Cellular and Molecular Biology of Inflammation and Repair

描述(由申请人提供)：这是一项继续研究参与损伤早期炎症反应的细胞和分子生物学的建议。需要检验的假设是，中性粒细胞(PMN)通过调节巨噬细胞的表型表达，并引导巨噬细胞沿着替代或抗炎的激活途径，在无菌炎症中扮演指导细胞的角色。该假说预测，PMN对巨噬细胞炎症表型的抑制在组织损伤部位局部明显，在对出血或长骨骨折等刺激的反应中也是全身性的。该项目的最新进展为这一假设提供了支持。在这方面，中性粒细胞减少的小鼠伤口含有比正常对照组更多的肿瘤坏死因子-α和白介素6，而从中性粒细胞减少的伤口分离的巨噬细胞含有和释放过量的肿瘤坏死因子-α和白介素6。体外证据表明，PMN释放的可溶性因子(S)分子质量小于3000 Da，不是腺苷、NO或COX的产物，它诱导巨噬细胞表达一种抗炎表型，包括细胞因子/趋化因子基因和蛋白表达的改变，以及抑制超氧化物的产生。与这些观察结果相关联的是，人类PMN培养上清液抑制了内毒素刺激的人单核细胞来源的巨噬细胞释放肿瘤坏死因子-a。全身注射脂多糖使中性粒细胞减少的动物血清中的肿瘤坏死因子-α浓度比对照组高15倍，肝脏和脾组织中的肿瘤坏死因子-α的mRNA含量也增加。为了验证这一假设，该提案被构建为四个不重叠的特定目标，旨在：i)a.描述伤口PMN的表型。确定由中性粒细胞释放的抑制巨噬细胞活化的因子(S)的分子同一性。2)建立中性粒细胞因子抑制巨噬细胞产生肿瘤坏死因子的机制(S)。定义伤口巨噬细胞的表型。B)调查PMN的影响。以及PMN抑制产物在急性无菌炎症中对巨噬细胞表型和基因表达的抑制作用，以及IV)确定PMN和PMN分泌产物在全身损伤反应中的作用。这些研究的完成将在分子、细胞、组织和系统水平上表征PMN的抗炎活性，并扩大目前对PMN在确定巨噬细胞在炎症中的表型中的作用的了解。

DESCRIPTION (provided by applicant): This is a proposal to continue the study of the cellular and molecular biology of cells participating in the early inflammatory response to injury. The hypothesis to be tested is that neutrophils (PMN) act as instructive cells in sterile inflammation by regulating macrophage phenotypic expression and directing macrophages along the alternative or anti-inflammatory activation pathway. The hypothesis predicts that the suppression of macrophage inflammatory phenotype by PMN is evident locally at sites of tissue injury, and systemically during the response to stimuli such as hemorrhage or long bone fracture. Recent progress on this project provides support for the hypothesis. In this regard, wounds in neutropenic mice contain more TNF-a and IL-6 than in normal controls, and macrophages isolated from neutropenic wounds contain and release excess TNF-a and IL-6. Evidence garnered in vitro demonstrated PMN release soluble factor(s) less than 3000 Da in molecular mass that are not adenosine, NO, or products of COX, that induce the expression of an anti- inflammatory phenotype consisting of alterations in cytokine/chemokine mRNA and protein expression, and the suppression of superoxide production in macrophages. Added relevance to these observations is given by the finding that human PMN culture supernatants suppress TNF-a release from LPS-stimulated human monocyte-derived macrophages. Systemically, injection of LPS into neutropenic animals results in serum TNF-a concentrations 15-fold higher than those in controls, and in increased hepatic and splenic TNF-a mRNA content. In order to test the hypothesis, the proposal is structured in four non-overlapping Specific Aims designed to: I) A. Characterize the wound PMN phenotype. B. Define the molecular identity of the factor(s) released by PMN that suppress macrophage activation. II) Establish the mechanism of suppression of macrophage TNF-a production by PMN factor(s). Ill) A. Define the phenotype of the wound macrophage. B) Investigate the impact of PMN. and PMN inhibitory products on macrophage phenotype and gene expression in acute sterile inflammation, and IV) Determine the role of PMN and PMN secretory products in the systemic response to injury. Completion of the proposed studies will characterize the anti-inflammatory activity of PMN at the molecular, cellular, tissue, and systemic levels, and expand current understanding of a previously unrecognized role of PMN in determining the phenotype of macrophages in inflammation.