Mechanisms of Endothelial Cell Dysfunction in Critically Ill Children

危重儿童内皮细胞功能障碍的机制

• 批准号: 10204090
• 负责人: RICHARD W PIERCE
• 金额: $ 16.99万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-07-01 至 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10204090
• 关键词: Adherens Junction; Bioinformatics; Biological Assay; Biological Models; Biology; Blood; Blood Vessels; Blood capillaries; CRISPR/Cas technology; Candidate Disease Gene; Capillary Endothelial Cell; Capillary Leak Syndrome; Capillary Permeability; Cardiopulmonary; Cell Culture Techniques; Cell Line; Cells; Cessation of life; Child; Childhood; Clinical; Critical Care; Critical Illness; Critically ill children; Cultured Cells; Data; Dermal; Deterioration; Development; Edema; Electrical Resistance; Endothelial Cells; Endothelium; Etiology; Event; Extramural Activities; Failure; Fellowship; Functional disorder; Funding; GTPase-Activating Proteins; Gene Expression; Gene Mutation; Genes; Genetic; Genetic Techniques; Genetic Transcription; Goals; Guanine Nucleotide Exchange Factors; Guanosine Triphosphate Phosphohydrolases; Human; Immunologic Techniques; Immunology; Individual; Inflammatory; Jordan; Laboratories; Liquid substance; Lung; Mediating; Mediator of activation protein; Medicine; Mentors; Mentorship; Methodology; Methods; Modeling; Modification; Monomeric GTP-Binding Proteins; Morbidity - disease rate; Multiple Organ Failure; Mutation; Nutrient; Organ; Pathologic; Pathway interactions; Patients; Permeability; Pharmacology; Play; Process; Protocols documentation; Regulation; Research; Research Methodology; Research Personnel; Research Project Grants; Research Support; Resolution; Resources; Role; Scientist; Signal Pathway; Signal Transduction; Skin; Syndrome; TNF gene; Testing; Tight Junctions; Time; Training; Umbilical vein; Vascular Permeabilities; Venous; Work; cell type; clinically significant; cytokine; endothelial dysfunction; experience; gene product; improved; instructor; interstitial; knock-down; lymphotoxin beta; medical schools; mortality; multidisciplinary; novel; novel therapeutic intervention; overexpression; pediatric patients; prevent; professor; programs; response; single-cell RNA sequencing; skills; symposium; targeted treatment; therapy development; tool; transcriptome

Project Summary Candidate. I am an Instructor in pediatric critical care medicine at Yale School of Medicine and will be promoted to Assistant Professor with academic promotion per Yale School of Medicine's Clinician Scholar track in July 2018. My K08 application will allow me to acquire additional mentored research experience so that I may become an independent investigator with expertise in endothelial cell signaling and its disruption in critically ill children. This research will build on my fellowship training that focused on the signals that regulate changes in vascular permeability and their associated clinical consequences and will further develop my laboratory skills required to successfully conduct vascular research, such as genetic modification of human cells, interrogation of intracellular signaling pathways, and bioinformatic analysis. I will take graduate courses on vascular biology, bioinformatics, and immunology research methods and will participate in national conferences. I have an outstanding mentor, Dr. Jordan Pober, a world-class vascular biologist, along with advisors Dr. Mustafa Khokha, expert in genetics, and Dr. Vince Faustino, expert biostatistician and clinical trialist. This multidisciplinary mentorship team, along with the resources available through the VBT program at Yale, will allow me to develop the necessary skills to function as an extramurally funded clinician scientist focused on improving the care of critically ill children. Research Project. Endothelial cells (ECs) actively regulate vessel permeability that is essential for organ function and patient survival. During cardiopulmonary failure, loss of EC permselectivity results in capillary leak that contributes to morbidity and mortality in children. The unique regulation of the signaling events that produce permeability changes in capillary EC is largely unknown. Likewise, how critical illness induces EC dysfunction remains unknown, due in large part to our inability to directly assess EC changes in critically ill patients. I recently demonstrated the importance of the regulation of the small GTPase RhoB in a pediatric patient with systemic capillary leak syndrome caused by a single gene mutation. In Aim 1, I test the hypothesis that pathological capillary permeability is regulated by small GTPases and that these pathways are influenced by transcriptional changes induced by inflammatory cytokines. I will interrogate intracellular signaling pathways in capillary ECs cultured from different organs (skin and lung) to determine the effect of their modulation on trans-endothelial electrical resistance. In Aim 2, I will test the hypothesis that ECs undergo transcriptional changes in response to critical illness. I will analyze the entire transcriptome of individual venous ECs collected directly from critically ill children using single cell RNA sequencing. I will assess the functions of identified candidate gene products using pharmacologic, genetic, and immunologic techniques to manipulate and analyze their effect on the intracellular signaling pathways that control the initiation and resolution of capillary leak. Better understanding the pathways involved in the development and resolution of capillary leak is the first step in developing therapies for this common condition associated with serious morbidity and mortality.

项目摘要 候选人我是耶鲁大学医学院儿科重症监护医学讲师， 7月，根据耶鲁大学医学院的临床医生学者轨道， 2018.我的K08应用程序将允许我获得额外的指导研究经验，使我可以成为 他是一位独立的研究者，擅长内皮细胞信号传导及其在危重儿童中的破坏。 这项研究将建立在我的奖学金培训，重点是调节血管变化的信号， 渗透性及其相关的临床后果，并将进一步发展我的实验室技能， 成功地进行了血管研究，如人类细胞的遗传修饰，细胞内 信号通路和生物信息学分析。我将修血管生物学，生物信息学， 和免疫学研究方法，并将参加全国会议。我有一位杰出的导师， 博士世界级的血管生物学家乔丹·波伯，沿着顾问遗传学专家穆斯塔法·霍卡博士， 文斯·福斯蒂诺博士，生物统计学家和临床试验专家。这个多学科的导师团队，沿着 通过耶鲁大学的VBT项目提供的资源，将使我能够培养必要的技能， 作为一个校外资助的临床科学家，专注于改善重症儿童的护理。 研究项目。内皮细胞（EC）积极调节血管通透性，这是器官功能所必需的。 功能和患者生存率。在心肺衰竭期间，EC选择性渗透的丧失导致毛细血管渗漏 导致儿童发病率和死亡率。产生信号事件的独特调节 毛细管EC的渗透性变化在很大程度上是未知的。同样，危重病如何导致EC功能障碍 目前尚不清楚，这在很大程度上是由于我们无法直接评估危重患者的EC变化。我最近 证明了在全身性哮喘的儿科患者中调节小GTdR RhoB的重要性。 毛细血管渗漏综合征由单一基因突变引起。在目标1中，我检验了病理性的假设， 毛细血管通透性是由小GTP酶调节的，这些途径受转录因子的影响。 由炎性细胞因子引起的变化。我将研究毛细血管内皮细胞的细胞内信号通路 从不同的器官（皮肤和肺）培养，以确定它们对跨内皮细胞的调节作用。 电阻在目标2中，我将测试EC在响应于 病危我将分析直接从重症患者中收集的单个静脉内皮细胞的整个转录组， 儿童使用单细胞RNA测序。我将评估已确定的候选基因产物的功能， 药理学、遗传学和免疫学技术来操纵和分析它们对细胞内 控制毛细血管渗漏的起始和消退的信号通路。更好地理解这些途径 参与毛细血管渗漏的发展和解决是开发治疗方法的第一步。 与严重发病率和死亡率相关的常见病症。