Radiation induced DNA damage and damage repair in human tumor and fibroblast cell lines assessed by histone H2AX phosphorylation

Mahrhofer, Hartmut; Bürger, Susann; Oppitz, Ulrich; Flentje, Michael; Djuzenova, Cholpon S.

doi:10.1016/j.ijrobp.2005.09.037

Cited by 69 publications

(46 citation statements)

References 21 publications

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“…This phosphorylated form of H2AX, termed gH2AX, can be visualized by immunofluorescence analysis and forms discrete nuclear foci, which reflect sites of DSBs. gH2AX-foci analysis is a highly sensitive technique to detect DSBs, and the kinetics of gH2AX-foci loss strongly correlate with the time course of DSB repair (19,20).…”

mentioning

confidence: 99%

DNA Double-Strand Break Repair of Blood Lymphocytes and Normal Tissues Analysed in a Preclinical Mouse Model: Implications for Radiosensitivity Testing

Rübe

Grudzenski²,

Kühne³

et al. 2008

Clinical Cancer Research

144

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Purpose: Radiotherapy is an effective cancer treatment, but a few patients suffer severe radiation toxicities in neighboring normal tissues.There is increasing evidence that the variable susceptibility to radiation toxicities is caused by the individual genetic predisposition, by subtle mutations, or polymorphisms in genes involved in cellular responses to ionizing radiation. Doublestrand breaks (DSB) are the most deleterious form of radiation-induced DNA damage, and DSB repair deficiencies lead to pronounced radiosensitivity. Using a preclinical mouse model, the highly sensitive gH2AX-foci approach was tested to verify even subtle, genetically determined DSB repair deficiencies known to be associated with increased normal tissue radiosensitivity. Experimental Design: By enumerating gH2AX-foci in blood lymphocytes and normal tissues (brain, lung, heart, and intestine), the induction and repair of DSBs after irradiation with therapeutic doses (0.1-2 Gy) was investigated in repair-proficient and repair-deficient mouse strains in vivo and blood samples irradiated ex vivo. Results: gH2AX-foci analysis allowed to verify the different DSB repair deficiencies; even slight impairments caused by single polymorphisms were detected similarly in both blood lymphocytes and solid tissues, indicating that DSB repair measured in lymphocytes is valid for different and complex organs. Moreover, gH2AX-foci analysis of blood samples irradiated ex vivo was found to reflect repair kinetics measured in vivo and, thus, give reliable information about the individual DSB repair capacity. Conclusions: gH2AX analysis of blood and tissue samples allows to detect even minor genetically defined DSB repair deficiencies, affecting normal tissue radiosensitivity. Future studies will have to evaluate the clinical potential to identify patients more susceptible to radiation toxicities before radiotherapy.

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mentioning

confidence: 99%

DNA Double-Strand Break Repair of Blood Lymphocytes and Normal Tissues Analysed in a Preclinical Mouse Model: Implications for Radiosensitivity Testing

Rübe

Grudzenski²,

Kühne³

et al. 2008

Clinical Cancer Research

144

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“…Taneja et al [40] also found a correlation between radiosensitivity and residual amounts of γ-H2AX. By contrast, no correlation between residual foci and radiosensitivity in some tumor cell lines was observed by Mahrhofer et al and Yoshikawa et al [41,42]. A difference in study setup might explain these different findings, as they only studied survival of cells plated immediately after irradiation and correlated this with numbers of foci.…”

Section: Discussionmentioning

confidence: 90%

Decay of γ-H2AX foci correlates with potentially lethal damage repair and P53 status in human colorectal carcinoma cells

