Reversible mitochondrial swelling in cultured rat hepatocytes exposed to 1,2-dimethylhydrazine

Abstract

The early structural changes of F344 rat hepatocytes exposed to the hepatocarcinogen 1,2-dimethylhydrazine (DMH) were characterized in short-term monolayer cultures. Continuous exposure of monolayers to DMH (2–16 m M) caused cytoplasmic vacuoles visible by phase-contrast microscopy in all hepatocytes within 6 hr of exposure. These changes preceded maximal release of lactate dehydrogenase (LDH) which occurred after 48 hr of continuous exposure to cytocidal concentrations of DMH (8–16 m M). Ultrastructurally, hepatocytes exposed to DMH (4 m M, 6 hr) showed a twofold increase in mitochondrial diameter from 340 ± 70 nm in control hepatocytes to 800 ± 140 nm in DMH-exposed cells. Hepatocyte monolayers exposed to DMH (4 m M, 6 hr) with subsequent removal of DMH attained normal phase-contrast appearance within 6 hr. Ultrastructural studies showed no significant differences when compared with control hepatocytes and mitochondrial diameters (330 ± 70 nm) were comparable with control hepatocytes. Pretreatment of hepatocytes with depletors of cellular reduced glutathione concentration, including 1,3-bis(2-chloroethyl)-1-nitrosourea (40 μ M) and diethyl maleate (160 μ M), did not potentiate hepatocellular vacuolation nor release of LDH from hepatocytes exposed to DMH (0–16 m M, 48 hr). These studies demonstrate a distinctive form of reversible high-amplitude mitochondrial swelling that can be monitored by phase-contrast microscopy of cultured hepatocytes in monolayers. Since DMH-induced mitochondrial swelling and its progression to irreversible injury are not potentiated by depletors of reduced thiols, this response appears distinct from prelethal mitochondrial swelling in hepatocytes subjected to oxyradical-mediated mechanisms of injury.