| Objective:Rheumatoid arthritis(RA)is an inflammatory autoimmune disease characterized by polyarticular synovitis and pannus formation,and its pathogenesis remains obscure.Studies have shown that serum amyloid A(SAA)as an acute phase protein may be related to the pathogenesis of RA.In this study,a three-dimension in vitro model of angiogenesis was constructed to explore the role of SAA induced p38 mitogen-ativated protein kinase(MAPK)signaling pathway in angiogenesis mechanisms of RA.Furthermore,a variety of immunological techniques were used to analyze the expression of SAA in serum,synovial fluid and synovium in RA patients.Correlations of SAA serum levels with disease activity[Disease Activity Score for 28 joints(DAS28)],erythrocyte sedimentation rate(ESR),C-reactive protein(CRP),and autoantibodies were assessed,respectively.The aim of this study is to lay the foundation for the explorations of the effects of SAA in the pathogenesis of RA and find new targets for diagnosis and therapy.Methods:1.The activation and expression of p38 MAPK in human umbilical vein endothelial cell(HUVECs)were analyzed by western blot.2.The role of p38MAPK signaling pathway in SAA induced HUVECs proliferation,migration and angiogenesis were analysed by MTT assay,cell migration assay,and in vitro model of angiogenesis,respectively.3.Serum SAA levels were measured by ELISA in patients with established RA,systemic lupus erythematosus(SLE),other autoimmune disease(others),osteoarthritis(OA),and healthy controls(HC).SAA levels in synovial fluid(SF)were also measured by ELISA in patients with RA and patients with OA.4.Expression of SAA in synovium was detected by immunohistochemistry.5.Correlations of serum SAA levels with disease activity(DAS28),ESR and CRP were assessed,respectively.Serum SAA levels were also detected in RA patients whose RF-IgA,RF-IgG,and anti-CCP antibodies were positive and negative.Results:1.In the stimulation of SAA,p38MAPK signaling pathway was activated and the expression of phospho-p38MAPK was obviously strengthened.2.With the inhibitor of p38MAPK,proliferations of HUVECs were inhibited in a dose dependent.SAA can stimulate the migration and HUVECs tube formation,but these effects were significantly reduced by adding p38MAPK inhibitor.3.Serum SAA levels in RA[(6.15±3.27)mg/l]was significantly higher(P<0.05)compared with those in SLE[(2.54±0.31)mg/l],OA[(1.42±0.97)mg/l],others[(1.55±0.98)mg/l]and HC[(1.45±0.72)mg/l].SAA levels in SF were significantly higher(P<0.001)in RA[(7.63±3.39)mg/l]compared to that in OA[(1.54±0.4 mg/1)].4.Pathology showed that SAA was observed in endothelial cells,synovial fibroblasts,macrophages and perivascular areas in RA synovium.In OA,SAA was observed in perivascular areas and synovial fibroblasts.5.Significant correlations were observed between serum SAA levels and DAS28(R2=0.6147,P<0.001),ESR(R2=0.4422,P<0.001),and CRP(R2=0.3919,P<0.001)in RA.Patients with RA who were positive for RF-IgA and RF-IgG had increased levels of SAA compared to seronegative patients but with no significance(mean 6.53±3.33 mg/1 and 5.41±3.00)mg/1,P= 0.127;6.50±3.17 mg/1 and 5.29±3.37 mg/1,P= 0.116).RA patients who were positive for anti-CCP had significantly increased SAA levels compared to seronegative patients(mean 6.88±3.36 mg/1 and 4.20±2.02 mg/I,P<0.001).Anti-CCP had no correlation with DAS28(R2=0.0154,P=0.250).Conclusions:1.P38MAPK signaling pathway was involved in SAA induced HUVECs proliferation,migration and angiogenesis,which indicates that p38MAPK signaling pathway may be involved in pannus formation and bone and cartilage damage in RA.2.Serum SAA levels were increased in RA compared with controls,which suggests that SAA may play a role in inflammation in RA.3.A significant correlation was observed between serum SAA levels and DAS28 in RA.It may be a new serum biomarker in the assessment of disease severity... |
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