Determining the role of oxysterols in lymphocyte homing to lymph nodes in homeostasis and inflammation

确定氧甾醇在淋巴细胞归巢至淋巴结的稳态和炎症中的作用

基本信息

批准号：
10677408
负责人：
KEVIN Y CHEN
金额：
$ 3.98万
依托单位：
UNIVERSITY OF CALIFORNIA, SAN FRANCISCO
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-03-01 至 2027-02-28
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10677408
关键词：
25-hydroxycholesterol Adhesions Adoptive Transfer Antibody Response Antigens Area Autoimmunity Automobile Driving B-Cell Activation B-Lymphocytes BLR1 gene Basal lamina Binding Biological Assay Blood Blood Circulation Blood capillaries CCL19 gene CCL21 gene CD4 Positive T Lymphocytes CXCR4 gene Capillary Endothelial Cell Carbohydrates Cells Cellular Immunity Chemotactic Factors Cholesterol Chronic Circulation Complement Complement 3d Receptors Coupled Data Defect Dendritic Cells Dependence Development Disease Drops Endothelium Enzymes Exhibits Extravasation Fibroblasts G-Protein-Coupled Receptors Generations Genes Genetic Transcription Goals High Endothelial Venule Homeostasis Homing Hour Human Herpesvirus 4 Hydroxylation Immune Immunization Immunologic Surveillance Immunotherapy In Vitro Infection Inflammation Interferons Invaded Knock-out Knockout Mice Knowledge Ligands Lymphatic Lymphocyte Lymphocytic Infiltrate Lymphoid Tissue Malignant Neoplasms Mediating Memory Migration Assay Mixed Function Oxygenases Mus Organ Peripheral Peyer&apos s Patches Process Property Publishing Reporting Role Scanning Signal Transduction Site Skin Stains Stimulus Stromal Cells Surface T-Lymphocyte Testing Toll-like receptors Virus Diseases Visualization Work acute infection adaptive immune response antigen test cell motility cell type chemokine design draining lymph node experimental study fMet-Leu-Phe receptor improved in vivo intravital imaging lymph nodes lymphatic circulation migration novel novel therapeutics pathogen postcapillary venule protein expression receptor reconstitution recruit selective expression sialomucins trafficking tumor tumorigenesis two photon microscopy vascular addressins

项目摘要

PROJECT SUMMARY/ABSTRACT Lymphocyte entry into lymph nodes (LNs) from blood is a key homeostatic process for efficient initiation of an adaptive immune response to pathogens. Naïve B and T cells traffic through LNs to scan for foreign antigens delivered to and concentrated in these organs from diverse sites of potential infection. If no antigens are encountered, lymphocytes exit LNs into lymphatic circulation before returning to the blood and beginning the cycle again. This constant lymphocyte recirculation requires large scale extravasation into lymphoid tissue under non-inflammatory conditions, and the specialized vessels supporting this process in LNs are called high endothelial venules (HEVs). HEVs express and luminally present several vascular ‘addressins’ and chemokines that together support a multi-step adhesion cascade for lymphocyte entry into LNs. The signals promoting lymphocyte arrest on HEV walls are well-elucidated, but the chemoattractants driving transmigration across the endothelium are not fully understood. Elucidating the ligands and receptors that mediate this fundamental step of entry is essential for understanding and manipulating immune cell trafficking in diseases such as cancer and autoimmunity. The enzyme Ch25h, which produces 25-hydroxycholesterol (25HC) from cholesterol, is highly and selectively expressed in HEVs compared to capillary endothelium in LNs. Furthermore, Cyp7b1, which hydroxylates 25HC to generate 7𝛼𝛼,25-dihydroxycholesterol (7,25HC), is expressed in stromal cells surrounding LN HEVs. 7,25HC is a potent ligand for EBI2, a GPCR that helps guide activated B cell movement within LNs and is also highly expressed in naïve B and T cells. The capability for oxysterol synthesis by HEVs suggests EBI2 and 7,25HC may play a role in mediating lymphocyte entry into LNs. We hypothesize that a gradient of 7,25HC across HEVs supports post-adhesion transendothelial migration of naïve B and T cells. In preliminary studies, EBI2 KO B cells and CD4 T cells displayed a recruitment defect to LNs in adoptive transfers, and a similar homing defect was evident in mice lacking Ch25h or Cyp7b1. Furthermore, dependency on EBI2 for lymphocyte entry into LNs increased in a viral infection setting. We hypothesize this is due to elevated 7,25HC levels and reduced chemokine levels in inflamed LNs. From these results, we aim to determine the step of LN entry that EBI2 directs via a combination of in vivo homing assays, intravital imaging, and in vitro migration assays (Aim 1). In addition, we will examine EBI2’s role in maintaining lymphocyte homing in inflamed LNs by characterizing the changes, and signals mediating these changes, in LN oxysterol and chemokine levels during infection (Aim 2). Overall, this work will define a novel chemoattractant driving lymphocyte migration into LNs and advance our understanding of the transmigration process. As well as their role in LNs, HEVs form in tumor associated tertiary lymphoid tissues and at sites of chronic inflammation. Our findings could inform development of novel therapeutics that modulate lymphocyte infiltration in these disease settings.

