Experimental Therapeutics

British Journal of Cancer (2003) 89, 1802–1811. doi:10.1038/sj.bjc.6601299 www.bjcancer.com
Published online 28 October 2003

p16 gene transfer increases cell killing with abnormal nucleation after ionising radiation in glioma cells

S Hama1, S Matsuura2, H Tauchi2, F Yamasaki1, Y Kajiwara1, K Arita1, H Yoshioka1, Y Heike3, K Mandai4 and K Kurisu1

  1. 1Department of Neurosurgery, Hiroshima University School of Medicine, Kasumi 1-2-3, Minami-ku, Hiroshima 734-8551, Japan
  2. 2Department of Radiation Biology, Research Institute for Radiation Biology and Medicine, Hiroshima University, Kasumi 1-2-3, Minami-ku, Hiroshima 734-8553, Japan
  3. 3Division of Clinical Research, National Shikoku Cancer Center Hospital, Horinouchi 13, Matsuyama, Ehime 790-0007, Japan
  4. 4Division of Pathology, National Shikoku Cancer Center Hospital, Horinouchi 13, Matsuyama, Ehime 790-0007, Japan

Correspondence: Dr S Hama, E-mail: ml-hns@hiroshima-u.ac.jp

Received 16 June 2003; Revised 31 July 2003; Accepted 5 August 2003.

Top

Abstract

It is well established that cells synchronised at the G1–S phase are highly radiosensitive. In this study, p16-null human glioma cell lines were induced into G1 cell cycle arrest by adenovirus-mediated p16 gene transfer, and examined for radiation-induced cell killing. Clonogenic analysis and trypan blue extraction test showed that the p16 gene transfer enhanced radiation-induced cell killing in p16-null glioma cell lines. TUNEL assays and pulse-field gel electrophoresis confirmed that the radiation-induced cell killing of p16-transfected cells could be caused by a nonapoptotic mechanism. Gimsa staining demonstrated that irradiation alone or Ax-mock infection plus irradiation results in a slight increase in the frequency of cells with abnormal nucleus, compared to unirradiated uninfected or Ax-mock infected cells. However, Ax-hp16 or Ax-hp21 infection alone modestly increased the frequency of cells with abnormal nucleus (especially bi- and multinucleation), and 4-Gy irradiation of Ax-hp16 or Ax-hp21 infected cells substantially enhanced this frequency. These results suggest that there exists some unknown interaction between radiation and p16 in cytoplasm/membranes, which decreases cytokinesis and promotes abnormal nucleation. Thus, p16 expression prevented radiation-induced apoptosis by promoting abnormal nucleation, thereby leading to another mode of cell death.

Keywords:

p16, ionising radiation, abnormal nucleation, apoptosis, human glioma cell line