• Exposure to benzo[a]pyrene of Hepatic Cytochrome P450 Reductase Null (HRN) and P450 Reductase Conditional Null (RCN) mice: Detection of benzo[a]pyrene diol epoxide-DNA adducts by immunohistochemistry and 32P-postlabelling.

      Arlt, Volker M.; Poirier, Miriam C.; Sykes, Sarah E.; John, Kaarthik; Moserova, Michaela; Stiborova, Marie; Wolf, C. Roland; Henderson, Colin J.; Phillips, David H. (2012-09-03)
      Benzo[a]pyrene (BaP) is a widespread environmental carcinogen activated by cytochrome P450 (P450) enzymes. In Hepatic P450 Reductase Null (HRN) and Reductase Conditional Null (RCN) mice, P450 oxidoreductase (Por) is deleted specifically in hepatocytes, resulting in the loss of essentially all hepatic P450 function. Treatment of HRN mice with a single i.p. or oral dose of BaP (12.5 or 125mg/kg body weight) resulted in higher DNA adduct levels in liver (up to 10-fold) than in wild-type (WT) mice, indicating that hepatic P450s appear to be more important for BaP detoxification in vivo. Similar results were obtained in RCN mice. We tested whether differences between hepatocytes and non-hepatocytes in P450 activity may underlie the increased liver BaP-DNA binding in HRN mice. Cellular localisation by immunohistochemistry of BaP-DNA adducts showed that HRN mice have ample capacity for formation of BaP-DNA adducts in liver, indicating that the metabolic process does not result in the generation of a reactive species different from that formed in WT mice. However, increased protein expression of cytochrome b(5) in hepatic microsomes of HRN relative to WT mice suggests that cytochrome b(5) may modulate the P450-mediated bioactivation of BaP in HRN mice, partially substituting the function of Por.
    • Detection of acetaldehyde derived N(2)-ethyl-2'-deoxyguanosine in human leukocyte DNA following alcohol consumption.

      Singh, Rajinder; Gromadzinska, Jolanta; Mistry, Yogita; Cordell, Rebecca; Juren, Tina; Segerback, Dan; Farmer, Peter B. (2012-09-01)
      Epidemiological studies have shown an association between alcohol (ethanol) consumption and increased cancer risk. The effect of alcohol consumption on the levels and persistence of N(2)-ethylidene-2'-deoxyguanosine (N(2)-ethylidene-dG) formed by acetaldehyde, the oxidative metabolite of ethanol, in human leukocyte DNA was investigated. DNA was isolated from venous blood samples obtained from 30 male non-smoking individuals before consumption of alcohol (0h) and subsequently at 3-5h following the consumption of 150mL of vodka (containing 42% pure ethanol). Additional samples were collected 24h and 48h post-alcohol consumption. The levels of N(2)-ethyl-2'-deoxyguanosine (N(2)-ethyl-dG) in the DNA were determined following reduction of N(2)-ethylidene-dG with sodium cyanoborohydride using a liquid chromatography-tandem mass spectrometry selected reaction monitoring method. A slight time-dependent trend showing an increase and decrease in the levels of N(2)-ethyl-dG was observed following consumption of alcohol compared to time 0h, however, the differences were not statistically significant. The average levels of N(2)-ethyl-dG observed at 0h, 3-5h, 24h and 48h time points following ingestion of alcohol were 34.6±21.9, 35.1±21.0, 36.8±20.7 and 35.6±21.1 per 10(8) 2'-deoxynucleosides, respectively. In conclusion, alcohol consumption that could be encountered under social drinking conditions, does not significantly alter the levels of the acetaldehyde derived DNA adduct, N(2)-ethyl-dG in human leukocyte DNA from healthy individuals.
    • STrengthening the Reporting of OBservational studies in Epidemiology: Molecular Epidemiology STROBE-ME. An extension of the STROBE statement.

