Mechanism Of Triptolide Regulating Th17/Treg In The Treatment Of Adjuvant Arthritis In Rats

Objective: To observe the anti-inflammatory effect of triptolide(triptolide,TP)on adjuvant arthritis(adjuvant arthritis,AA)rats and its effect on the balance of joint synovium Th17/Treg,and to explore the anti-inflammatory mechanism of triptolide in the treatment of rheumatoid arthritis(rheumatoid arthritis,RA),so as to provide theoretical basis for clinical application of TP in the treatment of RA.Methods: 50 SD rats were randomly divided into 5 groups according to their body weight:normal control group,adjuvant arthritis model group,low,middle and high dose triptolide treatment group,10 rats in each group.Except for the normal group,the AA model was induced by intradermal injection of 0.1ml Freund’s complete adjuvant into the right hind paw of rats in the other groups,and the normal group was injected with the same amount of normal saline.After 14 days,TP was injected intraperitoneally(0.1mg/kg in low dose TP group,0.2 mg/kg in middle dose TP group,and 0.4mg/kg in high dose TP group)once a day for 10 days.The normal group and the model group were injected with the same amount of 4% propanediol solution.The degree of plantar swelling,thermal pain threshold and mechanical pain threshold were measured 1 day before modeling,14 days after modeling and 10 days after treatment.The rats were killed 10 days after treatment,and the blood and right ankle joint samples were extracted.The histopathological changes of synovium were observed by HE staining.The expression of RORγt,Foxp3 and TGF-β1 protein in synovial tissue was detected by immunohistochemistry.The expression of IL-17 and IL-10 in serum was detected by enzyme linked immunosorbent assay(ELISA).The expression of IL-17,IL-10,RORγt and Foxp3 m RNA in synovial tissue was detected by real-time fluorescence quantitative PCR.Results: 1.Compared with the normal group,the swelling degree of foot plantar in AA model rats was significantly increased(P<0.01),and the thermal pain threshold and mechanical pain threshold were significantly decreased(P<0.01).After treatment,compared with the model group,the swelling degree and mechanical pain threshold of AA rats in the middle and high dose TP group were significantly decreased(P<0.05),while the thermal pain threshold of AA rats in the high dose TP group was significantly increased(P<0.05).2.HE staining results showed that the structure of ankle joint was clear and normal in the normal group.Synovial hyperplasia and thickening,infiltration of proinflammatory cells,severe erosion of articular cartilage and inferior bone and pannus formation were observed in the model group.After TP treatment,the above pathological phenomena were significantly better than those in the model group.3.Compared with the normal group,the expressions of IL-17 in serum,RORγt and TGF-β1 protein in synovium,IL-17 and RORγt m RNA in synovium in model group were significantly increased(P<0.01),while the expressions of Foxp3 protein,m RNA and IL-10 m RNA in synovium were significantly decreased(P<0.01).Compared with the model group,the expression of IL-17 in serum,RORγt and TGF-β1 protein in synovium,IL-17 and RORγt m RNA in synovium in middle and high dose TP groups decreased significantly after treatment(P<0.05),while the expression of Foxp3 protein and m RNA in synovium increased significantly(P<0.01).High dose TP group significantly decreased the expression of IL-10 m RNA(P<0.05).Conclusion: 1.TP can relieve joint redness and swelling,increase pain threshold,reduce synovitis and reduce bone damage in AA rats.in AA rats.It has obvious anti-inflammatory,analgesic and osteoprotective effects.2.TP can down-regulate the protein and m RNA expression of Th17 cell-related cytokine IL-17 and specific transcription factor RORγt in AA rats,up-regulate the protein and m RNA expression of Treg cell-related cytokine IL-10 and specific transcription factor Foxp3,and down-regulate the abnormal expression of TGF-β1 in articular synovium.3.TGF-β1 can induce the expression of RORγt and Foxp3 respectively and participate in the differentiation of Th17/Treg,while TP down-regulates the high expression of TGF-β1 in the synovium of AA rats,suggesting that TP may regulate the balance of Th17 and Treg cells by regulating the expression of TGF-β1.4.Regulating the balance of synovial Th17/Treg and then regulating the expression of cytokines secreted by TP may be one of the mechanisms by which TP plays an anti-inflammatory role in the inflammatory synovium of RA.