Study On Immunomodulatory Effect And Mechanism Of Fufang Shatai Heji

Objective: This study was to investigate the regulation of Fufang Shatai Heji(STHJ)on the immune status of cyclophosphamide-induced immunosuppression and immune disorders caused by rheumatoid arthritis,and to explore its mechanism.Contents: This study was focused on the following issues:(1)To establish an immunocompromised mouse model by intraperitoneal injection of cyclophosphamide(CTX)and then study the immunomodulatory effects of STHJ on immunocompromised mice;(2)Different rheumatoid arthritis models in two strains of mice and select the most appropriate model;(3)To establish the collagen-induced arthritis(CIA)in the context of DBA/1 mice,and study the protective effect on mouse joints after intragastric administration of STHJ.Methods:(1)The immunocompromised model was established on the Balb/c mice by intraperitoneal injection of CTX.After the model was established successfully,STHJ was continuously administered by gavage for 30 days.After administration,the mice were sacrificed by extracting eyeball and then blood and spleen were obtained.Serum was obtained after centrifugation of blood,and the expression levels of inflammatory factors(TNF-?,IL-2,IL-6,and IL-10)in the serum were measured by ELISA.The spleen lymphocytes were obtained after grinding and other operations,and the percentage of the relevant lymphocyte subsets was analyzed by flow cytometry.(2)Rheumatoid arthritis models were induced in DBA/1 and C57BL/6 mice by immunization.In this study,we mainly studied the modeling of collagen-induced arthritis(CIA)and antigen-induced arthritis(AIA)in different species.An appropriate modeling method would be selected to study the protective effect of STHJ on RA after analyzing and comparing the modeling.(3)The CIA model was established in DBA/1 mice by immunization.STHJ was administered continuously for 30 days after booster immunization,and the body weight and paw swelling were monitored regularly.After administration,blood was obtained by extracting eyeball and then the spleen,legs and femoral head was collected.Histopathological analysis of the joints was performed to evaluate the protective effect of STHJ on joint destruction.The femoral head was tested by RT-PCR to analyze the changes of related genes in the cartilage after administration.The changes in spleen structure and the expression of related inflammatory factors in serum were analyzed to study the regulation of STHJ on the immune status of CIA mice.Results:(1)The thymus and spleen index can be significantly increased after the administration of STHJ on CTX-induced immunosuppressive mice.Through flow cytometry analysis of spleen lymphocytes,STHJ can increase the activity of B cells and NK cells,and decrease the activity of CD8+ T,CD8+CD122+ T,natural killer T(NKT)and ??T cells.In addition,STHJ can up-regulate the expression of IL-2,IL-6 and TNF-? and down-regulate the expression of IL-10 in serum of CTX-induced immunosuppressive mice.(2)Analysis of the modeling in the background of DBA/1 and C57BL/6 mice found that the successful modeling in the background of DBA/1 was significantly better than in the background of C57BL/6.Modeling in the background of DBA/1,we found CIA mice have a high success rate and long duration of maintenance.Modeling in the background of C57BL/6,CIA mouse has a low success rate,and the AIA has a high success rate but a short maintenance time.(3)After administration of STHJ in CIA mice,joint swelling was significantly reduced.By analysising joint histopathology we also found that STHJ can reduce synovial hyperplasia and cartilage destruction after treatment.Immunohistochemical analysis of mouse cartilage revealed that the expression of MMP-9,MMP-13 and ADAMTS-5 in STHJ-treated mouse was significantly reduced.RT-PCR analysis of cartilage also found that m RNA expression of MMP-9,ADAMTS-4,ADAMTS-5,TIMP-1 and Type X collagen in mouse cartilage was significantly decreased after administration of STHJ,while m RNA expression of aggrecan was significantly increased.After the treatment of STHJ,the trend of structural deformation of the spleen was improved,and the proliferation of the red pulp was controlled.The fibrous capsule surrounding the spleen also became thicker,and there was no obvious appearance of iron-colored particles.In addition,high expression of IL-17 a in serum of CIA mice was also effectively inhibited.Conclusions: The above results indicated that STHJ can effectively ameliorate CTX-induced immunosuppression by regulating the expression of relevant lymphocytes and inflammatory factors.In CIA mice,STHJ can significantly improve arthritis symptoms and immune status,and has a protective effect on cartilage destruction.Therefore,STHJ has the function of regulating immunity in both immunosuppression and immune disorders.