Genomic Instability Induced by Low Dose Irradiation

The goal of this project was to determine if genomic instability could be initiated by poorly repaired DNA damage induced by low doses of ionizing radiation leading to a mutator phenotype. Human cells were irradiated, then transfected with an unirradiated reporter gene at various times AFTER exposure. The vector carried an inactive GFP gene that fluoresced when the gene was activated by a delayed mutation. Fluorescent cells were measured in the interval of 50 hours to four days after transfection. The results showed that delayed mutations occurred in these cells after exposure to relatively low doses (0.3-1.0 Gy) of low or high ionizing radiation, as well as after treatment with hyrodgen peroxide (30-100 micromolar). The occurrence was both dose and time dependent, often decreasing at higher doses and later times. No marked difference was observed between the response of mis-match repair-proficient and -deficient cell lines. Although the results were quite reproducible within single experiments, difficulties were observed from experiment to experiment. Different reagents and assays were tested, but no improvement resulted. We concluded that this method is not sufficiently robust or consisent to be useful in the assay of the induction of genomic instability by low doses of radiation, at least in these cell lines under our conditions

Relationships between expression of BCS1L, mitochondrial bioenergetics, and fatigue among patients with prostate cancer

Introduction: Cancer-related fatigue (CRF) is the most debilitating symptom with the greatest adverse side effect on quality of life. The etiology of this symptom is still not understood. The purpose of this study was to examine the relationship between mitochondrial gene expression, mitochondrial oxidative phosphorylation, electron transport chain complex activity, and fatigue in prostate cancer patients undergoing radiotherapy (XRT), compared to patients on active surveillance (AS). Methods: The study used a matched case-control and repeated-measures research design. Fatigue was measured using the revised Piper Fatigue Scale from 52 patients with prostate cancer. Mitochondrial oxidative phosphorylation, electron-transport chain enzymatic activity, and BCS1L gene expression were determined using patients' peripheral mononuclear cells. Data were collected at three time points and analyzed using repeated measures ANOVA. Results: The fatigue score was significantly different over time between patients undergoing XRT and AS (P<0.05). Patients undergoing XRT experienced significantly increased fatigue at day 21 and day 42 of XRT (P<0.01). Downregulated mitochondrial gene (BC1, ubiquinol-cytochrome c reductase, synthesis-like, BCS1L, P<0.05) expression, decreased OXPHOS-complex III oxidation (P<0.05), and reduced activity of complex III were observed over time in patients with XRT. Moreover, increased fatigue was significantly associated with downregulated BCS1L and decreased complex III oxidation in patients undergoing XRT. Conclusion: Our results suggest that BCS1L and complex III in mitochondrial mononuclear cells are potential biomarkers and feasible therapeutic targets for acute XRT-induced fatigue in this clinical population.National Institute of Nursing Research, National Institutes of Health [K01 NR015246]; Oncology Nursing Society Foundation, Pittsburg, PA [RES125833]; Clinical and Translational Science Collaborative Core Utilization Pilot Grant; Center for Mitochondrial Disease, Case Western Reserve UniversityOpen access journalThis item from the UA Faculty Publications collection is made available by the University of Arizona with support from the University of Arizona Libraries. If you have questions, please contact us at [email protected]

High-Density Genomic Scan with 50K SNP Arrays Reveals Existence of Cryptic Chromosomal Lesions and Germ Line Allelic Polymorphism That Might Determine Predisposition to MDS

Abstract Chromosomal damage is a hallmark of MDS. Inability to detect abnormal karyotype in a portion of MDS patients is consistent with the theory that large lesions may represent the extreme of possible DNA damage. Predisposition to MDS may be due to inherited defects; possibly, genes coding for detoxifying enzymes, DNA repair genes and immunogenetic factors could be involved and the risk can be multifactorial. To date studies dealing with complex pathogenesis of MDS have been based on empiric approaches allowing for very limited insights into the possibly complex genetic traits and DNA changes during evolution of the disease. Gene array technology facilitates detailed genomic analysis. We have applied 50K SNP Affymetrix arrays to the analysis of the MDS genome. The goal of our study was to establish the feasibility of this technology to study 2 separate aspects: i) to detect acquired cryptic chromosomal damage, ii) to investigate germ line allelic variants in MDS that could constitute predisposition factors. Our MDS cohort included 22 patients 15 RA/RS, 7 RAEB; 11 with typical cytogenetic abnormalities and 11 normal karyotypes. Using a threshold value 6 -log10(pval) we confirmed previously known changes in 8/11 patients. The remaining 3 showed multiple scattered lesions throughout affected chromosomes; false negatives were likely due to dilution by non-clonal cells. However, using a high density SNP scan, novel previously cryptic lesions contiguous over various chromosomal portions were found in 5/11 patients with normal metaphase cytogenetics and some patients with previously established defects. Chromosomes 5, 7 and 8 showed a higher number of smaller defects consistent with the pathogenesis of MDS; deletions within 7p14, 7p12.2 and 7q21.3 were found in 4 patients without obvious mononosomy 7. Similarly, deletions of various size within 5q12.3–35.5 were seen in 4/15 patients with low-grade MDS. Analysis of such defects within phenotypically defined subsets of patients may reveal possible consensus lesions or commonly affected genes. We also investigated the presence of putatively MDS-specific genotypes. Overrepresentation of otherwise rare SNP may suggest existence of cryptic changes involving adjacent portions of the genome. Globally SNP array allowed for classification of 55953 SNP in 22 patients. 28% were heterozygous, while 36% were homozygous for either variant. Complex analysis of such a large number of genotypes may allow for determination of potential associations between genetic makeup and clinical sub-entities. Based on the allelic frequencies of individual SNP in MDS group and controls, we have identified significantly overrepresented SNPs occurring in either homo- or heterozygous form. For example, FOXL1 SNP occurring at a frequency of 2.4% in homo- and heterozygote form in normals was found in 25% of patients, and 10% of patients were homozygous for a Semaphorin SNP present in only 0.015% of controls. Similarly, PARP variant showed an allelic frequency of 2.7 vs. 14% in controls and patients, respectively. In general, our study show the potential value of high density SNP arrays in precise analysis of clonal genomic lesions and/loss heterozygozity as well as complex genotypic profiles that may potentially contribute to inherited predisposition traits

