Role of integrin alpha 5 in the formation of the pharyngeal arch arteries

整合素α5在咽弓动脉形成中的作用

• 批准号: 10514523
• 负责人: Michael Warkala
• 金额: $ 4.1万
• 依托单位: RBHS-NEW JERSEY MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-04-01 至 2023-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10514523
• 关键词: 3-Dimensional; Address; Affect; Apoptosis; Arteries; Blood; Blood Vessels; Branchial arch structure; Cardiac; Cell Count; Cell Nucleus; Cell Proliferation; Cell Surface Receptors; Cells; Clinical; Confocal Microscopy; Congenital Abnormality; Contralateral; Data; Defect; Detection; Development; Ectoderm; Embryo; Embryonic Development; Endoderm; Endothelial Cells; Ensure; Etiology; Extracellular Matrix; Frequencies; Genes; Heart; ITGA5 gene; Immunofluorescence Immunologic; Immunohistochemistry; Integrins; Invaded; Lead; Left; Legal patent; Live Birth; Mediating; Mus; Nuclear; Operative Surgical Procedures; Organ; Pattern; Population; Prevention; Reporter; Role; Severities; Side; Signal Transduction; Stains; Structure; Testing; Time; aortic arch; cell motility; conditional knockout; confocal imaging; congenital heart disorder; experimental study; insight; migration; mouse model; mutant; neonate; novel; prevent; reconstruction; recruit; response; stem cell survival; stem cells; vasculogenesis

PROJECT SUMMARY The aberrant formation of the Pharyngeal Arch Arteries (PAAs) results in severe and lethal forms of congenital heart disease. The PAAs form through vasculogenesis, where an endothelial cell (EC) plexus invades the pharyngeal arches before coalescing into patent PAAs. Our lab has shown that a majority of the ECs that make up the PAAs derives from a population of mesodermal progenitor cells known as the Second Heart Field (SHF). Additionally, our lab has shown that Integrin ɑ5 (Itgɑ5) expression within the Isl1-lineages, encompassing the second heart field and pharyngeal endoderm and ectoderm, regulates the formation of the PAAs through unknown mechanisms. Integrins are cell surface receptors that organize the ECM and relay ECM signals to elicit cellular responses, including cell proliferation, survival, migration, and differentiation. Preliminary data suggests Itgɑ5 regulates the number of SHF-derived ECs that give rise to the PAAs. We hypothesize that integrin ɑ5 expression in the Isl1-lineages regulates the number of SHF-derived ECs in the pharyngeal arches by mediating SHF-progenitor cell survival and/or proliferation, migration, and/or differentiation. In Specific Aim I, we will investigate this hypothesis using Itgɑ5f/- ; Isl1Cre mutant and Itgɑ5f/+ ; Isl1Cre control embryos, which carry the Rosa26nTnG reporter. This reporter marks cells derived from the Isl1-lineages with the expression of nuclear GFP. Utilizing this conditional knockout mouse model and immunohistochemistry, we will investigate the proliferation and apoptosis frequencies, migration patterns, and differentiation capacity, of the Isl1-derived cells that give rise to the ECs of the PAAs. My preliminary data additionally shows SHF-derived ECs crossing the midline in the developing pharyngeal region within our mutants. During vertebrate development, cells in general do not cross the midline ensuring proper left-right patterning. The ECM organization and programmed cell death response at the midline create a barrier that stops cells from migrating across the midline. Because we know Itgɑ5 regulates ECM organization we hypothesize that Itgɑ5 expression in the Isl1-lineages regulates SHF-derived EC patterning through mediating the ECM organization and the programmed cell death response at the midline. We will investigate this hypothesis in Specific Aim II, utilizing our conditional mouse model and immunohistochemistry. We will compare the ECM organization and apoptosis frequency at the midline in control and mutant embryos. Additionally, we will investigate the distribution of Isl1-derived cells at the midline during early embryogenesis because if Isl1-derived cells are able to cross the midline in our mutants, the Isl1-derived cells that give rise to the PAAs could be directed to the wrong side, resulting in aberrant PAA formation. The completion of the experiments outlined in this proposal will provide insights into mechanisms involved in the development of the PAAs and will therefore increase our understanding of how related congenital heart diseases arise, creating new possibilities for the clinical detection, prevention, and treatments of these congenital heart diseases.

项目摘要 咽弓动脉（PAA）的异常形成导致严重和致命形式的先天性 心脏病PAA通过血管发生形成，其中内皮细胞（EC）丛侵入血管， 咽弓在合并成未闭的肺动脉瘤之前。我们的实验室已经表明， 这些PAA来源于一群被称为第二心脏区域（SHF）的中胚层祖细胞。 此外，我们的实验室已经表明，整合素β 5（Itg β 5）表达的Isl 1谱系，包括 第二心脏领域和咽内胚层和外胚层，通过调节PAA的形成， 未知的机制。整合素是细胞表面受体，其组织ECM并中继ECM信号以引起细胞凋亡。 细胞反应，包括细胞增殖、存活、迁移和分化。初步数据提示 Itg β 5调节产生PAA的SHF衍生的EC的数量。我们假设整合素β 5 Isl 1-谱系中的表达通过介导 SHF-祖细胞存活和/或增殖、迁移和/或分化。在具体目标I中，我们将 使用Itg Δ 5 f/- ; Isl 1Cre突变体和Itg Δ 5 f/+ ; Isl 1Cre对照胚胎研究这一假设，这些胚胎携带 Rosa 26 nTnG记者。该报告基因标记来自Isl 1谱系的细胞，表达核GFP。 利用这种条件性基因敲除小鼠模型和免疫组化，我们将研究增殖 以及Isl 1衍生细胞的凋亡频率、迁移模式和分化能力， 到临时行政区的选举委员会。我的初步数据还显示，SHF衍生的EC穿过中线， 我们变异人的咽部在脊椎动物的发育过程中，细胞一般不交叉， 中线确保正确的左右图案。ECM组织和程序性细胞死亡反应 中线形成阻止细胞迁移穿过中线的屏障。因为我们知道Itg β 5调节着 ECM组织我们假设Isl 1谱系中Itg β 5的表达调节SHF衍生的EC模式 通过介导ECM组织和中线的程序性细胞死亡反应。我们将 在Specific Aim II中，利用我们的条件性小鼠模型和免疫组织化学研究这一假设。 我们将比较对照和突变胚胎中线处的ECM组织和凋亡频率。 此外，我们将研究早期胚胎发生期间Isl 1衍生细胞在中线的分布 因为如果Isl 1衍生的细胞能够穿过我们突变体的中线，那么Isl 1衍生的细胞就会产生 PAA可能被引导到错误的一侧，导致异常PAA形成。的完成 在这项建议中概述的实验将提供深入了解参与发展的机制， 因此，PAAs将增加我们对相关先天性心脏病如何发生的理解， 这些先天性心脏病的临床检测，预防和治疗的可能性。