Oorschot

Oei

Nuijens

et al. 2014

Cellular and Molecular Biology Letters

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Abbreviations used: BrdUrd -bromodeoxyuridine; Cs -cesium; DAPI -4',6-diamidino-2-phenylindole; dp -delayed plating; DSB -double-strand breaks; FACS -fluorescent activated cell sorter; FCS -fetal bovine calfserum; HEPES -2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid; HPV -human papilloma virus; ip -immediate plating; PBSphosphate buffered saline; PLD -potentially lethal damage; PLDR -potentially lethal damage repair; PVDF -polyvinylidene fluoride; RIPA -radio immunoprecipitation assay; SDS PAGE -sodium dodecyl sulfate polyacrylamide gel electrophoresis; TNBS -PBS containing 0.1% Triton X-100 and 1% FCS Abstract:The influence of p53 status on potentially lethal damage repair (PLDR) and DNA double-strand break (DSB) repair was studied in two isogenic human colorectal carcinoma cell lines: RKO (p53 wild-type) and RC10.1 (p53 null). They were treated with different doses of ionizing radiation, and survival and the induction of DNA-DSB were studied. PLDR was determined by using clonogenic assays and then comparing the survival of cells plated immediately with the survival of cells plated 24 h after irradiation. Doses varied from 0 to 8 Gy. Survival curves were analyzed using the linear-quadratic formula: S(D)/S(0) = exp-(αD+βD 2 ). The γ-H2AX foci assay was used to study DNA DSB kinetics. Cells were irradiated with single doses of 0, 0.5, 1 and 2 Gy. Foci levels were studied in non-irradiated control cells and 30 min and 24 h after irradiation. Irradiation was performed with gamma rays from a 137 Cs source, with a dose rate of 0.5 Gy/min. The RKO cells show higher survival rates after delayed plating than after immediate plating, while no such difference was found for the RC10.1 cells. Functional p53 seems to be a relevant characteristic regarding PLDR for cell survival. Decay of γ-H2AX foci after exposure to ionizing radiation is associated with DSB repair. More residual foci are observed in RC10.1 than in RKO, indicating that decay of γ-H2AX foci correlates with p53 functionality and PLDR in RKO cells.

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“…The discrepancies in these observations arise from the fact that many studies have used cell lines with differing degrees of radiosensitivity and have compared cells with different genetic backgrounds [32]. To elucidate whether DNA repair fidelity is involved in cellular radioresistance, we estimated the mutation frequency and analyzed the mutation spectrum at the HPRT locus in HepG2-8960-R and HepG2 after exposure to AR or FR.…”

Section: Discussionmentioning

confidence: 99%

X-Ray Induced Mutation Frequency at the <i>Hypoxanthine Phosphoribosyltransferase</i> Locus in Clinically Relevant Radioresistant Cells

Kuwahara¹,

Roudkenar²,

Urushihara³

et al. 2017

IJMPCERO

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To elucidate the molecular mechanisms underlying cellular radioresistance, clinically relevant radioresistant cell lines were established via long-term exposure to X-rays with stepwise dose escalation. Established cells continue to proliferate despite exposure to 2 Gy X-rays/day for more than 30 days, a standard protocol in cancer radiotherapy. DNA repair fidelity in radioresistant and the parental cells by evaluating the mutation frequency at the hypoxanthine phosphoribosyltransferase (HPRT) locus after exposure to X-rays was determined. Mutation spectrum at the HPRT locus was examined by multiplex polymerase chain reaction. Rejoining kinetics of X-ray-induced DNA double strand breaks (dsbs) was evaluated by the detection of phosphorylated histone H2AX (γH2AX) after X-irradiation. The fold increase in the HPRT mutation frequency due to acute radiation was similar between radioresistant and the parental cell lines. However, fractionated radiation (FR) consisting of 2 Gy X-rays/day increased the mutation frequency at the HPRT locus in parental but not in radioresistant cells. Analysis of the FR-induced mutations at the HPRT locus revealed a high frequency of deletion mutations (>70%) in parental but not in

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Radiation induced DNA damage and damage repair in human tumor and fibroblast cell lines assessed by histone H2AX phosphorylation

Cited by 69 publications

References 21 publications

DNA Double-Strand Break Repair of Blood Lymphocytes and Normal Tissues Analysed in a Preclinical Mouse Model: Implications for Radiosensitivity Testing

DNA Double-Strand Break Repair of Blood Lymphocytes and Normal Tissues Analysed in a Preclinical Mouse Model: Implications for Radiosensitivity Testing

Decay of γ-H2AX foci correlates with potentially lethal damage repair and P53 status in human colorectal carcinoma cells

X-Ray Induced Mutation Frequency at the <i>Hypoxanthine Phosphoribosyltransferase</i> Locus in Clinically Relevant Radioresistant Cells

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Radiation induced DNA damage and damage repair in human tumor and fibroblast cell lines assessed by histone H2AX phosphorylation

Cited by 69 publications

References 21 publications

DNA Double-Strand Break Repair of Blood Lymphocytes and Normal Tissues Analysed in a Preclinical Mouse Model: Implications for Radiosensitivity Testing

DNA Double-Strand Break Repair of Blood Lymphocytes and Normal Tissues Analysed in a Preclinical Mouse Model: Implications for Radiosensitivity Testing

Decay of γ-H2AX foci correlates with potentially lethal damage repair and P53 status in human colorectal carcinoma cells

X-Ray Induced Mutation Frequency at the &lt;i&gt;Hypoxanthine Phosphoribosyltransferase&lt;/i&gt; Locus in Clinically Relevant Radioresistant Cells

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X-Ray Induced Mutation Frequency at the <i>Hypoxanthine Phosphoribosyltransferase</i> Locus in Clinically Relevant Radioresistant Cells