[Effect of genistein on MAPK signal pathway in the collagen-induced arthritis fibroblast-like synoviocytes].

OBJECTIVE: To study the effect of genistein (Gen) on MAPK signal pathway in the CIA rat fibroblast-like synoviocytes (FLS). METHODS: The rat model of collagen-induced arthritis (CIA) was established. The cultured FLS of CIA rats were divided using randomized method. The effects of Gen (at the concentration of 50, 100, and 200 micromol/L, respectively) on the proliferation of FLS in CIA rats using methyl thiazolyl tetrazolium (MTT) assay. Effects of Gen (at the concentration of 50, 100, and 200 pmol/L, respectively) on the expressions of extracellular signal-regulated kinase (ERK) and phosphorylated extracellular signal-regulated kinase (p-ERK) in the FLS of CIA rats were detected. RESULTS: Gen could inhibit the proliferation of FLS in CIA rats. The FLS proliferation in the high dose Gen group at 72 h was only 1.10+/-0.04, significantly lower than that in the model group (2.12+/-0.03, P<0.01). Besides, after Gen's action on FLS, the expression of p-ERK was down-regulated. It was only 0.34+/-0.02 in the high dose Gen group, significantly lower than that in the model group (2.68+/-0.14, P<0.01). There was no change in the expression of ERK (P>0.05). CONCLUSIONS: Gen could inhibit the proliferation of FLS in CIA rats. Its mechanism of action was mainly correlated to down-regulating the tyrosine kinase of MAPK signal transduction pathway and inhibiting phosphorylation of ERK.

[Anti-angiogenic effects of genistein on synovium in a rat model of type II collagen-induced arthritis].

OBJECTIVE: To explore the anti-angiogenic effects of genistein on synovium in a rat model of type II collagen-induced arthritis (CIA). METHODS: Forty SD rats were randomly divided into normal group, model group, genistein group, methotrexate (MTX) group and Gen plus MTX group with 8 rats in each group. Arthritis in rats was induced by subcutaneous injection of type II collagen combined with complete Freund's adjuvant (CFA). On the second day after the injection, 1 mL of suspension liquid of genistein (30 mg/kg body weight, once daily) and MTX (0.2 mg/kg body weight, once a week) were administered by oral gavage respectively. The rats in normal group and model group were administered with normal saline in the same volume. Synovium of knee joints and peripheral serum were collected from the CIA rats. Microvessel density in synovium (MVD) was detected by immunohistochemical method and serum vascular endothelial growth factor (VEGF) and matrix metallopeptidase (MMP)-1, 2 and 9 levels were detected by using Western blotting. RESULTS: Arthritis index score, paw volume of rats in the model group were significantly higher than those in the normal group (P<0.05), which suggested that a model of CIA induced by injection of type II collagen and CFA was successfully constructed. The arthritis index scores of rats in the treatment groups were decreased compared with the model group. The results of Western blotting showed that genistein obviously attenuate the levels of VEGF and MMP-1, 2 and 9 in serum (P<0.05). Immunohistochemical method showed that MVDs in the treatment groups were reduced as compared with the model group. CONCLUSION: The expressions of VEGF and MMP-1, 2 and 9 are related to the synovial pannus formation in CIA rats. The anti-angiogenic activity of genistein may correlate to its inhibitory effect on the expressions of VEGF and MMP-1, 2 and 9 in serum of CIA rats; genistein plus MTX are superior to single agents in treating rheumatoid arthritis.

[Effects of genistein on interleukin-1beta and tumor necrosis factor-alpha secreted by fibroblast-like synoviocytes isolated from type II collagen-induced arthritis rats].

OBJECTIVE: To investigate the effects of genistein (Gen) on interleukin-1beta (IL-1beta) and tumor necrosis factor-alpha (TNF-alpha) secreted by fibroblast-like synoviocytes (FLSs) of rats with type II collagen-induced arthritis (CIA). METHODS: Type II collagen was injected to induce arthritis in rats, and arthritis index was applied to evaluate whether the arthritis was induced successfully. The primary FLSs were separated from synovial membranes by using type II collagenase digestion. Then the expression of vascular cell adhesion molecule-1 (VCAM-1) was estimated by flow cytometry (FCM). After addition of different concentrations of Gen into the FLSs, IL-1beta and TNF-alpha contents in the supernatants were measured by enzyme-linked immunosorbent assay. RESULTS: Three days after collagen injection, the rats started to develop arthrocele and the arthritis index increased gradually. The arthritis index of CIA group was significantly higher than that of the control group. The expression of VCAM-1 was up to 85.5% in the 4th generation FLSs, indicating that most part of the cultured cells were type-B synoviocytes. After administration of different concentrations of Gen (100, 200, 400 micromol/L), the contents of IL-1beta and TNF-alpha in supernatants of FLSs were decreased dose-dependently. CONCLUSION: Genistein can suppress the secretion of TNF-alpha and IL-1beta in FLSs dose-dependently, which may be one of the mechanisms for genistein in inhibiting the arthromeningitis of CIA rats.