Anti-rheumatoid Arthritis Effect And Mechanism Of Clematis Clematis

Objective: To investigate the effect of ethanol extract and different extraction fractions of Clematis florida var.plena（CF）on a collagen-induced arthritis（CIA）rat model,and to explore the possible mechanisms,so as to provide reference for the development and clinical application of Chinese medicinal materials of CF.Methods:（1）CF was extracted and separated with 70 % Et OH under percolation,the extract was concentrated,then the extract was suspended in water and successively partitioned with ethyl acetate（EA）and n-butanol（NB）.（2）The CIA was induced by intradermal injection of bovine type Ⅱ collagen,to investigate the effects of CF.The rats were divided into model group,Tripterygium Glycosides Tablet positive drug group（TG,20 mg/kg）,the low,medium and high dose groups of CF-TAE（0.5,1.0,2.0 g/kg）,the low,medium and high dose groups of CF-NB（0.2,0.4,0.8 g/kg）,the low,medium and high dose groups of CF-A（0.4,0.8,1.6 g/kg）after the CIA model is successfully established.To evaluate the effects of CF-TAE,CF-NB and CF-A on CIA rats by measuring the body weight,arthritis score,mean swelling widthness,thickness and volume of paw,HE,Micro-CT,the production of inflammatory cytokines in serum,the contents of MDA and SOD in the liver were detected by trace method.（3）In vitro culture experiment,TNF-α stimulate human MH7 A cell line,to observe the effect of CF-TAE treatment at different concentrations.CCK-8 assay was used to detect the anti-liferation of MH7 A cells at different concentrations of CF-TAE.ELISA assay was used to detect the levels of IL-1β、IL-6、IL-10 at different concentrations of CF-TAE.Western Blot was used to detect the effect of JAK2,STAT3,NF-κBp65 protein expression on proliferation after CF-TAE treatment.Results:（1）The n-butanol,ethylacetate and aqueous extractive of CF were obtained.（2）During D18 to D42,compared to normal group,the body weight,arthritis score,mean swelling widthness,thickness and volume of paw have significant difference in Model group（P<0.05）.Compared to Model group,the body weight of rats were elevated in TG,CF-TAE（0.5,1.0,2.0 g/kg）,CF-NB（0.2,0.4,0.8 g/kg）and CF-A（0.4,0.8,1.6 g/kg）groups,and the arthritis score,mean swelling widthness,thickness and volume of paw of rats were reduced.The bone erosion and inflammatory cell infiltration of the ankle joint,and the degree of joint damage were inhibited.The levels of pro-inflammatory cytokines Il-1β,TNF-α and Il-6 were decreased and the levels of anti-inflammatory cytokines Il-10 were increased in the drug administration groups.The content of MDA in the liver was decreased in the drug administration group,showing a significant difference（P<0.05）,the content of SOD was increased without significant difference.（3）Compared to Control group,different concentrations of CF-TAE could inhibit the proliferation of MH7 A cells,and all have a significant different.And CF-TAE significantly inhibited the levels of IL-1β、IL-6、IL-10 and reduced the protein level of JAK2,STAT3,NF-κBp65.Conclusion: CF-TAE has anti-arthritis and anti-oxidation effects on type Ⅱcollagen-induced arthritis in rats,the high-dose group（0.8 g/kg）of CF-NB and the low-dose group（0.4 g/kg）of CF-A had the best anti-RA activities,but there were no significant differences between the different fractions of CF in vivo.CF-TAE could suppress inflammatory reations and proliferation of MH7 A cells via inhibiting the activation of JAK2-STAT3-NF-κBp65 signaling pathway in vitro.