p16 Gene transfer increases cell killing with abnormal nucleation after ionising radiation in glioma cells.
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.
Hama
S
S
Department of Neurosurgery, Hiroshima University School of Medicine, Kasumi 1-2-3, Minami-ku, Hiroshima 734-8551, Japan. ml-hns@hiroshima-u.ac.jp
Matsuura
S
S
Tauchi
H
H
Yamasaki
F
F
Kajiwara
Y
Y
Arita
K
K
Yoshioka
H
H
Heike
Y
Y
Mandai
K
K
Kurisu
K
K
D000256:Adenoviridae / Q000235:genetics; D017209:Apoptosis / Q000528:radiation effects; D045744:Cell Line, Tumor; D002467:Cell Nucleus / Q000473:pathology / Q000528:radiation effects*; D016521:Electrophoresis, Gel, Pulsed-Field; D018014:Gene Transfer Techniques; D019942:Genes, p16 / Q000528:radiation effects*; D005822:Genetic Vectors; D005910:Glioma / Q000235:genetics* / Q000473:pathology*; D006801:Humans; D020287:In Situ Nick-End Labeling; D011839:Radiation, Ionizing; D014162:Transfection