Progression of subcellular changes during chemical hypoxia to cultured rat hepatocytes: A laser scanning confocal microscopic study |
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| Institution: | 1. College of Environmental Science and Engineering, Hebei University of Science and Technology, Pollution Prevention Biotechnology Laboratory of Hebei Province, Shijiazhuang 050018, China;2. Hebei Key Laboratory of Molecular Chemistry for Drug, Shijiazhuang 050018, China;3. State Key Laboratory of Environment Chemistry and Ecotoxicology, Research Center for Eco-Environment Sciences, Chinese Academy of Sciences, Beijing 100085, China;4. The First Affiliated Hospital of Shandong First Medical University, Jinan 250014, China;5. Shijiazhuang Center for Disease Control and Prevention, Shijiazhuang 050011, China;1. School of Life Science and Technology, Harbin Institute of Technology, Harbin, 150001, China;2. State Key Laboratory of Urban Water Resource and Environment, Harbin Institute of Technology, Harbin, 150006, China;3. Department of Medicine, Renal Electrolyte and Hypertension Division, Department of Genetics, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, 19019, USA;4. Faculty of Education, Wakayama University, Wakayama, 640-8441, Japan;1. School of Environmental Science and Engineering, Tianjin Polytechnic University, Tianjin, 300387, China;2. Department of Civil and Environmental Engineering, Shantou University, Shantou, 515063, China;3. Agro-Environmental Protection Institute, Ministry of Agriculture of China, Tianjin, 300191, China;1. Institute of Environmental Assessment and Water Research (IDAEA-CSIC), Barcelona, Catalonia, Spain;2. Institute for Advanced Chemistry of Catalonia (IQAC), Barcelona, Catalonia, Spain;3. Thermo Fisher Scientific, StradaRivoltana, 20090 Rodano, Milano, Italy;4. Catalan Institute for Water Research (ICRA), Girona, Catalonia, Spain;1. Department of Marine Environmental Engineering, National Kaohsiung University of Science and Technology, Kaohsiung, 81157, Taiwan;2. Department of Biotechnology, National Formosa University, Yunlin, 63208, Taiwan;1. School of Materials Science and Engineering, Nanyang Technological University, 50 Nanyang Avenue, Singapore 639798, Republic of Singapore;2. Institute of Molecular and Cell Biology (IMCB), Agency for Science, Technology and Research (A⁎STAR), Singapore 138667, Republic of Singapore;3. Department of Psychological Medicine, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 119228, Republic of Singapore;4. Genome Institute of Singapore, Agency for Science, Technology and Research (A⁎STAR), 60 Biopolis Street, Singapore 138672, Republic of Singapore;5. Translational Cardiovascular Imaging Group, Institute of Bioengineering and Bioimaging (IBB), Agency for Science, Technology and Research (A⁎STAR), Republic of Singapore;6. Laboratory of In Vivo Genetics & Gene Therapy, Genome Institute of Singapore, Republic of Singapore;7. Department of Environmental Toxicology and Chemistry, Korea Institute of Toxicology, Jinju 52834, South Korea;8. Department of Physiology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117593, Republic of Singapore |
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| Abstract: | The aim of this study was to evaluate changes in the subcellular organelles of cultured hepatocytes by laser scanning confocal microscopy during chemical hypoxia with cyanide and iodoacetate, inhibitors of mitochondrial respiration and glycolysis, respectively. Parameter-specific fluorophores used were calcein for cell topography and membrane permeability, rhodaminedextran for lysosomes, rhodamine 123 and tetramethylrhodamine methylester (TMRM) for mitochondrial membrane potential (ΓΨ) and propidium iodide for loss of cell viability. During the first 30 to 40 minutes of chemical hypoxia to cultured hepatocytes, numerous surface blebs formed and cell volume increased, but ΓΨ decreased relatively little. Subsequently, the nonspecific permeability of mitochondrial membranes increased, and mitochondria depolarized. These events were followed a few minutes later by disintegration of individual lysosomes. After a few more minutes, viability was lost as indicated by bleb rupture, gross plasma membrane permeability to calcein, and nuclear labeling with propidium iodide. Thus, the following sequence of intracellular events occurred during chemical hypoxia: adenosine triphosphate (ATP) depletion, bleb formation with cellular swelling, onset of a mitochondrial permeability transition, disintegration of lysosomes, plasma membrane failure from bleb rupture, and cell death. Any explanation of the pathophysiology of hypoxic injury must take into account this unique sequence of events. |
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