| Ankylosing spondylitis is a chronic inflammatory disease, one of the spA (Spondyloarthropathies) . Mainly affects the sacroiliac joints, spine, joints and around tendons, ligaments and other tissue, which is often develop inflammation secondary to fibrosis and calcification, the gradual loss of flexibility to the spine, will develop into serious as the "bamboo", than some patients as above can not be bent or stretched. The main pathological features of AS include: sacroiliitis, spinal inflammation, new bone formation in spine and joint fusion may occur also. The progression of AS has two main stages: first, inflammation state company with erosion of bone caused by inflammation, followed with osteophyte formation often affects ligament. In this study, we make a preliminary understanding of the inflammation and ossification related factors in AS by detecting the DKK-1 and TNF-αexpression levels of serum in AS, rheumatoid arthritis (rheumatoid arthritis, RA) and healthy control ,and then to know the meaning of expression ,to make sure if the different stage of disease in patients with ankylosing spondylitis serum factor have significant differences and provide a theoretical basis for the screening and identification of protein expression profile with differences in different disease states of serum in ankylosing spondylitisTNF-αis a major pro-inflammatory cytokines, and relative to activity levels of disease state and inflammation in RA patients and osteoarthritis animal models, which play a key role in the pathogenesis, but the pathogenesis of AS is unclear. Wnt signaling pathway is a representative of the bone related signaling pathways, DKK-1 as an inhibitor of Wnt signaling pathway has been confirmed; at the same time, we have found that blocking the DKK-1 can induce joint fusion in osteoarthritis mice models, indicating that DKK-1 in mice models of arthritis plays an important role about regulation of joint remodeling, but in human joint disease whether plays the same work remain unclear. Therefore, detecting the level of DKK-1 and TNF-αin AS patients, RA patients, healthy controls, and the expression level of different stages in AS patients, which can give a contribution to understand the pathogenesis of DKK-1 and TNF-αin AS, and then provide a theoretical basis for the next analysis of serum proteomics in ankylosing spondylitis which maybe help to find some protein molecules are related to the specificity of AS, more ,can provide some new molecular targets for drug design or some new markers for the diagnosis of early ankylosing spondylitis.Our laboratory detected the expression levels of DKK-1 and TNF-αin serum of different stages of ankylosing spondylitis, found that some factors are relevant with inflammation and ossification show difference in expression of different stages of ankylosing spondylitis. Then we take the combination of proteomics technology and bioinformation analysis, find other cytokines which are associated with ossification or inflammation and is verified by ELISA.Chapter One: The expression of DKK-1 and its significance in serum of patients with ankylosing spondylitisObjectives: To detect the expression of Dickkopf-1 (DKK-1) and tumor necrosis factor (TNF-α) in serum of ankylosing spondylitis with different stages in order to study the relationship of bone-related factors and AS.Methods: The expression of DKK-1 and TNF-αwas detected by enzyme-linked immunosorbent assay (ELISA) in 47 cases of AS patients (22 cases of early stage and 25 cases of late stage), 20 cases of rheumatoid arthritis (RA) patients and 20 cases of healthy control.Results: The expression of DKK-1 and TNF-αin AS and RA were significantly higher than control, but AS and RA were not significantly different. The expression of DKK-1 in late stage was significantly decreased with early stage, TNF-αalso decreased in late stage patients, but not significant. Spearman analysis suggests that DKK-1 have positive correlative with TNF-αin AS.Conclusion: With the progress of ossification in AS, the expression of DKK-1 in a down regulated state, suggesting that the level of DKK-1 may be involved in ossification of AS, DKK-1 may be a potential marker which is associated with the degree of ossification in serum of AS.Chapter two: Comparative Proteomic of ankylosing spondylitis serum at Differential Clinical StagesObjective: ankylosing spondylitis clinical stages associated proteins would be found by comparing differential expressed proteins from differential clinical stage ankylosing spondylitis serum.Methods: Total proteins from ankylosing spondylitis serum was extracted; differential proteome profiles were established and analysed by means of immobilized pH gradient based two dimensional polyacrylamide gel Electrophoresis (2D-PAGE) and matrix assisted laser desorption/ ionization time of flight mass spectrometry ( MALDI-TOF-MS) .Results: Well resolved reproducible 2-DE profiles of human ankylosing spondylitis serum were obtained. By analysis with mass spectrometry and bioinformatics, 8 of them were well characterized. 6 proteins were overexpressed in early stage group, including Complement factor B,Complement factor I,Alpha-2-macroglobulin,Haptoglobin,Clusterin,Prothrombin; 2 proteins were overexpressed in medium advanced stage, including Hemoglobin subunit beta,Complement C3.Conclusion: Differential expressed proteins exist in clinical stage of ankylosing spondylitis, which would be biomarkers for diagnosis and prediction of prognosis.
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