Aberrant gene expression in CD4+CD28-T cells: mechanisms

描述(申请人提供)：衰老的T细胞亚群，特征是缺乏CD28，随着年龄的增长而发展，在类风湿性关节炎(RA)等慢性炎症性疾病中，在急性冠脉综合征中。CD4+亚群具有促炎性和细胞毒性，在心肌梗塞死亡患者的炎症滑膜和动脉粥样硬化斑块中发现，表明它们参与了自身免疫性疾病的过程。CD4+亚群异常表达杀伤细胞免疫球蛋白样受体(KIR)基因和穿孔素，通常在NK细胞上发现，并过度表达干扰素-γ、LFA-1和CD70。基因表达异常的原因尚不清楚。在初步研究中，我们发现正常T细胞具有KIR、穿孔素、干扰素-γ、LFA-1和CD70表达所必需的转录因子，但这些基因的表达被关键调控元件的甲基化抑制，这些元件的去甲基化足以促进转录。此外，我们发现，随着年龄的增长和RA患者，总T细胞DMA去甲基化，部分原因是DNA甲基转移酶(MTase)减少，抑制T细胞ERK和JNK信号以模拟随着年龄增加而减少的信号，会降低DNA MTase的表达并使DNA去甲基化。最后，我们发现CD28提供了共刺激信号，增加了DNA MTase的表达。我们推测，由JNK和/或ERK信号减弱引起的DNA低甲基化导致KIR、穿孔素、干扰素-γ、LFA-1和CD70在衰老和类风湿关节炎患者的CD4+亚群中过表达，从而发挥促炎和细胞毒作用。这将在3个具体目标中进行测试。我们将：1)比较调节元件的甲基化状态和候选基因在CD4+和CD4中的表达？类风湿关节炎患者和老年受试者外周血中的T细胞，以及外周血中的T细胞亚群。2)比较相同T细胞亚群中DNA MTase的水平，然后选择性地降低正常T细胞中DNA MTase的水平，并检测其对候选基因甲基化和表达的影响；3)比较相同T细胞亚群中ERK和JNK通路的信号，检测信号减少对DNA MTase表达和候选基因甲基化、表达和功能的影响。这些研究将确定改变这个病理亚群中基因表达的机制，并提出恢复这些细胞中正常基因表达的方法。

DESCRIPTION (provided by applicant): A senescent T cell subset, characterized by the absence of CD28, develops with aging, in chronic inflammatory diseases like rheumatoid arthritis (RA), and in acute coronary syndromes. The CD4+ subset is proinflammatory and cytotoxic, and is found in inflamed synovium as well as atherosclerotic plaques of patients dying from myocardial infarction, suggesting they participate in autoimmune disease processes. The CD4+ subset aberrantly expresses killer-cell immunoglobulin-like receptor (KIR) genes and perforin, normally found on NK cells, and overexpresses IFN-y, LFA-1, and CD70. The reason for the aberrant gene expression is unknown. In preliminary studies we found that normal T cells have the transcription factors necessary for KIR, perforin, IFN-y, LFA-1 and CD70 expression, but expression of these genes is suppressed by methylation of crucial regulatory elements, and demethylation of these elements is sufficient to promote transcription. Further, we found that total T cell DMA demethylates with age and in RA, due in part to decreases in DNA methyltransferases (Mtase), and that inhibiting T cell ERK and JNK signaling to mimic the decreased signaling that occurs with aging decreases DNA Mtase expression and demethylates DNA. Finally, we found that CD28 provides costimulatory signals increasing DNA Mtase expression. We hypothesize that DNA hypomethylation, caused by decreased JNK and/or ERK signaling, results in the overexpression of KIR, perforin, IFN-y, LFA-1 and CD70 in the CD4+ subset in aging and RA, conferring pro-inflammatory and cytotoxic functions. This will be tested in 3 specific aims. We will: 1) Compare methylation status of regulatory elements and expression of the candidate genes in CD4+ and CD4? T cells from RA patients and elderly subjects, and to CD4? cells in controls, 2) Compare DNA Mtase levels in the same T cell subsets, then selectively decrease DNA Mtase levels in normal T cells and examine the effects on candidate gene methylation and expression, and 3) Compare ERK and JNK pathway signaling in the same T cell subsets, and examine the effects of decreased signaling on DNA Mtase expression and candidate gene methylation, expression and function. These studies will identify mechanisms modifying gene expression in this pathologic subset, and suggest ways to restore normal gene expression in these cells.