      Gallo, Valentina; Egger, Matthias; McCormack, Valerie; Farmer, Peter B.; Ioannidis, John P. A.; Kirsch-Volders, Micheline; Matullo, Giuseppe; Phillips, David H.; Schoket, Bernadette; Stromberg, Ulf; et al. (2012-09)
      Advances in laboratory techniques have led to a rapidly increasing use of biomarkers in epidemiological studies. Biomarkers of internal dose, early biological change, susceptibility, and clinical outcomes are used as proxies for investigating the interactions between external and/or endogenous agents and the body components or processes. The need for improved reporting of scientific research led to influential statements of recommendations such as STrengthening Reporting of Observational studies in Epidemiology (STROBE) statement. The STROBE initiative established in 2004 aimed to provide guidance on how to report observational research. Its guidelines provide a user-friendly checklist of 22 items to be reported in epidemiological studies, with items specific to the three main study designs: cohort studies, case-control studies and cross-sectional studies. The present STrengthening the Reporting of OBservational studies in Epidemiology - Molecular Epidemiology (STROBE-ME) initiative builds on the STROBE Statement implementing 9 existing items of STROBE and providing 17 additional items to the 22 items of STROBE checklist. The additions relate to the use of biomarkers in epidemiological studies, concerning collection, handling and storage of biological samples; laboratory methods, validity and reliability of biomarkers; specificities of study design; and ethical considerations. The STROBE-ME recommendations are intended to complement the STROBE recommendations.
    • Inter-laboratory variation in DNA damage using a standard comet assay protocol.

      Forchhammer, Lykke; Ersson, Clara; Loft, Steffen; Moller, Lennart; Godschalk, Roger W. L.; van Schooten, Frederik J.; Jones, George D. D.; Higgins, Jennifer A.; Cooke, Marcus; Mistry, Vilas; et al. (2012-07-27)
      There are substantial inter-laboratory variations in the levels of DNA damage measured by the comet assay. The aim of this study was to investigate whether adherence to a standard comet assay protocol would reduce inter-laboratory variation in reported values of DNA damage. Fourteen laboratories determined the baseline level of DNA strand breaks (SBs)/alkaline labile sites and formamidopyrimidine DNA glycosylase (FPG)-sensitive sites in coded samples of mononuclear blood cells (MNBCs) from healthy volunteers. There were technical problems in seven laboratories in adopting the standard protocol, which were not related to the level of experience. Therefore, the inter-laboratory variation in DNA damage was only analysed using the results from laboratories that had obtained complete data with the standard comet assay protocol. This analysis showed that the differences between reported levels of DNA SBs/alkaline labile sites in MNBCs were not reduced by applying the standard assay protocol as compared with the laboratory's own protocol. There was large inter-laboratory variation in FPG-sensitive sites by the laboratory-specific protocol and the variation was reduced when the samples were analysed by the standard protocol. The SBs and FPG-sensitive sites were measured in the same experiment, indicating that the large spread in the latter lesions was the main reason for the reduced inter-laboratory variation. However, it remains worrying that half of the participating laboratories obtained poor results using the standard procedure. This study indicates that future comet assay validation trials should take steps to evaluate the implementation of standard procedures in participating laboratories.
    • Rotating night shift work and mammographic density.

      Peplonska, Beata; Bukowska, Agnieszka; Sobala, Wojciech; Reszka, Edyta; Gromadzinska, Jolanta; Wasowicz, Wojciech; Lie, Jenny Anne; Kjuus, Helge; Ursin, Giske (2012-07)
      An increased risk of breast cancer has been observed in night shift workers. Exposure to artificial light at night and disruption of the endogenous circadian rhythm with suppression of the melatonin synthesis have been suggested mechanisms. We investigated the hypothesis that rotating night shift work is associated with mammographic density.
    • Development and validation of a direct sandwich chemiluminescence immunoassay for measuring DNA adducts of benzo[a]pyrene and other polycyclic aromatic hydrocarbons.

