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Combined low dose ionizing radiation and green tea-derived epigallocatechin-3-gallate treatment induces human brain endothelial cells death

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Abstract

The microvasculature of brain tumors has been proposed as the primary target for ionizing radiation (IR)-induced apoptosis. However, the contribution of low dose IR-induced non-apoptotic cell death pathways has not been investigated. This study aimed to characterize the effect of IR on human brain microvascular endothelial cells (HBMEC) and to assess the combined effect of epigallocatechin-3-gallate (EGCg), a green tea-derived anti-angiogenic molecule. HBMEC were treated with EGCg, irradiated with a sublethal (≤10 Gy) single dose. Cell survival was assessed 48 h later by nuclear cell counting and Trypan blue exclusion methods. Cell cycle distribution and DNA fragmentation were evaluated by flow cytometry (FC), cell death was assessed by fluorimetric caspase-3 activity, FC and immunoblotting for pro-apoptotic proteins. While low IR doses alone reduced cell survival by 30%, IR treatment was found more effective in EGCg pretreated-cells reaching 70% cell death. Analysis of cell cycle revealed that IR-induced cell accumulation in G2-phase. Expression of cyclin-dependent kinase inhibitors p21(CIP/Waf1) and p27(Kip) were increased by EGCg and IR. Although random DNA fragmentation increased by approximately 40% following combined EGCg/IR treatments, the synergistic reduction of cell survival was not related to increased pro-apoptotic caspase-3, caspase-9 and cytochrome C proteins. Cell necrosis increased 5-fold following combined EGCg/IR treatments while no changes in early or late apoptosis were observed. Our results suggest that the synergistic effects of combined EGCg/IR treatments may be related to necrosis, a non-apoptotic cell death pathway. Strategies sensitizing brain tumor-derived EC to IR may enhance the efficacy of radiotherapy and EGCg may represent such a potential agent.

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Abbreviations

EC:

endothelial cells

EGCg:

epigallocatechin-3-gallate

GBM:

glioblastoma multiforme

HBMEC:

human brain microvascular endothelial cells

HUVEC:

human umbilical vein endothelial cells

IR:

ionizing radiation

VEGF:

vascular endothelial growth factor

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Acknowledgments

B.A. holds a Canada Research Chair in Molecular Oncology from the Canadian Institutes of Health Research. R.B. holds an Institutional UQAM Research Chair in Cancer Prevention and Treatment and the Claude Bertrand Chair of Neurosurgery (CHUM). This research was supported by the NSERC.

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Authors and Affiliations

  1. Laboratoire de Médecine Moléculaire, Centre de Cancérologie Charles-Bruneau, Hôpital Sainte-Justine-UQAM, Succ. centre-ville, C.P. 8888, H3C 3P8, Montréal, Québec, Canada

    Nancy McLaughlin, Marie-Paule Lachambre & Richard Béliveau

  2. Surgery Department, Neurosurgery Service, Centre Hospitalier de l'Université de Montréal, Montreal, Quebec, Canada

    Nancy McLaughlin & Robert Moumdjian

  3. Laboratoire d'Oncologie Moléculaire, Département de Chimie, Centre BioMed, Université du Québec à Montréal, Montreal, Quebec, Canada

    Borhane Annabi

  4. Division of Pediatrics Infectious Diseases, The John Hopkins University School of Medicine, Baltimore, MD, USA

    Kwang Sik Kim

  5. Radio-oncology Department, Centre Hospitalier de l'Université de Montréal, Montreal, Quebec, Canada

    Jean-Paul Bahary

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  1. Nancy McLaughlin
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  2. Borhane Annabi
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  3. Marie-Paule Lachambre
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  4. Kwang Sik Kim
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  5. Jean-Paul Bahary
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  6. Robert Moumdjian
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  7. Richard Béliveau
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Correspondence to Richard Béliveau.

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McLaughlin, N., Annabi, B., Lachambre, MP. et al. Combined low dose ionizing radiation and green tea-derived epigallocatechin-3-gallate treatment induces human brain endothelial cells death. J Neurooncol 80, 111–121 (2006). https://doi.org/10.1007/s11060-006-9171-8

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  • DOI: https://doi.org/10.1007/s11060-006-9171-8

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