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Objective: To observe the collagen-induced arthritis (collagen-induced arthritis, CIA) synovial cell β-arrestin1, β-arrestin2 expression and TGP (total glucosides of paeony, TGP) administered in vivo, Paeoniflorin (paeoniflorin , Pae) in vitro after administration of β-arrestin1, β-arrestin2 expression changes; human synovial cells were treated with IL-1β stimulation of β-arrestin1, β-arrestin2 expression changes and Pae role; and to further explore β-arrestin2 and extracellular signal-regulated kinase (extracellular signal-regulated kinase, ERK) relationship to the depth of rheumatoid arthritis (rheumatoid arthritis, RA) pathogenesis and treatment of RA TGP pharmacological mechanism. Methods: CC Ⅱ toes, tail root and intradermal multi-point injection and a week after the booster immunization France CIA rat model, TGP (25,50,100 mg · kg-1 · d-1), TWP ( glucosides of tripterygium wifordii, GTW) (40 mg · kg-1 · d-1) ig (intragastric injection administration, ig) (d14 ~ d28), detection of paw swelling; serum C Ⅱ antibody concentration; MTT method detection of synovial cell proliferation; Western blot assay β-arrestins expression in the disease process variations and different doses of TGP in vivo administration of β-arrestins expression. d28 CIA synovial tissue were cultured, take the 3rd generation of synovial cells Pae (10-5,10-6,10-7,10-8,10-9 mol · L-1) in vitro administration, Western blot analysis of different concentrations Pae administration of β-arrestins in vitro expression. Collection of patients undergoing hip arthroplasty synovial tissue, after informed consent, the use of tissue culture of human synovial cells were cultured synovial cells taken three generations, IL-1β stimulation observed at different time points after stimulation with cAMP, PKA, Changes of β-arrestins Pae role; using ERK inhibitor U0126 observation of the β-arrestin2 expression. Results: 1. CIA synovial tissue of β-arrestin1 and β-arrestin2 expression of CIA rats after sensitization, synovial tissue expression levels of β-arrestin1 was no significant change; while β-arrestin2 expression levels in sensitized significantly increased after d14, d28 expression reached a peak, d35 level is still higher than the normal group, and consistent with the development of inflammation, suggesting that β-arrestin2 in the pathogenesis of RA has played an important role in the process. (2) human synovial cells after stimulation with IL-1β at different time points cAMP, PKA, β-arrestin2 changes take three generations synovial cells in IL-1β stimulated 2h, 4h, 8h, 16h, 24h, detect cAMP levels, PKA activity, 4h cAMP significantly increased when, 8h and then decreased, 24h was significantly reduced; PKA activity was significantly elevated at 4h, and then gradually returned to normal levels. Take three generations of synovial cells in IL-1β stimulated 0,3 h, 6h, 12h, 24h, 48h detection?-Arrestin2 expression, the expression began to increase at 12h, 24h peak. Take three generations synovial cells by adding U0126 (25μM, 50μM) or solvent cultured 2h after adding IL-1β (final concentration of 10ng · ml-1) for 24 hours, U0126 (25μM, 50μM) can significantly reduce the β-arrestin2 expression. 3. TGP therapeutic effect on rat CIA and TGP and Pae on β-arrestins expression TGP (25,50,100 mg · kg-1, ig, d14 ~ d28) can significantly reduce the CIA rats polyarthritis ratings . CIA serum CII antibody levels were significantly increased, TGP (25,50,100 mg · kg-1, ig, d14 ~ d28) can significantly reduce the CII antibody concentration. CIA rats showed obvious pathological articular inflammation, TGP three dose groups may be different degrees of inhibition of arthritis. CIA rat fibroblast-like synovial cells (fibroblast-like synoviocytes, FLS) proliferative response enhancement, TGP (50,100 mg · kg-1, ig, d14 ~ d28) can inhibit the proliferative response of FLS. Take the TGP sensitized d28 CIA group and the treatment group synovial tissue, Western blot assay β-arrestin1 and β-arrestin2 expression levels were found TGP (50,100 mg · kg-1, ig, d14 ~ d28) can significantly reduce inflammation after β-arrestin2 expression of β-arrestin1 expression levels had no significant effect. d28 CIA rats were killed after cultured synovial cells, synovial cells taken three generations detected β-arrestin2 expression was significantly increased, Pae (10-5,10-6,10-7 mol · L-1) in vitro administration can significantly reduce the expression of β-arrestin2. Human synovial cells were subcultured synovial cells taken after three generations, adding Pae (10-5,10-6,10-7,10-8,10-9 mol · L-1) after adding the role of IL-1β stimulation 24h, compared with the untreated group, Pae (10-5,10-6,10-7 mol · L-1) can significantly reduce the expression of β-arrestin2. Conclusions: 1. CIA synovial tissue of β-arrestin2 expression levels increased with the duration of the development, and consistent with the development of inflammation. (2) human synovial cells stimulated by IL-1β, β-arrestin2 upregulation, which may be caused by G protein-coupled receptor signal transduction excessive desensitization, one of the mechanisms of synovial cell proliferation; ERK inhibitor U0126 (25μM, 50μM) to reduce the IL-1β stimulated the expression of β-arrestin2, which may facilitate the activation of ERK expression of β-arrestin2. 3. TGP multiple CIA rats significantly reduced arthritis index, lower CII antibody concentrations, inhibit proliferation of synovial cells, synovial tissue could down abnormally elevated expression of β-arrestin2; paeoniflorin vitro drug also can decrease the CIA rats and IL-1β stimulated synovial cells posterity expression of β-arrestin2. Reduced the expression of β-arrestin2 improve G protein-coupled receptor signaling transduction inhibition TGP excessive desensitization may be one of the mechanisms of synovial cell proliferation.
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