      Georgiadis, Panagiotis; Kovacs, Katalin; Kaila, Stella; Makedonopoulou, Paraskevi; Anna, Livia; Poirier, Miriam C.; Knudsen, Lisbeth E.; Schoket, Bernadette; Kyrtopoulos, Soterios A. (2012-05-18)
      We have developed and validated a sandwich chemiluminescence immunoassay (SCIA) which measures polycyclic aromatic hydrocarbon (PAH)-DNA adducts combining high throughput and adequate sensitivity, appropriate for evaluation of adduct levels in human population studies. Fragmented DNA is incubated with rabbit antiserum elicited against DNA modified with r7,t8-dihydroxy-t-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene (BPDE) and subsequently trapped by goat anti-rabbit IgG bound to a solid surface. Anti-single-stranded (ss) DNA antibodies binds in a quantity proportional to the adduct levels and is detected by chemiluminescence. The BPDE-DNA SCIA has a limit of detection of 3 adducts per 10(9) nucleotides with 5 μg DNA per well. We have validated the BPDE-DNA SCIA using DNA modified in vitro, DNA from benzo[a]pyrene (BP)-exposed cultured cells and mice. The levels of adduct measured by SCIA were lower (30-60%) than levels of bulky DNA adducts measured in the same samples by (32)P-postlabelling. The BPDE-DNA SCIA also detected adducts produced in vivo by PAHs other than BP. When blood DNA samples from maternal/infant pairs were assayed by BPDE-DNA SCIA, the adduct levels obtained were significantly correlated. However, there was no correlation between (32)P-postlabelling and SCIA values for the same samples. The SCIA can be extended to any DNA adduct and is expected to provide, when fully automated, a valuable high-throughput approach in large-scale population studies.
    • Progress in high-throughput assays of MGMT and APE1 activities in cell extracts.

      Georgiadis, Panagiotis; Polychronaki, Nektaria; Kyrtopoulos, Soterios A. (2012-05-17)
      DNA repair activity is of interest as a potential biomarker of individual susceptibility to genotoxic agents. In view of the current trend for exploitation of large cohorts in molecular epidemiology projects, there is a pressing need for the development of phenotypic DNA repair assays that are high-throughput, very sensitive, inexpensive and reliable. Towards this goal we have developed and validated two phenotypic assays for the measurement of two DNA repair enzymes in cell extracts: (1) O(6)-methylguanine-DNA-methyltransferase (MGMT), which repairs the O(6)-alkylguanine-type of adducts induced in DNA by alkylating genotoxins; and (2) apurinic/apyrimidinic endonuclease 1 (APE 1), which participates in base excision repair (BER) by causing a rate-limiting DNA strand cleavage 5' to the abasic sites. The MGMT assay makes use of the fact that: (a) the enzyme works by irreversibly transferring the alkyl group from the O(6) position of guanine to a cystein residue in its active site and thereby becomes inactivated and (b) that the free base O(6)-benzylguanine (BG) is a very good substrate for MGMT. In the new assay, cell extracts are incubated with BG tagged with biotin and the resulting MGMT-BG-biotin complex is immobilized on anti-MGMT-coated microtiter plates, followed by quantitation using streptavidin-conjugated alkaline phosphatase and a chemiluminescence-producing substrate. A one-step/one-tube phenotypic assay for APE1 activity has been developed based on the use of a fluorescent molecular beacon (partially self-complementary oligonucleotide with a hairpin-loop structure carrying a fluorophore and a quencher at each end). It also contains a single tetrahydrofuran residue (THF) which is recognized and cleaved by APE1, and the subsequently formed single-stranded oligomer becomes a fluorescence signal emitter. Both assays are highly sensitive, require very small amounts of protein extracts, are relatively inexpensive and can be easily automated. They have been extensively validated and are being used in the context of large-scale molecular epidemiology studies.
    • DNA repair gene expression level in peripheral blood and tumour tissue from non-small cell lung cancer and head and neck squamous cell cancer patients.

      Schena, Marina; Guarrera, Simonetta; Buffoni, Lucio; Salvadori, Angelica; Voglino, Floriana; Allione, Alessandra; Pecorari, Giancarlo; Ruffini, Enrico; Garzino-Demo, Paolo; Bustreo, Sara; et al. (2012-04-01)
      The nucleotide excision repair pathway is crucial for cellular DNA integrity and the ERCC1 helicase is also potentially involved in resistance to platinum-based chemotherapy, and high levels of ERCC1 mRNA in tumours have been associated with cisplatin resistance in different human cancers. The aim of this work was to investigate the correlation between DNA repair gene expression levels in tumour tissue, normal tissue and peripheral blood samples from patients with two common human cancers, non-small cell lung cancer (NSCLC) and squamous cell carcinoma of the head and neck (HNSCC), to test if blood gene expression could be a proxy for tumour tissue gene expression to predict response to platinum-based chemotherapy.
    • Biologically relevant oxidants and terminology, classification and nomenclature of oxidatively generated damage to nucleobases and 2-deoxyribose in nucleic acids.

