Effect of X-Irradiation at Different Stages in the Cell Cycle on Individual Cell–Based Kinetics in an Asynchronous Cell Population

Tsuchida, Eri; Kaida, Atsushi; Pratama, Endrawan; Ikeda, Masa‐Aki; Suzuki, Keiji; Harada, Kiyoshi; Miura, Masahiko

doi:10.1371/journal.pone.0128090

Cited by 17 publications

(19 citation statements)

References 36 publications

(58 reference statements)

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“…This observation indicates that the duration of G2 arrest was strongly dependent on distance from the Ir-192 source, demonstrating that it was primarily influenced by absorption dose. Previous work showed that the duration of G2 arrest is proportional to absorption dose [ 15 , 18 ]; however, in this study, the use of HeLa-Fucci cells allowed us to clearly temporo-spatially visualize this phenomenon, even in the steep dose gradient of an Ir-192 HDR-RALS.…”

Section: Discussionmentioning

confidence: 88%

“…7b ). Because this difference was longer than the normal red phase duration (~8 h) [ 15 ] for cells at 5 mm, the phenomenon described above could be due to cells irradiated in red phase entering M phase faster than those irradiated in green phase. On the other hand, such cases were not clearly observed at 20 mm, because the difference was much smaller than ~8 h. We also analyzed the number of entries into M phase as a function of fluorescence color at irradiation.…”

Section: Resultsmentioning

confidence: 99%

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Temporo-spatial cell-cycle kinetics in HeLa cells irradiated by Ir-192 high dose-rate remote afterloading system (HDR-RALS)

et al. 2016

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BackgroundIntracavitary irradiation plays a pivotal role in definitive radiotherapy for cervical cancer, and the Ir-192 high dose-rate remote afterloading system (HDR-RALS) is often used for this purpose. Under this condition, tumor tissues receive remarkably different absorption doses, with a steep gradient, depending on distance from the radiation source. To obtain temporo-spatial information regarding cell-cycle kinetics in cervical cancer following irradiation by Ir-192 HDR-RALS, we examined HeLa cells expressing the fluorescence ubiquitination-based cell cycle indicator (Fucci), which allowed us to visualize cell-cycle progression.MethodsHeLa-Fucci cells, which emit red and green fluorescence in G1 and S/G2/M phases, respectively, were grown on 35-mm dishes and irradiated by Ir-192 HDR-RALS under normoxic and hypoxic conditions. A 6 French (Fr) catheter was used as an applicator. A radiation dose of 6 Gy was prescribed at hypothetical treatment point A, located 20 mm from the radiation source. Changes in Fucci fluorescence after irradiation were visualized for cells from 5 to 20 mm from the Ir-192 source. Several indices, including first green phase duration after irradiation (FGPD), were measured by analysis of time-lapse images.ResultsCells located 5 to 20 mm from the Ir-192 source became green, reflecting arrest in G2, in a similar manner up to 12 h after irradiation; at more distant positions, however, cells were gradually released from the G2 arrest and became red. This could be explained by the observation that the FGPD was longer for cells closer to the radiation source. Detailed observation revealed that FGPD was significantly longer in cells irradiated in the green phase than in the red phase at positions closer to the Ir-192 source. Unexpectedly, the FGPD was significantly longer after irradiation under hypoxia than normoxia, due in large part to the elongation of FGPD in cells irradiated in the red phase.ConclusionUsing HeLa-Fucci cells, we obtained the first temporo-spatial information about cell-cycle kinetics following irradiation by Ir-192 HDR-RALS. Our findings suggest that the potentially surviving hypoxic cells, especially those arising from positions around point A, exhibit different cell-cycle kinetics from normoxic cells destined to be eradicated.Electronic supplementary materialThe online version of this article (doi:10.1186/s13014-016-0669-8) contains supplementary material, which is available to authorized users.

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Section: Discussionmentioning

confidence: 88%

Section: Resultsmentioning

confidence: 99%

Temporo-spatial cell-cycle kinetics in HeLa cells irradiated by Ir-192 high dose-rate remote afterloading system (HDR-RALS)

et al. 2016

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“…DNA content analysis by flow cytometry revealed that cells were released later from G2 arrest after 6 Gy irradiation than after 2 Gy irradiation; however, when irradiation was combined with KPU-300 treatment, no useful information was obtained to explain the difference ( Fig 7Ba ). The elongation of the green phase represents G2 arrest [ 41 ]; therefore, we concluded that G2 arrest was more strongly induced during KPU-300 treatment following 6 Gy irradiation than following 2 Gy irradiation ( Fig 7Ba ). Consequently, the ratio of mitotic cells released from G2 arrest was significantly higher in cells exposed to 2 Gy than in those exposed to 6 Gy ( Fig 7Bb , Fig 7Bc )( S6 Table ).…”

