Endothelial Regulation of Vascular Calcification

血管钙化的内皮调节

• 批准号: 10363955
• 负责人: Kristina I Bostrom
• 金额: $ 54.09万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-01-01 至 2025-12-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10363955
• 关键词: Angiogenesis Inhibitors; Aorta; Appearance; Area; Arteries; Blood capillaries; CD34 gene; Cardiovascular Diseases; Cartilage; Cell Lineage; Cell Maintenance; Cells; Chondrocytes; Clinical; Complication; Development; Disease; Endothelial Cells; Endothelium; Gene Deletion; Gene Expression Profile; Glycoproteins; Goals; Human; In Vitro; Intervention; Knock-out; Lead; Location; Mesenchymal; Modeling; Morbidity - disease rate; Mouse Protein; Mus; Operative Surgical Procedures; Osteogenesis; PECAM1 gene; Pathology; Peripheral; Play; Population; Prevention; Public Health; RNA analysis; Regulation; Role; Sampling; Severities; Side; Signal Transduction; Supporting Cell; Testing; Tunica Adventitia; Vascular Diseases; Vascular Endothelial Growth Factors; Vascular Endothelium; Vascular calcification; angiogenesis; bone; calcification; endothelial-specific sialomucin; experimental study; in vivo; inhibitor; matrix Gla protein; mortality; notch protein; novel; osteogenic; osteoprogenitor cell; postnatal; receptor; recruit; single-cell RNA sequencing; stem cells; therapy development; transcription factor; transcriptome sequencing; treatment strategy

PROJECT SUMMARY: Vascular calcification (VC) frequently complicates cardiovascular disease. It increases the morbidity and mortality and constitutes a significant obstacle in interventions and surgeries. The vascular endothelium plays an important role in VC. The intimal (luminal) endothelial cells (ECs) contribute to VC by providing osteoprogenitor cells through endothelial-mesenchymal transitions (EndMTs). The adventitial ECs are known to contribute to neo-angiogenesis in diseased vascular wall, but it is unknown whether such ECs support VC and what defines them. A subset of ECs with high expression of the EC marker CD31 and the glycoprotein Endomucin (Emcn) has been found in bone to support bone formation. It is possible that adventitial or other peripheral ECs are recruited to diseased areas to promote calcification. Preliminary experiments, using the Matrix Gla Protein null (Mgp-/-) mouse as a VC model, showed extensive EC involvement in the calcified aorta. We identified two subtypes of ECs in the adventitial vs. the intimal endothelium (referred to as a-ECs and i- ECs). The a-ECs were CD31+Emcm+ and correlated with the severity of the VC, whereas the i-ECs were CD31+Emcn-. The two ECs had distinct transcriptional profiles with stem cell and osteogenic markers in the i- ECs vs. enhanced Notch expression in the a-ECs. Endothelial deletion of Notch1 reduced the a-ECs and limited VC while promoting cartilage formation and survival in the Mgp-/- mice. The bone transcription factor Osterix was expressed in both types of ECs. We hypothesize that a-ECs are distinct from i-ECs, recruited to nascent VC, and susceptible to Notch disruption. We also hypothesize that Osterix is protective of EC lineage. In Aim 1, we will characterize the a-ECs (CD31+Emcn+) and compare to the i-ECs (CD31+Emcn-) in the Mgp-/- model, and correlate with severity of VC and marker expression. We will identify unique markers for the respective ECs, with comparison to bone, using transcriptional profiles from single cell RNA sequencing (scRNAseq). We will test the concept that angiogenesis is required for VC using angiogenic inhibitors. In Aim 2, we will determine the effect of loss or gain of endothelial Notch signaling on the EC subtypes and VC. We will examine the distribution of Notch components in relation to VC, and generate Mgp-/- mice with endothelial- specific loss of Notch1 or the Notch receptor inactivator Fbxw7. We will use the mice to determine the effect on the appearance of the endothelial subtypes, calcification and transcriptional profiles by scRNAseq. We will also apply loss and gain of Notch to ECs in vitro and identify novel Notch targets and networks. In Aim 3, we will determine if Osterix helps maintain EC lineage or promotes calcification in vitro and in vivo using human aortic ECs and inducible endothelial-specific Osterix gene deletion in Mgp-/- mice. We will compare the transcriptional profiles of ECs with and without Osterix by scRNAseq in order to clarify the effect on EC lineage, a-ECs vc. I- ECs, and involved signaling networks. Our results may have a significant impact on the field of VC, in particular on the understanding of the endothelial pathology and involvement in VC.

项目摘要： 血管钙化（VC）经常使心血管疾病复杂化。它增加了发病率和 死亡率和构成干预和手术的重要障碍。血管内皮播放 在VC中的重要作用。内膜（腔）内皮细胞（EC）通过提供对VC的贡献 骨基因细胞通过内皮间充质转变（ENDMTS）。外来EC是已知的 在病态血管壁中有助于新血管生成，但尚不清楚这种ECS是否支持VC 以及定义它们的原因。 EC标记CD31和糖蛋白具有高表达的EC的子集 在骨骼中发现了内核素（EMCN）以支撑骨形成。可能存在外在或其他 外围EC被招募到患病区域以促进钙化。初步实验，使用 矩阵GLA蛋白NULL（MGP - / - ）小鼠作为VC模型，显示出广泛的EC参与到钙化的主动脉中。 我们在外来膜和内皮内皮中确定了EC的两个亚型（称为A-EC和I- ECS）。 A-EC是CD31+ EMCM+，与VC的严重程度相关，而I-EC是 CD31+EMCN-。这两个EC在I-中具有与干细胞和成骨标记的不同转录曲线 ECS与A-EC中增强的Notch表达。 Notch1的内皮缺失减少了A-EC和 在MGP - / - 小鼠中促进软骨形成和存活的同时，有限的VC。骨转录因子 在两种类型的EC中都表达了Osterix。我们假设A-EC与I-EC不同，被招募到 新生的VC，容易受到缺口干扰。我们还假设Osterix保护了EC谱系。 在AIM 1中，我们将表征A-EC（CD31+EMCN+），并与MGP - / - 中的I-EC（CD31+EMCN-）进行比较。 模型，与VC和标记表达的严重程度相关。我们将确定独特的标记 与骨骼相比，相应的EC使用单细胞RNA测序的转录曲线 （scrnaseq）。我们将测试使用血管生成抑制剂VC需要血管生成的概念。目标 2，我们将确定内皮缺口信号传导对EC亚型和VC的损失或增益的影响。我们 将检查与VC相关的Notch组件的分布，并与内皮生成MGP - / - 小鼠 Notch1或Notch受体灭活剂FBXW7的特异性损失。我们将使用小鼠确定对 Scrnaseq的内皮亚型，钙化和转录曲线的外观。我们也会 将Notch的损失和增益应用于ECS的体外，并确定新颖的Notch目标和网络。在AIM 3中，我们将 确定Osterix是否有助于维持EC谱系或使用人主动脉在体外和体内促进钙化 MGP - / - 小鼠中的EC和可诱导的内皮特异性Osterix基因缺失。我们将比较转录 为了阐明对EC谱系的影响A-ECS VC，带有和不带Osterix的EC的概况。我- EC和涉及信号网络。我们的结果可能对VC领域有重大影响 特别是了解内皮病理学和参与VC的理解。