Reverse Tissue-Manufacturing of the Multicellular Sinoatrial Node Organoids

项目摘要/摘要 最近，窦房结功能障碍的患病率、严重程度和医疗负担不断增加 强调有必要对SA结节功能进行更详细的研究，以便有效地治疗和治疗 预防SA节点功能障碍。起搏器功能障碍的主要机制是起搏器功能受损。 细胞在其传导过程中诱发自发节律(自律性)和逆向重塑 周围心房组织(SA传导)。然而，目前的SA节点或起搏器型号已经 仅限于理论模型和孤立的单个细胞类型的细胞或细胞团，为建模留下了空白 自主性心脏收缩和心律，以及自律性和窦性心动传导功能障碍。此外， 目前的单细胞型起搏器在体内整合一个月期间心律稳定性变差， 这限制了其作为临床可行的生物起搏器的应用，该起搏器能够产生强大的起搏器 和传导性。 为了解决目前SA节点模型的局限性，本方案旨在开发一种三维的SA节点模型 通过复制人SA节点的多细胞组织结构和故障安全的多细胞SA节点有机体 机械装置。与单细胞型生物起搏器不同的是，人的SA结是一种天然的器官，具有 精心设计的绝缘建筑和不同种类的蜂窝结构。此外，人工SA节点是 配备多余的起搏器部位和传导通路，以保护心律免受不利影响 变时性刺激。因此，受SA节点结构和故障安全机制的启发，我们的目标是增强 自动化和SA传导的健壮性：首先，我们将专注于增强SA的自动性 结节类有机物通过鉴定起搏膜和钙钟蛋白的表达，细胞 构图和形状(目标1)。第二，我们将集中精力改善SA结点有机体的导电性 通过协调多个起搏器部位和传导通路(目标2)。最后，我们将评估 体外环境和体内房室传导阻滞啮齿动物模型中SA结器的稳健性(目标3)。 这些研究将确定组织级架构和多细胞组成是否中介SA节点的健壮性 起搏和传导，并可能揭示高保真的组织级生物起搏器作为一种新的 SA结节功能障碍的治疗策略。所提出的有机化合物将适用于人类临床前研究 加速药物开发，解剖患者特有的SA结节疾病病理生理学的测试分析， 以及植入式生物起搏器的开发。

Project Summary/Abstract The recent increasing prevalence, severity, and healthcare burden of sinoatrial (SA) node dysfunction emphasize the need for more detailed studies of SA node functions that allow for effective therapy to treat and prevent SA node dysfunction. The major mechanisms of the dysfunction are the impaired ability of pacemaker cells to induce spontaneous rhythm (automaticity) and adverse remodeling in their electric conduction to surrounding atrial tissues (SA conduction). However, the current SA node or pacemaker models have been limited to theoretical models and isolated single cell-type cells or cell clusters, leaving a gap to model the autonomous cardiac contraction and heart rhythm and dysfunctions in automaticity and SA conduction. Moreover, the current single cell-type pacemakers worsened heart rhythm stability during one-month in vivo integration, which limits its application as a clinically viable biological pacemaker capable of generating robust pacemaking and conduction. To address the current limitation of SA node models, this proposal aims to develop a three-dimensional multicellular SA node organoid by reproducing human SA node’s multicellular tissue structure and fail-safe mechanisms. In contrast to the single cell-type biological pacemakers, human SA node is a natural organoid with elaborate insulated architecture and heterogeneous cellular composition. Moreover, the human SA node is equipped with redundant pacemaker sites and conduction pathways to protect the rhythm against adverse chronotropic stimulations. Thus, inspired by SA node’s structure and fail-safe mechanism, we aim at enhancing robustness in both automaticity and SA conduction: First, we will focus on enhancing automaticity of SA node organoids by identifying the expression of pacemaker membrane and calcium clock proteins, cell composition, and shape (Aim 1). Second, we will concentrate on improving conduction of SA node organoids by coordinating multiple pacemaker sites and conduction pathways (Aim 2). Last, we will evaluate the robustness of the SA node organoids in in vitro setting and in vivo atrioventricular block rodent model (Aim 3). These studies will define if tissue-level architecture and multicellular compositions mediate SA node’s robust pacemaking and conduction and may reveal a high-fidelity tissue-level biological pacemaker as a novel therapeutic strategy for SA node dysfunctions. The proposed organoids will be suitable for human preclinical testing assays to accelerate drug development, for dissecting patient-specific SA node disease pathophysiology, and for the development of implantable biological pacemakers.