Use Of Microarrays and Epigenetics In Gene Expression Of Uveitis & AMD Patients

微阵列和表观遗传学在葡萄膜炎基因表达中的应用

• 批准号: 8737625
• 负责人: Robert Nussenblatt
• 金额: $ 36.4万
• 依托单位: NATIONAL EYE INSTITUTE
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/8737625
• 关键词: Acetylation; Adrenal Cortex Hormones; Affect; Age related macular degeneration; Anti-Inflammatory Agents; Anti-inflammatory; Apoptosis; Autoimmune Process; Autoimmune Responses; Autoimmunity; B-Lymphocytes; Biological Markers; Blindness; CD4 Positive T Lymphocytes; Cells; Characteristics; Choroid; Chromatin; DNA Methylation; DNA Sequence; Deacetylation; Development; Diagnosis; Disease; Epigenetic Process; Etiology; Exposure to; Functional RNA; Gene Chips; Gene Expression; Genes; Genetic Transcription; Immune; Immune system; Inflammatory; Inflammatory Response; Interleukin-10; Interleukin-17; Investigation; Length; Leukocytes; Memory; Messenger RNA; Methylation; MicroRNAs; Modification; Molecular; Molecular Profiling; Natural Killer Cells; Pathogenesis; Pathway interactions; Patients; Pattern; Pharmaceutical Preparations; Phenotype; Play; Population; Promoter Regions; Proteins; RNA; Refractory; Refractory Disease; Regulation; Reporting; Retina; Role; Siblings; Signal Pathway; Small Interfering RNA; Steroids; Subgroup; Techniques; Telomere Shortening; Th1 Cells; Therapeutic Intervention; Time; Twin Multiple Birth; Uveitis; active control; cytokine; demethylation; genetic association; genome wide association study; histone modification; interest; interleukin-22; monocyte; novel; peripheral blood; promoter; response; transcriptomics

Ocular inflammatory diseases, including uveitis, cause significant visual loss. Using a pathway specific gene chip with genes which are known to be involved in focused signaling pathways, e.g. inflammatory and autoimmune pathways, we found that there exist 4 distinct molecular gene expression profiles when comparing those from uveitis patients to those from normal donors. We termed those profiles molecular signatures for uveitis. Among those genes evaluated 9 genes had not been reported to be involved in uveitis. Of particular interest is the identification of IL-22. The expression of IL-22 has been recently associated with Th17 cells. We showed for the first time that IL-22 resulted in apoptosis in cultured primary RPE cells, possibly by decreasing the phosphorylated-Bad level. In addition, we saw increased IL-17 activity in the immune cells of patients with age related macular degeneration. This past year we have particularly concentrated on epigenetic changes associated with disease. The current understanding of epigenetics is the study of mechanisms that control somatically heritable gene expression status without changes in the underlying DNA sequence, including 1) DNA methylation/demethylation 2) Histone modification (Acetylation/deacetylation) 3). Chromatin structural modification and 4) Control of transcription by non-coding RNAs (siRNA, miRNA). We have initiated a long term investigation on the involvement of DNA methylation in the immune system, focusing on cell subpopulations and gene specific DNA methylation patterns and its involvement in autoimmunity and intraocular inflammatory disease. DNA methylation has been shown to participate in the control of hematopoeitic cell development. Comprehensive studies on DNA methylation in controlling cytokine expression in other immune cells, e.g., monocytes, NK cells and B cells, and genes with anti-inflammatory effect, e.g., IL-10 gene, have been lacking We have evaluated patients with age related macular degeneration, starting with twins who have disparate disease, then siblings and then a wider population of AMD patients. Although recent studies have demonstrated strong genetic associations between AMD and SNPs in a number of genes, other modes of regulation are also likely to play a role in the etiology of this disease. We identified a significantly decreased level of methylation on the IL17RC promoter in AMD patients. Furthermore, we showed that hypomethylation of the IL17RC promoter in AMD patients led to an elevated expression of its protein and messenger RNA in peripheral blood as well as in the affected retina and choroid, suggesting that the DNA methylation pattern and expression of IL17RC may potentially serve as a biomarker for the diagnosis of AMD and likely plays a role in disease pathogenesis. In addition, patients with steroid refractory uveitis have a characteristic subpopulation of steroid refractory CD4+ T cells in their peripheral blood. Previously studies have demonstrated that this steroid refractory phenotype is restricted to the central memory pool of CD4+ cells which have the capacity to generate IL-17. We therefore compared transcriptomic responses of Th1 and Th17 cells to corticosteroids in order to identify novel biomarkers and targets for therapeutic intervention in steroid refractory disease. Steroid referactory patients have a greater propensity than sensitive patients to generate Th17 cells, but Th17 cells from either group of patients have a similarly restricted change in gene expression following exposure to Dex compared with Th1 cells. Using additional techniques we have identified that a subgroup of uveitis patients have markedly shortened telomere length.

