Endothelial cell injury by cytolytic lymphocytes

溶细胞淋巴细胞损伤内皮细胞

• 批准号: 6883159
• 负责人: Mitzi Nagarkatti
• 金额: $ 4.33万
• 依托单位: VIRGINIA COMMONWEALTH UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-04-01 至 2005-08-15
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6883159
• 关键词: CD44 molecule; CD95 molecule; cell cell interaction; cellular immunity; cytolysins; cytotoxic T lymphocyte; cytotoxicity; electron microscopy; enzyme linked immunosorbent assay; flow cytometry; genetically modified animals; high performance liquid chromatography; histochemistry /cytochemistry; interleukin 2; laboratory mouse; ligands; lymphokine activated killer cell; major histocompatibility complex; natural killer cells; polymerase chain reaction; pore forming protein; tissue /cell culture; vascular endothelium

DESCRIPTION (provided by applicant): Endothelial cell (EC) injury has been widely documented in certain infections, autoimmunity, allograft rejection, graft versus host disease and following immunotherapy with cytokines or immunotoxins. It is becoming increasingly clear that ECs are primary targets of immunologic attack, although the precise mechanism remains unclear. We have used interleukin-2 (IL-2)-induced vascular leak syndrome (VLS) as a model to study EC injury. IL-2 is the only FDA approved systemic therapy for treating metastatic renal cell carcinoma. It is also being used to treat metastatic melanomas, immunodeficiencies and viral infections including AIDS. Despite much success, the efficacy is limited by severe, life-threatening toxicity resulting from EC injury leading to VLS. Using CD44 knockout (KO), perforin KO and FasL mutant mice, we have demonstrated that IL-2 activates cytotoxic lymphocytes and such cells use CD44 to kill EC by triggering perforin and FasL. More recently, we have demonstrated that CD44v7 exon KO mice also exhibit decreased VLS and EC killing. Based on these studies, we will test the hypothesis that IL-2 treatment up regulates CD44 variant isoforms on cytolytic lymphocytes of which v7 isoform plays a key role in EC injury and induction of VLS. The specific aims are 1) To test the susceptibility of EC isolated from various organs to cytotoxicity mediated by IL-2 activated T/NK/NKT cells from WT, pefforin KO, Fas and FasL mutant mice. Using NK cell-deficient (NKCD-Tg) mice, the role of NK cells in IL-2 induced VLS will also be investigated. 2) To examine whether down regulation of killer cell inhibitory receptor (KIR)-like molecules and/or up regulation of stimulatory receptors on NK cells is responsible for CD44-mediated EC injury. Specifically, the expression of KiR-like molecules, including Ly49 and CD94/NKG2 present on NK cells, and the classical and non-classical MHC ligands on Ecs, will be studied. 3) Identification of the CD44 isoforms involved in EC injury caused by cytotoxic lymphocytes. 4) To address the mechanism by which CD44v7KO mice exhibit increased resistance to VLS. 5) Use of mimetics of CD44 or its ligand including mAbs against CD44s and CD44v isoforms, Pep-l(an inhibitor of HA), CD44Rg and CD44MutRg fusion proteins to prevent EC injury and vascular leak. Together, the current study should provide novel insights into the mechanism of immune cell-mediated EC injury and development of strategies to prevent vascular leak.

描述（由申请人提供）：内皮细胞（EC）损伤已在某些感染、自身免疫、同种异体移植排斥、移植物抗宿主病和细胞因子或免疫毒素免疫治疗后广泛记录。越来越清楚的是，内皮细胞是免疫攻击的主要靶点，尽管确切的机制仍不清楚。我们使用白细胞介素-2（IL-2）诱导的血管渗漏综合征（VLS）作为模型来研究EC损伤。IL-2是唯一FDA批准的用于治疗转移性肾细胞癌的全身疗法。它也被用于治疗转移性黑色素瘤，免疫缺陷和病毒感染，包括艾滋病。尽管取得了很大成功，但由于EC损伤导致VLS而产生的严重、危及生命的毒性，其功效受到限制。使用CD 44敲除（KO），穿孔素KO和FasL突变小鼠，我们已经证明，IL-2激活细胞毒性淋巴细胞和这些细胞使用CD 44通过触发穿孔素和FasL杀死EC。最近，我们已经证明，CD 44 v7外显子KO小鼠也表现出降低的VLS和EC杀伤。基于这些研究，我们将检验IL-2处理上调溶细胞淋巴细胞上的CD 44变体同种型的假设，其中v7同种型在EC损伤和VLS诱导中起关键作用。1）检测从不同器官分离的EC对来自WT、pefforin KO、Fas和FasL突变小鼠的IL-2激活的T/NK/NKT细胞介导的细胞毒性的敏感性。使用NK细胞缺陷型（NKCD-Tg）小鼠，还将研究NK细胞在IL-2诱导的VLS中的作用。2)研究NK细胞上杀伤细胞抑制性受体（KIR）样分子的下调和/或刺激性受体的上调是否是CD 44介导的EC损伤的原因。具体而言，将研究KiR样分子的表达，包括NK细胞上存在的Ly 49和CD 94/NKG 2，以及Ecs上的经典和非经典MHC配体。3)鉴定参与细胞毒性淋巴细胞引起的EC损伤的CD 44亚型。4)阐明CD 44 v7 KO小鼠对VLS的抗性增强的机制。5)CD 44或其配体的模拟物，包括针对CD 44 ε和CD 44 v同种型、Pep-I（HA的抑制剂）、CD 44 Rg和CD 44 MutRg融合蛋白的mAb，用于预防EC损伤和血管渗漏的用途。总之，目前的研究应该提供新的见解免疫细胞介导的EC损伤的机制和发展的战略，以防止血管渗漏。