Regulation of brain endothelial integrity by angiotensin receptors as treatment for cerebral malaria

血管紧张素受体调节脑内皮完整性治疗脑型疟疾

• 批准号: 10350640
• 负责人: ANA RODRIGUEZ
• 金额: $ 32.2万
• 依托单位: NEW YORK UNIVERSITY SCHOOL OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 2018
• 资助国家: 美国
• 起止时间: 2018-06-01 至 2024-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10350640
• 关键词: AGTR2 gene; Affect; Agonist; Angiotensin Receptor; Angiotensins; Antimalarials; Apoptosis; Binding; Biological Assay; Blood; Blood - brain barrier anatomy; Brain; Brain Diseases; Cardiovascular system; Cause of Death; Cells; Cerebral Malaria; Cessation of life; Characteristics; Complication; Concentration Camps; Development; Disease model; Edema; Electrical Resistance; Embryo; Endothelial Cells; Endothelium; Erythrocytes; Fingerprint; Foundations; Genetic Models; Goals; Hemorrhage; Human; Immunofluorescence Immunologic; Impaired cognition; In Vitro; Individual; Intercellular Junctions; Knock-out; Knockout Mice; Life; Maintenance; Malaria; Maps; Mediating; Modeling; Mus; Outcome; Parasites; Pathology; Patients; Permeability; Pharmacology; Plasmodium falciparum; Pre-Clinical Model; Predisposition; Proteins; Receptor Signaling; Regulation; Renin-Angiotensin System; Role; Second Messenger Systems; Sequence Homology; Signal Transduction; Signaling Molecule; Small Interfering RNA; Syndrome; Testing; Therapy Clinical Trials; Type 2 Angiotensin II Receptor; Virulent; Wild Type Mouse; angiotensin I (1-7); antagonist; arm; beta catenin; blood-brain barrier disruption; brain endothelial cell; brain tissue; endothelial dysfunction; experimental study; in vitro Assay; in vitro Model; in vivo; irbesartan; kidney cell; peptide analog; pre-clinical; prevent; protective effect; receptor; response; treatment effect

Summary. Cerebral malaria (CM) remains the most virulent and deadliest manifestation of malaria. It is caused by Plasmodium falciparum infected erythrocytes (iRBC) adhering to host brain endothelial cells and compromising the blood brain barrier. While available anti-malarial drugs are effective at clearing parasites from the blood, they do not have specific effects against CM. We have found that P. falciparum-iRBC induced disruption of human brain microvascular endothelial cell junctions was prevented by the activation of the angiotensin (Ang) II receptor type 2 (AT2), achieving protection of endothelial integrity. We also found that AT2 agonists are protective against experimental CM, and mice deficient in the AT2 receptor are more susceptible to this syndrome. However, over the last years, we have expanded the renin-angiotensin system by identifying additional receptors (Mas and MrgD) that are activated by agonists similar or equal to AT2 agonists, such as C21, Ang-(1-7) and Ang-(1-9). We could also show that these compounds are protective against P. falciparum-induced disruption of endothelial integrity. Thus, we hypothesize that activation of one, two, or all three receptors of the beneficial arm of the renin- angiotensin system protects brain endothelial cells from parasite-induced disruption of their barrier function. To identify which receptor(s) are key in the protection of endothelial integrity, we will first quantify the effect of Ang-(1-9), Ang-(1-7) and C21 in receptor-transfected HEK293 cells and in human and murine brain endothelial cells on intracellular signaling molecules, known to be regulated by the agonists and important in the maintenance of endothelial barrier integrity. Receptor blockers targeting different angiotensin receptors will be tested for their ability to block the intracellular signaling induced by the protective Ang peptides/analogues. Primary endothelial cells will be also targeted with siRNA against AT2 and/or Mas and/or MrgD to identify the responsible receptor(s) for the signaling effects. Finally, primary brain endothelial cells isolated from mice deficient in one, two or three receptors will be used as a genetic model to confirm the conclusions made from the pharmacological and siRNA experiments. We will then identify the receptors involved in the protection mediated by Ang-(1-9), Ang-(1-7), and C21 of brain endothelial cells from P. falciparum-induced endothelial disruption. We will use a pharmacological approach with the receptor antagonists and siRNA against the receptors to determine their effects on endothelial activation and junction integrity and on selected second messengers. Finally, we will test how treatment with Ang-(1-9), Ang-(1-7) and C21 affects the outcome of CM in wild-type and genetically deficient mice in one of the three receptors as well as double and triple knockouts. The main goal of this project is to identify the angiotensin receptor(s) that mediate protection of endothelial integrity during CM. Our results will lay the foundation for the development of agonists against this receptor(s) as adjunct therapy for CM and potentially for other diseases where brain endothelial integrity is compromised.

摘要脑型疟疾（CM）仍然是最致命和最致命的疟疾表现。是 由恶性疟原虫感染的红细胞（iRBC）粘附于宿主脑内皮细胞引起， 危及血脑屏障虽然现有的抗疟疾药物在清除寄生虫方面是有效的， 从血液中分离出来，它们对CM没有特异性作用。 我们已经发现恶性疟原虫-iRBC诱导人脑微血管内皮细胞的破坏 血管紧张素（Ang）II受体2型（AT 2）的激活阻止了连接， 保护内皮完整性。我们还发现AT 2激动剂对实验性CM具有保护作用， 而缺乏AT 2受体的小鼠更容易患上这种综合征。然而，在过去的几年里，我们 已经通过鉴定额外的受体（Mas和MrgD）扩展了肾素-血管紧张素系统， 由类似或等同于AT 2激动剂的激动剂如C21、Ang-（1-7）和Ang-（1-9）激活。我们也可以 表明这些化合物对恶性疟原虫诱导的内皮完整性破坏具有保护作用。 因此，我们假设，激活一个，两个，或所有三个受体的有益臂的肾素-血管紧张素转换酶- 血管紧张素系统保护脑内皮细胞免受寄生虫诱导的屏障功能破坏。 为了鉴定哪种受体在保护内皮完整性中是关键的，我们将首先量化 受体转染的HEK 293细胞以及人和小鼠脑内皮细胞中的Ang-（1-9）、Ang-（1-7）和C21 细胞内信号分子，已知由激动剂调节，并在 维持内皮屏障的完整性。靶向不同血管紧张素受体的受体阻断剂将是 测试它们阻断由保护性Ang肽/类似物诱导的细胞内信号传导的能力。 还将用针对AT 2和/或Mas和/或MrgD的siRNA靶向原代内皮细胞，以鉴定内皮细胞的功能。 负责信号传导作用的受体。最后，从小鼠分离的原代脑内皮细胞 缺乏一个，两个或三个受体将被用作遗传模型，以证实从 药理学和siRNA实验。然后我们将识别出参与保护的受体 由恶性疟原虫诱导的脑内皮细胞的Ang-（1-9）、Ang-（1-7）和C21介导的 破坏我们将使用受体拮抗剂和siRNA的药理学方法来对抗 受体，以确定其对内皮活化和连接完整性以及对所选第二 使者最后，我们将测试用Ang-（1-9）、Ang-（1-7）和C21治疗如何影响CM的结果。 在野生型和遗传缺陷小鼠中，三种受体之一以及双重和三重敲除。 本项目的主要目标是鉴定介导内皮细胞保护的血管紧张素受体。 CM期间的完整性。我们的研究结果将为开发抗该受体的激动剂奠定基础 作为CM的辅助治疗，并可能用于脑内皮完整性受损的其他疾病。