Strong induction of p73 protein in vivo coincides with the onset of apoptosis in rat liver after treatment with the hepatocarcinogen N-nitrosomorpholine (NNM)

Treatment of rats with genotoxic hepatocarcinogens such as N-nitrosomorpholine (NNM) causes severe hepatotoxicity associated with apoptosis of hepatocytes beginning after 12 h. Previously, we reported that after a single administration of high NNM dose p53 protein level increased in liver but not in testis and that the first wave of apoptosis preceded the induction of p53 indicating that apoptosis in liver was driven by a p53-independent pathway. We now show a pronounced upregulation of p73 protein, a p53-related gene product. The increase of p73 a and 0 occurred already 6 h after NNM administration and preceded the onset of apoptosis by 6 h. Very strong p73 signals appeared 20 and 40 h post-treatment and persisted for a few days, whereas p53 was induced only transiently at 20 and 40 h post-treatment. Immunohistochemical analysis revealed that unlike p53, p73 was detected in the nuclei of hepatocytes undergoing apoptosis. Following the upregulation of p73 levels, the products of several genes regulating DNA repair, e.g., GADD-45 and p53R2 and mediating apoptosis such as apoptosis inducing factor (AIF) were rapidly induced, whereas transient elevation of MDM-2 protein was delayed and coincided temporary with activation of p53 protein. Interestingly, NF-KB another transcription factor responding to cellular stress was activated at 20 h after NNM administration and reached a maximum after an additional 20 h. Our data indicate that activated p73 protein may positively affect the induction and execution of apoptosis in response to genotoxic action of NNM. (C) 2003 Wiley-Liss, Inc.