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Survival analysis of F98 glioma rat cells following minibeam or broad-beam synchrotron radiation therapy.

Gil S, Sarun S, Biete A, Prezado Y, Sabés M - Radiat Oncol (2011)

Bottom Line: A dose escalation study was performed in order to delimit the range of doses where a therapeutic effect could be expected.These results will help in the design and optimization of the forthcoming in vivo studies at the ESRF.In addition, flow cytometric analysis pointed at a larger effectiveness for minibeams, due to the higher proportion of early apoptotic cells.

View Article: PubMed Central - HTML - PubMed

Affiliation: Centre d'Estudis en Biofísica, Faculty of Medicine, Autonomous University of Barcelona, Cerdanyola del Vallès, Spain. silvia.gild@campus.uab.cat

ABSTRACT

Background: In the quest of a curative radiotherapy treatment for gliomas new delivery modes are being explored. At the Biomedical Beamline of the European Synchrotron Radiation Facility (ESRF), a new spatially-fractionated technique, called Minibeam Radiation Therapy (MBRT) is under development. The aim of this work is to compare the effectiveness of MBRT and broad-beam (BB) synchrotron radiation to treat F98 glioma rat cells. A dose escalation study was performed in order to delimit the range of doses where a therapeutic effect could be expected. These results will help in the design and optimization of the forthcoming in vivo studies at the ESRF.

Methods: Two hundred thousand F98 cells were seeded per well in 24-well plates, and incubated for 48 hours before being irradiated with spatially fractionated and seamless synchrotron x-rays at several doses. The percentage of each cell population (alive, early apoptotic and dead cells, where either late apoptotic as necrotic cells are included) was assessed by flow cytometry 48 hours after irradiation, whereas the metabolic activity of surviving cells was analyzed on days 3, 4, and 9 post-irradiation by using QBlue test.

Results: The endpoint (or threshold dose from which an important enhancement in the effectiveness of both radiation treatments is achieved) obtained by flow cytometry could be established just before 12 Gy in the two irradiation schemes, whilst the endpoints assessed by the QBlue reagent, taking into account the cell recovery, were set around 18 Gy in both cases. In addition, flow cytometric analysis pointed at a larger effectiveness for minibeams, due to the higher proportion of early apoptotic cells.

Conclusions: When the valley doses in MBRT equal the dose deposited in the BB scheme, similar cell survival ratio and cell recovery were observed. However, a significant increase in the number of early apoptotic cells were found 48 hours after the minibeam radiation in comparison with the seamless mode.

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Related in: MedlinePlus

(A-D) - Flow cytometry results. Normalized histograms obtained by flow cytometry 48 hours after irradiation at several doses, with MBRT (left) and BB (right). The plots correspond to: (A) the unstained surviving cells, (B) the positive green fluorescence cells, corresponding to those cells undergoing apoptosis in an early stage, (C) double stained cells, with propidium iodide and annexin, indicating as late apoptotic as necrotic dead cells, and (D) the early (in green) and late (in green and red) apoptotic cells, in order to represent altogether the different death processes.
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Figure 2: (A-D) - Flow cytometry results. Normalized histograms obtained by flow cytometry 48 hours after irradiation at several doses, with MBRT (left) and BB (right). The plots correspond to: (A) the unstained surviving cells, (B) the positive green fluorescence cells, corresponding to those cells undergoing apoptosis in an early stage, (C) double stained cells, with propidium iodide and annexin, indicating as late apoptotic as necrotic dead cells, and (D) the early (in green) and late (in green and red) apoptotic cells, in order to represent altogether the different death processes.

Mentions: Normalized data for unstained surviving cells (Table 3), and plotted in histograms in Figure 2A, indicated the same dose-response for both kind of irradiations. Common observations are: i) a significant decrease in survival for cells irradiated up to 12 Gy, and ii) the same percentage of living cells for doses higher than 12 Gy (Table 3). For these data, the endpoint could be established around 12 Gy (by flow cytometry) after both irradiation modes. The same conclusion can be reached by evaluating altogether apoptotic and necrotic cells as a function of dose (Figure 2-D), where a significant increase in cell death is achieved from doses ≥ 12 Gy.


