Cleavage of poly(ADP-ribose) transferase during p53-independent apoptosis in rat liver after treatment with N-nitrosomorpholine and cyproterone acetate

Mol Carcinog. 1999 Apr;24(4):263-75. doi: 10.1002/(sici)1098-2744(199904)24:4<263::aid-mc4>3.0.co;2-8.

Abstract

The aim of this work was to study the role of the tumor suppressor p53 and of poly(ADP-ribose) transferase (pADPRT) in the control of hepatocyte apoptosis in two different in vivo models, i.e., during the process of tumor initiation by the genotoxin and cytotoxin N-nitrosomorpholine (NNM) and after withdrawal of the hepatomitogen cyproterone acetate (CPA). Treatment with NNM induces apoptosis followed by necrosis and regenerative DNA synthesis. At the first wave of apoptosis 12 h after NNM application, no p53 expression could be detected by immunohistochemical analysis and immunoblotting. However, 24 h after treatment, numerous p53-positive hepatocyte nuclei were detected, whereas hepatocytes in early and later stages of apoptosis were always negative. Simultaneously with the increased p53 levels, p21 protein was induced. This was accompanied by a block in replicative DNA synthesis, as detected by proliferating-cell nuclear antigen immunostaining. Concomitantly with the increase in apoptosis, dramatic degradation of the nuclear enzyme pADPRT was observed, as evidenced by immunoblotting and activity blotting. The decrease in pADPRT enzymatic activity observed 12 h after treatment coincided with the greatest extent of pADPRT cleavage. One prominent cleavage product was 64 kDa, suggesting that granzyme B was involved in pADPRT degradation. In the second in vivo model we used, i.e., withdrawal of treatment with the hepatomitogen CPA, apoptosis of excessive hepatocytes but no necrosis occurs. Again, no induction of p53 expression could be detected in the liver even at the maximum level of apoptosis, whereas a strong correlation between induction of apoptosis and cleavage of pADPRT to a 64-kDa fragment was observed. These results from whole-animal experiments strongly suggest that the induction of apoptosis in rat liver after genotoxic and cytotoxic damage and during regression of hyperplasia is driven by a p53-independent pathway but is accompanied by cleavage of pADPRT.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Apoptosis / physiology*
  • Carcinogens / toxicity*
  • Cyclin-Dependent Kinase Inhibitor p21
  • Cyclins / biosynthesis
  • Cyclins / genetics
  • Cyproterone Acetate / pharmacology*
  • DNA Replication
  • Female
  • Gene Expression Regulation / drug effects
  • Genes, p53
  • Glycoside Hydrolases / metabolism
  • Granzymes
  • Hyperplasia
  • Liver / drug effects*
  • Liver / pathology
  • Liver Neoplasms, Experimental / chemically induced
  • Liver Neoplasms, Experimental / pathology
  • Male
  • Mitogens / pharmacology*
  • Necrosis
  • Nitrosamines / toxicity*
  • Poly(ADP-ribose) Polymerases / metabolism*
  • Rats
  • Rats, Wistar
  • Recombinant Fusion Proteins / biosynthesis
  • Serine Endopeptidases / physiology
  • Specific Pathogen-Free Organisms
  • Testis / drug effects
  • Testis / pathology
  • Tumor Suppressor Protein p53 / biosynthesis
  • Tumor Suppressor Protein p53 / physiology*

Substances

  • Carcinogens
  • Cdkn1a protein, rat
  • Cyclin-Dependent Kinase Inhibitor p21
  • Cyclins
  • Mitogens
  • Nitrosamines
  • Recombinant Fusion Proteins
  • Tumor Suppressor Protein p53
  • N-nitrosomorpholine
  • Cyproterone Acetate
  • Poly(ADP-ribose) Polymerases
  • Glycoside Hydrolases
  • poly ADP-ribose glycohydrolase
  • Granzymes
  • Gzmb protein, rat
  • Serine Endopeptidases