Prolonged Sublethal Exposure to the Protein Phosphatase Inhibitor Microcystin-LR Results in Multiple Dose-Dependent Hepatotoxic Effects

Abstract

The purpose of this study was to relate dose-dependent hepatotoxicity stemming from prolonged exposure to sublethal concentrations of the cyclic heptapeptide microcystin-LR (Mcyst) to hepatic Mcyst concentrations and protein phosphatase activity. Mcyst is a potent inhibitor of protein phosphatase types 1 and 2A (PP1 and PP2A). Twenty male Sprague-Dawley rats were infused continuously with 0, 3, 6, or 9 micrograms Mcyst/day for 28 days using intraperitoneal mini-osmotic pumps containing highly purified toxin or saline. At the end of 28 days, dose-dependent increases in several serum biochemical tests including sorbitol dehydrogenase, aspartate aminotransferase, gamma-glutamyl transferase, alkaline phosphatase, and bile acids had occurred. Serum albumin decreased in a dose-dependent fashion. Liver activity of both PP1 and PP2A decreased in a dose-dependent manner, but with a relatively greater effect on PP2A than PP1. Liver cytosol Mcyst concentrations, measured by direct competitive ELISA, also increased in a dose-dependent manner, although at a higher rate than would be predicted from the incremental increase in dose given. This disproportional increase is suggestive of the bioaccumulation of Mcyst with increasing dose. Histopathological abnormalities included hepatocellular apoptosis and cytosolic vacuolation of principally zone 3 hepatocytes. Immunohistochemical stains revealed Mcyst predominantly within pericanalicular regions of zone 3 hepatocytes. It was concluded that prolonged exposure to sublethal concentrations of Mcyst results in multiple dose-dependent hepatotoxic effects that correspond to decreased hepatic serine/threonine protein phosphatase activity and increasing cytosolic Mcyst concentrations. The disproportional increase of hepatic Mcyst concentrations observed may suggest the bioaccumulation of toxin and an increasing relative risk of hepatotoxicity with increasing dose.

Hepatic Oxidative Stress Following Prolonged Sublethal Microcystin LR Exposure

“…The presence of clear cytoplasmic vacuoles in pericentral areas was reported previously (40). In this study, oilred-O-stained sections demonstrated the presence of lipid within these vacuoles.…”

“…Histologic lesions in the liver of rats exposed to prolonged sublethal doses of MCLR were examined to determine whether they were consistent with those induced by oxidative damage, hepatic concentrations of the lipid peroxidation product, malondialdehyde, were correlated with dose of MCLR given, and liver slices from MCLRexposed rats were tested for susceptibility to the oxidizing agent tert-butyl hydroperoxide. gain was seen in a previous study (40) were treated with 1 mM tert-butyl hydroperoxide for 2 hr.…”

“…In the few studies reported to date, pathologic effects induced by chronic exposure include tumor promotion, hepatocellular degeneration, single-cell necrosis, fibrosis, and neutrophil infiltration (11,13,34). In a recent study, rats exposed to MCLR for 28 days developed dose-dependent apoptosis and hepatocellular vacuolization of zone 3 hepatocytes and increased serum levels of sorbitol dehydrogenase, aspartate aminotransferase (AST), bile acids, alkaline phosphatase, and gamma-glutamyl transferase (40). Although changes were correlated with the degree of protein phosphatase inhibition, the exact mechanism of chronic toxicity has yet to be fully determined.…”

Intraperitoneal injection of extracted microcystins results in hypovolemia and hypotension in crucian carp (Carassius auratus)

“…analogous to those in mammals Li et al, 2005;Solter et al, 1998;Tencalla and Dietrich, 1997;Råbergh et al, 1991). Li et al (2005) reported that intrahepatic hemorrhage occurred in bighead carp (Aristichthys nobilis) injected intraperitoneally with crude MC extracts.…”

“…Problems related to toxigenic cyanobacteria seem to be of increasing importance due to their potential risk to animal and human health, and in a recent study by Chen et al (2009), microcystins were identified for the first time in the serum of a chronically exposed human population (fishermen at Lake Chaohu, China) together with indication of hepatocellular damage. Extracts of toxic cyanobacteria given orally, intraperitoneally, or intravenously in acutely and subacutely toxic doses may cause hepatotoxicity or even death of fish and mammals (Ernst et al, 2006;Falconer et al, 1988;LeClaire et al, 1995;Li et al, 2005;Lovell, 1989;Miura et al, 1989;Solter et al, 1998).…”