Project description:Children are considered more sensitive to radiation-induced cancer than adults, yet any differences in genomic alterations associated with age at exposure and their underlying mechanisms remain unclear. We analyzed the mutation of key tumor suppressor genes in T-cell lymphomas arising after weekly irradiation of female B6C3F1 mice with 1.2 Gy X-rays for 4 consecutive weeks starting during infancy (1 week old), adolescence (4 weeks old) or as young adults (8 weeks old). T-cell lymphoma incidence did not show any statistical differences among three irradiated groups; however, the frequency of loss of heterozygosity on chromosomes 11 and 19 showed opposing correlations with age at exposure. Ikaros on chromosome 11 was mutated frequently in the young adultâ??irradiation group (63%; 5/8), frequently incurring multiple independent hits (including deletions and mutations) or suffering a single hit predicted to result in a dominant negative protein (such as those lacking exon 4, an isoform we have designated Ik11, which lacks two of the four zincâ??finger domains important for DNA binding). Pten on chromosome 19 was mutated more frequently in tumors from infant- (67%; 10/15) compared with young adult-irradiated mice (13%; 1/8, P = 0.041). Despite the presence of homozygous Pten mutations, DNA copy number was unchanged in the infant-irradiation group, suggesting duplication of the mutated allele by chromosome mis-segregation or mitotic recombination. Taken together, our findings demonstrate that while deletions on chromosome 11 affecting the Ikaros locus are a prominent feature of young adult irradiation-induced T-cell lymphoma, tumors arising after infant irradiation suffer a second hit in the Pten gene by chromosome mis-segregation or recombination. This is the first report showing age-at-exposure associated radiation-induced genomic alterations in two key tumor suppressor genes linked to T-cell lymphomagenesis. These data are important for considering the risks associated with childhood exposure to radiation. 38 T-cell lymphomas arising after weekly irradiation of female B6C3F1 mice with 1.2 Gy X-rays for 4 consecutive weeks starting at different ages (Infant: 1 week old (15 tumors), Adolescent: 4 weeks old (13 tumors), Young adult: 8 weeks old (10 tumors)) were analyzed.

Project description:Genomic radiation signature illuminates low-dose effects with sharply reflected transcriptome in Ptch1-deficient medulloblastomas. Cancer risks of low-dose radiation are of great concern especially in relation to rapidly increasing medical exposures; however, their accurate assessments cope with many challenges and difficulties, partly due to the inability to distinguish radiation-induced tumors from spontaneous ones. Here, we analyzed the dose-dependent effect of radiation on medulloblastoma development in Ptch1 heterozygous mice on C3B6F1 background. The incidence and latency of medulloblastoma increased and shortened with increasing radiation dose, respectively. Amazingly, radiation contributed to tumorigenesis even at 50 mGy and 100% of mice got medulloblastoma with 1.5 Gy. Loss of heterozygosity (LOH) analysis on a total of 164 tumors indicated that spontaneous tumors showed LOH in broad regions on chromosome 13, including Ptch1 and distally-extending telomeric portion (S-type). In contrast, tumors developed after 3 Gy irradiation exhibited interstitial losses around Ptch1 (R-type). A clear dose-dependent increase in the proportion of R-type tumor at intermediate doses suggested R-type to be a reliable radiation signature. Array-CGH analysis indicated the R-type-specific copy-number reduction around Ptch1 and LOH-type-independent frequent gains of whole chromosome 6. Integrated expression microarray analysis indicated that expression levels of many genes within the altered genomic regions faithfully reflected the genomic copy-number changes. Furthermore, it was also suggested that these expression changes in turn influenced on many other genes, such as Tgfb2 and Tgfb3, on widespread genomic regions. This is the first demonstration that radiation-induced tumors developed after low-dose irradiation can be characterized quite precisely by interstitial deletion of Ptch1 and by associated gene expression profile. Three medulloblastomas were analyzed by array-CGH method.