      Cadet, Jean; Loft, Steffen; Olinski, Ryszard; Evans, Mark D.; Bialkowski, Karol; Richard Wagner, J.; Dedon, Peter C.; Moller, Peter; Greenberg, Marc M.; Cooke, Marcus S. (2012-04)
      A broad scientific community is involved in investigations aimed at delineating the mechanisms of formation and cellular processing of oxidatively generated damage to nucleic acids. Perhaps as a consequence of this breadth of research expertise, there are nomenclature problems for several of the oxidized bases including 8-oxo-7,8-dihydroguanine (8-oxoGua), a ubiquitous marker of almost every type of oxidative stress in cells. Efforts to standardize the nomenclature and abbreviations of the main DNA degradation products that arise from oxidative pathways are reported. Information is also provided on the main oxidative radicals, non-radical oxygen species, one-electron agents and enzymes involved in DNA degradation pathways as well in their targets and reactivity. A brief classification of oxidatively generated damage to DNA that may involve single modifications, tandem base modifications, intrastrand and interstrand cross-links together with DNA-protein cross-links and base adducts arising from the addition of lipid peroxides breakdown products is also included.
    • Harmonising measurements of 8-oxo-7,8-dihydro-2'-deoxyguanosine in cellular DNA and urine.

      Moller, Peter; Cooke, Marcus S.; Collins, Andrew; Olinski, Ryszard; Rozalski, Rafal; Loft, Steffen (2012-04)
      Levels of oxidatively damaged cellular DNA and urinary excretion of damaged 2'-deoxyribonuclosides are widely measured in biomonitoring studies examining the role of oxidative stress induced by environmental exposures, lifestyle factors and development of disease. This has promoted efforts to harmonise measurements of oxidised guanine nucleobases by the variety of analytical approaches for DNA and urinary levels of damage, in multi-laboratory trials that are centred in Europe. The large inter-laboratory variation reported of values of oxidatively damaged DNA is reduced by harmonising assay protocols. Recent attention on optimal conditions for the comet assay may lead to better understanding of the most critical steps in procedure, which generate variation in DNA damage levels between laboratories. Measurements of urinary excretion of oxidatively generated 8-oxo-7,8-dihydro-2'-deoxyguanosine also show large differences between different methods, where chromatographic techniques generally show more reliable results than antibody-based methods. In this case, standardising calibrants is aimed at improving within technique agreement.
    • Vitamins at physiological levels cause oxidation to the DNA nucleoside deoxyguanosine and to DNA--alone or in synergism with metals.

      Bergstrom, Therese; Ersson, Clara; Bergman, Jan; Moller, Lennart (2012-03-30)
      Vitamins with antioxidant properties have the ability to act as pro-oxidants, inducing oxidative damage and oxidative stress as opposed to preventing it. While vitamin supplements are commonly consumed, the scientific evidence for their health beneficial effects is inconclusive. In fact, even harmful effects have been reported. The present study aimed to investigate and compare pro-oxidant properties of different antioxidants and vitamins commonly found in dietary supplements, at concentrations of physiological relevance, alone or in combination with metals also found in supplements. Focus was on damages related to DNA. The vitamins' chemical oxidation potencies were studied by measuring the amount of the oxidation product 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG) formed from the DNA nucleoside deoxyguanosine (dG) after vitamin exposure, using a high-performance liquid chromatography system with electrochemical and ultraviolet detection. To study the vitamins' ability to cause DNA damage to cultured cells, promyelocytic leukemia cells (HL-60) were exposed to vitamins, and strand breaks, alkali-labile sites and oxidative DNA lesions, i.e. formamido pyrimidine DNA glycosylase-sensitive sites, were detected using the comet assay. Vitamins A and C chemically induced oxidation of dG, alone and in synergism with iron or copper, whereas only vitamin C and copper induced DNA damage in cultured cells. Contrary, vitamins B1, B2, B3, B6 and B12, β-carotene, folic acid, α-tocopherol, δ-tocopherol or γ-tocopherol did not induce oxidative damage to dG, while lycopene induced a weak dose-response increase. Taken together, vitamin C and copper stood out with the strongest oxidative potency, which is of potential concern since both substances are commonly found in multivitamins.
    • DNA damage induced by micro- and nanoparticles--interaction with FPG influences the detection of DNA oxidation in the comet assay.

