Implementation of Mass Cytometry as a Tool for Mechanism of Action Studies in Inflammatory Bowel Disease.
Implementation of Mass Cytometry as a Tool for Mechanism of Action Studies in Inflammatory Bowel Disease.
复制标题
实施质谱细胞术作为炎症性肠病作用机制研究的工具。
DOI：
10.1093/ibd/izy214
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发表时间：
2018
影响因子：
4.9
通讯作者：
Rive
中科院分区：
文献类型：
--
作者：
Tyler,ChristopherJ;Pérez-Jeldres,Tamara;Ehinger,Erik;Capaldo,Brian;Karuppuchamy,Thangaraj;Boyer,JoshuaD;Patel,Derek;Dulai,Parambir;Boland,BrigidS;Lannigan,Joanne;Eckmann,Lars;Ernst,PeterB;Sandborn,WilliamJ;Ho,SamuelB;Rive
炎症性肠病(IBD)的新疗法正在开发中，但缺乏组织水平的机械性读数。评估肠道免疫成分的技术可能是新药作用机制(MOA)研究的宝贵工具。质量细胞术能够以前所未有的分辨率分析肠道炎症细胞浸润物和相应的分子指纹。在这里，我们的目标是优化从肠道组织中分离和冷冻保存细胞的方法，以便在MOA研究中潜在地实施质量细胞术。方法我们使用健康人的肠道活组织和外周血，研究了关键技术问题，包括最低组织要求、细胞分离方案和细胞存储。结果二硫苏糖醇和机械分离减少了上皮细胞的污染，在胶原酶消化20分钟后，可以从2~4例结肠或回肠活检组织中分离出足够数量的细胞(6×104±2×104)，从而能够可靠地检测大多数主要免疫细胞亚群。活检组织和抗体标记的单个核细胞可以被冷冻保存，以供以后的处理和获取(存活率为70%；P<；0.05)。结论质量细胞术是一种独特的免疫表型分析工具。这项技术通过识别特定的细胞亚群及其分子特征，有可能促进药物在靶组织中的作用分析。它的广泛实施不仅可能影响IBD的研究，还可能影响炎症细胞在发病机制中发挥作用的其他胃肠道疾病。
BackgroundNovel therapeutics for inflammatory bowel disease (IBD) are under development, yet mechanistic readouts at the tissue level are lacking. Techniques to assess intestinal immune composition could represent a valuable tool for mechanism of action (MOA) studies of novel drugs. Mass cytometry enables analysis of intestinal inflammatory cell infiltrate and corresponding molecular fingerprints with unprecedented resolution. Here, we aimed to optimize the methodology for isolation and cryopreservation of cells from intestinal tissue to allow for the potential implementation of mass cytometry in MOA studies.MethodsWe investigated key technical issues, including minimal tissue requirements, cell isolation protocols, and cell storage, using intestinal biopsies and peripheral blood from healthy individuals. High-dimensional mass cytometry was employed for the analyses of biopsy-derived intestinal cellular subsets.ResultsDithiothreitol and mechanical dissociation decreased epithelial cell contamination and allowed for isolation of adequate cell numbers from 2 to 4 colonic or ileal biopsies (6 × 104±2 × 104) after a 20-minute collagenase digestion, allowing for reliable detection of most major immune cell subsets. Biopsies and antibody-labeled mononuclear cells could be cryopreserved for later processing and acquisition (viability > 70%;P< 0.05).ConclusionsMass cytometry represents a unique tool for deep immunophenotyping intestinal cell composition. This technique has the potential to facilitate analysis of drug actions at the target tissue by identifying specific cellular subsets and their molecular signatures. Its widespread implementation may impact not only IBD research but also other gastrointestinal conditions where inflammatory cells play a role in pathogenesis.