Nodularin-triggered apoptosis and hyperphosphorylation of signaling proteins in cultured rat hepatocytes.

The hepatotoxic cyclic peptide nodularin (NOD) is produced by the cyanobacterium Nodularia spumigena in fresh and brackish water lakes, ponds and rivers throughout the world and occurs in variable structures resulting in eight different NODs. NOD has been shown to elicit its hepatotoxicity via inhibition of protein phosphatases (PP) 1 and 2A leading to hyperphosphorylation of vital cellular proteins. This mechanism of action is thought to be associated with the tumour promoting action of NOD in rodent liver. Here, we report on the possible consequences of the inhibition of PP1 and PP2A by NOD to stimulate a dualistic cell response linked to induction of apoptosis and to signals needed for cell proliferation. We determined the concentration-dependent induction of apoptosis and DNA damage after 24 h of NOD treatment in primary rat hepatocytes in culture. NOD significantly increased caspase activities at ⩾100 nM. Furthermore, nuclear apoptosis characterized by condensed, fragmented or crescent-shaped cell nuclei in NOD-treated primary rat hepatocytes after 24 h at NOD concentrations from 10 nM upwards and DNA fragmentation at 100 and 200 nM NOD could be shown. Furthermore, we demonstrated a time- and concentration-dependent early increase in phosphorylated ERK1/2, p90RSK, p85S6K, p70S6K as well as p38 and a late induction of the anti-apoptotic Bcl-xL. Consequently, these in vitro data suggest that NOD is able to cause acute cell death and to activate both signals involved in ambivalent apoptosis/proliferation crossroads and late anti-apoptotic events possibly involved in liver tumor promotion under conditions of sustained exposure.