| Objective:In this experiment,the model rats of wind-cold-dampness were used to observe the effects of heat-reinforcing acupuncture on ankle swelling,arthritis index score and synovial histopathology of rheumatoid arthritis model rats,to investigate the effect of heat-reinforcing needling on RA and its possible mechanism,the levels of Th1/Th2 related cytokines(IFN-γ,IL-2,IL-4,IL-10)in serum and synovium of rats were detected,to provide reliable experimental basis for further clinical application.Methods:Forty SPF SD rats were randomly divided into blank group(N=8)and model group(N=32),in this study,complete Freund’s adjuvant and chicken type Ⅱ collagen were used to induce rheumatoid arthritis(RA)in the wind-cold-damp model,on the 21st day after the first immunization,the success of modeling was evaluated according to the AI score.After successful modeling,the model group was randomly divided into model group,Western medicine group,Ping Bu Ping Xie Group and re bu acupuncture group,8 rats in each group.The Rats in the Blank Group,the Model Group and the acupuncture group were grabbed and fixed at the same time without acupuncture intervention,while the rats in the western medicine group were injected intraperitoneally with 0.1 mg 100g methotrexate,the Rats in the acupuncture group were treated with ping-bu-ping-reducing needling method and re-bu-shen needling method respectively at Zusanli for 30 min,once a day for 7 days.During the experiment,the general conditions of diet,action,spirit,hair and luster of the rats in 5 groups were observed every day The AI index and the diameter of the left ankle joint of the experimental rats were observed and measured before modeling,after modeling and after treatment,after the rats were anesthetized,the blood samples were collected by centrifuge,and all the rats were killed by cervical dislocation,and the synovium of the ankle joint was stored in 4%paraformaldehyde,the pathological changes of ankle joint synovium were observed by HE staining,and the levels of IFN-γ,IL-2,IL-4 and I1-10 in serum were measured by Elisa.The levels of IFN-γ,IL-2,IL-4 and 11-10 in the synovium of ankle joint were detected by immunohistochemistry.Results:1.The general condition of the rats:The Rats in the blank group were in good mental condition,had normal food and water intake,body hair was white and shiny,skin temperature was normal,moved freely,reacted sensitively,defecated regularly,fecal quality was normal,paws were not inflamed and swollen,compared with the blank group,the rats in the model group were in a poor mental state,and their food intake and water intake were significantly reduced,their body hair became yellow and lustrous,their activities were obviously limited,and they even developed claudication,after modeling,the hind feet became red and swollen,some of them could touch the rheumatic nodules at the tail root,and the swelling of the bind feet was most obvious on the 14th day after immunization Compared with the model group,the rats in the Western medicine group,the Pingbupingxie Group and the heat-reinforcing acupuncture group had better mental status,basically normal food and drinking water,gradually improved activity limitation,reduced joint swelling and recovered skin color.2.Ankle diameter:before modeling,there was no significant difference in ankle diameter among the three groups(P>0.05),but after modeling,there was significant difference in ankle diameter between the two groups(p<0.01),after treatment,the diameter of ankle joint in the Western medicine group,the Ping Bu Ping Xie group and the hot bu Zhen group were smaller than that in the model group(p<0.01),and the difference was significant(p<0.01),the results showed that the symptoms of RA rats were obviously improved after treatment,and there was no significant difference between the Western medicine group and the heat-reinforcing acupuncture group(P>0.05),and the diameter of ankle joint in the heat-reinforcing acupuncture group was smaller than that in the ping-reinforcing-reducing group,the difference was statistically significant(p<0.01),indicating that the hot reinforcing needling method was superior to the flat reinforcing and reducing method in reducing the ankle joint diameter.3.Arthritis index:Before Intervention,the AI scores of the model group were all over 8,indicating the success of the model,after intervention,the AI scores of each treatment group were decreased compared with the model group,the difference was statistically significant(p<0.01);Compared with the Western medicine group,the AI score of the heat-reinforcing acupuncture group was higher(p<0.05),and the AI score of the heat-reinforcing acupuncture group was lower(p<0.05).4.Synovial pathology of the ankle joint:the synovium of the blank group showed normal structure,regular arrangement of synoviocytes,smooth surface and no inflammatory reaction.In the model group,there were a lot of inflammatory cells infiltrating in synovium,the surface of synovium was irregular,accompanied with pannus formation.Compared with the model group,the synovium infiltration of inflammatory cells in the pingbupingxie group was improved,and the synovium tissue structure of the ankle joint in the Western Medicine Group and the heat-reinforcing acupuncture group was basically normal,with only a small amount of synovium inflammatory cells infiltration,the proliferation of synovial cells was also less,and the degree of injury was obviously improved compared with the model group.5.Compared with the blank group,the levels of IFN-γ and IL-2 in the model group were significantly increased(p<0.01),while the levels of IL-4 and Il-10 were significantly decreased(p<0.01),compared with the model group,the levels of IFN-γ and IL-2 in the Western medicine group,the heat-reinforcing acupuncture group and the ping-bu-ping-xie group were significantly decreased(p<0.01),while the levels of IL-4 and Il-10 were significantly increased(p<0.05,p<0.01)Compared with the Western medicine group,the changes of serum IFN-γ and IL-2 levels in the heat-reinforcing acupuncture group had no statistical significance(p>0.05),while the levels of IL-4 and Il-10 in the heat-reinforcing acupuncture group were lower than those in the western medicine group(P>0.05)Compared with the Ping-bu-ping-xie group,the levels of IFN-γ and IL-2 in the heat-bu-acupuncture group were significantly lower(P<0.05),while the levels of IL-4 and Il-10 were significantly higher(P<0.05).6.The expression of IFN-γ and IL-2 in the synovium of the model group was significantly higher than that of the control group(P<0.01),the expression levels of IL-4 and IL-10 were significantly decreased(p<0.01),after treatment,compared with the model group,the expression levels of IFN-γ and IL-2 were significantly decreased(p<0.01),the expressions of IFN-γ and IL-2 in the synovium of the ankle joint of the rats in the Western medicine group,the heat-reinforcing acupuncture group and the ping-bu-ping-xie group were significantly decreased(P<0.01),the expressions of IL-10 were significantly decreased(p<0.01),and the expressions of IL-2 were significantly decreased in the synoviurn of ankle joint in the heat-reinforcing acupuncture group as compared with the Western medicine group(P<0.01),there was significant difference in the expression levels of IL-10(P<0.01),but the expression levels of IFN-γ and IL-2 in the synovium of the ankle joint of the rats in the heat-reinforcing acupuncture group were significantly lower than those in the ping-buping-reducing group(P<0.01),the expression levels of IL-4 and Il-10 were increased(P<0.01).Conclusions:1.The method of heat-reinforcing acupuncture can obviously reduce the diameter and swelling of the ankle joint of RA Rats,reduce the AI score of rat model,relieve the general inflammation of rats,improve the degree of activity limitation,improve the tissue structure of synovium,to some extent,it can inhibit synovial inflammatory reaction,reduce inflammatory cell infiltration and improve synovial proliferation.2.The experimental data showed that the heat-reinforcing needling method could effectively reduce the levels of IFN-γ and IL-2 secreted by Thl and increase the levels of IL-4 and IL-10 secreted by Th2 in serum and synovial tissue of rats,thus,the balance of Th1/Th2 cell immunity can be maintained,which provides a reliable basis for the treatment of the disease with the method of heat-reinforcing acupuncture. |
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