Wang et al. BMC Cancer 2010, 10:661 http://www.biomedcentral.com/1471-2407/10/661 apoptotic frequencies in BPA-BNCT groups were higher than those of the irradiated controls. Additionally, when the absorbed doses were identical, both the proliferation and the clonogenicity of glioma cells were suppressed by BNCT to an extent significantly greater than seen after exposure to a neutron beam. Thus, the RBE of BNCT is higher than that of either a neutron beam or g-rays. In the present study, the contribution of a boron- dependent dose to the total BNCT dose was low. Thus, with optimization of the reactor and improvement in the boron-dependent dose, the RBE of BNCT can be further increased. Previous studies have shown that the intrinsic apopto- tic pathway (involving the mitochondria) plays a key role in the regulation of apoptosis in response to irradia- tion [28]. Bcl-2 family proteins interact to permeabilize the mitochondrial membrane, thus playing an important role in irradiation-induced apoptosis [29-31]. Bcl-2 and Bax, the two main members of the Bcl-2 protein family, function as tumor anti-apoptotic and pro-apoptotic fac- tors, respectively [32-34]. Bax can cause release of cyto- chrome c after formation of the hetero-oligomer Bax/ Bak, whereas Bcl-2 can engage with activator proteins or Bax/Bak, thereby sequestering these proteins [28]. We detected expression of Bcl-2 and Bax in all of U87, U251, and SHG44 cells after BNCT treatment in efforts to understand the impact of BNCT on the mitochon- drial pathway of apoptosis. Down-regulation of Bcl-2 and up-regulation of Bax synthesis after BNCT showed that apoptosis induced by BNCT may be mediated through the mitochondrial pathway. A Bcl-2 antisense has been developed as a gene thera- peutic agent to treat cancer, such as: melanoma carci- noma, multiple myeloma, and small cell lung cancer [28]. Elimination of Bcl-2 induced by BNCT indicates that a directed Bcl-2-targeting strategy may be combined with BNCT, thus possibly leading to synergistic reduction of Bcl-2; this possibility deserves further investigation. Our investigation of BPA-BNCT therapy was con- ducted in vitro, and many constraints are involved in attempts to extrapolate in vitro data to the in vivo con- text. During BNCT, the normal brain tissue and tumors are non-specifically irradiated by reactor-generated mixed irradiation beam components, and the patient tolerance to BNCT could be impaired. When it is con- sidered that the RBE of BNCT is higher than that of g- rays, neutron beam, and X-rays, all of which are widely used as treatment options for patients with brain tumors, the absorbed dose from BNCT can be much lower than those strategies. This will limit irradiation damage to normal tissue. The boron level in blood and brain parenchyma also contributes to the irradiation dose to normal tissue. With the development of boron containing agents and the optimization of strategies of Page 7 of 9 delivery, [10B] can be highly accumulated in tumor cells and the level in normal cells becomes relatively lower [2,13,35,36]. Moreover, BNCT relies mainly on the use of boron compounds, and not on a neutron beam per se, to selectively irradiate a tumor [37]. Thus, the opti- mization of selective boron delivery to tumor cells is an important area of BNCT research. As known the BBB can impede the uptake of boron-containing compounds by tumor cells, with the invasive properties of glioma tumors, the BBB can be destroyed and the boron uptake by tumor cells could be increased. Drugs such as manni- tol that transiently compromise BBB integrity have been used to improve boron absorbance by tumor cells. Over- all, the use of BNCT in vivo will likely become effective, with a reduction in side effects and improvements in tumor-targeting boron compounds, following further development. Conclusions Altogether, induction of apoptosis contributes to the main therapeutic efficacy of BNCT to glioma cells in vitro. Activation of Bax and downregulation of Bcl-2 are involvement in the apoptosis triggered by BNCT. More- over, the RBE of BPA-BNCT is higher than that of g-ray and reactor neutron irradiation. Understanding the mechanisms involved in BNCT will ultimately contri- bute to the enhancement of the therapeutic effectiveness of this therapeutic modality. Acknowledgements This research was supported by the National Natural Science Foundation (No: 10375047), Beijing, P.R. China. Author details 1Department of Neurosurgery of Xijing Hospital, Fourth Military Medical University, Xi’an, Shaanxi 710032, P.R.China. 2Northwest Institute of Nuclear Technology, Xi’an, Shaanxi 710024, PR China. 3Department of Biochemistry and Molecular Biology, Fourth Military Medical University, Xi’an, Shaanxi 710032, PR China. Authors’ contributions Peng Wang, Haining Zhen, and Xinbiao Jiang carried out the cell culture and experiment of irradiation. Peng Wang and Xin Cheng carried out Proliferation assays, Hoechst staining, Electron microscopy assay, AnnexinV/PI analysis, Colony formation assay, and drafted the manuscript. Wei Zhang, Geng Guo, and Xinggang Mao participated in the design of the study and performed the statistical analysis. Xiang Zhang conceived of the study, and participated in its design and coordination. All authors read and approved the final manuscript. Competing interests The authors declare that they have no competing interests. Received: 8 February 2010 Accepted: 2 December 2010 Published: 2 December 2010 References 1. Barth RF, Joensuu H: Boron neutron capture therapy for the treatment of glioblastomas and extracranial tumours: as effective, more effective or less effective than photon irradiation? Radiother Oncol 2007, 82:119-122.