Pathologic Myeloid Activation in Pediatric Burn Injury

摘要 烧伤是儿童创伤的主要原因，导致终身的身体和心理疾病。 幼儿，特别是5岁以下的幼儿，是一个非常脆弱和面临风险的群体， 过度的全身宿主反应导致多器官系统功能障碍（MODS）。不幸的是， 我们目前的许多烧伤治疗策略都是从成人研究中推断出来的，或者广泛应用于 年龄谱。这些方法没有考虑到儿科患者独特的宿主免疫反应 在不同年龄阶段的儿童。了解年龄特异性免疫功能障碍导致的 儿科烧伤将解决开发新的个性化干预措施所需的关键知识缺口 为这一高度脆弱的人群。我们的总体假设是，烧伤诱导病理性骨髓 在儿科宿主中激活，导致髓源性抑制细胞（MDSC）扩增， 单核细胞功能、淋巴细胞减少和淋巴细胞功能障碍。此外，我们认为这种影响是年龄- 和烧伤大小依赖性，这样年轻的患者更常见地进入病理性骨髓炎状态， 活化与烧伤脓毒症和免疫抑制的易感性增加。我们有两个具体目标：1） 确定小儿烧伤是否导致病理性骨髓激活，增加循环 免疫抑制MDSC，产生功能障碍的单核细胞和减少淋巴细胞数量， 功能;和，2）检查小儿烧伤是否诱导独特的血液骨髓和淋巴样 转录组在年轻和老年小儿患者的大小烧伤，可以解释功能性 和表型变化。为了实现这些目标，我们打算招募两个不同的小儿烧伤患者队列， 年龄和烧伤面积我们打算比较1-5岁的儿童（n=30）与小（<15% TBSA， n=15）或更大（15-30% TBSA，n=15）烧伤，9-15岁儿童（n=30）的TBSA匹配烧伤尺寸 （每组n=15）和年龄匹配的未受伤对照组（n=30）。使用流式细胞术，我们将确定MDSC如何 随着时间的推移，数量和表型受到儿童年龄和烧伤面积的影响。同样，我们将进行调查 MDSC的数量和功能与T细胞增殖和IFNγ产生的易感性和丧失相关 （ELISpot）。此外，我们将确定血液单核细胞HLA-DR表达，单核细胞分布， 宽度（MDW）和定量离体刺激的单核TNF-α产生（ELISpot）。在其中的一个子集中， 患者，我们将进行单细胞RNA-seq/CITE-Seq（转录组和表位的细胞索引， 烧伤后早期（第4天）和晚期（第30天）血液骨髓和淋巴细胞的测序。 我们的最终目标是开发新的治疗模式，用个性化的方法治疗儿童 确保这一弱势群体“.充分发挥其健康生产生活的潜力”。

ABSTRACT Burn injury is a leading cause of trauma in children, resulting in life-long physical and psychological morbidities. Young children, particularly those under 5-years of age, represent a highly vulnerable and at-risk population for an exaggerated systemic host response leading to multiple organ system dysfunction (MODS). Unfortunately, much of our current burn treatment strategies are extrapolated from adult studies or are broadly applied across the age spectrum. These approaches fail to consider the unique host immune responses of pediatric patients across the different age stages of childhood. Understanding age-specific immune dysfunction resulting from pediatric burn injury will address a key gap of knowledge necessary to develop novel personalized interventions for this highly vulnerable population. Our overarching hypothesis is that burn injury induces pathologic myeloid activation in the pediatric host, resulting in expansion of myeloid-derived suppressor cells (MDSCs), suboptimal monocyte function, lymphopenia and lymphocyte dysfunction. Additionally, we believe this effect is both age- and burn size-dependent, such that younger patients more commonly enter a state of pathologic myeloid activation with increased susceptibility to burn sepsis and immunosuppression. We have two specific aims: 1) to determine whether pediatric burn injury leads to pathologic myeloid activation, increasing circulating immunosuppressive MDSCs, engendering dysfunctional monocyte and decreasing lymphocyte numbers and function; and, 2) to examine whether pediatric burn injury induces a unique blood myeloid and lymphoid transcriptome in young and older pediatric patients with small and larger burns that can explain the functional and phenotypic changes. To achieve these goals, we intend to enroll two pediatric burn patient cohorts differing in their age and burn size. We intend to compare children 1–5 years old (n=30) with either small (<15% TBSA, n=15) or larger (15-30% TBSA, n=15) burns, with children 9-15 years old (n=30) with TBSA-matched burn sizes (n=15 each), and age-matched non-injured controls (n=30). Using flow cytometry, we will determine how MDSC numbers and phenotypes are influenced by pediatric age and burn size over time. Similarly, we will investigate MDSC numbers and function in relation to susceptibility to and loss of T-cell proliferation and IFNγ production (ELISpot). Additionally, we will determine changes in blood monocyte HLA-DR expression, monocyte distribution width (MDW) and quantify ex vivo stimulated mononuclear TNF-α production (ELISpot). In a subset of these patients, we will perform single cell RNA-seq/CITE-Seq (Cellular Indexing of Transcriptomes and Epitopes by Sequencing) of blood myeloid and lymphoid cells in the early (day 4) and later (day 30) periods after burn injury. Our ultimate goal is to develop novel treatment paradigms that treat children with a personalized approach ensuring that this vulnerable population “…achieve(s) their full potential for healthy productive lives.”