title: Unique spectral signatures of the nucleic acid dye acridine orange can distinguish cell death by apoptosis and necroptosis.
abstract: Cellular injury and death are ubiquitous features of disease, yet tools to detect them are limited and insensitive to subtle pathological changes. Acridine orange (AO), a nucleic acid dye with unique spectral properties, enables real-time measurement of RNA and DNA as proxies for cell viability during exposure to various noxious stimuli. This tool illuminates spectral signatures unique to various modes of cell death, such as cells undergoing apoptosis versus necrosis/necroptosis. This new approach also shows that cellular RNA decreases during necrotic, necroptotic, and apoptotic cell death caused by demyelinating, ischemic, and traumatic injuries, implying its involvement in a wide spectrum of tissue pathologies. Furthermore, cells with pathologically low levels of cytoplasmic RNA are detected earlier and in higher numbers than with standard markers including TdT-mediated dUTP biotin nick-end labeling and cleaved caspase 3 immunofluorescence. Our technique highlights AO-labeled cytoplasmic RNA as an important early marker of cellular injury and a sensitive indicator of various modes of cell death in a range of experimental models.
authors.affiliation: Department of Clinical Neurosciences and the Hotchkiss Brain Institute, University of Calgary, Calgary, AB T2N 4N1, Canada.
authors.forename: Jason R
authors.identifier: 0000-0003-1385-1464
authors.initials: JR
authors.lastname: Plemel
authors.affiliation: Department of Clinical Neurosciences and the Hotchkiss Brain Institute, University of Calgary, Calgary, AB T2N 4N1, Canada.
authors.forename: Andrew V
authors.initials: AV
authors.lastname: Caprariello
authors.affiliation: Department of Clinical Neurosciences and the Hotchkiss Brain Institute, University of Calgary, Calgary, AB T2N 4N1, Canada.
authors.forename: Michael B
authors.identifier: 0000-0001-5361-3327
authors.initials: MB
authors.lastname: Keough
authors.affiliation: Department of Clinical Neurosciences and the Hotchkiss Brain Institute, University of Calgary, Calgary, AB T2N 4N1, Canada.
authors.forename: Tyler J
authors.initials: TJ
authors.lastname: Henry
authors.affiliation: Department of Clinical Neurosciences and the Hotchkiss Brain Institute, University of Calgary, Calgary, AB T2N 4N1, Canada.
authors.forename: Shigeki
authors.initials: S
authors.lastname: Tsutsui
authors.affiliation: Department of Clinical Neurosciences and the Hotchkiss Brain Institute, University of Calgary, Calgary, AB T2N 4N1, Canada.
authors.forename: Tak H
authors.initials: TH
authors.lastname: Chu
authors.affiliation: Department of Anatomy and Neurosciences, VU University Medical Center, 1081 HV Amsterdam, Netherlands.
authors.forename: Geert J
authors.initials: GJ
authors.lastname: Schenk
authors.affiliation: Department of Anatomy and Neurosciences, VU University Medical Center, 1081 HV Amsterdam, Netherlands.
authors.forename: Roel
authors.identifier: 0000-0001-6506-6420
authors.initials: R
authors.lastname: Klaver
authors.affiliation: Department of Clinical Neurosciences and the Hotchkiss Brain Institute, University of Calgary, Calgary, AB T2N 4N1, Canada.
authors.forename: V Wee
authors.initials: VW
authors.lastname: Yong
authors.affiliation: Department of Clinical Neurosciences and the Hotchkiss Brain Institute, University of Calgary, Calgary, AB T2N 4N1, Canada pstys@ucalgary.ca.
authors.forename: Peter K
authors.identifier: 0000-0003-0326-4194
authors.initials: PK
authors.lastname: Stys
mesh_terms: D000165:Acridine Orange / Q000378:metabolism*; D000818:Animals; D017209:Apoptosis / Q000502:physiology*; D053148:Caspase 3 / Q000378:metabolism; D016923:Cell Death / Q000502:physiology*; D002460:Cell Line; D003856:Deoxyuracil Nucleotides / Q000378:metabolism; D006801:Humans; D020287:In Situ Nick-End Labeling / Q000379:methods; D008810:Mice, Inbred C57BL; D009336:Necrosis / Q000378:metabolism / Q000473:pathology*; D009696:Nucleic Acids / Q000378:metabolism*; D012313:RNA / Q000378:metabolism
chemical_list: D003856:Deoxyuracil Nucleotides; D009696:Nucleic Acids; C027078:deoxyuridine triphosphate; D012313:RNA; D053148:Caspase 3; D000165:Acridine Orange