Precision-cut bladder slices: an enabling technology for urologic research

精密切割膀胱切片：泌尿学研究的支持技术

• 批准号: 10785220
• 负责人: Rosalyn M Adam
• 金额: $ 33.73万
• 依托单位: BOSTON CHILDREN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-09-20 至 2025-08-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10785220
• 关键词: Acceleration; Animal Model; Benign; Biochemical; Biological Assay; Biological Models; Biomimetics; Bladder; Bladder Dysfunction; Bladder Tissue; Calcium Signaling; Cell Communication; Cells; Cholinergic Agonists; Coupled; Cryopreservation; Development; Devices; Disease; Dissociation; Distal; Environment; Evaluation; Extracellular Matrix; Functional disorder; Generations; Genetically Engineered Mouse; Health; Heart; Histologic; Human; Image; In Vitro; Intestines; Laboratories; Lower urinary tract; Lung; Measurement; Microscopy; Molecular; Molecular Analysis; Mus; Muscle Contraction; Muscle function; Neurogenic Bladder; Organ; Organoids; Pharmacotherapy; Phenotype; Physiological; Physiology; Preparation; Proteomics; Recovery; Relaxation; Research; Signal Transduction; Slice; Smooth Muscle; Specimen; Stretching; Structure; System; Technology; Testing; Tissue Slice Technology; Tissues; Traction; Translational Research; Urologic Diseases; Validation; Wild Type Mouse; bioprinting; cell type; chromatin immunoprecipitation; experimental study; fluid flow; human disease; human tissue; insight; novel; novel therapeutic intervention; novel therapeutics; organ on a chip; pharmacologic; preservation; pressure; reconstitution; response; technology development; three dimensional cell culture; tool; translational study; urologic

Mechanistic evaluation of smooth muscle is essential to a rigorous understanding of lower urinary tract function in both health and disease, and to facilitate translational urologic disease research. Existing strategies for smooth muscle evaluation include whole bladder preparations or bladder strips from animal models as well as single cells from human tissues. However, with these approaches the ability to perform integrated analysis of contraction, stiffness and underlying signaling within a single experiment in specimens that capture the native tissue environment is limited. Precision-cut tissue slice technology has been applied to a number of hollow organs yielding novel insights into cell and tissue physiology and pathophysiology. Research from the Krishnan laboratory has demonstrated that precision-cut lung slices (PCLS) can be coupled with tissue traction microscopy (TTM) to enable quantitative assessment of physiologically relevant parameters in distal airways, including tissue contraction and stiffness. In this application we propose to develop the precision-cut bladder slice (PCBS) as a novel platform to interrogate contraction, relaxation and stiffness of the bladder in vitro, while preserving the native cellular microenvironment. The overall objective of this proposal is to demonstrate the utility of PCBS technology for evaluation of bladder contraction and stiffness in healthy murine and healthy and diseased human bladder tissues. We hypothesize that development of precision-cut bladder slices will enable quantitative assessment of smooth muscle function and signaling in vitro while preserving the native cellular environment. We will test this hypothesis with the following Specific Aims: Aim 1: Evaluate precision-cut bladder slices for measurement of smooth muscle contraction. Aim 2: Evaluate precision-cut bladder slices for assessment of bladder wall stiffness. We will use tissue traction microscopy to assess evoked contraction and relaxation in mouse and human bladder tissues. In addition, we will perform biochemical and immunohistochemical analysis to determine the extent to which PCBS retain viability, phenotypic and functional responses, including after cryopreservation. We will also apply a novel tissue stretching device to explore stiffness in PCBS from diseased versus healthy human bladders, enabling us to evaluate smooth muscle activity within the microenvironment of the extracellular matrix. At the end of the project period, we expect to have created a novel platform enabling measurement of physiologically relevant endpoints in bladder tissue, including contraction, relaxation and stiffness, and to have determined the extent to which PCBS can be modulated with pharmacological agents to enable mechanistic studies. Successful completion of these studies is expected to establish a new platform that can facilitate the validation of new therapeutic interventions for bladder dysfunction.

平滑肌的机制评估对于严格理解下尿路在健康和疾病中的功能，以及促进泌尿系统疾病的转化研究至关重要。现有的平滑肌评估策略包括动物模型的全膀胱制剂或膀胱条以及人体组织的单细胞。然而，通过这些方法，在捕获原生组织环境的标本中进行单一实验的收缩、刚度和潜在信号的综合分析的能力是有限的。