Axial Flow Effects in Proximal Tubule

近端小管的轴流效应

• 批准号: 7900388
• 负责人: Tong Wang Wang
• 金额: $ 38.99万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-05-01 至 2013-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7900388
• 关键词: Actins; Angiotensin II; Animal Model; Apical; Bicarbonates; Blood Pressure; Brush Border; Calcium; Calcium Signaling; Calmodulin; Cells; Chelating Agents; Cilia; Collaborations; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Cytoskeletal Modeling; Cytoskeleton; Data; Dopamine; Equilibrium; F-Actin; Fimbrin; Immunofluorescence Immunologic; In Vitro; Ions; Kidney; Knockout Mice; Length; Liquid substance; MDCK cell; Maintenance; Measures; Mechanics; Mediating; Membrane Transport Proteins; Microtubules; Modeling; Mus; Myosin ATPase; Na(+)-K(+)-Exchanging ATPase; Nephrons; Neurons; PKA inhibitor; Perfusion; Permeability; Physiological; Play; Proton-Translocating ATPases; Regulation; Relative (related person); Role; Second Messenger Systems; Signal Transduction; Sodium; Sodium Chloride; Structure; Sulpiride; Tight Junctions; Torque; Water; Western Blotting; Work; absorption; autocrine; cellular microvillus; data modeling; hormone regulation; inhibitor/antagonist; kidney cell; knockout animal; luminal membrane; mathematical model; public health relevance; research study; response; second messenger; sensor; shear stress; theories; trafficking; villin

DESCRIPTION (provided by applicant): The physiological importance of flow-activated salt and water transport in proximal tubules has been recognized for more than four decades, however the mechanism of this regulation is still not well defined. Recently we have demonstrated that a) Perfusion-absorption balance is present in the isolated perfused proximal tubule of the mouse. b) Both luminal membrane NHE3 and H-ATPase are regulated by flow; c) Changes in tight junction permeabilities do not play a role in flow-modulated transport. We have developed a theory for calculating the forces and torques on the microvilli, and demonstrated that flow-induced changes of proximal tubule absorption are torque dependent, and that an intact actin cytoskeleton is required to transduce the signal to the cell. Experimental data and modeling calculations provide strong evidence that brush border microvilli function as flow sensors in the proximal tubule. However, whether the primary cilium also functions as flow sensor and whether peritubular ion transporters can also be regulated by axial flow has not been examined. In the work proposed, in vitro microperfusion experiments, immunofluorescence, and Western blotting will be conducted with the following three aims: 1) To study the controversy of whether central cilia or microvilli are the flow sensors in proximal tubules by comparing model predictions with flow-induced changes on Na+ and HCO3- absorption in wild-type, and villin, fimbrin, myosins and cilia-deficient mice; 2) To study the role of flow-induced actin cytoskeletal reorganization in modulating transporter trafficking and function in mouse proximal tubules; 3) To examine the role of second messengers, calcium signals, cAMP- and PKA- modulated mechanisms and the role of dopamine in flow-dependent proximal tubule transport. The unique features of our proposed collaboration are: 1) the comparison of flow-dependent proximal tubule transport in intact tubules in mice and in knockout animals; 2) the representation of reabsorptive fluxes as a function of the hydrodynamic forces and torques on microvilli and cilia; and 3) the assessment of flow-induced actin cytoskeletal reorganization, ion transporter localization, and functional changes within a mathematical model of proximal tubule transport. These studies will provide new information on mechanisms of glomerulotubular balance (GTB) and aspects of renal fluid and HCO3- transport in physiological and pathophysiological conditions. PUBLIC HEALTH RELEVANCE: The maintenance of systemic blood pressure depends upon the rate of sodium reabsorption within the kidney, along the entire nephron, and about 2/3 of this occurs in the proximal tubule. The proposed studies will provide direct information on the regulation of proximal tubule sodium transport. Possible benefits include identification of target molecules, which may be blocked or modified in order to modulate sodium reabsorption by the kidney.

描述（由申请人提供）：近端小管中流动激活的盐和水运输的生理重要性已被认识了四十多年，但是这种调节的机制仍然没有明确定义。最近我们证明了 a) 小鼠离体灌注近端小管中存在灌注-吸收平衡。 b) 管腔膜NHE3和H-ATPase均受流量调节； c) 紧密连接渗透性的变化在流量调节运输中不起作用。我们开发了一种计算微绒毛上的力和扭矩的理论，并证明近端小管吸收的流动引起的变化是扭矩依赖性的，并且需要完整的肌动蛋白细胞骨架将信号转导到细胞。实验数据和建模计算提供了强有力的证据，证明刷状缘微绒毛在近端小管中充当流量传感器。然而，初级纤毛是否也起到流量传感器的作用以及管周离子转运蛋白是否也可以通过轴流调节尚未得到研究。在拟议的工作中，将进行体外微灌注实验、免疫荧光和蛋白质印迹，以达到以下三个目的：1）通过将模型预测与流量比较，研究中央纤毛或微绒毛是否是近端小管中的流量传感器的争议。诱导野生型和绒毛、纤维蛋白、肌球蛋白和纤毛缺陷小鼠的 Na+ 和 HCO3- 吸收变化； 2) 研究流诱导的肌动蛋白细胞骨架重组在调节小鼠近曲小管转运蛋白运输和功能中的作用； 3) 检查第二信使、钙信号、cAMP 和 PKA 调节机制的作用以及多巴胺在流量依赖性近端小管转运中的作用。我们提出的合作的独特之处在于：1）比较小鼠和基因敲除动物的完整肾小管中的流量依赖性近端小管运输； 2）重吸收通量作为微绒毛和纤毛上的水动力和扭矩的函数的表示； 3）评估流动诱导的肌动蛋白细胞骨架重组、离子转运蛋白定位以及近端小管转运数学模型内的功能变化。这些研究将提供有关肾小球小管平衡 (GTB) 机制以及生理和病理生理条件下肾液和 HCO3- 转运方面的新信息。公众健康相关性：全身血压的维持取决于肾脏内整个肾单位的钠重吸收率，其中约 2/3 发生在近端肾小管中。拟议的研究将提供有关近端小管钠转运调节的直接信息。可能的好处包括识别目标分子，可以阻断或修饰这些分子以调节肾脏对钠的重吸收。