Pacemaker cells and mechanism in the renal pelvis

肾盂起搏细胞及其机制

基本信息

批准号：
10397176
负责人：
KENTON M SANDERS
金额：
$ 47.77万
依托单位：
UNIVERSITY OF NEVADA RENO
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-03-01 至 2023-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10397176
关键词：
Address Affect Alpha Cell Anatomy Animal Model Automobile Driving Biological Markers Bladder Cells Chimeric Proteins Data Defect Development Diabetes Mellitus Distal Electrophysiology (science)Exhibits Fertilization Functional disorder Future Gene Expression Gene Expression Profile Gene Expression Profiling Generations Heart Histones Hydronephrosis Image In Situ Inflammation Intervention Investigation Ion Channel Kidney Kidney Failure Location Molecular Monitor Morphology Mouse Strains Mus Muscle Obstruction Organ Outcome Study PDGFRB gene Pacemakers Pathologic Periodicity Peristalsis Phenotype Physiological Platelet-Derived Growth Factor alpha Receptor Population Population Heterogeneity Pump Reagent Regulatory Pathway Renal pelvis Reporter Role Signal Pathway Signal Transduction Signaling Molecule Site Smooth Muscle Smooth Muscle Myocytes Techniques Testing Therapeutic Intervention Tissue Model Tissues Transgenic Mice Ureter Urinary tract Urine Visceral cell type clinical investigation follow-up interstitial interstitial cell kidney cell new technology nodal myocyte novel optogenetics pre-clinical preclinical study prevent promoter receptor renal damage sensor targeted treatment therapeutic evaluation

项目摘要

Project Summary The renal pelvis (RP), a smooth muscle organ that transports urine from the kidney to the ureter, generates regular rhythmic contractions that are vital for urine transport and bladder filling. Propagating contractions originate in the most proximal region of the RP, where specialized cells, called “atypical” smooth muscle cells (ASMCs)”, have been proposed to serve as the pacemaker cells. However, identification of ASMCs has been imprecise, and thus experimental findings regarding their phenotype and functions are controversial. Ambiguities arise from the lack of specific markers for ASMCs and have prevented understanding the role of ASMC in driving peristaltic contractions or the mechanism of pacemaker activity in the RP. Newer technologies can now identify specific cell-types within tissues composed of heterogeneous populations of cells. This study will employ strains of mice with cell-specific reporters and optogenetic sensors to clarify how the pacemaker and responder cells of the RP generate peristaltic contractions. Our preliminary data show that the specialized cells, ASMCs, express platelet-derived-growth-factor-receptor-alpha (PDGFRα), and using transgenic mice that express a histone 2B- eGFP fusion protein driven off the endogenous PDGFRα+ promoter, we can identify these cells unequivocally in intact tissues or in enzymatic dispersions of the tissues. We hypothesize that PDGFRα+ cells are pacemaker cells in the renal pelvis. The following Specific Aims will be addressed: 1. Test the hypothesis that PDGFRα+ are the primary pacemaker cells of the RP; 2. Investigate the dynamics of Ca2+ signaling in PDGFRα+ cells; 3. Elucidate the specific mechanisms of pacemaker generation and propagation. Because of their specialized function as pacemaker cells, PDGFRα+ cells are likely to have specialized gene expression patterns that encode essential ionic conductances and other signaling molecules to facilitate pacemaker activity. This will be investigated by analysis of gene expression in FACS-enriched populations PDGFRα+ cells isolated from the proximal RP of the reporter strain of mice. Mice expressing a genetically encoded Ca2+ indicator (GECI) in PDGFRα + cells will be used to image Ca2+ signaling in PDGFRα+cells in situ. The ion channels activated by Ca2+ dynamics will be determined and the consequences of this conductance in RP peristalsis will be evaluated. Preliminary data show that PDGFRα+ cells exhibit dynamic Ca2+ signaling in situ and express the Ca2+-activated- Cl- channel, Ano1, making this a prime candidate for the pacemaker conductance. This project will serve as a basis for future studies to understand developmental or pathological problems affecting RP function. OMB No. 0925-0001/0002 (Rev. 01/18 Approved Through 03/31/2020) Page Continuation Format Page

项目概要肾盂 (RP) 是一种平滑肌器官，负责将尿液从肾脏输送到输尿管，产生有规律的节律性收缩对于尿液输送和膀胱充盈至关重要。起源于 RP 的最近端区域，其中有称为“非典型”平滑肌细胞的特殊细胞（ASMC）”，已被提议用作起搏细胞，然而，ASMC 的鉴定已被证实。不精确，因此有关其表型和功能的实验结果存在争议。其原因是缺乏 ASMC 的特异性标记物，并且阻碍了对 ASMC 在驱动中的作用的理解。现在可以识别 RP 中的蠕动收缩或起搏器活动机制。本研究将使用由异质细胞群组成的组织内的特定细胞类型。具有细胞特异性生产者和光遗传学传感器的小鼠，以阐明起搏器和反应细胞如何我们的初步数据表明，特化细胞 ASMC 会产生蠕动收缩。血小板衍生生长因子受体-α (PDGFRα)，并使用表达组蛋白 2B- 的转基因小鼠 eGFP 融合蛋白驱动内源性 PDGFRα+ 启动子，我们可以明确地识别这些细胞我们认为 PDGFRα+ 细胞是起搏器。将解决以下具体肾脏目标： 1. 检验 PDGFRα+ 的假设。是 RP 的主要起搏细胞；2. 研究 PDGFRα+ 细胞中 Ca2+ 信号传导的动态；阐明起搏器产生和传播的具体机制，因为它们具有特殊性。作为起搏细胞，PDGFRα+细胞可能具有编码的特殊基因表达模式促进起搏器活动的重要离子电导和其他信号分子。通过分析从 FACS 富集的群体中分离出的 PDGFRα+ 细胞的基因表达来进行研究表达基因编码的 Ca2+ 指示剂 (GECI) 的小鼠报告品系的近端 RP。 PDGFRα + 细胞将用于对 PDGFRα+ 细胞中的 Ca2+ 信号传导进行原位成像由 Ca2+ 激活的离子通道。将确定动力学并评估 RP 蠕动中的这种电导的后果。初步数据表明，PDGFRα+ 细胞在原位表现出动态 Ca2+ 信号传导，并表达 Ca2+ 激活的 Cl- 通道，Ano1，使其成为起搏器电导的主要候选者。该项目将充当起搏器电导的主要候选者。为未来研究了解影响 RP 功能的发育或病理问题奠定基础。 OMB 编号 0925-0001/0002（修订版 01/18 批准至 03/31/2020）页面延续格式页面