Poly(ADP-ribose) synthesis: a useful parameter for identifying apoptotic cells

Histochem J. Nov-Dec 1997;29(11-12):831-7. doi: 10.1023/a:1026485622824.

Abstract

Cell death by apoptosis was analysed in HeLa cells either treated with the antitumoral drug bleomycin or depleted of growth factors by long-term culture without medium change. The interference of apoptosis with normal cell cycle progression was followed by flow cytometry in cells stained with propidium iodide and with antibody to S-phase-related PCNA protein. Bleomycin-treated cells showed a net accumulation in G2/M phase paralleled by the appearance of material with a subdiploid DNA content. Cells with a subdiploid DNA content were also present in growth factor-depleted cultures and were shown to derive from all the cell cycle phases. To identify apoptotic features in HeLa cell cultures, we applied a recently developed assay based on the simultaneous analysis in the single cell of three parameters, namely chromatin condensation, DNA degradation and poly(ADP-ribose) synthesis. Apoptotic cells were visualized by sequential reactions: Hoechst staining, terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick-end labelling assay and immunoreaction with anti-poly(ADP-ribose) monoclonal antibody. Positive reactions were obtained for cells at different stages of the apoptotic programme showing condensed nuclei, fragmented chromatin and apoptotic bodies.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Apoptosis / physiology*
  • Bisbenzimidazole
  • Bleomycin / pharmacology
  • Cell Cycle
  • Chromatin / metabolism
  • Culture Media
  • DNA Fragmentation
  • Flow Cytometry
  • HeLa Cells
  • Humans
  • Poly Adenosine Diphosphate Ribose / biosynthesis*
  • Propidium

Substances

  • Chromatin
  • Culture Media
  • Bleomycin
  • Poly Adenosine Diphosphate Ribose
  • Propidium
  • Bisbenzimidazole