Role of Sialidase in Pulmonary Host Defenses and Acute Lung Injury

唾液酸酶在肺宿主防御和急性肺损伤中的作用

• 批准号: 7477660
• 负责人: Alan S. Cross
• 金额: $ 45.63万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-08-01 至 2011-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7477660
• 关键词: Accounting; Acids; Acute Lung Injury; Address; Adherence; Adhesions; Adult Respiratory Distress Syndrome; Affect; Affinity; Alveolar; Animal Model; Aspirate substance; Bacteria; Binding; Blood capillaries; Breathing; Cathepsin G; Cell Adhesion; Cell Adhesion Molecules; Cell Communication; Cell Surface Receptors; Cell surface; Cells; Cessation of life; Charge; Complex; Condition; Coupled; Data; Defense Mechanisms; Diffuse; Elastases; Embryo; Endopeptidases; Endothelial Cells; Endothelium; Environment; Epithelium; Equilibrium; Event; Excision; Free Radicals; Gases; Glycoconjugates; Host Defense; Human; ICAM1 gene; ITGAM gene; ITGB2 gene; Immune response; In Vitro; Infection; Infiltration; Inflammation; Inflammatory; Injury; Invaded; Lead; Length of Stay; Leukocyte Elastase; Leukocyte Trafficking; Leukostasis; Ligands; Lung; Mammalian Cell; Mediating; Mediator of activation protein; Microbe; Modeling; Modification; Molecular; Mus; N-Acetylneuraminic Acid; Neuraminidase; Organ; PECAM1 gene; Pancreatic Elastase; Pathway interactions; Pattern recognition receptor; Peptide Hydrolases; Peptides; Permeability; Phosphorylation; Principal Investigator; Process; Production; Protein Isoforms; Protein Overexpression; Proteins; Range; Reactive Oxygen Species; Regulation; Role; Sialic Acids; Sialyltransferases; Signal Transduction; Site; Stimulus; Surface; Syndrome; Testing; Tissues; Toll-like receptors; Tyrosine Phosphorylation; base; cadherin 5; capillary; collagenase; cytokine; in vivo; in vivo Model; inhibitor/antagonist; interstitial; kidney cell; killings; lung injury; macromolecule; microbial; migration; neutrophil; novel therapeutics; programs; response; restoration; sialylation; toll-like receptor 4; trafficking

DESCRIPTION (provided by applicant): Polymorphonuclear leukocytes (PMN) protect the lung against microbial infection; however, an excessive or prolonged influx of PMNs may cause tissue damage. Acute lung injury is characterized by widespread inflammatory changes, including diffuse PMN infiltration, with injury to both lung epithelium and endothelium. We established that the removal of sialyl residues from the surface of PMNs and endothelial cells (EC) by an endogenous PMN sialidase is critical to normal PMN trafficking. Further, we find that desialylation of the Toll-like receptor (TLR) 4 complex enhances LPS-induced cytokine production, while restoration of sialyl residues by sialyltransferases (ST) may restore the basal state. The overall hypothesis to be tested is that sialidases and STs regulate innate immune responses in the lung, and the balance achieved will dictate whether or not the innate immune response develops into acute lung injury. To address this central hypothesis, we propose the following: In Specific Aim 1 we will define which form(s) of PMN sialidase(s) (neul and neu3) regulates adherence to and migration across human lung microvascular EC by overexpression and knockdown of specific neu proteins. In Specific Aim 2 we will define the substrates for PMN sialidase in PMNs and ECs and assess how their sialylation status affects PMN adhesion in vitro and trafficking in vivo. Since PMNs induce signaling cascades in ECs, in Specific Aim 3 we will examine whether an "appropriate" level of PMN sialidase-induced tyrosine phosphorylation of EC proteins opens the EC-EC paracellular pathway sufficiently for transendothelial migration without protein leak, while at levels that exceed this threshold, if there will be loss of EC barrier function. A "two-hit" model of acute lung injury has been proposed whereby a combination of LPS and chemotactic peptide promotes lung injury. In Specific Aim 4, we will examine the role of sialidase in sensitizing the pulmonary vasculature to circulating LPS and lung injury. The molecular mechanism(s) by which desialylation promotes signaling through the TLR4/MD2/CD14 complex will be examined, as well as the ability of STs to downregulate this response. The ability of sialidase inhibitors to reverse acute lung injury will be studied in vivo in a murine model of pulmonary leukostasis. Since inhibitors of sialidase are commercially available, these studies will provide the experimental rationale for testing a new therapeutic approach to the treatment of lung injury.

描述(申请人提供)：多形核白细胞(PMN)保护肺免受微生物感染；然而，过多或长时间的PMN流入可能会导致组织损伤。急性肺损伤的特点是广泛的炎性改变，包括弥漫性PMN浸润，对肺上皮和内皮细胞均有损伤。我们证实，内源性PMN唾液酸酶从PMN和内皮细胞(EC)表面清除唾液酸残基是正常PMN运输的关键。此外，我们还发现，去除Toll样受体(TLR)4复合体可以增强内毒素诱导的细胞因子的产生，而唾液酸基转移酶(ST)恢复唾液酸残基可能会恢复基础状态。有待检验的总体假设是唾液酸酶和STS调节肺内的先天免疫反应，而所达到的平衡将决定先天免疫反应是否发展为急性肺损伤。为了解决这一中心假设，我们提出如下建议：在特定的目标1中，我们将定义哪种形式(S)的中性粒细胞唾液酸酶(S)(neu1和neu3)通过过度表达和敲除特定的neu蛋白来调节与人肺微血管内皮细胞的黏附和迁移。在特定的目标2中，我们将定义PMN和ECs中PMN唾液酸酶的底物，并评估它们的唾液酸化状态如何影响PMN在体外的黏附和体内的运输。由于中性粒细胞在内皮细胞中诱导信号级联反应，在特定的目标3中，我们将研究在适当水平的PMN唾液酸酶诱导的EC蛋白酪氨酸磷酸化是否足以打开EC-EC旁细胞途径而不会发生蛋白质泄漏，如果超过这一阈值，EC屏障功能将会丧失。已经提出了一种“两次打击”的急性肺损伤模型，在这种模型中，内毒素和趋化肽的组合促进了肺损伤。在特定的目标4中，我们将研究唾液酸酶在肺血管对循环内毒素和肺损伤的敏感性中的作用。我们将研究脱氢酶通过TLR4/MD2/CD14复合体促进信号传递的分子机制(S)，以及STS下调这一反应的能力。唾液酸酶抑制剂逆转急性肺损伤的能力将在肺白细胞停滞的小鼠模型中进行活体研究。由于唾液酸酶抑制剂已经上市，这些研究将为测试治疗肺损伤的新治疗方法提供实验依据。