| Background:Rheumatoid arthritis (RA) is one of the most prevalent rheumatic diseases affecting human health, characterized as joint synovial inflammation, pannus formation, cartilage destruction and bone lesion. Researches on drugs of treating RA stay in unceasingly implementation, which also have the obvious therapeutic effect. Studies have found that some RA patients have obvious extravasated blood symptoms and abnormal blood rheology, and conventional treatment is not very effective to relieve the symptoms.Objective:To detect effects of Tetramethylpyrazine (TMP) and Leflunomide (Lef) combination on the collagen induced arthritis (CIA) rat model and the possible mechanism of action for providing clinical treatment with experimental evidence.Method:After two immunization injection, Ⅱ type CIA rat model were established successfully. Then rats equally randomized into seven groups:the normal group (N group, n=8), the model group (M group, n=8), the Lef group (10g/kg/d, n=8), the low dose of TMP group (TL group,25mg/kg/d, n-8), the high dose of TMP group (TH group,50mg/kg/d, n=8), the TL+Lef group (25+10mg/kg/d, n=8), the TH+Lef group (50+10mg/kg/d, n=8). The drug intervention lasted for4weeks. During the period of the drug treatment, the general situation of rats was observed, rat body weight measured, AI changes judged and ankle swelling degree measured with the vernier caliper. After the end of treatment, the detection of CIA rats peripheral blood rheology and the expression levels of the inflammatory cytokines in serum by enzyme linked immunosorbent method such as TNF-α, IL-1β, IL-6, IL-10, VEGF-C and PAF were performed. Meanwhile, HE staining was executed to observe the histopathology changes of the ankle and immunohistochemical staining method carried out to detect the expression level of VEGF, PAF in local rat ankle joint. All of these were completed to evaluate the effect of combination and preliminary explore their mechanism. In addition, the detection of the serum level of biochemical indexes (AST, ALT, BUN, Cr) and the observation of the hep- atic and renal histopathology changes were executed to analyze the effects of drugs on the hepatic and renal function.Result:1Ankle joint swelling and histopathological results showed:TMP and Lef combination could significantly alleviate the degree of inflammation in model rats. During treatment, on day16, AI values of TH, Lef, TL+Lef and TH+Lef groups were lower than that of the M group (P<0.01, P<0.05); from the beginning of the20th day, AI value of each drug group was significantly was lower than that of the M group (P<0.01, P<0.05); on day28, compared with the Lef group, AI value of the TH+Lef group significantly decreased (P<0.05). After20days, the foot plantar thickness in the TH+Lef group was significantly lower than that in the M group (P<0.05). After16days, compared with the M group, the foot plantar swelling area in the TH+Lef group significantly decreased (P<0.05). The swelling inhibition rate of the ankle joint in the Lef, TH and TH+Lef groups gradually increased with the treatment time lasted from the12th day, and the TH+Lef group even presented the highest inhibition rate. On day28, the inhibition rate reached27%. Compared with the TL group,24days after the treatment, the TH group also had a relatively high inhibition rate, even higher than that of the TL+Lef group on day24. By pathology observation and analysis, in addition to the TL group, other drug groups had different degree relief in pathological changes of the ankle joint, especially the TH+Lef group.2The results of the blood rheology showed:After drug intervention, compared with the M group, TH and TH+Lef groups improved peripheral blood rheology abnormalities, reduced hematokrit (P<0.01); the content of fibrinogen in the Lef, TH, TL+Lef and TH+Lef groups was lower than that in the M group (P<0.01, P<0.05); compared with the M group, the TH and TL+Lef groups presented a reduction in platelet aggregation rate (P<0.01, P<0.05).3Cytokine levels according to the results showed:Compared with the N group, TNF-α, IL-1β, IL-6serum levels increased significantly (P<0.01, P<0.05). In addition to the TL group, TNF-α serum level of other each drug group decreased significantly, compared with the M group, there were significant differences (P<0.01, P<0.05). TNF-α serum level in the TH+Lef group was lower than that in the Lef group, but which was no statistical difference; Compared with the M group, IL-1β and IL-6serum levels significantly decreased (P<0.01); compared with the N group, IL-10serum level in the M group obviously decreased (P<0.01), while compared with the M group, IL-10serum level in the Lef and TH+Lef increased significantly (P<0.01, P<0.05). VEGF-C serum level in each drug group was lower than that in the M group (P<0.01), and VEGF-C serum level of the TH+Lef group was significantly lower than that of the Lef group (P<0.05); the serum content of PAF in the TH+Lef group was significantly lower than that in the M group (P<0.01), in other drug groups there were no significant differences; VEGF and PAF expression in TH, TL+Lef、TH+Lef groups decreased, and VEGF expression of the TH+Lef group was obviously lower than that of the Lef group (P<0.05).4Blood biochemical indexes and the hepatic and renal histopathological results showed:Blood biochemical index results also showed that compared to the M group, AST in the Lef group and Cr in the TL+Lef group serum levels obviously increased (P<0.05), but ALT and BUN had no obvious statistical differences in each drug group. Liver pathological staining results only in the Lef and TL+Lef groups met slight pathological changes, which were not seen in the rest of groups. From renal pathological dyeing results, there were no significant pathological changes in each group.Conclusion:1Each drug group has a certain therapeutic effect on joint swelling of the CIA model rats and TMP50mg/kg in combination with Lef group presents the most significant therapeutic effect.2.TMP and Lef combination may have the therapeutic effect on joint swelling of the CIA model rats through improving hemorheology indexes, regulating inflammatory cytokines TNF-α, IL-1β, IL-6, IL-10and inhibiting angiogenesis.3TMP has no abnormal effects on the hepatic and renal function of the CIA model rats when combining with Lef... |
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