Dna Methylation Level Of Transcriptional And Inflammatory Factors Of CD4+T Cell Subset In Vogt-koyabagi-harada Syndrome And Behcet's Disease

Background:Uveitis is a common intraocular inflammatory disease,which mainly involves in uveal,retina and retinal vascular and vitreous body.Vogt-Koyanagi-Harada(VKH)syndrome and Behcet's disease(BD)are two common uveitis entities in Asians.VKH syndrome is a multi-systemic autoimmune disease characterized by bilateral granulomatous panuveitis and other systemic involvements,such as vitiligo,auditory and central nervous system damage.BD is an autoinflammatory disease and its clinical characters include recurrent uveitis,oral ulcers,genital ulcers,and skin lesions.Nowadays,immunosuppressive drugs and corticosteroids are extensively used in the treatment of uveitis.The main regulation mechanisms included in epigenetic variation are DNA methylation,histone modification,and RNA-related silence.Among these,DNA methylation is an important epigenetic mechanism,which has been proven to encode inheritable information that can affect gene function without changing the DNA sequence.DNA methylation is involved in the regulation of the immune response and cytokine expression during T-cell differentiation.Early studies have shown that as a na?ve CD4+ T cell differentiated into a Th1 cell,increased IFN-? expression was associated with reduced DNA methylation level of the IFN-? promoter region,and that IL-4 silencing was related to DNA methylation of GATA3 and IL-4 genes.In recent years,more and more attention has been focused on the role of epigenetic alterations in the development of autoimmune disease.A growing body of literature has demonstrated the relationship between impaired T cell DNA methylation and some forms of autoimmune disease.Even though the etiology and pathogenesis of uveitis remain unclear,T-cell-driven cellular immune responses have been recognized to play an important role in the pathogenesis of VKH syndrome and BD.Various studies have found that abnormal expression of cytokines and aberrant frequency of T cell subsets are crucial in the development of uveitis.Early studies have demonstrated that patients with active VKH or BD with IFN-?,IL-17 A,TBX21,ROR?t abnormal expression,suggesting that immune dysfunction of T cell subsets can participate in the development of uveitis.Based on the above research background,we present the following problem: Whether DNA methylation at the promoter region of inflammatory and transcriptional factors in CD4+T cells is associated with the pathogenesis of uveitis?Part I DNA methylation level of transcriptional andinflammatory factors of CD4+T cell subset inVogt-Koyanagi-Harada syndromePurposeIn this study,we investigate the role of promoter methylation of transcriptional and inflammatory factors,including TBX21,GATA3,ROR?t,FOXP3,IFN-?,IL-4,IL-17 A and TGF-? in the development of Vogt-Koyanagi-Harada(VKH)disease.Methods1 The promoter methylation levels were detected by the Sequenom MassARRAY system in CD4+ T cells which were separated from healthy individuals and VKH patients.2 The mRNA expression level of GATA3,IL-4 and TGF-? in CD4+ T cells was analyzed by real-time polymerase chain reaction(RT-PCR).3 The correlation between promoter methylation and mRNA expression of GATA3,IL-4,and TGF-? was analyzed by Pearson correlation test.Results1 The promoter methylation levels of GATA3,IL-4,and TGF-? were significantly increased in active VKH patients compared to that observed in healthy controls(P< 0.05).2 A decreased mRNA expression of GATA3 and TGF-? was found in active VKH patients and the mRNA expression of GATA3,IL-4 and TGF-? was negatively correlated with their DNA methylation.3 Treatment with systemic corticosteroid and Cyclosporin A(Cs A)decreased the methylation level of GATA3 and TGF-? in inactive VKH patients and increased their mRNA expression.ConclusionsOur findings suggest that promoter hypermethylation of GATA3 and TGF-? in CD4+ T cells can reduce the mRNA expression and confers risk to VKH disease in Han Chinese.The therapeutic effect of CsA and corticosteroids may be associated with the reduction of the methylation level of GATA3 and TGF-? promoter,increase of the mRNA expression and decrease of the disease activity.Part II DNA methylation level of transcriptional and inflammatory factors of CD4+T cell subset in Behcet's diseasePurposeTo investigate the association between the promoters methylation level of transcriptional and inflammatory factors in CD4+T cell(including TBX21,GATA3,ROR?t,FOXP3,IFN-?,IL-4,IL-17 A and TGF-?)and Behcet's disease.Methods1 Matrix-assisted laser desorption/ionization-time of-flight mass spectrometer(MALDI-TOF-MS)was used to detect the promoter methylation levels in CD4+ T cells from BD patients and healthy individuals.2 Real-time PCR was applied to measure the mRNA expression level of GATA3,IL-4 and TGF-? in CD4+ T cells.3 Pearson correlation test was used to analyze the correlation between promoter methylation and mRNA expression of GATA3,IL-4,and TGF-?.Results1 The promoter methylation levels of GATA3,IL-4,and TGF-? were significantly higher in active BD patients than that in healthy individuals(P<0.05).2 The GATA3 and TGF-? mRNA expression were markedly reduced in active BD patients compared to healthy controls,and the DNA methylation level of GATA3,IL-4,and TGF-? were negatively correlated with their mRNA expression.3 Methylation levels of GATA3 and TGF-? were all significantly decreased in inactive BD patients compared to that observed in active.ConclusionsOur findings suggest that promoter hypermethylation of GATA3 and TGF-? in CD4+ T cells confers risk to BD in Han Chinese...