A Standardized Method for the Analysis of Liver Sinusoidal Endothelial Cells and Their Fenestrations by Scanning Electron Microscopy
A Standardized Method for the Analysis of Liver Sinusoidal Endothelial Cells and Their Fenestrations by Scanning Electron Microscopy
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DOI：
10.3791/52698
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发表时间：
2015-04-01
影响因子：
1.2
通讯作者：
Le Couteur, David G.
中科院分区：
文献类型：
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作者：
Cogger, Victoria C.;O'Reilly, Jennifer N.;Le Couteur, David G.
肝窦内皮细胞是通往肝脏的门户，其跨细胞的开窗允许小的和溶解的物质从血液不受阻碍地转移到肝实质中进行代谢和加工。开窗是动态结构-它们的大小和/或数量可以响应于各种生理状态、药物和疾病而改变，使它们成为调节的重要靶标。要了解LSEC形态如何受到各种疾病、毒性和生理状态的影响，以及这些变化如何影响肝功能，需要准确测量开窗的大小和数量。在本文中，我们描述了在我们的实验室中使用的扫描电子显微镜固定和处理技术，以确保可重复的标本制备和准确的解释。方法包括灌注固定、二次固定脱水、扫描电镜制备和分析。最后，我们提供了一个一步一步的标准化图像分析方法，这将有利于该领域的所有研究人员。
Liver sinusoidal endothelial cells are the gateway to the liver, their transcellular fenestrations allow the unimpeded transfer of small and dissolved substances from the blood into the liver parenchyma for metabolism and processing. Fenestrations are dynamic structures - both their size and/or number can be altered in response to various physiological states, drugs, and disease, making them an important target for modulation. An understanding of how LSEC morphology is influenced by various disease, toxic, and physiological states and how these changes impact on liver function requires accurate measurement of the size and number of fenestrations. In this paper, we describe scanning electron microscopy fixation and processing techniques used in our laboratory to ensure reproducible specimen preparation and accurate interpretation. The methods include perfusion fixation, secondary fixation and dehydration, preparation for the scanning electron microscope and analysis. Finally, we provide a step by step method for standardized image analysis which will benefit all researchers in the field.