Cancer Res Treat.  2015 Apr;47(2):221-241. 10.4143/crt.2013.159.

Radiation-Induced Autophagy Contributes to Cell Death and Induces Apoptosis Partly in Malignant Glioma Cells

Affiliations
  • 1Specific Organs Cancer Branch, National Cancer Center, Goyang, Korea.
  • 2Department of Neurosurgery, University of Texas, MD Anderson Cancer Center, Houston, TX, USA.
  • 3Department of Microbiology, Ajou University School of Medicine, Suwon, Korea.
  • 4Cancer Cell and Molecular Biology Branch, National Cancer Center, Goyang, Korea.
  • 5Neuro-oncology Clinic, National Cancer Center, Goyang, Korea. nsghs@ncc.re.kr

Abstract

PURPOSE
Radiation-induced autophagy has been shown to play two different roles, in malignant glioma (MG) cells, cytocidal or cytoprotective. However, neither the role of radiation-induced autophagy for cell death nor the existence of autophagy-induced apoptosis, a well-known cell-death pathway after irradiation, has been verified yet.
MATERIALS AND METHODS
We observed both temporal and dose-dependent response patterns of autophagy and apoptosis to radiation in MG cell lines. Additionally, we investigated the role of autophagy in apoptosis through knockdown of autophagy-related proteins.
RESULTS
Autophagic activity measured by staining of acidic vesicle organelles and Western blotting of LC-3 protein increased in proportion to radiation dose from day 1 to 5 after irradiation. Apoptosis measured by annexin-V staining and Western blotting of cleaved poly(ADP-ribose) polymerase demonstrated relatively late appearance 3 days after irradiation that increased for up to 7 days. Blocking of pan-caspase (Z-VAD-FMK) did not affect apoptosis after irradiation, but silencing of Atg5 effectively reduced radiation-induced autophagy, which decreased apoptosis significantly. Inhibition of autophagy in Atg5 knockdown cells was shown to be beneficial for cell survival. Stable transfection of GFP-LC3 cells was observed after irradiation. Annexin-V was localized in cells bearing GFP-LC3 punctuated spots, indicating autophagy in immunofluorescence. Some of these punctuated GFP-LC3 bearing cells formed conglomerated spots and died in final phase.
CONCLUSION
These findings suggest that autophagy appears earlier than apoptosis after irradiation and that a portion of the apoptotic population that appears later is autophagy-dependent. Thus, autophagy is a pathway to cell death after irradiation of MG cells.

Keyword

Apoptosis; Autophagy; Cell death; Glioma; Radiation
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