| Objective:Rheumatoid Arthritis(RA) is the most common inflammatory arthritis,a kind of chronic, systemic and autoim -mune disease, and also a major cause of disability. Its histopathologic feature is synonitis,formed pannus and changing of many kinds of cytokines, antigen-antibody and stiky molecule correlated with serum and tissues.Among the total, tumor necrosis factor-α(TNF-α) and interleukin-1β(IL-1β) as inflammatory cytokines play a center role in arthritis of RA and impairs,and macrophage migration inhibitory factor (MIF) as the upstream regular of TNF-αand IL-1βmay play a key role in onset of RA,today it is payed more attentions by international. MIF has become a new target of drug treatments of RA. Triptolide is isolated from Tripterygium wilfordii Hook.f, which is used in immune suppression and anti-inflammatory effect as the best activity of chemical compound. The collagen induced arthritis(CIA)of rats are widely considered having a similar pathologic change in arthritis compared to RA, so our study is to use this model to research the expression of MIF in the serum and synovium of CIA in rats from the whole to cell of multistrata angles and discuss the possible effective mechanism. Methods:⑴Fouty-five male Wistar rats were randomly divided into 9 groups.Except for one normal group,the other 8 different expreriment groups were injected with 0.2ml collagen from chicken sternal cartilage typeⅡin the tail,right hind limb and back together, and boosted at the 14th day to make manufa -cture the model of CIA.⑵Medicines were given from the 21th day to the 35th day once a day to observe its effect on arthritides. CIA was assessed by measuring arthritis index(AI), paw swell volume, general status and weight.⑶The levels of TNF-αand IL-1βin the serum and supernate fluid of spleen were measured by radioimmunoassay.⑷The level of MIF in the serum was measured by ELISA.⑸The general synovial tissues were acquired and HE staining was done to observe the pathologic change by light microscope.⑹Immunohistochemistry was used to detect the expression of MIF in synovialtissues and their intensity were analyzed by computerized image system.⑺The best concentration of drug-serum's effect on the cell proliferation inhibitory action of synovial fibroblasts by MTT method.⑻The level of MIF in the supernate fluid of synovial fibroblasts was measured by ELISA.⑼Immunohistochemistry was used to detect the gene expression of MIF.⑽SPSS11.5 statistical software was used to analyze the above-mentioned data.Results1 The rats of CIA appeared symptom in the 10th day post-modal,achieved peak in 14th day,and obviously after booster, weights of model and medication groups decreased in the 21th day of post-model,there was significant difference compared to normal group (P<0.05);Gradually, AI and claw swell volume increased, there was significant difference compared to normal group (P<0.05);After administrated for 14 days,those of medication group decreased,and there was significant difference compared to model group(P<0.05); Tri+LEF group was the best,and there was significant difference compared to model group(P<0.05);There was significant difference between model and normal group(P<0.05).2 The levels of MIF,IL-1βand TNF-αin the serum and supernate fluid of spleen of model group increased,and there was significant difference compared to normal group(P<0.01); The levels of those cytokine of medication groups decreased more rapidly than model group, and there was significant difference compared to model group(P<0.05); Matching groups decreased furthermore(P<0.01).3 Analysis of correlation between serum MIF and AI,serum IL-1βand TNF-αof the CIA rats showed that the level of serum MIF was positively correlated with those of the three latters (r=0.498,0.903,0.771, P<0.01).4 Pathologicals results showed that there were a lot of macrophages and T cells in synovial tissues.There were pannus formattds in synovial tissue.Remarks of model group were higher than those of normal group(P<0.01); Remarks of medication groups were lower than those of model group (P< 0.05); Remarks of Tri+LEF group were the lowest.5 Immunohistochemistry showed that the expression of MIF in model group was higher than normal group(P<0.01); those of medication groups were lower than that of model group (P<0.01); those of matching groups were lower than that of single medication group(P<0.05).6 MTT method showed that drug-serum had obvious effection on cell proliferation inhibitory action of synovial fibroblasts, and there was significant difference compared to control group(P<0.05);The best concentration of effect was that of 10% of single traditional Chinese medicine or the former combined with that of 30% LEF; there was significent difference between matching groups and single medication group(P<0.05).7 Supernate fluid of synovial fibroblasts by ELISA show that the levels of MIF of medication groups decreased, and there was significant difference compared to control group(P<0.01).8 Expression of MIF protein in the synovial fibroblasts were detected by Immunohistochemistry. The LI (%) of MIF of control group was 45.30±4.42, and that of Tri group was 33.45±2.27, and that of Tri+LEF group was 20.30±2.25, and there was significant difference compared to control group(P< 0.05);We conclude that Tri can cut down expression of MIF.Conclusions1 MIF can express significantly in the serum and synovim tissues of CIA in rats, and analysis of correlation between serum MIF and AI,serum IL-1β,TNF-αshowed that the level of serum MIF was positively correlated with those of the three latters,all of which can play a key role on onset of RA2 Triptolide has a good therapeutic effect on rats of CIA,the effective mechanism may correlate with that Triptolide decrease the expressions of MIF,TNF-αand IL-1β, and effect on cell proliferation inhibitory action of synovial fibroblasts.3 That Triptolide combined with Leflunomide show evident cooperation or synergistic effect in the respect of treating CIA in rats, suppressing synoviocyte proliferation in vitro culture and the expression of synoviocyte. This study can provide experimental evidence for association with effective ingredient of the traditional Chinese medicine and chemicals in treating RA.
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