Project description:Genomic radiation signature illuminates low-dose effects with sharply reflected transcriptome in Ptch1-deficient medulloblastomas. Cancer risks of low-dose radiation are of great concern especially in relation to rapidly increasing medical exposures; however, their accurate assessments cope with many challenges and difficulties, partly due to the inability to distinguish radiation-induced tumors from spontaneous ones. Here, we analyzed the dose-dependent effect of radiation on medulloblastoma development in Ptch1 heterozygous mice on C3B6F1 background. The incidence and latency of medulloblastoma increased and shortened with increasing radiation dose, respectively. Amazingly, radiation contributed to tumorigenesis even at 50 mGy and 100% of mice got medulloblastoma with 1.5 Gy. Loss of heterozygosity (LOH) analysis on a total of 164 tumors indicated that spontaneous tumors showed LOH in broad regions on chromosome 13, including Ptch1 and distally-extending telomeric portion (S-type). In contrast, tumors developed after 3 Gy irradiation exhibited interstitial losses around Ptch1 (R-type). A clear dose-dependent increase in the proportion of R-type tumor at intermediate doses suggested R-type to be a reliable radiation signature. Array-CGH analysis indicated the R-type-specific copy-number reduction around Ptch1 and LOH-type-independent frequent gains of whole chromosome 6. Integrated expression microarray analysis indicated that expression levels of many genes within the altered genomic regions faithfully reflected the genomic copy-number changes. Furthermore, it was also suggested that these expression changes in turn influenced on many other genes, such as Tgfb2 and Tgfb3, on widespread genomic regions. This is the first demonstration that radiation-induced tumors developed after low-dose irradiation can be characterized quite precisely by interstitial deletion of Ptch1 and by associated gene expression profile. Twelve medulloblastomas were analyzed by array-CGH method.

Project description:Genomic radiation signature illuminates low-dose effects with sharply reflected transcriptome in Ptch1-deficient medulloblastomas. Cancer risks of low-dose radiation are of great concern especially in relation to rapidly increasing medical exposures; however, their accurate assessments cope with many challenges and difficulties, partly due to the inability to distinguish radiation-induced tumors from spontaneous ones. Here, we analyzed the dose-dependent effect of radiation on medulloblastoma development in Ptch1 heterozygous mice on C3B6F1 background. The incidence and latency of medulloblastoma increased and shortened with increasing radiation dose, respectively. Amazingly, radiation contributed to tumorigenesis even at 50 mGy and 100% of mice got medulloblastoma with 1.5 Gy. Loss of heterozygosity (LOH) analysis on a total of 164 tumors indicated that spontaneous tumors showed LOH in broad regions on chromosome 13, including Ptch1 and distally-extending telomeric portion (S-type). In contrast, tumors developed after 3 Gy irradiation exhibited interstitial losses around Ptch1 (R-type). A clear dose-dependent increase in the proportion of R-type tumor at intermediate doses suggested R-type to be a reliable radiation signature. Array-CGH analysis indicated the R-type-specific copy-number reduction around Ptch1 and LOH-type-independent frequent gains of whole chromosome 6. Integrated expression microarray analysis indicated that expression levels of many genes within the altered genomic regions faithfully reflected the genomic copy-number changes. Furthermore, it was also suggested that these expression changes in turn influenced on many other genes, such as Tgfb2 and Tgfb3, on widespread genomic regions. This is the first demonstration that radiation-induced tumors developed after low-dose irradiation can be characterized quite precisely by interstitial deletion of Ptch1 and by associated gene expression profile. Gene expression in 3 normal cerebellum tissues and 12 medulloblastomas was measured.

Project description:While modern radiotherapy technologies can precisely deliver higher doses of radiation to tumors; thus, reducing overall radiation exposure to normal tissues, moderate dose and normal tissue toxicity still remains a significant limitation. The present study profiled the global effects on transcript and miR expression in Human Coronary Artery Endothelial Cells (HCAECs) using single-dose irradiation (SD, 10Gy) or multi-fractionated irradiation (MF, 2Gy x 5) regimens. Longitudinal timepoints were collected after a SD or final dose of MF irradiation for analysis using Agilent Human Gene Expression and miRNA microarray platforms. Compared to SD, the exposure to MF resulted in robust transcript and miR expression changes in terms of the number and magnitude. For data analysis, statistically significant mRNAs (2-fold) and miRs (1.5-fold) were processed by Ingenuity Pathway Analysis (IPA) to uncover miRs associated with target transcripts from several cellular pathways post-irradiation. Interestingly, MF radiation induced a cohort of mRNAs and miRs that coordinate the induction of immune response pathway under tight regulation. Additionally, mRNAs and miRs associated with DNA replication, recombination and repair, apoptosis, cardiovascular events and angiogenesis were revealed. Human Coronary Artery Endothelial Cells (HCAECs) were irradiated in a PANTAK high frequency X-ray generator (Precision X-ray Inc., N. Bedford, CT), operated at 300kV and 10MA. The dose rate was 1.6 Gy per min. Cells were plated into T75cm2 flasks (1-1.5 x 10^6 for single dose radiation and 0.6-0.8 x 10^6 for fractionated radiation). After 24h, cells were exposed to a total of 10 Gy radiation administered either as a single-dose radiation (SD), or as multi-fractionated radiation of 2 Gy x 5 (MF). These non-isoeffective doses were selected to simulate clinical hypofractionated and conventionally fractionated radiotherapy regimens. For the MF protocol, cells were exposed to 2 Gy radiation twice a day, at 6h interval. The cells were approximately 90% confluent at the time of harvesting. For both protocols, radiation-induced changes were analyzed at 6h and 24h after a SD and 6h and 24h after the final dose of fractionated irradiation. Separate controls were maintained for SD and MF radiation protocols.