包括葡萄膜炎在内的眼部炎症性疾病会导致严重的视力丧失。使用具有已知参与聚焦信号传导途径（例如炎症和自身免疫途径）的基因的途径特异性基因芯片，我们发现当比较来自葡萄膜炎患者的那些与来自正常供体的那些时，存在4种不同的分子基因表达谱。我们将这些特征称为葡萄膜炎的分子标记。在这些基因中，有9个基因尚未报道与葡萄膜炎有关。特别感兴趣的是IL-22的鉴定。IL-22的表达最近与Th 17细胞相关。我们首次发现IL-22可能通过降低磷酸化Bad水平导致培养的原代RPE细胞凋亡。 另外我们 在年龄相关性黄斑变性患者的免疫细胞中观察到IL-17活性增加。 在过去的一年里，我们特别关注与疾病相关的表观遗传变化。目前对表观遗传学的理解是研究控制体细胞遗传基因表达状态而不改变潜在DNA序列的机制，包括1）DNA甲基化/去甲基化2）组蛋白修饰（乙酰化/去乙酰化）3）。染色质结构修饰和4）通过非编码RNA（siRNA，miRNA）控制转录。 我们已经启动了一项关于DNA甲基化在免疫系统中的参与的长期研究，重点是细胞亚群和基因特异性DNA甲基化模式及其在自身免疫和眼内炎性疾病中的参与。DNA甲基化已被证明参与造血细胞发育的控制。关于DNA甲基化在控制其他免疫细胞中细胞因子表达方面的综合研究，例如，单核细胞、NK细胞和B细胞，以及具有抗炎作用的基因，例如，IL-10基因，一直缺乏 我们评估了年龄相关性黄斑变性患者，从患有不同疾病的双胞胎开始，然后是兄弟姐妹，然后是更广泛的AMD患者人群。尽管最近的研究已经证明AMD和许多基因中的SNP之间存在强烈的遗传关联，但其他调节模式也可能在这种疾病的病因学中发挥作用。我们发现AMD患者IL 17 RC启动子甲基化水平显著降低。此外，我们发现AMD患者中IL 17 RC启动子的低甲基化导致其蛋白和信使RNA在外周血以及受影响的视网膜和脉络膜中的表达升高，这表明IL 17 RC的DNA甲基化模式和表达可能潜在地用作诊断AMD的生物标志物，并且可能在疾病发病机制中起作用。 此外，类固醇难治性葡萄膜炎患者在其外周血中具有类固醇难治性CD 4 + T细胞的特征性亚群。先前的研究已经证明，这种类固醇难治性表型仅限于具有产生IL-17能力的CD 4+细胞的中央记忆库。因此，我们比较了Th 1和Th 17细胞对皮质类固醇的转录组反应，以确定新的生物标志物和类固醇难治性疾病的治疗干预靶点。类固醇激素依赖患者比敏感患者有更大的倾向产生Th 17细胞，但与Th 1细胞相比，来自两组患者的Th 17细胞在暴露于Dex后的基因表达变化相似。 使用额外的技术，我们已经确定，一个亚组的葡萄膜炎患者有显着缩短端粒长度。