Survival analysis of F98 glioma rat cells following minibeam or broad-beam synchrotron radiation therapy.

Gil S, Sarun S, Biete A, Prezado Y, Sabés M - Radiat Oncol (2011)

(A-D) - Flow cytometry results. Normalized histograms obtained by flow cytometry 48 hours after irradiation at several doses, with MBRT (left) and BB (right). The plots correspond to: (A) the unstained surviving cells, (B) the positive green fluorescence cells, corresponding to those cells undergoing apoptosis in an early stage, (C) double stained cells, with propidium iodide and annexin, indicating as late apoptotic as necrotic dead cells, and (D) the early (in green) and late (in green and red) apoptotic cells, in order to represent altogether the different death processes.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3094367&req=5

Figure 2: (A-D) - Flow cytometry results. Normalized histograms obtained by flow cytometry 48 hours after irradiation at several doses, with MBRT (left) and BB (right). The plots correspond to: (A) the unstained surviving cells, (B) the positive green fluorescence cells, corresponding to those cells undergoing apoptosis in an early stage, (C) double stained cells, with propidium iodide and annexin, indicating as late apoptotic as necrotic dead cells, and (D) the early (in green) and late (in green and red) apoptotic cells, in order to represent altogether the different death processes.
Mentions: Normalized data for unstained surviving cells (Table 3), and plotted in histograms in Figure 2A, indicated the same dose-response for both kind of irradiations. Common observations are: i) a significant decrease in survival for cells irradiated up to 12 Gy, and ii) the same percentage of living cells for doses higher than 12 Gy (Table 3). For these data, the endpoint could be established around 12 Gy (by flow cytometry) after both irradiation modes. The same conclusion can be reached by evaluating altogether apoptotic and necrotic cells as a function of dose (Figure 2-D), where a significant increase in cell death is achieved from doses ≥ 12 Gy.

Bottom Line: A dose escalation study was performed in order to delimit the range of doses where a therapeutic effect could be expected.These results will help in the design and optimization of the forthcoming in vivo studies at the ESRF.In addition, flow cytometric analysis pointed at a larger effectiveness for minibeams, due to the higher proportion of early apoptotic cells.

View Article: PubMed Central - HTML - PubMed

Affiliation: Centre d'Estudis en Biofísica, Faculty of Medicine, Autonomous University of Barcelona, Cerdanyola del Vallès, Spain. silvia.gild@campus.uab.cat

ABSTRACT

Background: In the quest of a curative radiotherapy treatment for gliomas new delivery modes are being explored. At the Biomedical Beamline of the European Synchrotron Radiation Facility (ESRF), a new spatially-fractionated technique, called Minibeam Radiation Therapy (MBRT) is under development. The aim of this work is to compare the effectiveness of MBRT and broad-beam (BB) synchrotron radiation to treat F98 glioma rat cells. A dose escalation study was performed in order to delimit the range of doses where a therapeutic effect could be expected. These results will help in the design and optimization of the forthcoming in vivo studies at the ESRF.

Methods: Two hundred thousand F98 cells were seeded per well in 24-well plates, and incubated for 48 hours before being irradiated with spatially fractionated and seamless synchrotron x-rays at several doses. The percentage of each cell population (alive, early apoptotic and dead cells, where either late apoptotic as necrotic cells are included) was assessed by flow cytometry 48 hours after irradiation, whereas the metabolic activity of surviving cells was analyzed on days 3, 4, and 9 post-irradiation by using QBlue test.

Results: The endpoint (or threshold dose from which an important enhancement in the effectiveness of both radiation treatments is achieved) obtained by flow cytometry could be established just before 12 Gy in the two irradiation schemes, whilst the endpoints assessed by the QBlue reagent, taking into account the cell recovery, were set around 18 Gy in both cases. In addition, flow cytometric analysis pointed at a larger effectiveness for minibeams, due to the higher proportion of early apoptotic cells.

Conclusions: When the valley doses in MBRT equal the dose deposited in the BB scheme, similar cell survival ratio and cell recovery were observed. However, a significant increase in the number of early apoptotic cells were found 48 hours after the minibeam radiation in comparison with the seamless mode.

Show MeSH
Related in: MedlinePlus