Characterizing the Role of ATF3 in Regulating Adipogenesis During Age-Associated Thymic Involution

表征 ATF3 在年龄相关胸腺退化过程中调节脂肪生成的作用

基本信息

批准号：
9912068
负责人：
Angel Edgardo Flores
金额：
$ 6.84万
依托单位：
UNIVERSITY OF GEORGIA
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-04-15 至 2022-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9912068
关键词：
Adaptive Immune System Address Adipocytes Age Age-Months Aging Bacteria Biological Process Blood Vessels Cancerous Cell Lineage Cells Data Dendritic Cells Disease Elderly Engineering Epidemiology Epithelial Epithelium Face Fatty acid glycerol esters Female Flow Cytometry Freezing Gene Deletion Genes Genetic Goals Health Human Image Immunohistochemistry Immunology Incidence Inflammatory Investigation Island Label Laboratories Lipids Mesenchyme Molecular Mus Output PPAR gamma PTPRC gene Parasites Pathway interactions Pattern Population Process Production Proteins Regulation Reporter Research Research Personnel Risk Role Self Tolerance Specificity Stromal Cells T-Lymphocyte TACSTD1 gene Techniques Testing Thymic Tissue Thymic epithelial cell Thymus Gland Tissues Tomatoes Virus Wild Type Mouse activating transcription factor 3 adaptive immunity aged base capsule cell type cytokine design emerging adult experimental study fighting functional loss functional restoration immune function immunocytochemistry immunosenescence improved lipid biosynthesis male mutant novel pathogen sex therapy design transcriptome sequencing

项目摘要

Immunosenescence caused by age-associated involution of the thymus poses a significant risk to the aging human population. During age-associated thymic involution, a reduction and disorganization of the functional regions of the thymus, along with an increase in adipogenic and other undesirous cell types, results in a lower capacity of the thymus to generate functional T-cells required for adaptive immunity. Currently, the cellular origins of thymic adipocytes remain obscure, with some investigators suggesting that adipocytes infiltrate the thymus during involution and others indicating that they differentiate directly from thymic stromal cells. Our lab has identified a potential regulator of adipogenesis in the thymus, the Activating Transcription Factor 3 (ATF3), in which deletion of the Atf3 gene results in an increased presence of lipid-laden thymic stromal cells at 2- months of age in mice when assessed by both flow cytometry and immunohistochemistry. At 10-months of age, FACs analysis of adipogenic cells in Atf3 mutants and wild-type controls show that they are similar quantitatively. However, qualitative analysis by immunohistochemistry reveals differences in adipogenic cell types in Atf3 homozygous and heterozygous mutants, which warrants further investigation. We have also conducted a lineage trace study using Atf3null/null; Foxn1Cre/+; Rosa26Tom/+ mice to identify if a subset of thymic epithelial cells (TECs) undergo adipogenesis by using imaging flow cytometry. We identified four major classes of TEC-derived adipogenic cells (LipidTox+EpCAM+Ly51+, LipidTox+EpCAM+Ly51-, LipidTox+EpCAM- Ly51-, and LipidTox+EpCAM-Ly51+ cells), which include lipid-laden cTECs and mTECs. In frozen thymic sections, we have identified lipid-laden cTECs that express PPARy starting at 6, 7, and 10-months of age. In addition, we identified adipogenic mTECs that express FSP1, a marker for EMT, when we looked at 10-month old mice. These findings suggest that although both cTECs and mTECs become adipogenic, each may be regulated by different molecular mechanisms. We have also looked at tissue from a PPARγ-tdTomato reporter mouse engineered by the laboratory of Diane Mathis at Harvard. Using the reporter mice, we have identified adipogenic vascular-associated cells and the presence of globular fat cells within the lumen of the vasculature that appear to be infiltrating the thymus. Overall, we have begun to characterize the different classes of thymic adipocytes more fully, and we are beginning to understand more about the genes that may be governing the process. We have optimized techniques to study thymic adipocytes using standard and imaging flow cytometry and immunocytochemistry. We have access to a novel mouse line, the PPARy-tdTomato line, that we plan on using for our research purposes. These data suggest that both cTECs and mTECs give rise to a subpopulation of thymic adipocytes and that ATF3 is a likely repressor of adipogenesis during thymic involution. In Aim 1, we will identify the cell type in which ATF3 acts to regulate thymic adipogenesis. In Aim 2, we will identify the mechanisms by which ATF3 regulates adipogenesis.

由年龄相关的胸腺退化引起的免疫衰老对衰老构成重大风险人类群体在与年龄相关的胸腺退化过程中，功能减少和瓦解。胸腺区域，以及脂肪生成和其他不良细胞类型的增加，导致较低的胸腺产生适应性免疫所需的功能性 T 细胞的能力。胸腺脂肪细胞的起源仍然不清楚，一些研究人员认为脂肪细胞浸润退化过程中的胸腺和其他表明它们直接从胸腺基质细胞分化。已经确定了胸腺中脂肪生成的潜在调节因子，即激活转录因子 3 (ATF3)，其中 Atf3 基因的缺失导致 2- 载脂胸腺基质细胞的存在增加通过流式细胞术和免疫组织化学对小鼠月龄进行评估。 Atf3 突变体和野生型对照中的脂肪形成细胞的 FAC 分析表明它们相似然而，免疫组织化学的定性分析揭示了脂肪形成细胞的差异。 Atf3 纯合子和杂合子突变体的类型，这值得我们进一步研究。使用 Atf3null/null 的 Rosa26Tom/+ 小鼠进行谱系追踪研究，以确定胸腺的子集；通过使用成像流式细胞术，我们确定了四种主要的上皮细胞（TEC）发生脂肪形成的过程。 TEC 衍生的脂肪形成细胞类别（LipidTox+EpCAM+Ly51+、LipidTox+EpCAM+Ly51-、LipidTox+EpCAM- Ly51- 和 LipidTox+EpCAM-Ly51+ 细胞），其中包括冷冻胸腺中的脂质负载 cTEC 和 mTEC。在切片中，我们鉴定出负载脂质的 cTEC，这些 cTEC 在 6、7 和 10 个月龄时开始表达 PPARy。此外，当我们观察 10 个月时，我们发现了表达 FSP1（EMT 标志物）的成脂 mTEC 这些发现表明，虽然 cTEC 和 mTEC 都具有脂肪生成作用，但它们都可能具有脂肪形成作用。我们还研究了 PPARγ-tdTomato 报告基因的组织。哈佛大学黛安·马西斯实验室利用报告小鼠培育出的小鼠，我们已经鉴定出。脂肪形成血管相关细胞和脉管系统管腔内存在球状脂肪细胞总体而言，我们已经开始描述胸腺的不同类别。脂肪细胞更全面，我们开始更多地了解可能控制脂肪细胞的基因我们优化了使用标准和成像流式细胞术研究胸腺脂肪细胞的技术。我们已经获得了一个新的小鼠品系，即我们计划开发的 PPARy-tdTomato 品系。用于我们的研究目的。这些数据表明 cTEC 和 mTEC 都会产生一个亚群。在目标 1 中，我们发现 ATF3 可能是胸腺退化过程中脂肪生成的抑制因子。将鉴定 ATF3 在其中调节胸腺脂肪生成的细胞类型。 ATF3 调节脂肪生成的机制。