Fulminant hepatocyte apoptosis in vivo following microcystin-LR administration to rats

Microcystin-LR (MCLR) is a cyanobacterial toxin responsible for human and livestock deaths worldwide. MCLR has also been implicated as a contributing factor in hepatocellular carcinoma. Following absorption, MCLR is taken up via a hepatocyte-specific bile acid carrier. Inside hepatocytes, MCLR selectively binds to protein phosphatases I and ZA, resulting in rapid, massive Liver damage. However, the apoptotic nature of this toxicosis in rats has not been fully characterized as such at appropriate time points utilizing Light and electron microscopy, terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL), and electrophoresis of hepatic DNA. Rats were administered intraperitoneal saline or MCLR at 500 mu g/kg (0.5 mu mol/kg) and necropsied at 3 or 9 hours. Light microscopy at 3 hours revealed massive, widespread apoptotic necrosis of the majority of hepatocytes. Hepatocytes were rounded and disassociated, with cell shrinkage, increased eosinophilia, and margination of nuclear chromatin or pyknosis. The apoptotic index increased from 0.03% +/- 0.02% in controls to 205% +/- 12% in MCLR-treated animals (p less than or equal to 0.0001). At 3 hours, transmission electron microscopy revealed hepatocellular changes typical of apoptotic necrosis: rounding and disassociation of hepatocytes, toss of microvilli, and margination and condensation of nuclear chromatin. Laddering of hepatic DNA by electrophoresis and widespread TUNEL staining of hepatocytes were consistent with apoptosis. These results demonstrate chat in rats, hepatic damage caused by MCLR is due to extremely rapid induction and progression of apoptosis in virtually every hepatocyte in the liver. This model of fulminant hepatic necrosis should be useful for increased characterization and understanding of the relationship between protein phosphatase inhibition and apoptosis.