项目概要/摘要淋巴细胞从血液进入淋巴结 (LN) 是有效启动免疫反应的关键稳态过程。对病原体的适应性免疫反应通过 LN 扫描外来抗原。如果没有抗原，则从不同的潜在感染部位输送并集中在这些器官中。遇到这种情况时，淋巴细胞会离开淋巴结进入淋巴循环，然后返回血液并开始淋巴循环。这种持续的淋巴细胞再循环需要大规模外渗到淋巴组织中。非炎症性疾病，并且 LN 中支持这一过程的特殊血管被称为高内皮小静脉 (HEV) 表达并在管腔内呈递多种血管“地址素”和趋化因子。它们共同支持淋巴细胞进入 LN 的多步粘附级联。淋巴细胞在 HEV 壁上的滞留已得到很好的阐明，但驱动跨膜轮回的化学引诱剂内皮细胞尚未完全阐明介导这一基本步骤的配体和受体。进入对于理解和操纵癌症等疾病中的免疫细胞贩运至关重要 Ch25h 酶可从胆固醇产生 25-羟基胆固醇 (25HC)，具有高度的自身免疫性。与 LN 中的毛细血管内皮相比，Cyp7b1 在 HEV 中选择性表达。羟基化 25HC 生成 7𝛼𝛼,25-二羟基胆固醇 (7,25HC)，在周围基质细胞中表达 LN HEV 是 EBI2 的有效配体，EBI2 是一种 GPCR，有助于引导激活的 B 细胞在 LN 内运动。 HEV 合成氧化甾醇的能力表明它在幼稚 B 细胞和 T 细胞中也高度表达。 EBI2 和 7,25HC 可能在介导淋巴细胞进入 LN 中发挥作用。初步研究表明，HEV 中的 7,25HC 支持初始 B 细胞和 T 细胞的粘附后跨内皮迁移。研究发现，EBI2 KO B 细胞和 CD4 T 细胞在过继转移中表现出 LN 募集缺陷，并且类似的归巢缺陷在缺乏 Ch25h 或 Cyp7b1 的小鼠中也很明显。此外，对 EBI2 的依赖性。在病毒感染环境中，淋巴细胞进入淋巴结的数量增加，我们发现这是由于 7,25HC 升高所致。从这些结果中，我们的目标是确定 LN 的步骤。 EBI2 通过体内归巢、活体成像和体外迁移测定相结合来引导进入此外，我们将通过以下方法检查 EBI2 在维持炎症 LN 中淋巴细胞归巢方面的作用。描述 LN 氧化甾醇和趋化因子水平的变化以及介导这些变化的信号总体而言，这项工作将定义一种驱动淋巴细胞迁移到 LN 的新型趋化剂。并加深我们对迁移过程及其在 LN、HEV 在肿瘤中形成的作用的理解。我们的研究结果可以为发育提供信息。调节这些疾病中淋巴细胞浸润的新疗法。