      Kain, J.; Karlsson, H. L.; Moller, L. (2012-03-23)
      Reliable methods for evaluation of toxicity from particles, such as manufactured nanoparticles, are needed. One promising tool is the comet assay, often used to measure DNA breaks (strand breaks and alkali-labile sites) as well as oxidatively damaged DNA, the latter by addition of specific DNA repair enzymes such as formamidopyrimidine DNA glycosylase (FPG). The aim of this study was to investigate the use of the comet assay for analysis of DNA oxidation by a range of micro- and nanoparticles in the lung cell lines A549 and BEAS-2B and to test the hypothesis that nanoparticles present in the cells during the assay performance may interact with FPG. This was done by investigating the ability of micro- and nanoparticles (stainless steel, subway particles, MnO(2), Ag, CeO(2), Co(3)O(4), Fe(3)O(4), NiO and SiO(2)) to induce DNA breaks, oxidatively damaged DNA (FPG sites, dominantly 8-oxoguanine), intracellular production of reactive oxygen species (ROS) and non-cellular oxidation of the DNA base guanine, as well as by studying interactions of the particles and their released ions with FPG. Several particles caused DNA breaks, but low levels of FPG sites. The ability of FPG to detect DNA oxidation induced by a photosensitiser was however shown. An oxidative capacity of the particles was indicated by increased levels of intracellular ROS, and especially Ag and subway particles caused non-cellular oxidation of guanine. Incubation of FPG with the particles led to less FPG activity, particularly with nanoparticles of Ag but also with CeO(2), Co(3)O(4) and SiO(2). Further investigations of these particles revealed that for Ag, the decreased activity was mainly due to released Ag ions, whereas for CeO(2) and Co(3)O(4), FPG interactions were due to the particles. We conclude that measurement of oxidatively damaged DNA in cells exposed to nanoparticles may be underestimated in the comet assay due to interactions with FPG.
    • Association between total number of deaths, diabetes mellitus, incident cancers, and haplotypes in chromosomal region 8q24 in a prospective study.

      Guarrera, Simonetta; Ricceri, Fulvio; Polidoro, Silvia; Sacerdote, Carlotta; Allione, Alessandra; Rosa, Fabio; Voglino, Floriana; Critelli, Rossana; Russo, Alessia; Vineis, Paolo; et al. (2012-03-15)
      The 8q24 region is a gene desert, although chromosomal aberrations and somatic amplification involving this region, including translocations involving the protooncogene c-MYC, have been frequently reported in people with cancer. To investigate the role of variants in 8q24 region, the authors analyzed data from a prospective study (n = 10,372 participants who were followed for 11 years) in which a large number of health events (>1,500) occurred (1993-1998). They genotyped all subjects for 5 candidate single nucleotide polymorphisms (rs672888, rs1447295, rs9642880, rs16901979, and rs6983267) that were identified in previous genome-wide scans. Although significant associations with individual single nucleotide polymorphisms were small in magnitude, the authors observed higher increases in the risks of different types of cancer with specific haplotypes, particularly when subjects were homozygous for the haplotype: for breast cancer and homozygotes for haplotype CAGCT, hazard ratio = 3.40, 95% confidence interval: 1.24, 9.21; for prostate cancer and grouped rare haplotypes, hazard ratio = 7.43, 95% confidence interval: 3.00, 18.37; and for brain cancer and homozygotes for haplotype CGGCT, hazard ratio = 13.48, 95% confidence interval: 3.00, 59.53. Significant associations were also observed between haplotypes and deaths from cardiovascular diseases and cerebrovascular diseases; the most stable association was between homozygotes for haplotypes CGTCG and CAGCT and total deaths in men (hazard ratio = 3.5, 95% confidence interval: 1.8, 6.9, and hazard ratio = 2.8, 95% confidence interval: 1.3, 6.4, respectively). In conclusion, the authors have observed a strong pleiotropic effect of the 8q24 region in a large prospective study. This observation can shed light on the mechanisms underlying reported associations between 8q24 variants and disparate chronic diseases.
    • Urinary excretion of 8-oxo-7,8-dihydroguanine as biomarker of oxidative damage to DNA.

