The Effect Of Combination Methotrexate And Cyclophosphamide On Synovial Cell Apoptosis Of Collagen-Induced Arthritis Rats On The Basis Of Cell Cycle

Objective To investigate the effect of combination low dosage methotrexate(MTX) and Cyclophosphamide according cell cycle on synovial cell apoptosis in collagen-induced arthritis Rats.Investigate its possible synergistic effect and evaluate the possible mechanisms.Methods1.After established CIA on female Wistar rats successfully.They were randomly divided into CIA model group,Low-dose MTX treatment group(0.9mg/kg/w),Low-dose CTX treatment group(24mg/kg/21d),and Low-dose MTX+Low-dose CTX treatment group(MTX 0.9mg/kg/w,CTX 24mg/kg/21 d).2.All of the rats were sacrificed 24 weeks after the first immunization.Daily clinical evaluations were assessed by arthritis index.Treatment effects were assessed clinically, radiologically,and histologically.3.At the end of the experiment(24w),the specimens of ankle joints were exposed,fixed, decalcified,wrapped and cut into slices.All slices were stained with hematoxylin and eosin to evaluate synovial inflammation and bone erosive changes.The rats of synoviocyte apoptosis were detected with terminal deoxynucleotidyl transferees-mediated nick end labeling(TUNEL) method.Results1.CIA models were established in 66.67%of rats,presenting clinical,radiological,and histological characters of RA.A chronic inflammatory arthritis developed in 60 of 90 rats with typeⅡcollagen.2.24 weeks after the frist immunization,21 weeeks after the treatment,the improvement of arthritis index of combination treatment group was higher than other groups,which showed statistically significant difference versus CIA model group, Low-dose MTX group,Low-dose CTX group(P＜0.05).3.Compared to CIA control group,HE staining showed that the layers of synoviocytes of the CIA group were increased to 6～10,and the arrangement of synoviocytes was disordered and heavy inflammatory of cell infiltration was found in the CIA group,In the single treatment group, Low-dose MTX group and Low-dose CTX group,the layers of synoviocytes were increase to 3～4,and medium amount of inflammatory of cell infiltration was found.In the combination treatment group,the layers of synoviocytes were nearly normal,and light amount of inflammatory of cell infiltration was found.4.Compared to CIA control group,synoviocyte apoptosis in the treatment groups were increased markedly.The combination treatment group was higher than other groups,which showed statically significant(P＜0.05).Conclusions1.There were lots of synovial cells proliferated extremely in the model CIA rats.2.The results suggest combination MTX and CTX may be synergistic in the RA.They can inhibit synoviocyte differentiation and induce synoviocyte apoptosis.3.There maybe have important cooperant mechanisms of combination MTX and CTX treatment according to cell cycle in treating RA.