To Explore The Mechanism Of Xinfeng Capsule In Treating Pannus Of RA Based On Platelet Activation Regulation Of Angiogenesis

1 ObjectiveThrough animal experiments and clinical observation,the effects of TCM compound XX capsules on VEGF,PDGF,TSP-1 released after platelet activation,vascular endothelial cell marker CD31 and vascular smooth muscle cell markerαSMA were explored to observe the mechanism of inhibiting the synovial pus of RA,providing a new direction for the treatment of RA with traditional Chinese medicine.2 Methods2.1 Experimental StudyForty-five clean SD rats were randomly divided into three groups:normal group(group A),model group(group B)and XX capsule group(group C),with 15 rats in each group.In addition to the normal group,mold.The drug was administered for 30 days.The degree of toe joint swelling and arthritis index were observed.HE staining was used to detect the pathological morphology of synovium of knee joint.Microvascular pathology was observed by microscope.Immunofluorescence double staining CD31 andαSMA detect knee joint synovial vascular distribution;Immunohistochemistry was used to detect the expression levels of microvascular density(MVD),angiogenesis marker CD31,platelet activation product VEGF,PDGF and TSP-1 positive cells in joint synovium tissue,and to observe the influence of XX capsule on them.2.2 Clinical StudyTwenty-five patients with RA were selected in this study.All patients were treated with XX capsules(3 times a day,3 capsules once a time),and celeoxib of the same type and dose(0.2g/tablet,manufactured by Qingdao Baiyang Pharmaceutical)could be added according to the needs of the disease.The application of other symptomatic drugs remained unchanged.The changes of disease activity score,ESR,hs-CRP,RF,anti-CCP and platelet parameters were observed before and after treatment.ELISA was used to detect the changes of VEGF,PDGF and TSP-1.3 The Results3.1 Experimental Research Results3.1.1 Effect of XFC on Toe Swelling and Arthritis Index of AA Rats30 days after administration,compared with normal group,toe swelling and AI of AA rats in model group were significantly increased(P<0.01);Compared with model group,the toe swelling and AI of AA rats in XX capsule group were significantly decreased(P<0.01).3.1.2 Effect of XFC on Pathomorphology of Joints in AA RatsThe normal group of AA rats’synovium and synovium epithelial cells arranged regularly,no proliferation and protrusion,no obvious inflammatory cell infiltration.AA rats in the model group showed obvious joint pathological changes,including a large number of inflammatory cell infiltration in the local joint,synovial tissue hyperplasia and protrusion into the joint cavity,neovascularization and obvious pannus in the synovial tissue.XFC treatment significantly alleviated the histopathology of arthritis in AA rats.There were less inflammatory cell infiltration in joint synovium and blood vessel wall,relatively regular joint structure,relatively neat arrangement of synovium cells,synovium tissue hyperplasia and a small amount of blood vessel hyperplasia.3.1.3 Effect of XFC on Microvessel Density(MVD)Count in Synovium of Knee Joint of AA RatsAfter 30 days of continuous administration,a large number of neovascularization and MVD were significantly increased in the synovial tissue of the model group compared with the normal group(P<0.01).Compared with model group,neovascularization and MVD in synovial tissue of rats in XX capsule group were significantly decreased(P<0.01).3.1.4 Influence of XFC on positive expression of CD31,a vascular marker in the synovium of knee joint,in AA ratsAfter 30 days of continuous administration,compared with normal group,the expression of synovial blood vessel marker CD31 in model group was significantly increased(P<0.01);Compared with model group,the positive expression of vascular marker CD31 in the synovial tissue of AA rats in XX capsule group was significantly decreased,and the average optical density DOI was significantly decreased(P<0.01).3.1.5 Influence of XFC on synovial vascular distribution in AA ratsAfter 30 days of continuous administration,the positive staining of immature blood vessels(CD31~++αSMA~-)and mature blood vessels(CD31~++αSMA~-)increased significantly in model group compared with normal group,and the number of total blood vessels increased significantly.Compared with model group,the positive staining of immature blood vessels(CD31~++αSMA~-)and mature blood vessels(CD31~++αSMA~-)in XX capsule group was significantly reduced,and the total number of blood vessels decreased accordingly.3.1.6 Effect of XFC on Expression of VEGF,PDGF and TSP-1 in Synovium of AA RatsAfter 30 days of continuous administration,the positive expressions of VEGF,PDGF and TSP-1 in the synovial tissue of inflammatory joints of AA rats in model group were significantly increased compared with normal group(all P<0.01).Compared with model group,positive expressions of VEGF,PDGF and TSP-1 in the synovial tissue of inflammatory joints of AA rats in XX capsule group were significantly reduced(all P<0.01).3.1.7 Correlation Analysis of Platelet Activator and Vascular Marker CD31 with Toe Swelling and AI in AA RatsCorrelation analysis showed that platelet activator VEGF was positively correlated with toe swelling in AA rats(P<0.05),and PDGF was positively correlated with toe swelling in AA rats(P<0.05).Vascular marker CD31 was positively correlated with toe swelling(P<0.05).3.2 Clinical Research Results3.2.1 Changes in Clinical Indicators of RA Patients before and after TreatmentAfter treatment,the DAS-28 score,ESR,hs-CRP,RF and anti-CCP of RA patients were significantly improved(P<0.01).3.2.2 Changes of Platelet Parameters in RA Patients before and after TreatmentAfter treatment,PLT and PCT of RA patients decreased significantly(P<0.01);There was no significant difference in MPV and PDW(P<0.05).3.2.3 Changes of Platelet Activators in RA Patients before and after TreatmentAfter treatment,VEGF,PDGF and TSP-1 in RA patients were significantly decreased(P<0.01).3.2.4 Correlation Analysis of Platelet Activation Products and Clinical Indicators in RA PatientsCorrelation analysis showed that VEGF was positively correlated with DAS-28,RF,anti-CCP,ESR and hs-CRP in RA patients(P<0.01 or P<0.05).PDGF was positively correlated with DAS-28,RF and anti-CCP(P<0.01 or P<0.05).TSP-1 was negatively correlated with RF(P<0.05).3.2.5 Correlation Analysis of Platelet Activation Products and Platelet Parameters in RA PatientsCorrelation analysis showed that VEGF was positively correlated with PLT and MPV in RA patients(P<0.05).PDGF was positively correlated with PLT and PDW(P<0.05).TSP-1 was negatively correlated with PLT and PDW.4 ConclusionXFC has a significant clinical effect in the treatment of RA.It can significantly reduce the DAS-28 integral value of RA patients,and reduce the ESR,hs-CRP,RF,anti-CCP and other laboratory indicators.XFC can significantly improve the joint pathology,toe swelling and arthritis index of AA rats,suggesting that XFC can improve the formation of synovial pannus in AA rats.The mechanism of XFC improving the formation of synovial pannus in AA rats:XFC decreased the microvessel density of AA rats,significantly decreased the number of immature and mature vessels in the synovium of joints,and decreased the positive expression of CD31,an angiogenic index,suggesting that XFC can improve the formation of joint synovium pannus by inhibiting angiogenesis.XFC reduces the expression of platelet activators VEGF,PDGF and TSP-1 in RA patients and AA rats,and down-regulates platelet parameters,suggesting that XFC can improve the formation of pannus in RA patients by inhibiting platelet activation,regulating angiogenesis..