Pneumocyte-Derived Host Defence Lectins

肺细胞衍生的宿主防御凝集素

• 批准号: 6823503
• 负责人: ERIKA Christine CROUCH
• 金额: $ 20.5万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-09-01 至 2008-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6823503
• 关键词: SDS polyacrylamide gel electrophoresis; endopeptidases; enzyme activity; glycoproteins; inflammation; ion exchange chromatography; laboratory mouse; lectin; ligands; lipopolysaccharides; lung; lung development; lung injury; microorganism immunology; neutrophil; protein degradation; protein structure function; proteolysis; pulmonary surfactants; recombinant proteins; regeneration

Surfactant Protein D (SP-D) plays important roles in the defense against inhaled microorganisms, contributes to the pulmonary response to antigenic challenge, and participates in the regulation of surfactant lipid homeostasis. SP-D can directly interact with neutrophils, and can modulate the anti-viral, anti-bacterial, and anti-fungal functions of neutrophils in vitro. However, there is also evidence that neutrophils contribute to the degradation and clearance of SP-D in the setting of acute lung injury, suggesting a complex interplay between neutrophils and SP-D in vivo. We have recently observed that SP-D is specifically degraded by the three major human neutrophil serine proteases: neutrophil elastase (NE), cathepsin G (CG), and proteinase 3 (PR3) with the liberation of similar, high molecular weight, disulfide-crosslinked fragments; similar cleavage is mediated by murine neutrophils and whole neutrophil lysates. Given the above, we hypothesize the neutrophil-derived proteases specifically degrade SP-D within the functionally important lectin domains. We further hypothesize the neutrophil serine proteases contribute to the enhanced clearance of SP-D in the setting of LPS challenge, thereby contributing to a depletion of functional forms of this host defense protein in the interval prior to compensatory increases in SP-D production. Accordingly, we propose: 1) to characterize the effects of purified neutrophil-derived serine proteases on SP-D structure and biological activity; 2) to examine mechanisms of neutrophil-mediated proteolysis in vitro with emphasis on the potential roles of "quantal proteolysis" and membrane-associated serine proteases; and 3) to examine the potential roles of neutrophil proteases on SP-D degradation and clearance in vivo. The contributions of serine proteases will be assessed in vitro using protease-deficient murine neutrophils, and the potential roles of these enzymes in SP-D clearance and degradation will be examined in vivo using murine models of protease deficiency in combination with models of acute lung injury. Together, these studies should provide important new information relating to the mechanisms that could determine the amount and functional activity of SP-D in the setting acute lung injury.

肺表面活性物质蛋白D(SP-D)在抵抗吸入微生物、促进肺组织对抗原攻击的应答、参与肺表面活性物质脂质平衡的调节等方面发挥着重要作用。SP-D可直接与中性粒细胞相互作用，并在体外调节中性粒细胞的抗病毒、抗细菌和抗真菌功能。然而，也有证据表明，在急性肺损伤中，中性粒细胞有助于SP-D的降解和清除，这表明在体内中性粒细胞和SP-D之间存在复杂的相互作用。我们最近观察到，SP-D被三种主要的人中性粒细胞丝氨酸蛋白酶：中性粒细胞弹性蛋白酶(NE)、组织蛋白酶G(CG)和蛋白酶3(PR3)特异性地降解，释放出类似的高相对分子质量的二硫键交联片段；类似的切割是由小鼠中性粒细胞和整个中性粒细胞裂解产物介导的。鉴于上述情况，我们假设 中性粒细胞衍生的蛋白水解酶在重要的凝集素结构域中特异性地降解SP-D。我们进一步假设，中性粒细胞丝氨酸蛋白酶有助于在内毒素攻击时增强对SP-D的清除，从而在SP-D产生代偿性增加之前的一段时间内，有助于该宿主防御蛋白功能形式的耗尽。因此，我们建议：1)表征纯化的中性粒细胞来源的丝氨酸蛋白酶对SP-D结构和生物活性的影响；2)检测 中性粒细胞介导的体外蛋白分解的机制，重点是“量子蛋白分解”和膜相关丝氨酸蛋白酶的潜在作用；以及3)研究中性粒细胞蛋白水解酶在体内对SP-D降解和清除的潜在作用。丝氨酸蛋白酶的作用将在体外使用缺乏蛋白酶的小鼠中性粒细胞进行评估，这些酶在清除和降解SP-D方面的潜在作用将在体内使用蛋白酶缺乏的小鼠模型和急性肺损伤模型进行研究。总之，这些研究应该提供重要的新信息，这些信息与决定SP-D在急性肺损伤中的数量和功能活性的机制有关。