Effects of Compound Dahuang Powder combined with acupuncture on synovial ultrastructure,pathological angiogenesis and Ras/Raf signal of synovial cells in rats with rheumatoid arthritis
Objective To investigate the effects of external application of Compound Dahuang Powder combined with acupuncture on the synovial ultrastructure,pathological angiogenesis,and Ras/Raf signaling pathway in rheumatoid arthritis rats.Methods Totally 100 SPF grade SD rats were selected and divided into normal group,model group,acupuncture group,external application group,and combination group according to random number table method,with 20 rats in each group.Except for the normal group,Freund's complete adjuvant was injected into the bilateral plantars to establish a rheumatoid arthritis model.One week after modeling,the acupuncture group received acupuncture at"Yinlingquan(SP9)"and"Sanyinjiao(SP6)"for 30 minutes,with continuous intervention for 6 days and 7 days as one course of treatment,for a total of 3 courses of intervention.The external application group of rats applied compound Dahuang Powder on the right hind foot and ankle joint,10 g/time,twice a day,for 7 days as one course of treatment,with a total of 3 intervention courses;the combination group received a combined intervention of acupuncture and external application of compound Dahuang Powder.No intervention was performed in the normal group and model group.The levels of TNF-α and IL-1β in serum of rats in each group were detected using enzyme-linked immunosorbent assay.Transmission electron microscopy was used to observe the ultrastructure of synovial cells.HE staining was used to observe the morphology of synovial tissue.Immunohistochemistry was used to detect vascular endothelial growth factor(VEGF)and its receptor VEGFR2 in synovial tissue.Western blot and RT qPCR were used to detect Ras/Raf protein and mRNA in synovial tissue,respectively.Results Compared with model group,the levels of TNF-α and IL-1β in serum of rats in acupuncture group and external application group decreased(P＜0.05),the expressions of VEGF and VEGFR2 decreased(P＜0.05),and the expressions of Ras and Raf protein and mRNA in acupuncture group and external application group decreased(P＜0.05).Compared with acupuncture group and external application group,the levels of TNF-α and IL-1β in serum of rats in the combination group decreased(P＜0.05),the expressions of VEGF and VEGFR2 decreased(P＜0.05),and the expressions of Ras and Raf protein and mRNA in the combination group decreased(P＜0.05).The results of synovial cell ultrastructure showed that the nuclear membrane of synovial cells in normal group was good,and the nuclear membrane in model group was seriously damaged.The acupuncture group and external application group had different degrees of improvement,and the combination group had the most significant improvement.HE staining results showed that the joint synovial tissue of the normal group was normal,the joint synovial tissue of the model group had a large amount of inflammatory infiltration,the joint synovial injury of the acupuncture group and the external application group was slightly improved,and the pathological injury of the combined group was significantly reduced.Conclusion The combination of external application of compound Dahuang Powder and acupuncture can regulate angiogenesis related factors,regulate the Ras/Raf signaling pathway,and improve the ultrastructure of synovial cells in rheumatoid arthritis model rats.
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