Chinese Herbal Formula Ermiao Powder(二妙散) Regulates Cholinergic Anti-inflammatory Pathway in Rats with Rheumatoid Arthritis

Objective:To investigate the effect of Chinese herbal formula Ermiao Powder(二妙散,EMP)on the expression of cholinergic anti-inflammatory pathway in rats with rheumatoid arthritis(RA).Methods:Seventytwo rats were randomly divided into 6 groups according to body weight,including normal control group,collageninduced arthritis(CIA)group,three doses EMP groups,and methotrexate(MTX)group(n=12 per group).All of the rats except for those in the normal control group were given multipoint subcutaneous injection of bovine typeⅡcollagen to establish a CIA model.Three EMP groups received a high-(4.5 g/kg),medium-(3.0 g/kg),and low-(1.5 g/kg)doses of EMP by intragavage,respectively.MTX group was injected intraperitoneally MTX at 0.9 mg/kg once a week as the positive control.The administration was 3 consecutive weeks.Joint swelling,arthritis index,and body weight changes in different experimental groups of rats were tested.The joint damage was evaluated by masson staining.Quantitative real-time polymerase chain reaction,Western blot,and immunohistochemistry(IHC)were performed to evaluate the expression of CHRNA7,encodingα7 nicotinic acetylcholine receptor in the cholinergic anti-inflammatory pathway,in different tissues and their localization in the spleen and joints.Results:CHRNA7 expression levels were significantly higher in the joints and spleens of CIA group than those in normal control group(both P<0.05).Moreover,the CHRNA7 mRNA and protein levels in the spleen and joints of MTX and three doses of EMP groups were significantly lower than CIA group(all P<0.05).Compared with the MTX group,treatment with low-dose EMP resulted in significant reduction of CHRNA7 mRNA and protein expression levels(P<0.05 or P<0.01).IHC showed positive signals of CHRNA7 in the white pulp and red pulp of the spleens of rats;CHRNA7 was expressed on fibroblast-like synoviocytes,macrophages,and endothelial cells in the joints of rats,and the expression in the joints of low-dose EMP group was significantly lower than that in the CIA group(P<0.01).Conclusion:Cholinergic anti-inflammatory pathway was involved in the generation of the inflammatory reaction in CIA rats,and EMP exerted therapeutic effect on RA through cholinergic anti-inflammatory pathway.