      Loft, Steffen; Danielsen, Pernille; Løhr, Mille; Jantzen, Kim; Hemmingsen, Jette G.; Roursgaard, Martin; Karotki, Dorina Gabriela; Møller, Peter (2012-02-15)
      Oxidatively damaged DNA may be important in carcinogenesis. 8-Oxo-7,8-dihydroguanine (8-oxoGua) is an abundant and mutagenic lesion excised by oxoguanine DNA glycosylase 1 (OGG1) and measurable in urine or plasma by chromatographic methods with electrochemical or mass spectrometric detectors, reflecting the rate of damage in steady state. A common genetic OGG1 variant may affect the activity and was associated with increased levels of oxidized purines in leukocytes without apparent effect on 8-oxoGua excretion or major change in cancer risk. 8-OxoGua excretion has been associated with exposure to air pollution, toxic metals, tobacco smoke and low plasma antioxidant levels, whereas fruit and vegetable intake or dietary interventions showed no association. In rodent studies some types of feed may be source of 8-oxoGua in collected urine. Of cancer therapies, cisplatin increased 8-oxoGua excretion, whereas radiotherapy only showed such effects in experimental animals. Case-control studies found high excretion of 8-oxoGua in relation to cancer, dementia and celiac disease but not hemochromatosis, although associations could be a consequence rather than reflecting causality of disease. One prospective study found increased risk of developing lung cancer among non-smokers associated with high excretion of 8-oxoGua. Urinary excretion of 8-oxoGua is a promising biomarker of oxidatively damaged DNA.
    • Gastrointestinal conditions influence the solution behaviour of cereal β-glucans in vitro.

      Ulmius, Matilda; Adapa, Srimannarayana; Onning, Gunilla; Nilsson, Lars (2012-02)
      The solution behaviour of β-glucans in a gastrointestinal model was investigated in order to explore the mechanisms explaining the physiological effects of the soluble fibre. Asymmetrical flow field-flow fractionation was used to determine the molar mass distribution, and in-line calcofluor labelling allowed specific detection of β-glucans in complex samples. When dispersed in water, weight-average molar mass (Mw) was determined to 1 × 106 g/mol for pure oat and barley β-glucans, and 200 × 106 g/mol for β-glucans in oat bran, indicating that the β-glucans were aggregating. Samples from the gastric digestion displayed disrupted aggregates, while samples from the small intestinal digestion contained re-formed aggregates. Additionally, the aggregates from pure β-glucans were considerably denser after intestinal digestion. This may be construed as gel-formation in the small intestine, which should be tested for its relevance to health effects. Our results signal the difficulties in predicting β-glucan activity in the gastrointestinal tract purely from analysis of the fibre-rich product.
    • STrengthening the Reporting of OBservational studies in Epidemiology--Molecular Epidemiology (STROBE-ME): an extension of the STROBE statement.

