Cellular mechanism of immune dysfunction following burn injury

描述(申请人提供)：严重烧伤后死亡的主要原因是脓毒症和继发于严重的先天和适应性免疫功能障碍的多系统器官衰竭，尽管反应的控制机制尚不清楚。免疫活性的主要决定因素是天然的Toll样受体(TLR)和核苷酸结合寡聚化结构域(NOD)样受体(NLR)家族。它们由许多免疫细胞表达，其主要功能是感知微生物分子，导致细胞因子释放或细胞死亡。我们的总体假设是烧伤后先天免疫系统和获得性免疫系统的损害是由TLR和NLR信号直接介导的。拟议的实验旨在在烧伤后的两个免疫致病阶段验证这一假说，并研究先天受体阻断作为一种可能的免疫治疗方法来治疗烧伤患者面临的临床相关问题。首先，为了确定TLR和NLR信号控制创伤后先天免疫系统的机制：我们将利用烧伤动物模型测定MF和DC的表型和功能(细胞因子和细胞死亡)的变化。我们将TLR和NLR的表达与它们在烧伤后早期和晚期的功能变化联系起来。然后我们将使用TLR和NLR缺陷小鼠来验证改变的烧伤损伤介导的先天免疫反应是通过先天受体信号传递的。第二，探讨先天信号调控烧伤后获得性免疫反应的机制：检测烧伤后TLR和NLR在CD4+、CD8+T细胞及相关T细胞亚群上的表达和活化。我们将把TLR和NLR表达的变化与T细胞诱导适应性免疫反应的能力联系起来。为了确定直接(在T细胞上表达天然受体)和间接(在MF或DC上表达)天然受体表达在介导烧伤适应性反应中的相对重要性，我们将先天信号受体缺乏的T细胞或MF和DC转移到烧伤或假手术受者体内，检测T细胞免疫功能和存活率。第三，为了证明先天受体在体内烧伤后免疫功能紊乱反应中的关键作用：我们将利用TLR和NLR先天信号缺陷的小鼠来研究先天受体在介导同种异体皮肤移植排斥反应和防止烧伤后有效免疫耐受建立中的作用。我们将使用缺乏TLR和NLR信号的小鼠，研究先天信号通路在烧伤后早期和晚期介导铜绿假单胞菌敏感性的重要性。

DESCRIPTION (provided by applicant): The leading causes of death following serious burn injury are sepsis and multiple system organ failure secondary to profound innate and adaptive immune dysfunction, though the controlling mechanism for the response is unknown. Major determinants of immune activity are the innate Toll-like receptor (TLR) and nucleotide binding oligomerization domain (NOD)-like receptor (NLR) families. These are expressed by many immune cells and their primary function is to sense microbial molecules leading to cytokine release or cell death. Our overall hypothesis is that impairment of both the innate and adaptive immune systems after burn injury is mediated directly by TLR and NLR signaling. The proposed experiments are designed to test this hypothesis during both immunopathogenic phases after burn injury and investigate innate receptor-blockade as a possible immunotherapeutic approach for clinically relevant issues facing burn patients. First, to determine the mechanism by which TLR and NLR signaling controls the innate immune system after injury: We will determine phenotypic and functional (cytokine and cell death) changes of MF and DC using an animal model of burn injury. We will correlate TLR and NLR expression with changes in their function during early and late after burn injury. We will then use TLR and NLR deficient mice to validate that the altered burn injury-mediated innate immune response is mediated through innate-receptor signaling. Second, to determine the mechanism by which innate signaling controls the adaptive immune response after burn injury: We will examine the effect of burn injury on the expression and activation of TLR and NLR on CD4+ and CD8+ T cells and relevant T cell subsets. We will correlate changes in TLR and NLR expression with the ability of T cells to elicit an adaptive immune response. To determine the relative importance of both direct (expression of innate receptors on T cells) and indirect (expression on MF or DC) innate receptor expression in mediating the adaptive response to burn injury, we will transfer T cells or MF and DC, deficient in innate signaling receptors, into burn or sham recipients and test T cell immune function and survival. Third, to demonstrate that innate receptors are critical for the dysfunctional immunologic response to burn injury in vivo: We will use mice deficient in TLR and NLR innate signaling to examine the role of innate receptors in mediating skin allograft rejection and preventing establishment of effective immune tolerance after burn. We will examine the importance of innate signaling pathways in mediating Pseudomonas aeruginosa susceptibility early and late after burn injury using mice deficient in TLR and NLR signaling.