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Objective To study the relationship between DNA methylation inhibitor and histone deacetylase inhibitor (HDACI) with HLA-B27 gene expression. Methods Hela-HLA-B27 promoter stable cell line was developed as the first step and followed by detecting the effect of DNA methylation inhibitor(5-Aza-CdR) and HDACI. The synergetic effect of HDACI and cytokines as well as the effect of anti-TNF-α antibody, anti-IFN-β antibody, anti-IFN-γ antibody and Infiximab on HLA-B27 promoter activity were tested by measuring luciferase value. Then HLA-B27 mRNA expression level in CCL6-B27 genomic DNA stable cell line was measured by real-time PCR. Results Sodium butyrate(NaB) and valproic acid(VPA)could significantly up-regulate HLA-B27 promoter activity at 24 h (24.0±1.7) times, ( 17.4±2.2 ) times respectively,(P＜0.05). The synergetic effect between VPA with TNF-α, IFN-α, IFN-β, IFN-γ and PMA on HLA-B27 promoter activity was found. None of the antibody could inhibit the effect of HDACI. It also showed that NaB,TSA, VPA and 5-Aza-CdR could significantly increase HLA-B27 mRNA expression in CCL6-B27 stable cell line. Conclusion DNA methylation and HDACI can up-regulate HLA-B27 gene expression level.
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Objective To study the gene expression profile of peripheral blood and synovial fluid mononuclear cells (PBMC and SFMC) of patients with spondyloarthropathy (SpA) and try to identify genes which might be related to the pathogenesis of SpA.Methods Microarrays were carried out with 1 176 target gene filters using PBMC and SFMC of 5 patients with SpA,5 patients with rheumatoid arthritis (RA) and 6 healthy volunteers.More precise measurement was performed with semi quantitative PT PCR on 9 microarray positive genes.Results There was no significant difference in positive gene numbers between SpA and RA patients although the positive gene numbers were much higher in PBMC than in SFMC in both groups.It is noteworthy that in RA group there were 53 genes which expression in PBMC was much higher than in SFMC,while in SpA patients there were only 5 genes which expression in PBMC was much higher than in SFMC ( P
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AIM:To screen the effective chemicals,which can suppress the promoter activity of the HLA-B2705 gene as potential therapeutic agents.METHODS:The HeLa-HLA-B27,293T-HLA-B27 stable transfectants were used to monitor the effect of 12 264 chemicals through high throughput screening(HTS).Chemicals which regulates HLA-B*2705 promoter activity more than 150% or less than 60% were picked out for the further IC50/EC50 and cell viability detection.RESULTS:(1)The primary screening used by 293T-HLA-B27 stable transfectant yielded about 5.1% hits which either suppressed(556 chemicals)or enhanced(68 chemicals)the HLA-B*2705 promoter activity.(2)A reconfirmation screening was carried out with these 624 of the candidates using transfected HeLa-HLA-B27 cells.Seventy hits were confirmed.(3)Based on the bioinformatics of those positive hits,40 chemicals were selected for in-depth analysis by dose-response experiment and IC50/EC50 detection.Six suppressors showed potential pharmacological activities.Interestingly,two suppressors(celastrol and pristimerin)are derived from Leigongteng,a herbal medicine already used for several decades for treatment of immune regulatory and inflammatory diseases.Four active chemicals were computer designed with no relevance to the above structures.CONCLUSION:Chinese traditional herb Nansheteng and Leigongteng might be the potential drugs for HLA-B27 positive patients.These results provide new direction for research in both the therapeutics and the pathogenesis of spondyloarthritis.
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AIM: To determine the role of chemokine and its receptors in the pathogenesis of ankylosing spondylitis (AS). METHODS: Gene expression profiles of peripheral blood mononuclea r cells (PBMC) in 13 AS patients and 7 healthy volunteers were determined by cDN A microarray with 588 targeting gene filter. Differentiated expressed CXCR4 and its only ligand SDF-1 were confirmed by semi-quantitative RT-PCR, ELISA, immunoh istochemistry and FACS analysis using PBMC, synovial fluid mononuclear cells (SF MC) and synoviocytes. RESULTS: The gene expression profile of AS patients was signific antly different from those of healthy volunteers. Higher expression of CXCR4 in monocytes and CD8+ T lymphocytes from PBMC in AS patients were found with st atis tical significance (P