      Gallo, Valentina; Egger, Matthias; McCormack, Valerie; Farmer, Peter B.; Ioannidis, John P. A.; Kirsch-Volders, Micheline; Matullo, Giuseppe; Phillips, David H.; Schoket, Bernadette; Stromberg, Ulf; et al. (2012-01)
      Advances in laboratory techniques have led to a rapidly increasing use of biomarkers in epidemiological studies. Biomarkers of internal dose, early biological change, susceptibility and clinical outcomes are used as proxies for investigating interactions between external and / or endogenous agents and body components or processes. The need for improved reporting of scientific research led to influential statements of recommendations such as the STrengthening Reporting of OBservational studies in Epidemiology (STROBE) statement. The STROBE initiative established in 2004 aimed to provide guidance on how to report observational research. Its guidelines provide a user-friendly checklist of 22 items to be reported in epidemiological studies, with items specific to the three main study designs: cohort studies, case-control studies and cross-sectional studies. The present STrengthening the Reporting of OBservational studies in Epidemiology - Molecular Epidemiology (STROBE-ME) initiative builds on the STROBE statement implementing nine existing items of STROBE and providing 17 additional items to the 22 items of STROBE checklist. The additions relate to the use of biomarkers in epidemiological studies, concerning collection, handling and storage of biological samples; laboratory methods, validity and reliability of biomarkers; specificities of study design; and ethical considerations. The STROBE-ME recommendations are intended to complement the STROBE recommendations.
    • STrengthening the Reporting of OBservational studies in Epidemiology - Molecular Epidemiology (STROBE-ME): an extension of the STROBE statement.

      Gallo, Valentina; Egger, Matthias; McCormack, Valerie; Farmer, Peter B.; Ioannidis, John P. A.; Kirsch-Volders, Micheline; Matullo, Giuseppe; Phillips, David H.; Schoket, Bernadette; Stromberg, Ulf; et al. (2012-01)
      Advances in laboratory techniques have led to a rapidly increasing use of biomarkers in epidemiological studies. Biomarkers of internal dose, early biological change, susceptibility and clinical outcomes are used as proxies for investigating interactions between external and/or endogenous agents and body components or processes. The need for improved reporting of scientific research led to influential statements of recommendations such as the STrengthening Reporting of OBservational studies in Epidemiology (STROBE) statement. The STROBE initiative established in 2004 aimed to provide guidance on how to report observational research. Its guidelines provide a user-friendly checklist of 22 items to be reported in epidemiological studies, with items specific to the three main study designs: cohort studies, case-control studies and cross-sectional studies. The present STrengthening the Reporting of OBservational studies in Epidemiology -Molecular Epidemiology (STROBE-ME) initiative builds on the STROBE statement implementing nine existing items of STROBE and providing 17 additional items to the 22 items of STROBE checklist. The additions relate to the use of biomarkers in epidemiological studies, concerning collection, handling and storage of biological samples; laboratory methods, validity and reliability of biomarkers; specificities of study design; and ethical considerations. The STROBE-ME recommendations are intended to complement the STROBE recommendations.
    • Exposure to Polycyclic Aromatic Hydrocarbons Among Never Smokers in Golestan Province, Iran, an Area of High Incidence of Esophageal Cancer - a Cross-Sectional Study with Repeated Measurement of Urinary 1-OHPG in Two Seasons.

      Islami, Farhad; Boffetta, Paolo; Van Schooten, Frederik J.; Strickland, Paul; Phillips, David H.; Pourshams, Akram; Fazel-Tabar Malekshah, Akbar; Godschalk, Roger; Jafari, Elham; Etemadi, Arash; et al. (2012)
      Studies have suggested a possible role of polycyclic aromatic hydrocarbons (PAHs) in the etiology of esophageal cancer in Golestan Province, Iran, where incidence of this cancer is very high. In order to investigate the patterns of non-smoking related exposure to PAHs in Golestan, we conducted a cross-sectional study collecting questionnaire data, genotyping polymorphisms related to PAH metabolism, and measuring levels of 1-hydroxypyrene glucuronide (1-OHPG), a PAH metabolite, in urine samples collected in two seasons from the same group of 111 randomly selected never-smoking women. Beta-coefficients for correlations between 1-OHPG as dependent variable and other variables were calculated using linear regression models. The creatinine-adjusted 1-OHPG levels in both winter and summer samples were approximately 110 μmol/molCr (P for seasonal difference = 0.40). In winter, red meat intake (β = 0.208; P = 0.03), processed meat intake (β = 0.218; P = 0.02), and GSTT1-02 polymorphism ("null" genotype: β = 0.228; P = 0.02) showed associations with 1-OHPG levels, while CYP1B1-07 polymorphism (GG versus AA + GA genotypes: β = -0.256; P = 0.008) showed an inverse association. In summer, making bread at home (> weekly versus never: β = 0.203; P = 0.04), second-hand smoke (exposure to ≥3 cigarettes versus no exposure: β = 0.254; P = 0.01), and GSTM1-02 "null" genotype (β = 0.198; P = 0.04) showed associations with 1-OHPG levels, but GSTP1-02 polymorphism (CT + TT versus CC: β = -0.218; P = 0.03) showed an inverse association. This study confirms high exposure of the general population in Golestan to PAHs and suggests that certain foods, cooking methods, and genetic polymorphisms increase exposure to PAHs.
    • Sources of pre-analytical variations in yield of DNA extracted from blood samples: analysis of 50,000 DNA samples in EPIC.

