Role of Surfactant Protein D in Surfactant Homeostasis

表面活性剂蛋白 D 在表面活性剂稳态中的作用

• 批准号: 7093563
• 负责人: MACHIKO IKEGAMI
• 金额: $ 39.48万
• 依托单位: CINCINNATI CHILDRENS HOSP MED CTR
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-09-01 至 2007-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7093563
• 关键词: RNA; alveolar macrophages; confocal scanning microscopy; enzyme linked immunosorbent assay; flow cytometry; genetically modified animals; immunocytochemistry; laboratory mouse; lipid metabolism; lung; lung alveolus; microarray technology; phosphatidylcholines; phospholipids; protein structure function; pulmonary surfactants; respiratory epithelium; tissue /cell culture

DESCRIPTION (provided by applicant): This proposal seeks to continue funding of HL-63329 to determine the mechanisms by which surfactant protein D (SPD) regulates surfactant phospholipid homeostasis. SP-D is a 43 kd member of the collectin family of polypeptides that is expressed in bronchiolar and alveolar epithelial cells of the vertebrate lung. While SP-D plays an important rote in the mediation of innate host defenses against viral, bacterial and fungal pathogens, our findings in SP-D gene targeted mice demonstrated that SP-D played a critical role in the generation of 1) normal surfactant pool sizes, 2) the structure of tubular myelin and other alveolar lipids, 3) macrophage activation, oxidant and metalloproteinase production, and 4) maintenance of normal alveolar structure during development. Our preliminary data strongly support a model in which SP-D regulates surfactant metabolism, catabolism, and/or recycling by type II epithelial cells in the lung. The present aims are therefore designed to discern the mechanisms by which SP-D regulates surfactant homeostasis. The specific aims are designed to test two alternative hypothesis: 1) that SP-D contributes to the generation of unique surfactant forms whose uptake catabolism and routing degradation are perturbed leading to the accumulation of surfactant phospholipids in the airspaces and in type II ceils, and 2) the alternative and/or overlapping hypothesis that SP-D interacts directly with type II epithelial cells to alter surfactant homeostasis. In Specific Aim 1 we will determine the effects of SP-D on surfactant structure in experiments in which SP-D is restored in vivo and in vitro. Effects of the presence or absence of SP-D on ultrastructure, large aggregate/small aggregate ratios and uptake or catabolism of surfactant particles and surfactant-coated beads by type II cells will be assessed. SP-D(-/-) mice in which SP-D is conditionally expressed in the lung will be utilized. In Aim 2, the structure and function of chimeric mutant SP-D molecules will be assessed. SP-D and mutant SP-D molecules will be produced in vitro and in vivo. SP-D/SP-A mutant proteins will be reconstituted with surfactant phospholipids to discern the precise structural domains mediating the effects of SP-D on surfactant structure and its metabolism/catabolism in vitro. Function of site-specific SP-D mutant proteins will be tested in SP-D (-/-) mice in which the mutant SP-D proteins are expressed. Finally, in Specific Aim 3 we will utilize microarray analyses of lung and isolated type II cells from SP-D (-/-) mice to define the genomic responses to altered lipid pool sizes in the presence and absence of SP-D. SP-D plays a critical role in surfactant homeostasis and in defense of the lung. Elucidation of the critical roles of SP-D on surfactant homeostasis and host defense will enhance our understanding of the pathogenesis of a number of acute and chronic lung disorders including cystic fibrosis, acute bacterial infection and ARDS. The intended studies will provide fundamental insights into the role of SP-D in the protection of the lung, as well as into the basic mechanisms determining surfactant homeostasis.

描述（由申请人提供）：该提案旨在继续资助HL-63329，以确定表面活性剂D（SPD）调节表面活性剂磷脂稳态的机制。 SP-D是在脊椎动物肺的支气管和肺泡上皮细胞中表达的多肽的收集蛋白家族的43 KD成员。 尽管SP-D在针对病毒，细菌和真菌病原体的先天宿主防御措施中的调解中起着重要的作用发育过程中的正常牙槽结构。 我们的初步数据强烈支持SP-D调节表面活性剂代谢，分解代谢和/或通过肺中II型上皮细胞回收的模型。 因此，目前的目标旨在辨别SP-D调节表面活性剂稳态的机制。 具体目的旨在检验两个替代假设：1）SP-D有助于产生独特的表面活性剂形式的产生，其吸收分解代谢和降解降解会受到干扰，从而导致表面活性剂磷脂的积累在空间中的磷脂积累，并在II型ceils中与替代性和/或不同的ePotist sultherts sperf sulterfters speplatist sperferts spheptials，以及2）替代性和/或/或不同的效果。稳态。 在特定的目标1中，我们将在体内和体外恢复SP-D的实验中确定SP-D对表面活性剂结构的影响。 SP-D存在或不存在SP-D对超微结构，大骨料/小骨料比以及通过II型细胞对表面活性剂颗粒和表面活性剂涂层珠的摄取或分解代谢的影响。 将使用SP-D（ - / - ）小鼠在肺中有条件地表达SP-D的小鼠。 在AIM 2中，将评估嵌合突变体SP-D分子的结构和功能。 SP-D和突变体SP-D分子将在体外和体内产生。 SP-D/SP-A突变蛋白将用表面活性剂磷脂重构，以辨别介导SP-D对表面活性剂结构及其代谢/分解代谢的影响的精确结构结构域。 位点特异性SP-D突变蛋白的功能将在表达突变体SP-D蛋白的SP-D（ - / - ）小鼠中进行测试。 最后，在特定的目标3中，我们将利用SP-D（ - / - ）小鼠对肺和II型细胞进行微阵列分析，以定义在存在和不存在SP-D的情况下对脂质池的改变的基因组响应。 SP-D在表面活性剂稳态和防御肺中起着至关重要的作用。 SP-D在表面活性剂稳态和宿主防御上的关键作用阐明将增强我们对许多急性和慢性肺部疾病（包括囊性纤维化，急性细菌感染和ARD）的发病机理的理解。 预期的研究将提供有关SP-D在保护肺部的作用的基本见解，以及决定表面活性剂稳态的基本机制。

项目成果

Reversibility of pulmonary abnormalities by conditional replacement of surfactant protein D (SP-D) in vivo.

通过体内表面活性蛋白 D (SP-D) 的条件替代可逆转肺部异常。

• DOI: 10.1074/jbc.m206200200
• 发表时间: 2002
• 期刊: The Journal of biological chemistry
• 作者: Zhang,Liqian;Ikegami,Machiko;Dey,ChittaR;Korfhagen,ThomasR;Whitsett,JeffreyA
• 通讯作者: Whitsett,JeffreyA

Activity of pulmonary surfactant protein-D (SP-D) in vivo is dependent on oligomeric structure.

肺表面活性蛋白-D (SP-D) 的体内活性取决于寡聚结构。

• DOI: 10.1074/jbc.m010191200
• 发表时间: 2001
• 期刊: The Journal of biological chemistry
• 作者: Zhang,L;Ikegami,M;Crouch,EC;Korfhagen,TR;Whitsett,JA
• 通讯作者: Whitsett,JA