| Rheumatoid arthritis (RA) is a chronic autoimmune disease which characterized by chronic inflammation of the synovial tissues in multiple joints that leads to joint destruction. The pathologic characterizes of RA are chronic joint inflammation, inflammatiory cell infiltrate, cartilage and osseous tissue destruction, accompany by neovascularity hyperplasy and system inflammation, ultimum lead to joint deform and functional incapacitation. Although there are a lot of anti-inflammatory drugs showing effectives on treating RA, they do not alter the fundamental mechanisms of this disease procession or its progression. And their severe side effects call for new and more effective natural drugs.Chaenomeles speciosa (sweet) Nakai is one of the valuable traditional herbs used in treatment for RA with a long history in traditional Chinese medicine. Glucosides of Chaenomeles speciosa (GCS), extracted from the fructus of Chaenomeles speciosa, is an active compound. Catechin (Cat) is a constituent of Chaenomeles speciosa. Our previous studies showed that GCS has effects on inhibiting the inflammation of rat with carrageenan-induced and of rat with adjuvant arthritis (AA). GCS decreased mRNA expression of Gαi, increased mRNA expression Gαs of synoviocyte, increased cAMP level of synoviocytes so as to decrease the proliferation and secretion of synoviocytes. The effects of GCS on rat AA due to affecting G protein–AC–cAMP signal transduction pathway of synoviocytes. However, it is unknown whether or how Cat exerts its effect on the chronic autoimmune diseases such as RA. The present study was therefore designed to investigate the therapeutic effects of GCS and Cat on rat adjuvant arthritis (AA) and its relative mechanisms by which GCS and Cat affects the synovial cells function of AA, especially influence desensitization of prostaglandin E2 receptors in synoviocytes of rats with adjuvant arthritis.AIMAccording to the changes that secondary paw swelling, pain response, polyarthritis, and histological morphological of rats AA, this study tends to clarify the therapeutic effects of GCS and Cat on AA rats. Meanwhile, our study aimed to investigate the correlation of synoviocyte proliferation with level of intracellular cAMP. To confirm the actions of GCS and Cat, this study sought to indentify relationships between the effects of GCS and Cat on rats AA and its relative mechanisms by which GCS and Cat influence desensitization of prostaglandin E2 receptors in synoviocytes of rats AA.METHODSFreund's complete adjuvant (FCA) was used to induce AA in rats. Therapeutic treatment of interagastric administration GCS and Cat (30, 60, 120 mg·kg-1·d-1, d 17~d 24) were given after immunization. GTW (40mg·kg-1) were given as positive control. Secondary paw swelling of AA rats was measured with volume meter. The pain response and polyarthritis index were scored. Histopathological change of joint was observed by light microscope. Synovial cells from rat knees were excised and dispersed with sequential incubation of collagenase typeⅡand trypsin. The proliferation of fibroblast-like synoviocytes was assayed using the 3-(4, 5-dimethylthiazol-2-thiazolyl) -2,5-diphenyl-2H tetrazolium bromide (MTT) assay. Activity of IL-1 was measured by ConA-induced thymocyte proliferation of mice assay. PGE2, cAMP, TNFαwere measured by radioimmunoassay. The expression of EP2,GRK2,β-arrestin1 were detected by Western blot analysis.RESULTS1. GCS and Cat had therapeutic effects on AA ratsThe AA model in rats was induced by FCA. There was a marked secondary inflammatory response in this model similar characteristic to RA, which accompanied with paw edema, pain, polyarthritis, and the development of inflammatory lesions. The histological morphological of rats knee joints examination showed synovial tissue encompass infiltration by inflammatory cells, synovial lining hyperplasia, neoangiogenesis, pannus formation, severe articular inflammation and destruction of the cartilage, proliferation of collagen fribrils, the quantity of bone marrow cells decreased, and fatty changes in the synovial cells.Therapeutic treatment of GCS (30, 60, 120 mg·kg-1·d-1, d 17~d 24) and Cat ( 60, 120 mg·kg-1·d-1, d 17~d 24) suppressed significantly the secondary paw swelling and pain response, as well as down-regulated the index of polyarthritis in AA rats. The articular histopathological changes were also improved. This suggested that GCS and Cat had therapeutic effects on AA rats.2. GCS and Cat inhibited secretion of proimflammatory cytokines. GCS and Cat inhibited proliferation of fibroblast-like synoviocytes and increased the level of cAMP in synoviocytesGCS and Cat (30,60,120mg/kg, ig, d17-24) significantly reduced the elevated PGE2,IL-1 and TNFαof synoviocytes supernatants. GCS and Cat (30, 60,120mg /kg, ig, d17-24) significantly inhibited proliferation of fibroblast-like synoviocytes and increased the level of cAMP in synoviocytes. The experimental in vitro had the same results. The result of correlation analysis demonstrated that GCS and Cat inhibit the fibroblast-like synoviocytes hyperplasia through increasing the level of cAMP in synoviocytes. GCS (12.5~100μg·ml-1) and Cat (12.5~100μg·ml-1) inhibited the production of PGE2 in synoviocytes induced by IL-1α.3. GCS and Cat improved the expression of EP2The result of Western blot analysis demonstrated that GCS and Cat (60,120mg /kg, ig, d17-24) significantly improved the expression of EP2. These result suggested that recovering the EP-G protein-cAMP signal normal transduction is one of the most important mechanisims of GCS and Cat in preventing RA.4. GCS and Cat increased the expression of GRK2 andβ-arrestin1 in membrane The result of Western blot analysis demonstrated that GCS and Cat (50,100μg/ml) in vitro significantly increased the expression of GRK2 andβ-arrestin1 in membrane. These result suggested that GCS and Cat might recover the EP-G protein-cAMP signal normal transduction by regulating the expression and desensitization of EP2.CONCLUSIONS1. GCS and Cat had therapeutic effects on paw swelling, pain, polyarthritic symptoms and articular histopathological changes in rats with AA.2. GCS and Cat decreased the exorbitant secretion function of synoviocytes by inhibited secretion of proimflammatory cytokines. GCS and Cat inhibited proliferation of fibroblast-like synoviocytes by increased the level of cAMP in synoviocytes.3. GCS and Cat improved the expression of EP2. IL-1αmay influence the desensitization of EP2 by inhibiting the translocation of GRK2 andβ-arrestin1 to membrane. GCS and Cat influence the desensitization of EP2 by increasing the expression of GRK2 andβ-arrestin1 in membrane. These result suggested that GCS and Cat might prevent RA by recovering the EP-G protein-cAMP signal normal transduction.
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