A HUMAN IPSC-BASED ORGANOID PLATFORM FOR STUDYING MATERNAL HYPERGLYCEMIA-INDUCED CONGENITAL HEART DEFECTS

基于人体 IPSC 的类器官平台，用于研究母亲高血糖引起的先天性心脏缺陷

• 批准号: 10752276
• 负责人: Javier Contreras
• 金额: $ 4.13万
• 依托单位: RESEARCH INST NATIONWIDE CHILDREN'S HOSP
• 依托单位国家: 美国
• 财政年份: 2024
• 资助国家: 美国
• 起止时间: 2024-02-01 至 2029-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10752276
• 关键词: 3-Dimensional; Address; Adherent Culture; Adult; Affect; Animals; Apoptosis; Biological; Biological Assay; Blood Glucose; Cardiac; Cardiac Myocytes; Cardiovascular system; Cell Culture Techniques; Cell Differentiation process; Cell Lineage; Cell Maturation; Cells; Cellular Metabolic Process; Congenital Abnormality; Congenital Heart Defects; Culture Media; Development; Diabetic mother; Differentiated Gene; Disease; Dose; Embryonic Development; Endothelium; Environment; Environmental Risk Factor; Fluorescence; Gene Expression; Gene Expression Profiling; Generations; Genes; Genetic Risk; Gestational Diabetes; Glucose; Goals; Heart; Heart Abnormalities; Heterogeneity; Human; Hyperglycemia; Immunofluorescence Immunologic; Impairment; Incidence; Infant Mortality; Knowledge; Live Birth; Measures; Mesoderm; Mesoderm Cell; Metabolic; Metabolism; Mitochondria; Modeling; Mus; Myocardial; Organ; Organoids; Oxidative Stress; Pathogenesis; Phenotype; Physiological; Physiology; Population; Pregnancy; Pregnant Women; Production; Reactive Oxygen Species; Reporting; Respiration; Risk; Sampling; Signal Transduction; Specific qualifier value; Stains; Stem Cell Development; Structure; Supplementation; Translations; WNT Signaling Pathway; cardiogenesis; cell determination; cell injury; cell type; congenital heart disorder; differential expression; differentiation protocol; heart cell; high risk; human model; human stem cells; human tissue; induced pluripotent stem cell; insight; interest; maternal diabetes; maternal hyperglycemia; mitochondrial metabolism; mortality; mouse model; offspring; oxidative damage; response; single-cell RNA sequencing; stem cell differentiation; stem cells; therapeutic evaluation; transcriptome sequencing; transcriptomics; two-dimensional

Congenital heart defects (CHD) are the most common type of birth defect, with cardiac malformation resulting from abnormal heat development contributed by genetic and/or environmental risk factors. The association of maternal diabetes with increased offspring CHD incidence is proposed to be attributed to factors including glucose imbalance and metabolic dysregulation. Murine studies on hyperglycemia have been beneficial in uncovering the phenotypic effects of maternal hyperglycemia on embryonic development, with reports that high glucose conditions are associated with general suppression of WNT signaling, apoptosis, and excessive reactive oxygen species (ROS) production. However, little is known about the effects of hyperglycemia on human stem cells in regard to cardiomyocyte determination, metabolism, and functionality. Additionally, there is little reports of these alterations using induced pluripotent stemcells (iPSCs), which provide an opportunityto observe cardiac progenitor cell (CPC) heterogeneity, multicellular crosstalk, and differentiated cell functionality at key development timepoints. We hypothesize that high glucose environments interfere with normal cardiac differentiation to cause altered cell lineage determination in iPSCs and iPSC-derived cardiomyocytes (iPSC- CMs). We propose use of immunofluorescence assays and staining to quantify mitochondrial metabolism, mitochondrial ROS production, and apoptosis marker amounts. Our goal is to investigate the impact of high glucose on human cardiomyocyte differentiation and crosstalk signaling abnormalities during 3D stem cell differentiation that could lead to impaired cardiac development. We will investigate differentially expressed genes at the single-cell resolutionin early and late differentiation in responseto high glucose doses in iPSC populations. Should our transcriptional profiling comparisons yield iPSC and iPSC-CM expression differences, the resulting phenotypes would provide insight into cardiac cell determination during hyperglycemic conditions. Understanding the hyperglycemia-response mechanisms is important in the onset of maternal hyperglycemia- associated CHD. These mechanisms would be further investigated through looking at cell lineage population composition in organoids and analyze response per lineage to narrow down what structures of the mature heart are affected.

先天性心脏病（CHD）是最常见的出生缺陷类型，会导致心脏畸形 由遗传和/或环境风险因素导致的异常发热。该协会的 孕产妇糖尿病导致后代冠心病发病率增加的原因包括： 葡萄糖失衡和代谢失调。对高血糖的小鼠研究已有益于 揭示母体高血糖对胚胎发育的表型影响，有报道称高血糖 葡萄糖状况与 WNT 信号传导的普遍抑制、细胞凋亡和过度反应有关 氧物质（ROS）的产生。然而，人们对高血糖对人体干细胞的影响知之甚少。 心肌细胞的决定、代谢和功能。另外，相关报道还很少 使用诱导多能干细胞（iPSC）研究这些改变，这提供了观察心脏的机会 祖细胞 (CPC) 异质性、多细胞串扰和关键的分化细胞功能 开发时间点。我们假设高葡萄糖环境会干扰正常的心脏功能 分化导致 iPSC 和 iPSC 衍生心肌细胞 (iPSC- CM）。我们建议使用免疫荧光测定和染色来量化线粒体代谢， 线粒体 ROS 产生和凋亡标记物量。我们的目标是调查高风险的影响 葡萄糖对 3D 干细胞过程中人心肌细胞分化和串扰信号异常的影响 分化可能导致心脏发育受损。我们将研究差异表达基因 在 iPSC 群体中响应高葡萄糖剂量的早期和晚期分化中的单细胞分辨率。 如果我们的转录谱比较产生 iPSC 和 iPSC-CM 表达差异，则结果 表型将有助于深入了解高血糖条件下心肌细胞的测定。 了解高血糖反应机制对于孕产妇高血糖的发生非常重要 相关的冠心病。这些机制将通过观察细胞谱系群体来进一步研究 类器官的组成并分析每个谱系的反应，以缩小成熟心脏的结构范围 受到影响。