Progression of subcellular changes during chemical hypoxia to cultured rat hepatocytes: A laser scanning confocal microscopic study

doi:10.1016/0270-9139(95)90058-6

Hepatology

Volume 21, Issue 5, May 1995, Pages 1361-1372

https://doi.org/10.1016/0270-9139(95)90058-6 Get rights and content

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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^☆: Supported in part by grants no. AG07218, DK37034, AG13318, and DK34987 from the National Institutes of Health, Bethesda, MD.

^☆☆: A preliminary report of portions of this work was presented at the 42nd Annual Meeting of the American Association for the Study of Liver Diseases, Chicago, IL, November 2–5, 1991.

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Progression of subcellular changes during chemical hypoxia to cultured rat hepatocytes: A laser scanning confocal microscopic study☆,☆☆

Abstract

Cell Calcium

Biochem Biophys Acta

Biophys J

J Biol Chem

Biophys J

Biochem Biophys Res Commun

J Biol Chem

J Mol Cell Cardiol

Biochem Biophys Acta

J Mol Cell Cardiol

J Biol Chem

Blebbing, free Ca+- and mitochondrial membrane potential preceding cell death in hepatocytes

Nature

Extracellular acidosis delays onset of cell death in ATP-depleted hepatocytes

Am J Physiol

Cell surface changes and enzyme release during hypoxia and reoxygenation in the isolated, perfused rat liver

J Cell Biol

Blebbing, free Ca^+- and mitochondrial membrane potential preceding cell death in hepatocytes