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Visualization of cytoskeletal dynamics in podocytes using adenoviral vectors

机译：使用腺病毒载体可视化足细胞中的细胞骨架动力学

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摘要

Glomerular visceral epithelial cells (podocytes) play a key role in maintaining selective protein filtration in the kidney. Podocytes have a complex cell shape characterized by the presence of numerous actin-rich processes, which cover the surface of glomerular capillaries and are connected by specialized cell-cell adhesion complexes (slit diaphragms). Human genetic studies and experiments in knockout mouse models show that actin filaments and actin-associated proteins are indispensable for the maintenance of podocyte shape, slit diaphragm integrity, and normal glomerular filtration. The ability to examine cytoskeletal protein organization and dynamics in podocytes and to test the effects of disease-associated mutations on protein localization provides valuable information for researchers aiming to dissect the molecular mechanisms of podocyte dysfunction. We describe how adenovirus-mediated transduction of cultured podocytes with DNA constructs can be used to reliably introduce fluorescently tagged cytoskeletal markers for live cell imaging with high efficiency and low toxicity. This technique can be used to study the dynamic reorganization of the podocyte cytoskeleton and to test the effects of novel mutations on podocyte cytoskeletal dynamics.
机译：肾小球内脏上皮细胞（足细胞）在维持肾脏选择性蛋白质过滤中起关键作用。足细胞具有复杂的细胞形状，其特征是存在许多富含肌动蛋白的过程，该过程覆盖了肾小球毛细血管的表面，并通过专门的细胞-细胞粘附复合物（裂膜）相连。人类基因研究和基因敲除小鼠模型中的实验表明，肌动蛋白丝和肌动蛋白相关蛋白对于维持足细胞形状，裂隙膜完整性和正常肾小球滤过是必不可少的。检查足细胞中细胞骨架蛋白质组织和动力学以及测试疾病相关突变对蛋白质定位的影响的能力为旨在剖析足细胞功能障碍的分子机制的研究人员提供了有价值的信息。我们描述了如何用腺病毒介导的DNA载体培养的足细胞转导来可靠地引入荧光标记的细胞骨架标记物，用于活细胞成像，效率高，毒性低。该技术可用于研究足细胞细胞骨架的动态重组，并测试新突变对足细胞细胞骨架动力学的影响。

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