Pacemaker cells and mechanism in the renal pelvis

肾盂起搏细胞及其机制

• 批准号: 10116375
• 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
• 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/10116375
• 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

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

项目摘要 肾骨盆（RP）是一种光滑的肌肉器官，可将尿液从肾脏运输到输尿管 常规的节奏收缩对于尿液运输和膀胱填充至关重要。传播收缩 起源于RP最近端区域，其中专门的细胞称为“非典型”平滑肌细胞 （ASMC）”已被提议用作起搏器细胞。但是，ASMC的识别已是 不精确，因此有关其表型和功能的实验发现是有争议的。歧义 由于缺乏ASMC的特定标记而产生，并阻止了ASC在驾驶中的作用 RP中的蠕动收缩或起搏器活动的机制。较新的技术现在可以识别 由细胞的异质种群组成的组织中的特定细胞类型。这项研究将采用菌株 带有细胞特异性记者和光遗传传感器的小鼠的小鼠，以澄清起搏器和响应细胞如何 RP产生蠕动收缩。我们的初步数据表明，专门的细胞ASMC表达 血小板衍生的生长因子受体 - α（PDGFRα），并使用表达组蛋白2B-的转基因小鼠 EGFP融合蛋白从内源性PDGFRα+启动子驱动，我们可以在 完整的组织或组织的酶促分散体。我们假设PDGFRα+细胞是起搏器 肾脏骨盆中的细胞。将解决以下特定目标：1。检验PDGFRα+的假设 是RP的主要起搏器细胞； 2。研究PDGFRα+细胞中Ca2+信号传导的动力学； 3。 阐明起搏器产生和传播的特定机制。因为他们的专业 PDGFRα+细胞充当空间器细胞，可能具有编码的专业基因表达模式 基本的离子电导和其他信号分子，以促进起搏器活性。这将是 通过分析富含FACS的人群中的基因表达PDGFRα+细胞中分离的。 小鼠报告菌株的近端RP。表达一般编码的Ca2+指示器（GECI）的小鼠 PDGFRα+细胞将用于原位PDGFRα+细胞中的Ca2+信号传导。由Ca2+激活的离子通道 将确定动力学，并评估RP蠕动中该电导的后果。 初步数据表明，PDGFRα+细胞原位暴露了动态Ca2+信号传导，并表达Ca2+激活的 - CL渠道，ANO1，使其成为起搏器电导的主要候选人。这个项目将作为一个 未来研究了解影响RP功能的发展或病理问题的基础。 OMB No. 0925-0001/0002（修订版01/18通过03/31/2020批准）页面延续格式页面