Section: Resultsmentioning

confidence: 93%

KPU-300, a Novel Benzophenone–Diketopiperazine–Type Anti-Microtubule Agent with a 2-Pyridyl Structure, Is a Potent Radiosensitizer That Synchronizes the Cell Cycle in Early M Phase

et al. 2015

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KPU-300 is a novel colchicine-type anti-microtubule agent derived from plinabulin (NPI-2358). We characterized the effects of KPU-300 on cell cycle kinetics and radiosensitization using HeLa cells expressing the fluorescent ubiquitination-based cell cycle indicator (Fucci). Cells treated with 30 nM KPU-300 for 24 h were efficiently synchronized in M phase and contained clearly detectable abnormal Fucci fluorescence. Two-dimensional flow-cytometric analysis revealed a fraction of cells distinct from the normal Fucci fluorescence pattern. Most of these cells were positive for an M phase marker, the phosphorylated form of histone H3. Cells growing in spheroids responded similarly to the drug, and the inner quiescent fraction also responded after recruitment to the growth fraction. When such drug-treated cells were irradiated in monolayer, a remarkable radiosensitization was observed. To determine whether this radiosensitization was truly due to the synchronization in M phase, we compared the radiosensitivity of cells synchronized by KPU-300 treatment and cells in early M phase isolated by a combined method that took advantage of shake-off and the properties of the Fucci system. Following normalization against the surviving fraction of cells treated with KPU-300 alone, the surviving fractions of cells irradiated in early M phase coincided. Taken together with potential vascular disrupting function in vivo, we propose a novel radiosensitizing strategy using KPU-300.

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“…DSB repair pathways have overlapping roles in DNA repair and are highly dependent on cell cycle phase [ 31 ]. Differential cellular sensitivities to IR have been reported depending on the phase of the cell cycle at the time of irradiation [ 13 ]. Hence, cells are known to be more radioresistant in the early G1 and late S phases and show increased radio sensitivity in the early S phase [ 14 , 15 ].…”

Section: Discussionmentioning

confidence: 99%

“…Ropolo et al indicated that GSC lines display a significantly elongated cell cycle compared to non-stem cells in response to enhanced activities of the DNA damage checkpoint kinases Chk1 and Chk2 [ 12 ]. Several other studies have reported that the sensitivity of cells to IR varied widely depending on the phase of the cell cycle at the time of irradiation, and have also shown that cells in the G2 and M phases were approximately 3 times more sensitive than cells in the S phase [ 13 , 14 , 15 ].…”

Section: Introductionmentioning

confidence: 99%

Cell Cycle Changes after Glioblastoma Stem Cell Irradiation: The Major Role of RAD51

Tachon

Cortes

Guichet

et al. 2018

IJMS

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“Glioma Stem Cells” (GSCs) are known to play a role in glioblastoma (GBM) recurrence. Homologous recombination (HR) defects and cell cycle checkpoint abnormalities can contribute concurrently to the radioresistance of GSCs. DNA repair protein RAD51 homolog 1 (RAD51) is a crucial protein for HR and its inhibition has been shown to sensitize GSCs to irradiation. The aim of this study was to examine the consequences of ionizing radiation (IR) for cell cycle progression in GSCs. In addition, we intended to assess the potential effect of RAD51 inhibition on cell cycle progression. Five radiosensitive GSC lines and five GSC lines that were previously characterized as radioresistant were exposed to 4Gy IR, and cell cycle analysis was done by fluorescence-activated cell sorting (FACS) at 24, 48, 72, and 96 h with or without RAD51 inhibitor. Following 4Gy IR, all GSC lines presented a significant increase in G2 phase at 24 h, which was maintained over 72 h. In the presence of RAD51 inhibitor, radioresistant GSCs showed delayed G2 arrest post-irradiation for up to 48 h. This study demonstrates that all GSCs can promote G2 arrest in response to radiation-induced DNA damage. However, following RAD51 inhibition, the cell cycle checkpoint response differed. This study contributes to the characterization of the radioresistance mechanisms of GSCs, thereby supporting the rationale of targeting RAD51-dependent repair pathways in view of radiosensitizing GSCs.

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Effect of X-Irradiation at Different Stages in the Cell Cycle on Individual Cell–Based Kinetics in an Asynchronous Cell Population

Cited by 17 publications

References 36 publications

Temporo-spatial cell-cycle kinetics in HeLa cells irradiated by Ir-192 high dose-rate remote afterloading system (HDR-RALS)

Temporo-spatial cell-cycle kinetics in HeLa cells irradiated by Ir-192 high dose-rate remote afterloading system (HDR-RALS)

KPU-300, a Novel Benzophenone–Diketopiperazine–Type Anti-Microtubule Agent with a 2-Pyridyl Structure, Is a Potent Radiosensitizer That Synchronizes the Cell Cycle in Early M Phase

Cell Cycle Changes after Glioblastoma Stem Cell Irradiation: The Major Role of RAD51

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