Endothelial protease activity, organogenesis and birth defects

内皮蛋白酶活性、器官发生和出生缺陷

基本信息

批准号：
8318871
负责人：
SUNEEL S APTE
金额：
$ 7.85万
依托单位：
CLEVELAND CLINIC LERNER COM-CWRU
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-09-01 至 2013-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8318871
关键词：
ADAMTS9 gene Adult Affect Alleles Antibodies Area Blood Vessels Blood capillaries CD31 Antigens Candidate Disease Gene Cardiac Cells Cleft Palate Congenital Abnormality Congenital Heart Defects Cornea Data Defect Deposition Development Developmental Biology Developmental Process Differentiation and Growth Embryo Embryonic Development Endocardium Endothelial Cells Endothelium Extracellular Matrix Failure Generations Goals Heart Heart failure Histocompatibility Testing Histologic Human In Situ Invaded Knowledge Left Limb Development Mediating Mesenchyme Metalloproteases Modification Morphogenesis Mus Myocardial Myocardium Organ Organogenesis Palate Peptide Hydrolases Phenotype Pregnancy Process Proteins Proteoglycan Proteolysis Regulation Research Right-On Role Secondary Palate Staging Staining method Stains Structure Syndrome Tamoxifen Tumor Angiogenesis Vascular Endothelium Vascularization Work angiogenesis base behavior influence body system cadherin 5 capillary craniofacial endothelial-specific sialomucin gastrulation improved insight mouse development mouse model palatogenesis versican

项目摘要

DESCRIPTION (provided by applicant): The goal of this project is to elucidate the role of vascular endothelium in embryonic development and organogenesis. Microvascular endothelial cells (mEC), which line capillaries, have a well-known function in regulation of angiogenesis, but less is known about their role in organogenesis. Our preliminary data suggests that the evolutionarily conserved, secreted metalloprotease ADAMTS9, a product of microvascular endothelium, has an important role in normal organogenesis. Adamts9-/- mice die by 7.5 days of gestation (E7.5), prior to the onset of angiogenesis and organogenesis. Nevertheless, using mice lacking one allele of Adamts9, we have shown that ADAMTS9 influences both vascular and non-vascular developmental processes. Specifically, Adamts9 expressed by mEC worked cooperatively with Adamts20 expressed by craniofacial mesenchyme in closure of the mouse secondary palate, and also influenced development of the myocardium and angiogenesis. Moreover, ADAMTS9 proteolysis of versican, a large, developmentally critical proteoglycan, was crucial in both the palate and the heart. Since ADAMTS9 is expressed globally by mEC during organogenesis and because versican is widely expressed during embryogenesis, our intriguing observations suggest a broader impact of mEC-produced ADAMTS9 during development. Based on these findings, we hypothesize that through the proteolytic modification of secreted proteins, ADAMTS9 produced by mEC influences surrounding cells during organogenesis. To gain a complete understanding of the developmental impact of mEC-derived ADAMTS9, and to elucidate the underlying mechanisms, a conditional targeting approach is required. The specific aim of this RO3 proposal is to achieve conditional deletion of ADAMTS9 in mEC, and perform morphologic analysis of the resulting mouse phenotype, thereby resolving the overall impact on angiogenesis and organogenesis. This will lay the groundwork for continued work on the identified developmental defects, including their underlying mechanisms. Approach: We will delete Adamts9 specifically in mEC at the earliest developmental stages (E8.5) using a Tie2-Cre strain. In addition, we will undertake regulated conditional deletion in mEC at later developmental stages (E10.5 and later) using tamoxifen-induced Cre expression (via a VE-cadherin-Cre ERT2 mouse). Morphologic analysis of the resulting mouse phenotypes will identify effects on angiogenesis and organogenesis. Significance: In addition to the expected impact on angiogenesis, palatogenesis and myocardial development, it is foreseen that the proposed study will elucidate a widespread role for mEC-expressed ADAMTS9 in organogenesis. The proposed work has immediate significance for common human birth defects, such as cleft palate and cardiac anomalies, and may provide new mouse models for other developmental defects.

描述（由申请人提供）：该项目的目标是阐明血管内皮在胚胎发育和器官发生中的作用。毛细血管内的微血管内皮细胞（mEC）在调节血管生成方面具有众所周知的功能，但人们对其在器官发生中的作用知之甚少。我们的初步数据表明，进化上保守的分泌型金属蛋白酶 ADAMTS9（微血管内皮的产物）在正常器官发生中具有重要作用。 Adamts9-/- 小鼠在妊娠 7.5 天 (E7.5) 时死亡，在血管生成和器官发生开始之前。然而，通过使用缺乏 Adamts9 一种等位基因的小鼠，我们发现 ADAMTS9 影响血管和非血管发育过程。具体而言，mEC表达的Adamts9与颅面间充质表达的Adamts20协同作用，闭合小鼠次级腭，并影响心肌的发育和血管生成。此外，ADAMTS9 对多功能蛋白聚糖（一种对发育至关重要的大型蛋白多糖）的蛋白水解作用对于上颚和心脏都至关重要。由于 ADAMTS9 在器官发生过程中由 mEC 全面表达，并且由于多功能蛋白聚糖在胚胎发生过程中广泛表达，因此我们有趣的观察表明 mEC 产生的 ADAMTS9 在发育过程中具有更广泛的影响。基于这些发现，我们假设 mEC 产生的 ADAMTS9 通过分泌蛋白的蛋白水解修饰影响器官发生过程中的周围细胞。为了全面了解 mEC 衍生的 ADAMTS9 的发育影响，并阐明潜在机制，需要有条件的靶向方法。该RO3提案的具体目标是实现mEC中ADAMTS9的条件删除，并对所得的小鼠表型进行形态学分析，从而解决对血管生成和器官发生的整体影响。这将为继续研究已确定的发育缺陷（包括其潜在机制）奠定基础。方法：我们将使用 Tie2-Cre 菌株在最早的发育阶段 (E8.5) 专门删除 mEC 中的 Adamts9。此外，我们将使用他莫昔芬诱导的 Cre 表达（通过 VE-cadherin-Cre ERT2 小鼠）在 mEC 的后期发育阶段（E10.5 及更高版本）进行调节条件删除。对所得小鼠表型的形态学分析将确定对血管生成和器官发生的影响。意义：除了对血管生成、腭生成和心肌发育的预期影响外，预计拟议的研究还将阐明 mEC 表达的 ADAMTS9 在器官发生中的广泛作用。这项工作对于常见的人类出生缺陷（例如腭裂和心脏异常）具有直接意义，并可能为其他发育缺陷提供新的小鼠模型。