title: Metabolomic studies of radiation-induced apoptosis of human leukocytes by capillary electrophoresis-mass spectrometry and flow cytometry: adaptive cellular responses to ionizing radiation.
abstract: There is growing interest in the development of new methods for elucidating the biological effects of low-dose exposure to ionizing radiation (IR) on human health. Herein, we introduce a strategy for assessment of the impact of radiation-induced oxidative stress on the intra-cellular metabolism of human leukocytes. Untargeted metabolomic analyses were performed by CE-ESI-MS on irradiated leukocytes exposed to increasing doses of -radiation emitted from a Taylor source, which were subsequently incubated for 44 h to allow for cellular recovery. Flow cytometry with dual fluorescence staining revealed a major shift from early- to late-stage apoptosis associated with cell membrane permeability changes as radiation dosage was increased relative to the control, but with a significant attenuation measured at intermediate dose levels. CE-ESI-MS analysis of filtered white blood cell lysates was also performed to quantify changes associated with 22 intra-cellular metabolites, which were consistently measured in leukocytes across all radiation levels. Preliminary experiments demonstrated that there was an overall depletion in metabolites with extended exposure to IR; however, there was a non-linear upregulation of specific metabolites at the 4 Gy level relative to pre-irradiated levels, notably for arginine, glutamine, creatine, proline and reduced glutathione. Our studies demonstrate that leukocytes require a minimum threshold level of IR to induce a cytoprotective response in metabolism associated with antioxidant defense, energy homeostasis and cell signaling, which is relevant to improved understanding of the mechanisms of oxidative stress in radiobiology and cancer therapy.
authors.affiliation: Department of Chemistry and Chemical Biology, McMaster University, Hamilton, ON, Canada.
authors.forename: Richard
authors.initials: R
authors.lastname: Lee
authors.forename: Philip
authors.initials: P
authors.lastname: Britz-McKibbin
mesh_terms: D017209:Apoptosis / Q000528:radiation effects*; D002478:Cells, Cultured; D019075:Electrophoresis, Capillary / Q000379:methods; D005434:Flow Cytometry / Q000379:methods; D006801:Humans; D007962:Leukocytes / Q000166:cytology / Q000378:metabolism* / Q000528:radiation effects*; D013058:Mass Spectrometry / Q000379:methods; D055442:Metabolome / Q000528:radiation effects*; D055432:Metabolomics / Q000379:methods*; D018384:Oxidative Stress / Q000528:radiation effects; D011839:Radiation, Ionizing