An optimized approach and inflation media for obtaining complimentary mass spectrometry-based omics data from human lung tissue.
An optimized approach and inflation media for obtaining complimentary mass spectrometry-based omics data from human lung tissue.
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DOI：
10.3389/fmolb.2022.1022775
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发表时间：
2022
影响因子：
5
通讯作者：
中科院分区：
文献类型：
--
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人类疾病状态在生物分子上是多方面的，并且可以跨越表型状态，因此了解所有层面、跨细胞类型以及微解剖组织区室内部和之间的疾病非常重要。为了获得人肺内分子景观的准确且具有代表性的视图，必须对这种脆弱的组织进行充气和嵌入，以保持分子位置的空间保真度并最大限度地减少分子成像实验的分子降解。在这里，我们评估了琼脂糖膨胀和羧甲基纤维素包埋介质，并确定了用于执行基于体积和空间质谱的组学测量的有效组织制备方案。优化了质谱成像方法，以增加琼脂糖膨胀的肺样本中可注释分子的数量。这种优化的方案允许观察肺组织块中几个气道区域内独特的脂质分布。对这些气道区域进行激光捕获显微切割，然后进行高分辨率蛋白质组分析，使我们能够开始以空间分辨的方式将脂质组与蛋白质组联系起来，我们观察到专门定位于气道区域的高丰度蛋白质。我们还将质谱结果与使用其他两种膨胀/包埋介质保存的肺组织样本进行了比较，但我们发现使用这种保存方法的样本制备步骤存在一些缺陷。总的来说，我们证明了膨胀方法的多功能性，我们可以通过各种不同的实验开始揭示代谢组、脂质组和蛋白质组在人肺中以及跨疾病状态下如何在空间上连接。
Human disease states are biomolecularly multifaceted and can span across phenotypic states, therefore it is important to understand diseases on all levels, across cell types, and within and across microanatomical tissue compartments. To obtain an accurate and representative view of the molecular landscape within human lungs, this fragile tissue must be inflated and embedded to maintain spatial fidelity of the location of molecules and minimize molecular degradation for molecular imaging experiments. Here, we evaluated agarose inflation and carboxymethyl cellulose embedding media and determined effective tissue preparation protocols for performing bulk and spatial mass spectrometry-based omics measurements. Mass spectrometry imaging methods were optimized to boost the number of annotatable molecules in agarose inflated lung samples. This optimized protocol permitted the observation of unique lipid distributions within several airway regions in the lung tissue block. Laser capture microdissection of these airway regions followed by high-resolution proteomic analysis allowed us to begin linking the lipidome with the proteome in a spatially resolved manner, where we observed proteins with high abundance specifically localized to the airway regions. We also compared our mass spectrometry results to lung tissue samples preserved using two other inflation/embedding media, but we identified several pitfalls with the sample preparation steps using this preservation method. Overall, we demonstrated the versatility of the inflation method, and we can start to reveal how the metabolome, lipidome, and proteome are connected spatially in human lungs and across disease states through a variety of different experiments.
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影响因子：
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