Administrative support to R01 HL148104: Understanding Cardiac C-Looping Using Microscale In Vitro Models

R01 HL148104 的行政支持：使用微型体外模型了解心脏 C 环

• 批准号: 10630645
• 负责人: Leo Q. Wan
• 金额: $ 6.78万
• 依托单位: RENSSELAER POLYTECHNIC INSTITUTE
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-09-01 至 2025-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10630645
• 关键词: Address; African American; Anterior; Biochemical; Biological Sciences; Biomechanics; Biophysical Process; Cardiac; Cardiac Myocytes; Cardiac development; Cells; Chick; Chick Embryo; Clinical; Collecting Tube; Defect; Development; Dextrocardia; Disease; Double Outlet Right Ventricle; Embryo; Embryonic Development; Embryonic Heart; Event; Fetal Death; Funding; Grant; Handedness; Health; Heart; Human; In Vitro; Individual; Infant; Institutes; Knowledge; Left; Location; Morphogenesis; Morphology; Myocardium; Myosin Type II; N-Cadherin; Organ; Outcome; Pathway interactions; Pattern; Pharmaceutical Preparations; Protein Kinase C; Proteins; Publications; Research; Research Support; Role; Side; Signal Transduction; Surface; Testing; Travel; Tube; United States National Institutes of Health; Vertebrates; WNT Signaling Pathway; base; cardiogenesis; cost; genetic approach; graduate student; human disease; immunocytochemistry; in vitro Model; planar cell polarity; septal defect; simulation; small molecule

During normal cardiac development of vertebrates, the embryonic heart starts as a straight tube along the midline of the embryo. It subsequently transforms into a c-shaped heart loop reliably toward the right side of the body. This cardiac c-looping is the earliest evident event of left-right asymmetry breaking (also called chirality or handedness) of a human organ. The inversed lateralization (i.e., toward the left side) of cardiac looping often leads to severe clinical outcomes, including dextrocardia, septum defects, double outlet right ventricle, and even death of fetuses and infants. In the past, we have been able to recapitulate multicellular chiral morphogenesis in vitro and shown that cardiac cells from chick hearts right before looping have an intrinsic rightward bias. Also, protein kinase C activators, newly identified regulators of cardiac cell chirality, can reverse chick cardiac c-looping. In the funded R01 grant (HL148104), we proposed to investigate how intrinsic cell chirality determines asymmetric cardiac looping biochemically and biomechanically. We now request a research supplement to address a Reviewers’ concern (relevance to human diseases) by using human cardiac cells and investigating the role of the planar cell polarity (PCP; also called noncanonical wnt) signaling, a pathway known for cardiac laterality defects. We hypothesize that PCP regulates the scale of multicellular chiral morphogenesis and cardiac c-looping. To test the hypothesis, we propose to evaluate the expression of PCP proteins and assess the role of PCP proteins in forming 2D in vitro multicellular chiral patterns and ex ovo looping hearts in chick embryonic development. Specifically, we will first assess the polarity and chirality of PCP markers in patterned human cardiac cells. We will pattern cardiomyocytes in 2D, examine the location and morphology of the PCP markers, and determine how PCP signals biomechanically regulate chiral morphogenesis. Then, we will determine the role of PCP signaling in chick cardiac c looping. With the chick ex ovo culture, we will evaluate the localization of PCP proteins during the looping stages, interfere with PCP signaling to examine the effects of PCP, and use cell vertex-based numerical simulation to assess the biomechanical role of PCP signaling in the asymmetric heart looping. Overall, the proposed research will delineate the role of PCP signaling in chiral multicellular morphogenesis and in early cardiac asymmetric looping.

在脊椎动物正常的心脏发育过程中，胚胎心脏开始时是一个沿着 胚胎的中线。它随后转变成一个c形的心脏循环可靠地向右侧 身体这种心脏c环是最早的明显的左右不对称破坏事件（也称为 手性或手性）。反向偏侧化（即，向左） 成环常导致严重的临床结局，包括心绞痛、间隔缺损、右双出口 心室，甚至胎儿和婴儿死亡。在过去，我们已经能够概括多细胞 体外手性形态发生，并显示在循环之前，来自鸡心脏的心脏细胞具有 内禀偏置此外，蛋白激酶C激活剂，新发现的心脏细胞手性调节剂，可以 反向鸡心脏C环在资助的R 01补助金（HL 148104）中，我们建议调查内在的 细胞手性在生物化学和生物力学上决定了不对称的心脏循环。我们现在请求 研究补充，以解决审查员的关注（与人类疾病的相关性），通过使用人类心脏 细胞和调查的作用，平面细胞极性（PCP;也称为非典型wnt）信号， 已知的心脏偏侧缺陷的通路。我们假设PCP调节了多细胞 手性形态发生和心脏C环。为了验证这一假设，我们建议评估 PCP蛋白质，并评估PCP蛋白质在体外形成2D多细胞手性模式和体外形成2D多细胞手性模式中的作用。 鸡胚胎发育中的环状心脏。具体来说，我们将首先评估的极性和手性 图案化的人类心脏细胞中的PCP标记物。我们将在2D模式心肌细胞，检查位置 和PCP标记物的形态，并确定PCP信号如何生物力学调节手性 形态发生然后，我们将确定PCP信号在鸡心脏c循环中的作用。和那个小妞前男友 卵培养，我们将评估PCP蛋白在成环阶段的定位， 信号检查PCP的影响，并使用基于细胞顶点的数值模拟来评估 PCP信号在不对称心脏循环中的生物力学作用。总的来说，拟议的研究将 描述PCP信号传导在手性多细胞形态发生和早期心脏不对称中的作用 循环