MMP-8 as a novel therapeutic target in sepsis

描述（由申请人提供）：感染性休克是成人和儿童的主要公共卫生问题，因此需要开发新的靶点和治疗策略。基于以芯片为中心的临床研究和以动物为中心的后续实验，我们已经确定基质金属肽酶-8（MMP-8）作为脓毒症病理学的候选靶点。MMP-8最为人所知的是在细胞外基质周转期间切割1型胶原蛋白的嗜中性粒细胞产物。然而，现在已经变得明显的是，MMP-8也调节急性炎症，并可能在脓毒症病理学中发挥作用。该建议的总体假设是MMP-8的表达和活性在脓毒症病理学中起主要作用。我们现在建议通过四个互补的具体目标来检验这一假设。在具体目标1中，我们将检验抑制MMP-8活性改善与脓毒症病理学相关的多个终点的假设。该目的将使用MMP-8无效动物和MMP-8活性的药理学抑制剂。在具体目标2中，我们将检验发育年龄影响MMP-8在脓毒症中的作用的假设。该目的将使用适合于1周龄小鼠幼仔的脓毒症模型，并且还将使用MMP-8缺失小鼠和MMP-8活性的药理学抑制剂。在具体目标3中，我们将检验骨髓源性细胞是脓毒症中MMP-8基因调控的主要来源这一假设。用于该目的的体内实验将涉及MMP-8缺失骨髓向野生型小鼠的过继转移，反之亦然。为此目的的体外实验将系统地阐明导致中性粒细胞和单核细胞中MMP-8基因重新表达的信号传导机制。在特定目标4中，我们将检验1型胶原降解产物参与MMP-8调节炎症的机制的假设。为此目的的体外实验将测试条件培养基激活巨噬细胞的能力，所述条件培养基来源于用活化的MMP-8处理的1型胶原包被的组织培养板。用于该目的的体内实验将测量具有1型胶原突变的小鼠中对脓毒症的反应，所述1型胶原突变使其对MMP-8的切割具有抗性。该提案的主要成果包括确立MMP-8作为脓毒症的新治疗靶点，阐明发展如何影响MMP-8在脓毒症中的作用，阐明MMP-8在脓毒症中的主要细胞来源，全面了解炎症期间MMP-8表达的信号传导机制，以及MMP-8依赖性1型胶原降解产物作为先天免疫系统的危险相关分子模式的建立。

DESCRIPTION (provided by applicant): Septic shock is a major public health problem in both adults and children, and consequently there is a need to develop novel targets and therapeutic strategies. Based on microarray-centered clinical studies and follow up animal-centered experiments, we have indentified matrix metallopeptidase-8 (MMP-8) as a candidate target in sepsis pathology. MMP-8 is best known as a neutrophil product that cleaves collagen type 1 during extra-cellular matrix turnover. However, it has now become evident that MMP-8 also modulates acute inflammation and may play a role in sepsis pathology. The overarching hypothesis of this proposal is that MMP-8 expression and activity play a major role in sepsis pathology. We are now proposing to test this hypothesis via four complementary Specific Aims. In Specific Aim 1 we will test the hypothesis that inhibition of MMP-8 activity improves multiple endpoints relevant to sepsis pathology. This Aim will use MMP-8 null animals and a pharmacologic inhibitor of MMP-8 activity. In Specific Aim 2 we will test the hypothesis that developmental age influences the role of MMP-8 in sepsis. This Aim will use a sepsis model adaptable to 1 week old mouse pups, and will also make use of MMP-8 null mice and a pharmacologic inhibitor of MMP-8 activity. In Specific Aim 3 we will test the hypothesis that bone marrow-derived cells are the main source of MMP-8 gene regulation in sepsis. The in vivo experiments for this Aim will involve adoptive transfer of MMP-8 null bone marrow to wild-type mice, and vice versa. The in vitro experiments for this Aim will systematically elucidate the signaling mechanisms that lead to de novo MMP-8 gene expression in neutrophils and monocytes. In Specific Aim 4 we will test the hypothesis that collagen type 1 degradation products are involved in the mechanism by which MMP-8 regulates inflammation. The in vitro experiments for this Aim will test the ability of conditioned media, derived from collagen type 1 coated tissue culture plates treated with activated MMP-8, to activate macrophages. The in vivo experiments for this Aim will measure the response to sepsis in mice having a mutation of collagen type 1, which renders it resistant to cleavage by MMP-8. The major deliverables of this proposal include the establishment of MMP-8 as a novel therapeutic target in sepsis, the elucidation of how development influences the role of MMP-8 in sepsis, the elucidation of the major cellular source of MMP-8 in sepsis, a comprehensive understanding of the signaling mechanisms involved in MMP-8 expression during inflammation, and the establishment of MMP-8-dependent collagen type 1 degradation products as danger associated molecular patterns for the innate immune system.