhibits synoviocytes action may through modulating PGE2-EP-G protein-cAMP signal transduction pathway.