A Cross-Sectional Study of KLKB1 and PRCP Polymorphisms in Patients with Cardiovascular Disease

Plasma kallikrein formed from prekallikrein (PK) produces bradykinin from kininogens and activates factor XII. Plasma PK is activated by factors XIIa, XIIa or prolylcarboxypeptidase (PRCP). A cross-sectional investigation determined if there is an association of PRCP and KLKB1 polymorphisms with cardiovascular disease. DNA was obtained from 2243 individuals from the PEACE trial. Two PRCP SNPs, rs7104980 and rs2298668 and 2 KLKB1 SNPs, rs3733402 and rs3087505, were genotyped. Logistic regression models were performed for history of diabetes, MI, stroke, angina, angiographic coronary disease, CABG, intermittent claudication, PTCA, and TIA. The PRCP SNP rs7104980 increased the odds of having a history of PTCA by 21% [OR = 1.211; 95% CI = (1.008, 1.454)]; P= 0.041, but was non significant after Bonferroni correction. Alternatively, having the G allele for rs3733402 (KLKB1 gene) decreased the odds of having a history of angiographic coronary disease by 24% [OR = 0.759; 95% CI = (0.622, 0.927)]; P = 0.007 that was statistically significant (p<0.01) after Bonferroni correction for multiple hypothesis testing. When the best-fit model based on the Akaike information criterion (AIC) controlled for age, weight, gender, hypertension, and history of angina. the G allele of KLKB1 rs3733402 that is associated with less plasma kallikrein activity correlated with reduced history of cardiovascular diseas

Reduced expression of NFAT-associated genes in UCB versus adult CD4plus T lymphocytes during primary stimulation

The cellular and molecular mechanisms underlying the blunted allo-responsiveness of umbilical cord blood (UCB) T cells have not been fully elucidated. Protein expression of NFATc2 (nuclear factor of activated T cells c2), a critical transcription factor necessary for up-regulation of multiple cytokines known to amplify T-cell allogeneic responses, is reduced in UCB T cells. Affymetrix oligonucleotide microarrays were used to compare gene expression of primary purified CD4+ UCB T cells to adult peripheral blood CD4+ T cells (AB) at baseline, 6, and 16 hours of primary stimulation. NFAT-regulated genes exhibited lower expression in UCB CD4+ T cells including the following: granulocyte-macrophage colony-stimulating factor (GM-CSF), interferon-{gamma} (IFN-{gamma}), tumor necrosis factor-{alpha} (TNF-{alpha}), interleukin 3 (IL-3), IL-4, IL-5, IL-13, IL-2 receptor {alpha} (IL-2R{alpha}; CD25), CD40L, and macrophage inflammatory protein 1 {alpha} (MIP-1{alpha}). Transcription factors involved in the NFAT pathway including C/EBP{beta}, JunB, and Fosl1 (Fra-1), as well as Th1- and Th2-related transcription factors STAT4 (signal transducers and activators of transcription 4), T-bet, and c-maf showed reduced expression in UCB compared with AB during primary stimulation. Reduced cytokine, chemokine, and receptor expression was also found in UCB. Gene array data were confirmed using RNase protection assays, flow cytometry, and quantitative multiplexed cytokine measurements. Reduced global expression of NFAT-associated genes, as well as cytokines and chemokines, in UCB CD4+ T cells may contribute to the decreased graft-versus-host disease (GVHD) observed after UCB transplantation

Genetic Variation in 15-Hydroxyprostaglandin Dehydrogenase and Colon Cancer Susceptibility

<div><p>Background</p><p>15-Hydroxyprostaglandin dehydrogenase (15-PGDH) is a metabolic antagonist of COX-2, catalyzing the degradation of inflammation mediator prostaglandin E2 (PGE₂) and other prostanoids. Recent studies have established the 15-PGDH gene as a colon cancer suppressor.</p><p>Methods</p><p>We evaluated 15-PDGH as a colon cancer susceptibility locus in a three-stage design. We first genotyped 102 single-nucleotide polymorphisms (SNPs) in the 15-PGDH gene, spanning ∼50 kb up and down-stream of the coding region, in 464 colon cancer cases and 393 population controls. We then genotyped the same SNPs, and also assayed the expression levels of 15-PGDH in colon tissues from 69 independent patients for whom colon tissue and paired germline DNA samples were available. In the final stage 3, we genotyped the 9 most promising SNPs from stages 1 and 2 in an independent sample of 525 cases and 816 controls (stage 3).</p><p>Results</p><p>In the first two stages, three SNPs (rs1365611, rs6844282 and rs2332897) were statistically significant (p<0.05) in combined analysis of association with risk of colon cancer and of association with 15-PGDH expression, after adjustment for multiple testing. For one additional SNP, rs2555639, the T allele showed increased cancer risk and decreased 15-PGDH expression, but just missed statistical significance (p-adjusted = 0.063). In stage 3, rs2555639 alone showed evidence of association with an odds ratio (TT compared to CC) of 1.50 (95% CI = 1.05–2.15, p = 0.026).</p><p>Conclusions</p><p>Our data suggest that the rs2555639 T allele is associated with increased risk of colon cancer, and that carriers of this risk allele exhibit decreased expression of 15-PGDH in the colon.</p></div