Mechanisms of Endothelial Cell Dysfunction in Critically Ill Children

项目摘要 候选人。我是耶鲁医学院的一名儿科重症监护医学讲师，我将被提升 根据耶鲁医学院7月临床医生奖学金跟踪，授予助理教授学术晋升 2018年。我的K08申请将允许我获得更多的指导研究经验，这样我就可以成为 一位在危重儿童的内皮细胞信号及其干扰方面具有专业知识的独立研究员。 这项研究将建立在我的研究员培训的基础上，该培训侧重于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所需的 成功地进行血管研究，如人类细胞的基因改造，细胞内的询问 信号通路和生物信息学分析。我将选修血管生物学、生物信息学、 和免疫学研究方法，并将参加国家会议。我有一位杰出的导师， 世界级血管生物学家乔丹·波伯博士和遗传学专家穆斯塔法·霍卡博士的顾问， 以及文斯·福斯蒂诺博士，他是生物统计学家和临床试验专家。这个多学科的指导团队，以及 利用耶鲁大学VBT项目提供的资源，我将能够发展必要的技能 作为一名医疗外资助的临床科学家，专注于改善危重儿童的护理。 研究项目。血管内皮细胞(ECs)主动调节对器官至关重要的血管通透性 功能和患者生存。在心肺衰竭期间，EC通透性丧失会导致毛细血管渗漏 这导致了儿童的发病率和死亡率。对产生的信号事件的独特调节 毛细血管内皮细胞的通透性变化在很大程度上是未知的。同样，危重疾病如何导致EC功能障碍 仍然未知，很大程度上是因为我们无法直接评估危重患者EC的变化。我最近 证明调节小GTP酶RhoB在系统性红斑狼疮儿童患者中的重要性 由单基因突变引起的毛细血管渗漏综合征。在目标1中，我测试了病理性的假设 毛细血管通透性由小的GTP酶调节，这些途径受转录的影响 炎性细胞因子诱导的变化。我将询问毛细血管内皮细胞的细胞内信号通路 从不同器官(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，我将测试内皮细胞经历转录变化以响应 得了重病。我将分析直接从危重病人身上收集的单个静脉内皮细胞的全部转录组。 儿童使用单细胞RNA测序。我将评估识别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的影响 控制毛细管渗漏的开始和解决的信号通路。更好地理解这些途径 参与开发和解决毛细血管渗漏是开发治疗方法的第一步 与严重的发病率和死亡率相关的常见情况。

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.