| Objective To disscution the diagnostic value of flow cytometry(FCM) and sequence specific primer polymerase chain reaction(PCR-SSP) for detection of human leukocyte surface antigen-B27(HLA-B27) in patients with ankylosing spondylitis(AS); To observe the distribution of HLA-B27 allele subtypes in the region; To understand the mechanism of the immune system in the pathogenesis of AS.Methods Through case retrospective analysis, know the basic situation of the HLA-B27 detection of the population, to explore the value of the detection of HLA-B27 with FCM method to the diagnosis of AS; PCR-SSP method and FCM method were used to detect HLA-B27 in AS patients, suspected AS patients group and healthy control group, and HLA-B27 allele subtypes were identified, and the results were statistically analyzed using SPSS17.0 statistical software; The methods of two kinds of detection methods were evaluated by the receiver operating characteristic(ROC) curve; The absolute count of lymphocyte subsets and related cytokines were detected in the subgroup, using independent sample t test and variance analysis to analyze the data, and to explore the mechanism of the immune system in the pathogenesis of AS.Results1. 246 cases suspected of ankylosing spondylitis patients age mainly concentrated in between 21~55 years, 42 cases were diagnosed as patients, including 28 cases of HLA-B27 positive cases, accounting for number of confirmed 66.7%. FCM method was used to detect HLA-B27 areas under ROC curve(AUC) was 0.758(95% CI 0.659~0.856), obtained by the optimal cut-off value of 14.75. On this critical point,the sensitivity was 66.7%, specificity is89.7%, Youden index for 0.564.2. Packet data PCR-SSP method for detection of HLA-B27 positive cases in 45 cases,including 1 case of B*2701; 5 cases of type B*2702; B*2704 28 cases; B*2705 11 cases. the area under the ROC curve of PCR-SSP method and FCM method detection of HLA-B27 were0.913, 0.884; The sensitivity and specificity of PCR-SSP method were 94.1%, 84.0%; FCM assay sensitivity, specificity respectively for 64.7%, 88.0%.3. Ankylosing spondylitis group and the healthy control group compared to CD4+T lymphocyte absolute gauges numerical increased significantly; CD8+T lymphocytes and Blymphocyte absolute count value decreased, with statistically significant differences(P<0.05);group of ankylosing spondylitis is suspected as group compared to CD3+T lymphocytes,CD8+T lymphocyte absolute value less than suspected group, and the difference is statistically significant, the differences of the other indicators without statistical significance;suspected ankylosing spondylitis group and healthy group compare suspected CD3+T lymphocyte and CD4+T lymphocyte absolute value was significantly lower than the results of healthy controls, the difference has statistical significance, other index difference was not statistically significant.4. AS group TNF-?, IFN-?, IL-10, IL-17 expression levels were significantly higher than the healthy control group,the difference was statistically significant(P<0.05); The level of IL-4 was significantly lower than the healthy control group, the difference was statistically significant; IL-2 and TGF-?1 levels than healthy control group increased, but there was no statistical significance. AS group compared to suspected group, the levels of each cytokine were significantly higher in TNF-?, IFN-?, IL-17,and the difference was statistically significant(P<0.05), the other factor differences had no statistical significance. Suspected group compared with the healthy control group in addition to TNF-?, IFN-?, IL-10, IL-17 increased, IL-4 levels were reduced, and the difference was statistically significant(P<0.05),the other factor expression level had no significant difference.Conclusion1. The prevalence of ankylosing spondylitis in Inner Mongolia is basically the same as the whole country, The incidence of the population is mainly concentrated in young men;FCM method has good stability in the detection of HLA-B27. However, the high proportion of gray area samples that bringing some trouble to clinical inspection work.2. The diagnostic efficacy of PCR-SSP for AS was significantly better than that of FCM method in the detection of HLA-B27. PCR-SSP method to detect HLA-B27 is the future trend of development; The HLA-B27 allele in Inner Mongolia region is mainly dominated by B*2704 and B*2705.3. The pathogenesis of AS is closely related to the immune system, especially CD4+T lymphocytes play a leading role in the development of AS disease, which can be further studied.4. TNF-?, IFN-?, IL-4, IL-10 were significantly associated with the disease progression of AS; TGF-?1, IL-2 is not the main factor affecting the progression of the disease; IL-17 to participate in disease, the clinical judgement of disease development has certain guiding significance, In addition, IL-17 may provide help for the future treatment of AS. |
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