Reverse Tissue-Manufacturing of the Multicellular Sinoatrial Node Organoids

项目总结/摘要 最近窦房结（SA）功能障碍的患病率、严重程度和医疗负担增加 强调需要对SA结功能进行更详细的研究，以便进行有效的治疗， 防止SA结功能障碍。其主要机制是起搏器功能受损 细胞诱导自发节律（自律性）和不利的重塑，在他们的电传导， 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 局限于理论模型和孤立的单细胞型细胞或细胞簇，留下了一个空白，以模拟 自主性心脏收缩和心律以及自律性和SA传导功能障碍。此外，委员会认为， 目前的单细胞型起搏器在一个月的体内整合期间使心律稳定性恶化， 这限制了其作为能够产生稳健起搏的临床上可行的生物起搏器的应用 和传导。 为了解决SA节点模型的当前局限性，该提议旨在开发三维 通过复制人类SA结的多细胞组织结构和故障安全性， 机制等与单细胞型生物起搏器相比，人窦房结是一种天然的类器官， 精心设计的绝缘结构和异质的细胞组成。此外，人类的SA结是 配备冗余起搏器部位和传导通路，以保护心律不受不良影响 变时性刺激因此，受SA节点的结构和故障安全机制的启发，我们的目标是增强 自动性和SA传导的鲁棒性：首先，我们将专注于增强SA的自动性 通过鉴定起搏膜和钙时钟蛋白的表达， 结构和形状（目标1）。其次，我们将专注于改善SA结类器官的传导 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 SA结类器官在体外环境和体内房室传导阻滞啮齿动物模型中的稳健性（目的3）。 这些研究将确定是否组织水平的结构和多细胞组成介导SA节点的鲁棒性。 起搏和传导，并可能揭示高保真组织水平的生物起搏器作为一种新的 SA结功能障碍的治疗策略。所提出的类器官将适用于人类临床前研究。 测试分析以加速药物开发，用于剖析患者特异性SA结疾病病理生理学， 以及可植入生物起搏器的开发。

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.