Project description:Although ionizing radiation has been shown to influence gene transcription, little is known about the effects of radiation on gene translational efficiency. To obtain a genome-wide perspective of the effects of radiation on gene translation, microarray analysis was done on polysome-bound RNA isolated from irradiated human brain tumor cells; to allow for a comparison with the effects of radiation on transcription, microarray analysis was also done using total RNA. The number of genes whose translational activity was modified by radiation was f10-fold greater than those whose transcription was affected. The radiation-induced change in a gene’s translational activity was shown to involve the recruitment of existing mRNAs to and away from polysomes. Moreover, the change in a gene’s translational activity after irradiation correlated with changes in the level of its corresponding protein. These data suggest that radiation modifies gene expression primarily at the level of translation. In contrast to transcriptional changes, there was considerable overlap in the genes affected at the translational level among brain tumor cell lines and normal astrocytes. Thus, the radiation-induced translational control of a subset of mRNAs seems to be a fundamental component of cellular radioresponse. (Cancer Res 2006; 66(2): 1052-61) There are 32 samples in total in this study. Each sample has a technical duplicate (16samples x2). Comparison of radiation-induced gene expression profiles generated from microarray analysis of total and polysome-bound RNA isolated from three brain tumor cell lines U87, SF126, SF539 and normal astrocytes. Cells were irradiated at the dose of 7gy and collected 6 hours later for isolation of total cellar RNA or polysome-bound RNA. Gene expression was directly compared between irradiated and unirradiated control cells for total and polysome RNA. The array labeling is two-dye method. Labeling two dye samples are labeled using CY3 dye. All the human reference RNA from Stratagene are labeled using CY5 dye.

Project description:While modern radiotherapy technologies can precisely deliver higher doses of radiation to tumors; thus, reducing overall radiation exposure to normal tissues, moderate dose and normal tissue toxicity still remains a significant limitation. The present study profiled the global effects on transcript and miR expression in Human Coronary Artery Endothelial Cells (HCAECs) using single-dose irradiation (SD, 10Gy) or multi-fractionated irradiation (MF, 2Gy x 5) regimens. Longitudinal timepoints were collected after a SD or final dose of MF irradiation for analysis using Agilent Human Gene Expression and miRNA microarray platforms. Compared to SD, the exposure to MF resulted in robust transcript and miR expression changes in terms of the number and magnitude. For data analysis, statistically significant mRNAs (2-fold) and miRs (1.5-fold) were processed by Ingenuity Pathway Analysis (IPA) to uncover miRs associated with target transcripts from several cellular pathways post-irradiation. Interestingly, MF radiation induced a cohort of mRNAs and miRs that coordinate the induction of immune response pathway under tight regulation. Additionally, mRNAs and miRs associated with DNA replication, recombination and repair, apoptosis, cardiovascular events and angiogenesis were revealed. Human Coronary Artery Endothelial Cells (HCAECs) were irradiated in a PANTAK high frequency X-ray generator (Precision X-ray Inc., N. Bedford, CT), operated at 300kV and 10MA. The dose rate was 1.6 Gy per min. Cells were plated into T75cm2 flasks (1-1.5 x 10^6 for single dose radiation and 0.6-0.8 x 10^6 for fractionated radiation). After 24h, cells were exposed to a total of 10 Gy radiation administered either as a single-dose radiation (SD), or as multi-fractionated radiation of 2 Gy x 5 (MF). These non-isoeffective doses were selected to simulate clinical hypofractionated and conventionally fractionated radiotherapy regimens. For the MF protocol, cells were exposed to 2 Gy radiation twice a day, at 6h interval. The cells were approximately 90% confluent at the time of harvesting. For both protocols, radiation-induced changes were analyzed at 6h and 24h after a SD and 6h and 24h after the final dose of fractionated irradiation. Separate controls were maintained for SD and MF radiation protocols.

Dataset Information

Biallelic and complicated inactivation of Pten in radiation-induced mouse thymic lymphomas

Publications

Complicated biallelic inactivation of Pten in radiation-induced mouse thymic lymphomas.

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