Paradoxical ATP Elevation in Ischemic Penumbra Revealed by Quantitative Imaging Mass Spectrometry

Abstract

Local responses of energy metabolism during brain ischemia are too heterogeneous to decipher redox distribution between anoxic core and adjacent salvageable regions such as penumbra. Imaging mass spectrometry combined by capillary electrophoresis=mass spectrometry providing quantitative metabolomics revealed spatiotemporal changes in adenylates and NADH in a mouse middle-cerebral artery occlusion model. Unlike the core where ATP decreased, the penumbra displayed paradoxical elevation of ATP despite the constrained blood supply. It is noteworthy that the NADH elevation in the ischemic region is clearly demarcated by the ATPdepleting core. Results suggest that metabolism in ischemic penumbra does not respond passively to compromised circulation, but actively compensates energy charges. Antioxid. Redox Signal. 13, 1157-1167. Quantitative Imaging Mass Spectrometry as a Novel Tactics to Decipher Metabolic Dynamics of Brain IschemiaT o develop neuroprotective therapies for cerebrovascular diseases, it is necessary to characterize spatiotemporal changes in energy metabolism occurring at two functionally defined areas of ischemic brain: one is the ischemic core, which is unsalvageable, and another is its adjacent zone termed penumbra, which is salvageable by interventions. Such characterization requires technical breakthrough including simultaneous identification of multiple compounds comprising energy metabolic systems and quantitative analytical methods sensitive enough to detect low levels of metabolites in the heterogeneous regions of ischemic brain. To achieve these requirements, we combined two types of mass spectrometry (MS): matrix-assisted laser desorption ionization (MALDI)=MS and capillary electrophoresis=electrospray ionization (CE=ESI)=MS. Unlike conventional spectroscopic techniques with which chemical profiles are obtained from one selected volume at a time, MALDI=MS has strengths in visualizing multiple metabolites in discrete areas with a single laser ablation (10, 26, 32). However, it still requires further efforts to be supported for quantification. By contrast, CE=ESI=MS excels in quantification of metabolites (15,22,23) because ESI is efficient in transferring molecules from liquid phase to gas phase. Comparison of transcriptional expression profiles with CE=ESI= MS-based metabolomics previously led us to hypothesize the existence of novel metabolic pathways (33) and their regulatory mechanisms (15,22,23). However, it removes spatial distribution of molecules due to tissue homogenization to extract metabolites.Using imaging MS (IMS) combined with CE=ESI=MS, we herein constructed maps of adenine nucleotides whereby abundance of these metabolites was assigned in absolute terms, that is, mmol=g tissue. Such assignment of contents made it possible to directly compare patterns of biochemical derangements in and around the ischemic core at different time points during infarction. Our results suggest that, unlike the core, the penumbra displays paradoxical elevation of ATP despite the constrained b...

Energy Management by Enhanced Glycolysis in G1-phase in Human Colon Cancer Cells In Vitro and In Vivo

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“…Application of metabolomic analyses using capillary electrophoresis/electrospray ionization/mass spectrometry (CE/ESI/MS; refs. 21,22) allows us to determine fluxes of multiple metabolic pathways during cell-cycle progression in vitro (20,23,24). MALDI-IMS analyses of metastatic tumors have shown that the cancer cells in G 1 phase were more active in glycolysis than those in the other phases.…”

Development of an Imaging Mass Spectrometry Technique for Visualizing Localized Cellular Signaling Mediators in Tissues

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