Radiation dose determines the method for quantification of DNA double strand breaks

Bulat, Tanja; Keta, Otilija; Korićanac, Lela; Žakula, Jelena; Petrović, Ivana; Ristić-Fira, Aleksandra; Todorović, D. M.

doi:10.1590/0001-3765201620140553

Cited by 10 publications

(4 citation statements)

References 24 publications

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“…Histone H2AX phosphorylation (γH2AX) can be triggered by DNA double-strand breaks (DSBs) [21]. When cellular DSBs occur, H2AX is rapidly phosphorylated in the damaged chromatin, and this activity is localized in nuclear foci [22].…”

Section: Decreased Pak2 Causes the Dna Damage In Early Embryosmentioning

confidence: 99%

Pak2 reduction induces a failure of early embryonic development in mice

Zeng

Liu

Yang

et al. 2021

Reprod Biol Endocrinol

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Background The quality of the early embryo is vital to embryonic development and implantation. As a highly conserved serine/threonine kinase, p21-activated kinase 2 (Pak2) participates in diverse biologic processes, especially in cytoskeleton remodeling and cell apoptosis. In mice, Pak2 knock out and endothelial depletion of Pak2 showed embryonic lethality. However, the role of Pak2 in preimplantation embryos remains unelucidated. Methods In the present work, Pak2 was reduced using a specific small interfering RNA in early mouse embryos, validating the unique roles of Pak2 in spindle assembly and DNA repair during mice early embryonic development. We also employed immunoblotting, immunostaining, in vitro fertilization (IVF) and image quantification analyses to test the Pak2 knockdown on the embryonic development progression, spindle assembly, chromosome alignment, oxidative stress, DNA lesions and blastocyst cell apoptosis. Areas in chromatin with γH2AX were detected by immunofluorescence microscopy and serve as a biomarker of DNA damages. Results We found that Pak2 knockdown significantly reduced blastocyst formation of early embryos. In addition, Pak2 reduction led to dramatically increased abnormal spindle assembly and chromosomal aberrations in the embryos. We noted the overproduction of reactive oxygen species (ROS) with Pak2 knockdown in embryos. In response to DNA double strand breaks (DSBs), the histone protein H2AX is specifically phosphorylated at serine139 to generate γH2AX, which is used to quantitative DSBs. In this research, Pak2 knockdown also resulted in the accumulation of phosphorylated γH2AX, indicative of increased embryonic DNA damage. Commensurate with this, a significantly augmented rate of blastocyst cell apoptosis was detected in Pak2-KD embryos compared to their controls. Conclusions Collectively, our data suggest that Pak2 may serve as an important regulator of spindle assembly and DNA repair, and thus participate in the development of early mouse embryos.

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Section: Decreased Pak2 Causes the Dna Damage In Early Embryosmentioning

confidence: 99%

Pak2 reduction induces a failure of early embryonic development in mice

Zeng

Liu

Yang

et al. 2021

Reprod Biol Endocrinol

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“…According to Tanja Bulat’s study, foci was detected using Cellprofiler software, which automatically picks out foci within 6–30 pixels but did not include the foci over 30 pixels. 46 Thus, a high-resolution or confocal microscope could help to capture a more precise image, and proper counting strategies can provide more credible scores of γ-H2AX foci. 47 Irradiation has more than one way to induce DNA damage.…”

Section: Discussionmentioning

confidence: 99%

Moderate Dose Irradiation Induces DNA Damage and Impairments of Barrier and Host Defense in Nasal Epithelial Cells in vitro

Yang¹,

Liu²,

Liu³

et al. 2022

JIR

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Purpose Radiotherapy (RT) is the mainstay treatment for head and neck cancers. However, chronic and recurrent upper respiratory tract infections and inflammation have been commonly reported in patients post-RT. The underlying mechanisms remain poorly understood. Method and Materials We used a well-established model of human nasal epithelial cells (hNECs) that forms a pseudostratified layer in the air-liquid interface (ALI) and exposed it to single or repeated moderate dose γ-irradiation (1Gy). We assessed the DNA damage and evaluated the biological properties of hNECs at different time points post-RT. Further, we explored the host immunity alterations in irradiated hNECs with polyinosinic-polycytidylic acid sodium salt (poly [I:C]) and lipopolysaccharides (LPS). Results IR induced DNA double strand breaks (DSBs) and triggered DNA damage response in hNECs. Repeated IR significantly reduced basal cell proliferation with low expression of p63/KRT5 and Ki67, induced cilia loss and inhibited mucus secretion. In addition, IR decreased ZO-1 expression and caused a significant decline in the transepithelial electrical resistance (TEER). Moreover, hyperreactive response against pathogen invasion and disrupted epithelial host defense can be observed in hNECs exposed to repeated IR. Conclusion Our study suggests that IR induced prolonged structural and functional impairments of hNECs may contribute to patients post-RT with increased risk of developing chronic and recurrent upper respiratory tract infection and inflammation.

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“…However, CQ combined with zinc could prevent NF-κB/P65 translocation to the nucleus and decrease the expression of several proteins encoded by NF-κB dependent genes. IR could induce various forms of DNA damage include breaking the bases and cleavage of the DNA backbone, resulting in DNA single strand breaks (SSBs) and double strand breaks (DSBs) [40]. γH2AX is a highly specific and sensitive molecular marker for monitoring both DSBs initiation and resolution [41].We found that phosphorylation of histone γH2AX was enhanced by combination treatment with CQ combined with zinc and 4 Gy compared with IR alone, suggesting that CQ combined with zinc might be blocking DNA damage-repair pathways induced by IR in CNE-2s cells.…”

Section: Discussionmentioning

confidence: 99%

Radiosensitization of Clioquinol Combined with Zinc in the Nasopharyngeal Cancer Stem-like Cells by Inhibiting Autophagy in Vitro and in Vivo

Yuan¹,

Wu²,

Zeng³

et al. 2020

Int. J. Biol. Sci.

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Loco-regional recurrence of nasopharyngeal carcinoma (NPC) after radiation therapy is one of the main types of treatment failure. This study is aimed to explore the possible causes of inside-field recurrence of NPC patients in order to develop effective treatment methods. Our study indicated that CD44 and autophagy proteins in tumor tissues of patients with recurrent NPC are higher than that of the relapse free patients. The in vitro experiments further confirmed that cancer stem cells (CSCs) were more radioresistant with enhanced autophagy activity. Treatment with clioquinol (CQ) combined with zinc could obviously enhance the radiosensitivity of CNE-2s cells through autophagy inhibition, activation of the caspase system and impairment of DNA damage repair. The in vivo experiments have further consolidated our findings. Our results suggest that CSCs and enhanced autophagy activity may be involved in the inside-field recurrence of NPC, and CQ combined with zinc could be an important therapeutic approach for recurrent NPC.

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Radiation dose determines the method for quantification of DNA double strand breaks

Cited by 10 publications

References 24 publications

Pak2 reduction induces a failure of early embryonic development in mice

Pak2 reduction induces a failure of early embryonic development in mice

Moderate Dose Irradiation Induces DNA Damage and Impairments of Barrier and Host Defense in Nasal Epithelial Cells in vitro

Radiosensitization of Clioquinol Combined with Zinc in the Nasopharyngeal Cancer Stem-like Cells by Inhibiting Autophagy in Vitro and in Vivo

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