Molecular basis for defective pericyte-endothelial cell interactions regulating vascular malformations

调节血管畸形的有缺陷的周细胞-内皮细胞相互作用的分子基础

• 批准号: 10408085
• 负责人: George E Davis
• 金额: $ 38.03万
• 依托单位: UNIVERSITY OF SOUTH FLORIDA
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-07-01 至 2024-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10408085
• 关键词: 1-Phosphatidylinositol 3-Kinase; 3-Dimensional; Address; Affect; Antigens; Arteriovenous malformation; Automobile Driving; Basement membrane; Behavior; Biological; Blood Vessels; Blood capillaries; Brain hemorrhage; CCM1 gene; Cavernous Malformation; Cell Communication; Cell Culture Techniques; Cell Cycle; Cell Cycle Regulation; Cell Line; Cell Proliferation; Cells; Cerebrum; Clinical; Coupled; Cyst; Cystic Lesion; DTR gene; Data; Deposition; Development; Disease; Endothelial Cells; Endothelin-1; Event; Genes; Genetic; Growth; Human; In Vitro; Inflammation Mediators; Interleukin-1 beta; KRAS2 gene; Knock-out; Laboratories; Large T Antigen; Lead; Lesion; Link; Modeling; Molecular; Morphogenesis; Mus; Mutation; PIK3CA gene; Pathogenesis; Pathogenicity; Pathologic; Pathology; Pericytes; Phenotype; Platelet-Derived Growth Factor; Process; Reporting; Role; Signal Transduction; Small Interfering RNA; Specimen; Stains; System; TNF gene; TP53 gene; Thrombin; Tissues; Tube; Up-Regulation; Venous Malformation; Work; brain arteriovenous malformations; brain endothelial cell; cell immortalization; cell type; cerebral cavernous malformations; in vitro Model; in vivo; interest; malformation; mouse model; novel; novel therapeutics; postnatal; recruit; synergism

Capillaries are by far the most abundant blood vessels and are critically interfaced with tissue parenchymal cells to control both development and pathologic disease states. They consist of co-assembled endothelial cells (EC) tube networks with associated pericytes. For many years, our laboratory has been investigating the molecular basis for EC lumen and tube assembly, as well as the mechanisms and EC-derived molecules that control pericyte recruitment, proliferation, and capillary basement membrane deposition, a process that re- quires EC-pericyte interactions. Vascular malformations, such as arteriovenous malformations (AVMs) and cavernous malformations (CMs) (most often observed in cerebral tissue, termed CCMs), constitute a group of pathologies with marked abnormalities in EC tube morphogenesis coupled to deficiencies in mural cell interac- tions. A critical point is that there is a fundamental lack of understanding of the underlying molecular basis for the development of these malformations from either the EC or pericyte perspective. To address these issues, we have developed two novel in vitro models of vascular malformations, an AVM-like model using human ECs expressing a k-Ras activating mutation (i.e. k-RasV12) and a CM-like model using ECs expressing k-RasV12 and T antigen (TAg) (to dysregulate the cell cycle). In the AVM-like case, the ECs markedly accelerate tube formation compared to control ECs, however, pericyte recruitment and basement membrane deposition is strongly reduced compared to controls. In the CM-like case, the modified ECs form large cysts (with no sprout- ing behavior) with evident EC proliferation, while pericytes show responsiveness or no recruitment to the EC- lined cysts (strongly mimicking CMs in vivo). Thus, both of our in vitro models recapitulate what is observed in vivo with AVMs and CMs, and other preliminary data further supports these conclusions. To investigate and correlate in vitro with in vivo findings, we are utilizing mouse models of CCM disorder that delete CCM1 (selec- tively within ECs) in an inducible manner in postnatal mice with or without EC co-induction of activating muta- tions in k-Ras or PI3 kinase. Preliminary data suggests that such activating mutations can markedly enhance CCM development in vivo in conjunction with EC deletion of CCM1, which support our in vitro observations. We propose three specific aims to further investigate the underlying molecular basis for vascular mal- formations and to develop new therapeutic options for these diseases; and they are: Aim #1: Define how k-RasV12 expression in ECs results in accelerated EC tube formation, but reduced peri- cyte-EC interactions leading to arteriovenous-like malformations. Aim #2: Define how k-RasV12 expression in ECs coupled with loss of CCM genes and EC cell cycle regula- tion leads to cavernous-like malformations with markedly deficient pericyte recruitment. Aim #3: Define how pro-inflammatory mediators affect pericyte-EC interactions to regulate the formation or stability of k-RasV12-dependent arteriovenous-like and cavernous-like malformations.

Captured是迄今为止最丰富的血管，并与组织实质 细胞来控制发育和病理疾病状态。它们由共组装的内皮细胞 细胞（EC）与相关周细胞的管网。多年来，我们的实验室一直在研究 EC管腔和管组装的分子基础，以及 控制周细胞募集、增殖和毛细血管基底膜沉积，这是一个重新激活细胞的过程。 需要EC-周细胞相互作用。血管畸形，如动静脉畸形（AVM）和 海绵状血管畸形（CM）（最常在脑组织中观察到，称为CCM），构成了一组 EC管形态发生明显异常的病理学，与壁细胞相互作用的缺陷相结合， 选择。一个关键点是，人们对这种疾病的潜在分子基础缺乏基本的了解。 从EC或周细胞的角度来看这些畸形的发展。为了解决这些问题， 我们已经开发了两种新的血管畸形体外模型，一种是使用人类EC的AVM样模型， 表达k-Ras激活突变（即k-RasV 12）的EC和使用表达k-RasV 12的EC的CM样模型 和T抗原（TAg）（使细胞周期失调）。在AVM样情况下，EC显著加速管 然而，与对照EC相比， 与对照组相比大幅降低。在CM样的情况下，修饰的EC形成大的囊肿（没有芽， 具有明显的EC增殖，而周细胞显示对EC的反应性或不募集。 内衬囊肿（强烈模仿体内CM）。因此，我们的两种体外模型都概括了在 AVM和CM的体内试验以及其他初步数据进一步支持了这些结论。调查和 在体外与体内研究结果相关联，我们正在利用CCM障碍的小鼠模型，删除CCM 1（selec， 在有或没有EC共诱导的情况下，在出生后小鼠中以诱导方式激活穆塔- 在k-Ras或PI 3激酶中的作用。初步数据表明，这种激活突变可以显著增强 CCM在体内的发展与CCM 1的EC缺失相结合，这支持了我们的体外观察结果。 我们提出了三个具体的目标，以进一步研究血管畸形的潜在分子基础， 形成和开发新的治疗方案，这些疾病，他们是： 目的#1：定义EC中的k-RasV 12表达如何导致加速EC管形成，但减少细胞增殖。 细胞-EC相互作用导致动静脉样畸形。 目的#2：确定k-RasV 12在EC中的表达如何与CCM基因和EC细胞周期调节基因的缺失相结合。 导致海绵状畸形，周细胞募集明显不足。 目的#3：定义促炎介质如何影响周细胞-EC相互作用，以调节周细胞的形成或 k-RasV 12依赖性动静脉样和海绵样畸形的稳定性。