      Caboux, Elodie; Lallemand, Christophe; Ferro, Gilles; Hémon, Bertrand; Mendy, Maimuna; Biessy, Carine; Sims, Matt; Wareham, Nick; Britten, Abigail; Boland, Anne; et al. (2012)
      The European Prospective Investigation into Cancer and nutrition (EPIC) is a long-term, multi-centric prospective study in Europe investigating the relationships between cancer and nutrition. This study has served as a basis for a number of Genome-Wide Association Studies (GWAS) and other types of genetic analyses. Over a period of 5 years, 52,256 EPIC DNA samples have been extracted using an automated DNA extraction platform. Here we have evaluated the pre-analytical factors affecting DNA yield, including anthropometric, epidemiological and technical factors such as center of subject recruitment, age, gender, body-mass index, disease case or control status, tobacco consumption, number of aliquots of buffy coat used for DNA extraction, extraction machine or procedure, DNA quantification method, degree of haemolysis and variations in the timing of sample processing. We show that the largest significant variations in DNA yield were observed with degree of haemolysis and with center of subject recruitment. Age, gender, body-mass index, cancer case or control status and tobacco consumption also significantly impacted DNA yield. Feedback from laboratories which have analyzed DNA with different SNP genotyping technologies demonstrate that the vast majority of samples (approximately 88%) performed adequately in different types of assays. To our knowledge this study is the largest to date to evaluate the sources of pre-analytical variations in DNA extracted from peripheral leucocytes. The results provide a strong evidence-based rationale for standardized recommendations on blood collection and processing protocols for large-scale genetic studies.
    • STrengthening the Reporting of OBservational studies in Epidemiology--Molecular Epidemiology STROBE-ME: an extension of the STROBE statement.

      Gallo, Valentina; Egger, Matthias; McCormack, Valerie; Farmer, Peter B.; Ioannidis, John P. A.; Kirsch-Volders, Micheline; Matullo, Giuseppe; Phillips, David H.; Schoket, Bernadette; Stromberg, Ulf; et al. (2011-12)
      Advances in laboratory techniques have led to a rapidly increasing use of biomarkers in epidemiological studies. Biomarkers of internal dose, early biological change susceptibility and clinical outcomes are used as proxies for investigating the interactions between external and/or endogenous agents and body components or processes. The need for improved reporting of scientific research led to influential statements of recommendations such as the STrengthening Reporting of OBservational studies in Epidemiology (STROBE) statement. The STROBE initiative established in 2004 aimed to provide guidance on how to report observational research. Its guidelines provide a user-friendly checklist of 22 items to be reported in epidemiological studies, with items specific to the three main study designs: cohort studies, case-control studies and cross-sectional studies. The present STrengthening the Reporting of OBservational studies in Epidemiology -Molecular Epidemiology (STROBE-ME) initiative builds on the STROBE statement implementing 9 existing items of STROBE and providing 17 additional items to the 22 items of STROBE checklist. The additions relate to the use of biomarkers in epidemiological studies, concerning collection, handling and storage of biological samples; laboratory methods, validity and reliability of biomarkers; specificities of study design; and ethical considerations. The STROBE-ME recommendations are intended to complement the STROBE recommendations.