Abstract：
Highly specialized cells are fundamental for the proper functioning of complex organs. Variations in cell-type-specific gene expression and protein composition have been linked to a variety of diseases. Investigation of the distinctive molecular makeup of these cells within tissues is therefore critical in biomedical research. Although several technologies have emerged as valuable tools to address this cellular heterogeneity, most workflows lack sufficient in situ resolution and are associated with high costs and extremely long analysis times. Here, we present a combination of experimental and computational approaches that allows a more comprehensive investigation of molecular heterogeneity within tissues than by either shotgun LC-MS/MS or MALDI imaging alone. We applied our pipeline to the mouse brain, which contains a wide variety of cell types that not only perform unique functions but also exhibit varying sensitivities to insults. We explored the distinct neuronal populations within the hippocampus, a brain region crucial for learning and memory that is involved in various neurological disorders. As an example, we identified the groups of proteins distinguishing the neuronal populations of the dentate gyrus (DG) and the cornu ammonis (CA) in the same brain section. Most of the annotated proteins matched the regional enrichment of their transcripts, thereby validating the method. As the method is highly reproducible, the identification of individual masses through the combination of MALDI-IMS and LC-MS/MS methods can be used for the much faster and more precise interpretation of MALDI-IMS measurements only. This greatly speeds up spatial proteomic analyses and allows the detection of local protein variations within the same population of cells. The method\'s general applicability has the potential to be used to investigate different biological conditions and tissues and a much higher throughput than other techniques making it a promising approach for clinical routine applications.
摘要：
高度特化的细胞是复杂器官正常运作的基础。细胞类型特异性基因表达和蛋白质组成的变化与多种疾病有关。因此，研究这些细胞在组织内的独特分子组成在生物医学研究中至关重要。尽管几种技术已经成为解决这种细胞异质性的有价值的工具，大多数工作流程缺乏足够的原位分辨率，并且与高成本和极长的分析时间有关。这里,我们提出了一种实验和计算方法的组合，与单独使用shot弹枪LC-MS/MS或MALDI成像相比，可以更全面地研究组织内的分子异质性。我们把管道应用在老鼠的大脑上，它包含各种各样的细胞类型，不仅执行独特的功能，而且对侮辱表现出不同的敏感性。我们探索了海马体内不同的神经元群体，对学习和记忆至关重要的大脑区域，与各种神经系统疾病有关。作为一个例子,我们确定了在相同的大脑部分区分齿状回（DG）和玉米氨（CA）神经元群体的蛋白质组。大多数带注释的蛋白质与转录本的区域富集相匹配，从而验证该方法。由于该方法重现性高，通过MALDI-IMS和LC-MS/MS方法的组合来识别个体质量仅可用于MALDI-IMS测量的更快，更精确的解释。这大大加快了空间蛋白质组分析，并允许检测相同细胞群内的局部蛋白质变异。该方法的普遍适用性具有用于研究不同生物条件和组织的潜力，并且比其他技术具有更高的吞吐量，使其成为临床常规应用的有希望的方法。
