Table_1_A methodology to globally assess ectodomain shedding using soluble fractions from the mouse brain.xlsx

Ectodomain shedding (ES) is a fundamental process involving the proteolytic cleavage of membrane-bound proteins, leading to the release of soluble extracellular fragments (shed ectodomains) with potential paracrine and autocrine signaling functions. In the central nervous system (CNS), ES plays pivotal roles in brain development, axonal regulation, synapse formation, and disease pathogenesis, spanning from cancer to Alzheimer’s disease. Recent evidence also suggests its potential involvement in neurodevelopmental conditions like autism and schizophrenia. Past investigations of ES in the CNS have primarily relied on cell culture supernatants or cerebrospinal fluid (CSF) samples, but these methods have limitations, offering limited insights into how ES is modulated in the intact brain parenchyma. In this study, we introduce a methodology for analyzing shed ectodomains globally within rodent brain samples. Through biochemical tissue subcellular separation, mass spectrometry, and bioinformatic analysis, we show that the brain’s soluble fraction sheddome shares significant molecular and functional similarities with in vitro neuronal and CSF sheddomes. This approach provides a promising means of exploring ES dynamics in the CNS, allowing for the evaluation of ES at different developmental stages and pathophysiological states. This methodology has the potential to help us deepen our understanding of ES and its role in CNS function and pathology, offering new insights and opportunities for research in this field.

胞外域脱落（Ectodomain Shedding, ES）是一类基础生物学过程，指通过蛋白水解切割膜结合蛋白，进而释放具有潜在旁分泌与自分泌信号传导功能的可溶性细胞外片段（脱落胞外域）。在中枢神经系统（CNS）中，ES在脑发育、轴突调控、突触形成以及从癌症到阿尔茨海默病等多种疾病的发病机制中发挥关键作用。近期研究还表明，其可能参与孤独症、精神分裂症等神经发育疾病的发生发展。
既往针对中枢神经系统ES的研究主要依赖细胞培养上清液或脑脊液（CSF）样本，但这类方法存在局限性，难以完整揭示完整脑实质内ES的调控机制。本研究提出了一种可全局分析啮齿类动物脑样本中脱落胞外域的研究方法。通过组织生化亚细胞分离、质谱分析与生物信息学分析，我们证实脑内可溶性组分脱落组与体外神经元脱落组及脑脊液脱落组具有显著的分子与功能相似性。该方法为探索中枢神经系统内的ES动态变化提供了可靠途径，可用于评估不同发育阶段与病理生理状态下的ES水平。本研究方法有望帮助我们深化对ES及其在中枢神经系统功能与病理过程中作用的理解，为该领域的研究提供全新视角与机遇。