Burkholderia-mediated defective killing mechanisms in macrophages from cystic fibrosis (CF) patients

 描述(由申请人提供)：作为一名儿科肺科医生，我的长期职业目标是成为一名内科科学家，在囊性纤维化(CF)中微生物感染的免疫控制方面的专家。我的这个K08奖项的短期教育和研究计划集中在通过详细了解CF中伯克霍尔德氏菌感染的巨噬细胞控制获得科学独立性。我的住院医生和研究员研究培训使我能够在小鼠和人类CF模型中发现巨噬细胞自噬的缺陷，这对伯克霍尔德氏菌在CF中的毒力起着关键作用。新获得的数据表明，伯克霍尔德氏菌物种影响CF巨噬细胞氧化爆发，但这与CF中的自噬信号如何联系尚不清楚。对这些潜在的相互交织的途径的追求将产生必要的基础知识，以制定R01赠款提案，最终目标是未来针对宿主指导的新疗法的临床试验，以改进对CF急性和慢性感染的治疗。K08机制将使我能够将至少75%的精力集中在发展独立的翻译研究事业上。我的职业发展计划将采用有明确目标的结构化方法，使我能够过渡到独立的研究职业。我的培训目标包括：1)获得与病原体清除相关的自噬和NADPH氧化酶介导的途径研究方面的专业知识；2)培训巨噬细胞对病原体的信号反应的操纵；以及3)为临床前和临床试验的发展加强翻译研究设计技能。我将通过我的研究导师和指导顾问委员会的实践科学培训、相关的课程作业、参加专业人员公共卫生硕士课程和科学研讨会来实现这些目标。通过这种方法，我将广泛地和具体地提高我的研究技能，以便充分发展我的独立研究实验室和研究计划，以促进我通过完成K08奖项获得独立资金。该研究计划利用伯克霍尔德氏菌感染的人巨噬细胞模型来研究宿主免疫反应的逃避。我的初步数据表明，伯克霍尔德氏菌能够抑制CF中本已功能失调的氧化爆发，以避免宿主被杀。这也导致下游自噬小体的形成减少，进一步减少病原体的杀灭。我们假设，伯克霍尔德氏菌在CF中的存活是基于NADPH氧化酶的破坏和自噬小体的缺陷组装，NADPH氧化酶用于产生活性氧物种，而自噬小体的组装用于靶向杀灭细胞内的细菌。我们将分析氧化爆发的组装和产生，以及ROS和自噬启动之间的信号相互作用。这项工作的预期结果是，它将促进我们对潜在抗生素-- 药物输送和/或发现CF耐药伯克霍尔德氏菌感染的独立目标。这些研究的重要性反映在我们目前缺乏治疗这些致命的CF感染的金标准。国家儿童医院(RINCH)研究所的微生物致病中心和俄亥俄州立大学微生物界面生物学中心的环境非常出色，非常适合实现本研究计划和培训目标。我将利用一支熟练的指导团队来提供全面的研究指导和培训。拉里·施莱辛格博士是巨噬细胞生物学和巨噬细胞-病原体相互作用方面的世界领先者，有着长期而杰出的培训记录。他的领导能力与共同导师阿迈勒·阿梅尔博士在伯克霍尔德氏菌发病机制方面的专业知识相结合，包括自噬缺陷的新机制。此外，我的导师咨询委员会在临床、翻译和基础CF研究方面具有进一步的专业知识，此外还包括CF特定的细菌发病机制。我还有一个独特的好处，那就是在两个最先进的设施工作，并得到临床和基础科学研究核心专家的支持。来自RINCH的机构支持将使我能够继续将75%的时间用于拟议的培训和研究。通过这种跨学科的指导方法，我将大大提高我的科学进步，并在K08奖项的过程中成功竞争独立的R01级资金。

 DESCRIPTION (provided by applicant): As a pediatric pulmonologist, my long-term career goal is to become a physician-scientist who is an expert in the immunologic control of microbial infections in cystic fibrosis (CF). My short-term educational and research plans for this K08 award are focused on gaining scientific independence through a detailed understanding of the macrophage control of Burkholderia infections in CF. My resident and fellowship research training has allowed me to discover deficiencies in macrophage autophagy in both murine and human CF models that play a key role in Burkholderia virulence in CF. Newly acquired data suggest that Burkholderia species affect the CF macrophage oxidative burst, but it is unknown how this ties in with autophagy signaling in CF. The pursuit of these potentially intertwined pathways will generate the necessary fundamental knowledge for developing R01 grant proposals with the ultimate goal of future clinical trials of novel host-directed therapies for improving treatment of acute and chronic infections in CF. The K08 mechanism will allow me to focus at least 75% of my efforts on developing an independent translational research career. My career development plan will utilize a structured approach with specific objectives to allow me to transition to an independent research career. My training objectives include: 1) Gaining expertise in the study of autophagy and NADPH oxidase-mediated pathways relating to pathogen clearance; 2) Training in the manipulation of macrophage signaling responses to pathogens; and 3) Enhancing translational research design skills for the development of pre-clinical and clinical trials. I will achieve these objectives through hands-on scientific training ia my research mentors and mentoring advisory committee, relevant coursework, participation in a Master's in Public Health for Professionals program, and scientific seminars. Through this approach I will both broadly and specifically enhance my research skills in order to fully develop my independent research lab and research program to facilitate my achievement of independent funding by the completion of a K08 award. The research plan utilizes a human macrophage model of Burkholderia infection in CF to study evasion of host immune responses. My preliminary data suggests that Burkholderia are able to suppress an already dysfunctional oxidative burst in CF to avoid host killing. This also leads to a downstream reduction in autophagosome formation, further decreasing pathogen killing. We hypothesize that Burkholderia survival in CF is predicated upon both the disruption of the NADPH oxidase for generation of reactive oxygen species and the deficient assembly of autophagosomes for targeted intracellular bacterial killing. We will analyze both the assembly and generation of the oxidative burst, as well as signaling interactions between ROS and autophagy initiation. The anticipated outcome of this work is that it will advance our understanding of potential antibiotic- independent targets for drug delivery and/or discovery for antibiotic-resistant Burkholderia infections in CF. The significance of these studies is reflected in our current lack of a gold standard treatment for these deadly infections in CF. The environments of the Center for Microbial Pathogenesis at the Research Institute at Nationwide Children's Hospital (RINCH) and the Center for Microbial Interface Biology at The Ohio State University are outstanding and ideally suited for the achievement of this research plan and training objectives. I will utilize a skilled mentoring team to provide comprehensive research guidance and training. Dr. Larry Schlesinger is a world leader in macrophage biology and macrophage-pathogen interactions with a long and distinguished training record. His leadership is coupled with co-mentor Dr. Amal Amer's expertise in Burkholderia pathogenesis, including novel mechanisms of autophagy deficiency. Additionally, my mentor advisory committee features further expertise in clinical, translational, and basic CF research in addition to CF-specific bacterial pathogenesis. I also have the unique benefit of working in two state-of-the-art facilities and receiving support from experts in both clinical and basic science research cores. Institutional support from the RINCH will allow me to continue to devote 75% of my time towards the proposed training and research. Through this interdisciplinary mentoring approach I will greatly enhance my scientific progression and successful competition for independent R01 level funding during the course of this K08 award.