| Objective: In this study,rat rheumatoid arthritis was induced by complete Freund’s adjuvant,and the level of Ras and Raf1 protein in rat serum and protein expression of Ras and Raf1 in rat synovial tissue were observed by Zhuang Yao Long Zuan Tong Bi Formula.1.The role of H ras and Raf1 gene transcription levels in rat synovial tissue and the pathological changes of articular synovial tissue.To explore the mechanism of action of Zhuang Yao Long Zuan Tong Bi Formula on rheumatoid arthritis.Methods: 96 healthy male SD rats were selected.After 1 week of adaptive feeding,72 rats were randomly selected to replicate the RA model with the wind-cold-humidity plus biological factor composite modeling method,and the remaining 24 rats were classified as normal groups.On the 7th day after immunization,the right hind toe swelling degree was used to determine whether the modeling was successful.Rats with successful modeling were randomly divided into a model group,a Western medicine group,and a Zhuang medicine group,with 24 rats in each group.The Zhuang medicine group was given the Zhuang medicine Long Zuan Tong Bi Formula by intragastric administration,the methotrexate group was given the intragastric administration by the prescribed dose,and the model group and the normal group were fed with saline.Each rat was gavaged at a volum e of 1ml / 100 g.Each group measured the swelling of the right hind toe of the rats at four periods of 7d,14 d,21d,and one week after drug withdrawal(28d after the first dose).At the same time,sa mples were taken from each group.Six rats were taken fr om each group in each period.Collect rat serum and synovial tissue,use ELISA to determine the serum Ras,Raf1,p-raf1 protein content;use RT-q PCR to detect Hras,Raf1 gene transcription level in rat synovial tissue;take the degree of toe swelling as the therapeutic effect Indicators,sele ct the tissues from the best period of efficacy for Western Blot experiment,determine the relative expression of synovial tissue Ras,Raf1,p-raf1 protein;at the same time histopathological HE staining analysis of rat ankle.Results: 1.Swelling of toes of rats: On the 3rd day after immunization,the hind feet of rats began to appear red and swollen.On the 7th day after immunization,the joints of 72 model rats gradually became red,swollen,and deformed.The thickness of the toes was higher than that of the normal control group,indicating that the model was successfully replicated.On the 14 th day of administration,compared with the model group,the swelling of the toes in the western medicine group and the Zhuang medicine group both decreased,and the swelling degree of the two groups of rats fell to the minimum at the third week after the administration.2.Serum ELISA test results: There were no significant differences in serum Ras,Raf1,p-raf1 levels in normal group,model group,western medicine group and Zhuang medicine group in four different time periods(P> 0.05).3.RT-q PCR test results: Compared with the normal group,the relative expression levels of Hras and Raf1 m RNA in the animal synovial tissues of the model group were up-regulated in the four periods;compared with the model group,the strong drugs were 14 d,21d and 28 d after administration The relative expression of Hras and Raf1 m RNA in the Zhuang medicine group and the western medicine group was lower than that in the model group(P <0.05).There was no difference in the relative expression of Hras and Raf1 m RNA between the two groups in the Zhuang medicine group and the Western medicine group(P> 0.05).4.Western Blot test results: On the 21 st day of administration,the relative expression levels of Ras,Raf1,p-raf1 protein in the synovial tissue of the model group were up-regulated compared with the normal group,and the difference between the two groups was statistically significant(P < 0.05);Compared with the model group,the r elative expression of the synovial Ras,Raf1,p-raf1 protein of Zhuang Yao Long Zuan Tong Bi Fang granule and Western medicine group was down-regulated,and there was a difference in the down-regulation after statistical analysis(P <0.05).5.HE pathological staining results: By analyzing the ankle joint HE staining of RA rats on the 21 st day after administration,the pathological score of the model group rats was higher than that of the normal group(P <0.01).Compared with the model group,the total pathological score of rats in the Zhuang medicine group and the Western medicine group decreased(P <0.05),and there was no significant difference in the total pathology score between the Western medicine group and the Zhuang medicine group.Conclusion:The mechanism of Zhuang Yao Long Zuan Tong Bi Formula in the treatment of RA may be through down-regulation of the transcription level of Hras and Raf1 m RNA in the sy novium and down-regulation of the expression of Ras and Raf1 protein in the synovium,thereby reducing the synovial lesions and bone protection. |
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