S. aureus and Regulatory T cells in the Failure of Oral Tolerance

描述（由申请人提供）：这是西北范伯格医学院儿科过敏症和免疫学家Anne Marie Singh博士的K23奖申请。辛格博士是一名以患者为导向的食物过敏转化研究的年轻研究员。她之前对早期湿疹的表型表达和调节性T细胞在过敏中的作用的研究使她有资格研究耐受性丧失的早期机制。食物过敏影响了1200多万美国人，从1997年到2007年，儿童的患病率增加了18%。食物过敏引起的过敏反应往往危及生命，食物过敏是哮喘和变应性鼻炎未来发展的重要危险因素。尽管它的显著影响，很少知道关于细胞和免疫机制在儿童食物过敏的起始。食物过敏是由于口服对摄入抗原的耐受性丧失，导致病理性免疫反应。调节性T （Treg）细胞在平衡免疫反应中起重要作用，这些细胞的失调可能在食物过敏的发病机制中起重要作用。最近有研究表明，口服暴露于葡萄球菌肠毒素B （SEB）会破坏小鼠的口服耐受性，促进食物过敏相关反应。令人兴奋的新初步数据表明，食物过敏患者肠道中的金黄色葡萄球菌负荷增加，先前的工作也表明，在正式的鸡蛋过敏儿童中，随着他们获得耐受性，IL-10产生的Tregs增加。研究表明，局部金黄色葡萄球菌在特应性湿疹的皮肤炎症中起着至关重要的作用，金黄色葡萄球菌肠毒素B （SEB）激活CD25+ Treg细胞可消除其抑制性表型。综上所述，这些数据支持以下假设：金黄色葡萄球菌在儿童体内的定植会破坏Treg的功能，抑制口腔耐受，从而促进过敏性炎症和食物过敏的发生。为了验证这一假设，我们提出用流式细胞术对食物过敏儿童和非食物过敏儿童的Treg细胞进行表征，通过微生物组分析确定金黄色葡萄球菌在食物过敏和非食物过敏儿童中的定植和肠毒素产生，并确定食物抗原和微生物暴露对这些儿童Treg功能的体外影响。该建议侧重于Treg细胞和金黄色葡萄球菌在随之而来的耐受性丧失中的作用，并将开辟防止食物过敏发展的方法或增加抗原避免之外的治疗选择。通过这项拟议的研究，辛格博士将朝着她的长期目标努力，成为一名独立的转化医师兼研究者，研究早期生活暴露对特应性反应发展的细胞机制。短期目标是：(1)培养调节性T细胞分析、转录因子分析和下一代测序的研究技能；(2)培养分析微生物群数据集的生物信息学技能；(3)培养研究设计和实施的生物统计学和临床试验技能。完成该奖项后，Singh博士将接受培训，设计和实施关于细菌和环境暴露的免疫效应的研究，包括随着时间的推移获得微生物群以及微生物和其他环境暴露影响免疫反应调节的机制。为了实现这些目标，我们组建了一支一流的指导团队。主要导师Robert Schleimer博士是过敏症-免疫学（医学部）主任，在过敏症转化研究方面有着良好的记录，并成功培养了许多院士。共同导师包括Paul J. Bryce，博士，副教授，专长于食物过敏小鼠模型和调节性T细胞研究；Jacqueline Pongracic，医学博士，儿科过敏症-免疫学部门主任，拥有儿科过敏患者的多中心临床试验经验；Lewis Smith，医学博士，研究副主席，广泛的临床和实验室研究项目研究哮喘发病机制；Mitchell L. Sogin，博士，主任。海洋生物实验室，是细菌群落研究的先驱，包括人类微生物群的生物信息学分析。指导小组将监督辛格博士在研究金黄色葡萄球菌对食物过敏中调节性T细胞功能的影响方面的进展。

DESCRIPTION (provided by applicant): This is an application for a K23 award for Dr. Anne Marie Singh, a pediatric allergist-immunologist at Northwestern Feinberg School of Medicine. Dr. Singh is young investigator in patient-oriented translational research of food allergy. Her previous research studying the phenotypic expression of early eczema and role of regulatory T cells in allergy makes her uniquely qualified to study early mechanisms of loss of tolerance. Food allergy affects over 12 million Americans, and prevalence in children has increased by 18% from 1997-2007. Food allergy-induced anaphylaxis is often life threatening, and food allergy is an important risk factor for the future development of asthma and allergic rhinitis. Despite its significant impact, little is known regarding the cellular and immunologic mechanisms in the initiation of food allergy in children. Food allergy results from a loss of oral tolerance t ingested antigens, leading to the pathologic immune response. Regulatory T (Treg) cells are important in balancing immune responses, and dysregulation of these cells may play an important role in the pathogenesis of food allergy. It has recently been shown that oral exposure to Staphylcoccal enterotoxin B, SEB, breaks oral tolerance in mice, promoting food allergy-associated responses. Exciting new preliminary data demonstrate increased Staphylococcus aureus burden in the gut of patients with food allergy, and previous work has also illustrated increased IL-10 producing Tregs in formally egg allergic children as they acquire tolerance. It has been shown that topical S. aureus plays a crucial role in the cutaneous inflammation of atopic eczema and activation of CD25+ Treg cells with S. aureus enterotoxin B (SEB) abrogated their suppressive phenotype. Together, these data support the following hypothesis: colonization of children with Staphylococcus aureus abrogates Treg function, inhibiting oral tolerance, and thereby promotes allergic inflammation and the development of food allergy. To test this hypothesis we propose to characterize the Treg cells by flow cytometry in children with food allergy compared to non-food allergic children, determine the colonization and enterotoxin production of S. aureus in food allergic and non-food allergic children through microbiome analysis, and determine the effect of food antigen and microbial exposure on Treg function in vitro in these children. This proposal focuses on the role of Treg cells and S. aureus in the attendant loss of tolerance, and will open approaches to prevent the development of food allergy or increase treatment options beyond antigen avoidance. Through the proposed research, Dr. Singh will work towards her long-term goal of becoming an independent translational physician-investigator, studying cellular mechanisms of early life exposures on the development of atopy. Short term goals are to: (1) develop research skills of regulatory T cell analysis, transcription factor analysis and next-generation sequencing, (2) establish bioinformatics skills to analyze microbiota data sets and (3) develop biostatistical and clinical trial skills for study design and implementation. After completion of this award, Dr. Singh, will have had the training to design and implement studies on the immune effects of bacterial and environment exposures, including the acquisition of microbiota over time and the mechanisms that microbial and other environmental exposures impact the regulation of the immune response. To achieve these goals, a top-notch mentoring team has been assembled. The primary mentor, Robert Schleimer, PhD, Chief, Division of Allergy-Immunology (Department of Medicine) has a strong track record of translational research in allergy and has successfully trained many academicians. Co-mentors include Paul J. Bryce, PhD, Associate Professor, with expertise in mouse models of food allergy and regulatory T cell studies~ Jacqueline Pongracic, MD, Chief, Division of Allergy-Immunology (Department of Pediatrics) who has multicenter clinical trial experience in pediatric allergy patients~ Lewis Smith, MD, Associate Vice Chair for Research, with extensive clinical and laboratory-based research programs studying asthma pathogenesis~ and Mitchell L. Sogin, PhD, Director, Marine Biology Laboratory, a pioneer in studies of bacterial communities, including bioinformatics analysis of human microbiota. The mentoring team will oversee Dr. Singh's progress to study the effects of S. aureus on regulatory T cell function in food allergy.