Study On Association Of Seronegative Spondyloarthropathy (SpA) With HLA-B27 Alleles And Construction Of DNA Micro Array For HLA-B27 Genotyping

BackgroundAnkylosing Spondylitis (AS), Reactive Arthritis (ReA), Reiter's Syndrome (RS), Psoriatic Arthritis (PsA), Inflammatory Bowel Disease Arthritis (IBDA), Juvenile onset Spondyloar-thropathies (JspA), Whipple Disease, Synoriti Acne Pusrulosis Hyperostosis and Osteitis (SAPHO) Syndrome, Behcet's Disease (BD) and Undifferentiated Spondyloarthropathy (uSpA) form a group of inflammatory rheumatic diseases identified as the Seronegative Spondyloarthropathies/Spondyloarthropathy (SpA).These diseases have a common character which Rheumatoid Factor(RF) is negative expression. In the past twenty years, the relatively mechanical studies have been shown that inheritance is one of the key pathogenic factors, in which HLA-B27 is the most essential and important factor. It has been confirmed that HLA-B27 appears to play a direct role in the pathogenesis of SpA, although it has different pathogenic characteristics and clinical appearances. However, its pathogenic mechanism is still unclear.HLA-B27 is a serologic specificity that encompasses 26 different alleles that encode 24 different products (proteins)-HLA-B 2701 to B 2725, with the exclusion of B 2722, which was deleted as an official WHO allele in April 2002, with a note that the reference cell has shown to have the same sequence as B*2706. And these alleles are also called subtypes. These B27 subtypes are distinguished from each other by one or more amino acid substitutions, resulting from changes mostly in exons 2 and 3, which encode the alpha 1 and alpha 2 domains of the B27 molecule, respectively. Among these 24 alleles, B* 2705 is the most widespread subtype. B*2702, B*2704 and B*2705 have been reported to be clearly, associated with SpA, while there are two alleles have been confirmed to be not disease-associated: B* 2706 among Southeast Asian populations, and B* 2709 among Sardinians. The distinction between the disease-associated subtypes and those are not disease-associated may provide some clues to the actual role of HLA-B27 in disease pathogenesis. Therefore, a diagnosis of AS can virtually be excluded in the absence ofHLA-B27. Many techniques have been used for HLA-B27 typing, such as Microlymphocytotoxicity Test (MCLT), Flow Cytometry (FCM), PCR-Sequence Specific Primer (PCR-SSP) and PCR-Sequence Specific Oligonucleotide (PCR-SSO), et al. Compared with these methods, DNA micro array had many advantages such as higher discriminability, more stability, accuracy and higher speed. It has great significance to study HLA-B27 alleles and AS families. So the construction of a new technology of DNA micro array for HLA-B27 genotyping , analysis of B27 alleles among healthy, SpA patients, and AS families in the Han Nationality Chinese in Chongqing, not only will help to study the mechanism of association between HLA-B27 gene and SpA, but also will help the diagnosis, prognosis and prevention of SpA. Objectives1. Epidemiological study on the incidence of Seronegative Spondyloarthropathy (SpA) and the frequency of HLA-B27 antigens and gene expression among the Han Nationality Chinese in Chongqing , and the great variety pattern of HLA-B27 alleles expressed among health and SpA patients; meanwhile attempt to find out some new or infrequent B27 alleles which are different from that of other countries, regions and races. And exploration the role of HLA-B27 gene in the pathology of Ankylosing Spondylitis(AS) by detecting the positive frequency of HLA-B27 antigen expression and HLA-B27 allele expression in AS families, help the diagnosis, prognosis and prevention of AS.2. Detection and analysis genotyping of ACE in normal controls and AS patients, whose HLA-B27 antigens are positive expression, trying to figure out the association of ACE alleles with AS patients in whom HLA-B27 antigens are positive expression.3. Evaluation the methods of Microlymphocytotoxicity Test (MCLT), Flow Cytometry (FCM), PCR-Sequence Specific Oligonucleotide (SSO), PCR-Sequence Specific Primer (SSP), Sequence Based Typing(SBT) and the technology of DNA Micro array, which detect the...