Istituto per l'Endocrinologia e l'Oncologia Sperimentale-Consiglio Nazionale delle Ricerche (IEOS-CNR), Naples, Italy.
Unità di NeuroImmunologia, IRCCS Fondazione Santa Lucia, Rome, Italy.
Methods Mol Biol. 2021;2285:319-328. doi: 10.1007/978-1-0716-1311-5_24.
Metabolomics, lipidomics, and the study of cellular metabolism are gaining increasing interest particularly in the field of immunology, since the activation and effector functions of immune cells are profoundly controlled by changes in cellular metabolic asset. Among the different techniques that can be used for the evaluation of cellular metabolism, the Seahorse Extracellular Flux Analyzer allows the real time measurement of both glycolytic and mitochondrial respiration pathways in cells of interest, through the assessment of extracellular acidification and oxygen consumption rate. Metabolomics, on the other hand, is the high-throughput analysis of metabolites, i.e., the substrates, intermediates, and products of cellular metabolism, starting from biofluids, cells or tissues. The metabolome does not include lipids as their properties are different from water-soluble metabolites and are classified under the lipidome. Lipidomics analysis allows the identification and quantification of lipid species. Metabolomics and lipidomics are currently performed with mass-spectrometry coupled with liquid or gas chromatography (LC-MS or GC-MS) and/or nuclear-magnetic resonance (NMR). Here we describe the protocol for the evaluation of metabolic rate, metabolomics, and lipidomics in T cells, examining the detailed experimental approaches.
代谢组学、脂质组学和细胞代谢研究越来越受到关注，特别是在免疫学领域，因为免疫细胞的激活和效应功能受到细胞代谢资产变化的深刻控制。在可用于评估细胞代谢的不同技术中， Seahorse 细胞外通量分析仪可通过评估细胞外酸化率和耗氧量，实时测量感兴趣细胞中的糖酵解和线粒体呼吸途径。另一方面，代谢组学是对代谢物（即细胞代谢的底物、中间产物和产物）的高通量分析，从生物流体、细胞或组织开始。代谢组不包括脂质，因为它们的性质与水溶性代谢物不同，属于脂质组学范畴。脂质组学分析可用于鉴定和定量脂质种类。代谢组学和脂质组学目前与液质联用（LC-MS）或气质联用（GC-MS）和/或核磁共振（NMR）进行。在这里，我们描述了在 T 细胞中评估代谢率、代谢组学和脂质组学的方案，检查了详细的实验方法。
