tailieunhanh - Báo cáo khoa học: "The use of caspase inhibitors in pulsed-field gel electrophoresis may improve the estimation of radiation-induced DNA repair and apoptosis"

Tuyển tập các báo cáo nghiên cứu về y học được đăng trên tạp chí y học Radiation Oncology cung cấp cho các bạn kiến thức về ngành y đề tài: The use of caspase inhibitors in pulsed-field gel electrophoresis may improve the estimation of radiation-induced DNA repair and apoptosis. | Balart et al. Radiation Oncology 2011 6 6 http content 6 1 6 RADIATION ONCOLOGY METHODOLOGY Open Access The use of caspase inhibitors in pulsed-field gel electrophoresis may improve the estimation of radiation-induced DNA repair and apoptosis 1 1 1 2 3 1 Josep Balart Gemma Pueyo Lara I de Llobet Marta Baro Xavi Sole Susanna Marin Oriol Casanovas Ricard Mesia4 Gabriel Capella 1 Abstract Background Radiation-induced DNA double-strand break DSB repair can be tested by using pulsed-field gel electrophoresis PFGE in agarose-encapsulated cells. However previous studies have reported that this assay is impaired by the spontaneous DNA breakage in this medium. We investigated the mechanisms of this fragmentation with the principal aim of eliminating it in order to improve the estimation of radiation-induced DNA repair. Methods Samples from cancer cell cultures or xenografted tumours were encapsulated in agarose plugs. The cell plugs were then irradiated incubated to allow them to repair and evaluated by PFGE caspase-3 and histone H2AX activation gH2AX . In addition apoptosis inhibition was evaluated through chemical caspase inhibitors. Results We confirmed that spontaneous DNA fragmentation was associated with the process of encapsulation regardless of whether cells were irradiated or not. This DNA fragmentation was also correlated to apoptosis activation in a fraction of the cells encapsulated in agarose while non-apoptotic cell fraction could rejoin DNA fragments as was measured by gH2AX decrease and PFGE data. We were able to eliminate interference of apoptosis by applying specific caspase inhibitors and improve the estimation of DNA repair and apoptosis itself. Conclusions The estimation of radiation-induced DNA repair by PFGE may be improved by the use of apoptosis inhibitors. The ability to simultaneously determine DNA repair and apoptosis which are involved in cell fate provides new insights for using the PFGE methodology as functional assay. .