Novel imaging approach to study podocyte function in vivo

研究体内足细胞功能的新成像方法

• 批准号: 9097687
• 负责人: JANOS PETI-PETERDI
• 金额: $ 37.13万
• 依托单位: UNIVERSITY OF SOUTHERN CALIFORNIA
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-09-15 至 2018-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9097687
• 关键词: Actins; Acute; Address; Albumins; Albuminuria; Amplifiers; Angiotensin II; Animal Model; Applications Grants; Area; Binding; Blood Flow Velocity; Blood capillaries; Blood flow; Calcium; Caliber; Cells; Chronic Kidney Failure; Comorbidity; Cytoskeleton; Development; Diagnostic; Disease; Environment; FDA approved; Focal Segmental Glomerulosclerosis; Foot Process; Functional disorder; Genetic; Health; Histology; Homeostasis; Human; Image; Immunoglobulin G; In Vitro; Injury; Kidney; Kidney Diseases; Knock-out; Label; Maintenance; Mediating; Microscopy; Modeling; Morphology; Mus; Organ; P2Y2 receptor; Pathogenesis; Pathology; Permeability; Phenotype; Physiological; Purinergic P2 Receptors; Purinoceptor; Role; Sclerosis; Signal Transduction; Suramin; Testing; Therapeutic; Tissues; Tubular formation; base; capillary; cytotoxic; extracellular; glomerular filtration; glomerular function; glomerulosclerosis; hemodynamics; in vivo; innovation; microscopic imaging; mortality; mouse model; new therapeutic target; novel; novel therapeutic intervention; paracrine; podocyte; prevent; public health relevance; pyridoxal phosphate-6-azophenyl-2' ,4' -disulfonic acid; renal tubular transport; response; slit diaphragm

DESCRIPTION (provided by applicant): Glomerular dysfunction is a common basis for the development of chronic kidney disease (CKD), a condition with significant comorbidities and mortalities. Recent studies highlighted the role of podocyte actin cytoskeleton and the slit diaphragm in the maintenance of the glomerular filtration barrier (GFB), and the development of albuminuria (AU) and focal segmental glomerulosclerosis (FSGS). The key role of cytosolic calcium ([Ca2+]i) signaling in podocyte function and the above pathologies is established, however our mechanistic understanding of podocyte [Ca2+]i dynamics is limited to a few players (angiotensin II, TRPC5/6). Most P2 purinoceptors including P2Y2 that bind extracellular ATP signal via [Ca2+]i and have been implicated in a variety of (patho)physiological functions in many organs including paracrine cell-to-cell crosstalk, tissue injury and sclerosis. While the important function of the P2Y2 receptor in renal tubular transport and blood flow autoregulation is well established, its role in podocytes/glomerulus is largely unknown. Here we propose to study podocyte [Ca2+]i dynamics in vivo in the intact kidney in health and disease and to characterize the role and therapeutic relevance of a novel [Ca2+]i signaling mechanism in podocytes that is mediated by P2Y2 purinergic receptors. We hypothesize that P2Y2-mediated elevations in [Ca2+]i is a robust and key novel mechanism in primary podocyte injury, and also that its cell-to-cell propagation between podocytes results in amplified focal segmental dysfunction of the GFB and the development of AU and FSGS. The overall theme, to study the role of P2Y2 in podocyte/GFB injury and the effect of P2Y2 blockade on AU and FSGS, will be addressed by applying a novel imaging approach that employs intravital multiphoton microscopy (MPM) combined with new mouse models of fluorescent podocyte labeling and tagging. These studies may potentially change the current view of the pathogenesis of glomerular disorders and may also provide a clinically and immediately available, novel therapeutic approach for glomerular kidney diseases. The specific aims are to: (1) Characterize the role of P2Y2 signaling in podocyte [Ca2+]i dynamics and GFB/glomerular function in response to podocyte injury/disease. (2) Establish the amplifier function of P2Y2 signaling in the propagation of podocyte injury. (3) Test whether P2Y2 blockade can blunt/reduce the development of podocyte injury, AU and FSGS.

描述（由申请方提供）：肾小球功能障碍是慢性肾脏疾病（CKD）发展的常见基础，CKD是一种具有显著合并症和死亡率的疾病。最近的研究强调了足细胞肌动蛋白细胞骨架和狭缝隔膜在维持肾小球滤过屏障（GFB）、蛋白尿（Au）和局灶节段性肾小球硬化（FSGS）的发展中的作用。胞浆钙（[Ca 2 +]i）信号在足细胞功能和上述病理中的关键作用已经确立，但是我们对足细胞[Ca 2 +]i动力学的机制理解仅限于少数参与者（血管紧张素II，TRPC 5/6）。大多数P2嘌呤受体包括P2 Y2，其通过[Ca 2 +]i结合细胞外ATP信号，并且已经涉及许多器官中的多种（病理）生理功能，包括旁分泌细胞间串扰、组织损伤和硬化。虽然P2 Y2受体在肾小管转运和血流自动调节中的重要功能已得到充分证实，但其在足细胞/肾小球中的作用在很大程度上尚不清楚。在这里，我们建议研究足细胞[Ca 2 +]i在健康和疾病的完整肾脏在体内的动力学和表征的作用和治疗相关性的一种新的[Ca 2 +]i信号传导机制，在足细胞是由P2 Y2嘌呤受体介导的。我们假设P2 Y2介导的[Ca 2 +]i升高是原发性足细胞损伤的一种强大且关键的新机制，并且其在足细胞之间的细胞间传播导致GFB的局灶性节段性功能障碍放大以及Au和FSGS的发展。总体主题，研究P2 Y2在足细胞/GFB损伤中的作用以及P2 Y2阻断对Au和FSGS的影响，将通过应用一种新的成像方法来解决，该方法采用活体多光子显微镜（MPM）结合荧光足细胞标记和标记的新小鼠模型。这些研究可能会改变目前对肾小球疾病发病机制的看法，也可能为肾小球肾病提供一种临床上立即可用的新治疗方法。具体目标是：（1）表征P2 Y2信号传导在足细胞[Ca 2 +]i动力学和响应于足细胞损伤/疾病的GFB/肾小球功能中的作用。(2)建立P2 Y2信号在足细胞损伤传播中的放大器功能。(3)测试P2 Y2阻断是否可以减弱/减少足细胞损伤、Au和FSGS的发展。