Publications by authors named "Morena Trevisan"

27 Publications

  • Page 1 of 1

Associations between dietary amino acid intakes and blood concentration levels.

Clin Nutr 2021 Jun 27;40(6):3772-3779. Epub 2021 Apr 27.

International Agency for Research on Cancer, Nutrition and Metabolism Section, 69372, Lyon CEDEX 08, France.

Background And Aims: Emerging evidence suggests a role of amino acids (AAs) in the development of various diseases including renal failure, liver cirrhosis, diabetes and cancer. However, mechanistic pathways and the effects of dietary AA intakes on circulating levels and disease outcomes are unclear. We aimed to compare protein and AA intakes, with their respective blood concentrations in the European Prospective Investigation into Cancer and Nutrition (EPIC) cohort.

Methods: Dietary protein and AA intakes were assessed via the EPIC dietary questionnaires (DQ) and 24-h dietary recalls (24-HDR). A subsample of 3768 EPIC participants who were free of cancer had blood AA concentrations measured. To investigate how circulating levels relate to their respective intakes, dietary AA intake was examined in quintiles and ANOVA tests were run. Pearson correlations were examined for continous associations between intakes and blood concentrations.

Results: Dietary AA intakes (assessed with the DQ) and blood AA concentrations were not strongly correlated (-0.15 ≤ r ≤ 0.17) and the direction of the correlations depended on AA class: weak positive correlations were found for most essential AAs (isoleucine, leucine, lysine, methionine, threonine, tryptophan, and valine) and conditionally essential AAs (arginine and tyrosine), while negative associations were found for non-essential AAs. Similar results were found when using the 24-HDR. When conducting ANOVA tests for essential AAs, higher intake quintiles were linked to higher blood AA concentrations, except for histidine and phenylalanine. For non-essential AAs and glycine, an inverse relationship was observed. Conditionally-essential AAs showed mixed results.

Conclusions: Weak positive correlations and dose responses were found between most essential and conditionally essential AA intakes, and blood concentrations, but not for the non-essential AAs. These results suggest that intake of dietary AA might be related to physiological AA status, particularly for the essential AAs. However, these results should be further evaluated and confirmed in large-scale prospective studies.
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http://dx.doi.org/10.1016/j.clnu.2021.04.036DOI Listing
June 2021

Birthweight DNA methylation signatures in infant saliva.

Clin Epigenetics 2021 Mar 19;13(1):57. Epub 2021 Mar 19.

Cancer Epidemiology Unit, Department of Medical Sciences, University of Turin and CPO Piemonte, Via Santena 7, 10126, Turin, Italy.

Background: Low birthweight has been repeatedly associated with long-term adverse health outcomes and many non-communicable diseases. Our aim was to look-up cord blood birthweight-associated CpG sites identified by the PACE Consortium in infant saliva, and to explore saliva-specific DNA methylation signatures of birthweight.

Methods: DNA methylation was assessed using Infinium HumanMethylation450K array in 135 saliva samples collected from children of the NINFEA birth cohort at an average age of 10.8 (range 7-17) months. The association analyses between birthweight and DNA methylation variations were carried out using robust linear regression models both in the exploratory EWAS analyses and in the look-up of the PACE findings in infant saliva.

Results: None of the cord blood birthweight-associated CpGs identified by the PACE Consortium was associated with birthweight when analysed in infant saliva. In saliva EWAS analyses, considering a false discovery rate p-values < 0.05, birthweight as continuous variable was associated with DNA methylation in 44 CpG sites; being born small for gestational age (SGA, lower 10 percentile of birthweight for gestational age according to WHO reference charts) was associated with DNA methylation in 44 CpGs, with only one overlapping CpG between the two analyses. Despite no overlap with PACE results at the CpG level, two of the top saliva birthweight CpGs mapped at genes associated with birthweight with the same direction of the effect also in the PACE Consortium (MACROD1 and RPTOR).

Conclusion: Our study provides an indication of the birthweight and SGA epigenetic salivary signatures in children around 10 months of age. DNA methylation signatures in cord blood may not be comparable with saliva DNA methylation signatures at about 10 months of age, suggesting that the birthweight epigenetic marks are likely time and tissue specific.
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http://dx.doi.org/10.1186/s13148-021-01053-1DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7980592PMC
March 2021

Determination of saliva epigenetic age in infancy, and its association with parental socio-economic characteristics and pregnancy outcomes.

J Dev Orig Health Dis 2021 Apr 26;12(2):319-327. Epub 2020 May 26.

Cancer Epidemiology Unit, Department of Medical Sciences, University of Turin and CPO Piemonte, Turin, Italy.

Epigenetic age acceleration (AA) has been associated with adverse environmental exposures and many chronic conditions. We estimated, in the NINFEA birth cohort, infant saliva epigenetic age, and investigated whether parental socio-economic position (SEP) and pregnancy outcomes are associated with infant epigenetic AA. A total of 139 saliva samples collected at on average 10.8 (range 7-17) months were used to estimate Horvath's DNA methylation age. Epigenetic AA was defined as the residual from a linear regression of epigenetic age on chronological age. Linear regression models were used to test the associations of parental SEP and pregnancy outcomes with saliva epigenetic AA. A moderate positive association was found between DNA methylation age and chronological age, with the median absolute difference of 6.8 months (standard deviation [SD] 3.9). The evidence of the association between the indicators of low SEP and epigenetic AA was weak; infants born to unemployed mothers or with low education had on average 1 month higher epigenetic age than infants of mothers with high education and employment (coefficient 0.78 months, 95% confidence intervals [CIs]: -0.79 to 2.34 for low/medium education; 0.96, 95% CI: -1.81 to 3.73 for unemployment). There was no evidence for association of gestational age, birthweight or caesarean section with infant epigenetic AA. Using the Horvath's method, DNA methylation age can be fairly accurately predicted from saliva samples already in the first months of life. This study did not reveal clear associations between either pregnancy outcomes or parental socio-economic characteristics and infant saliva epigenetic AA.
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http://dx.doi.org/10.1017/S2040174420000380DOI Listing
April 2021

DNA methylation in repeat negative prostate biopsies as a marker of missed prostate cancer.

Clin Epigenetics 2019 10 30;11(1):152. Epub 2019 Oct 30.

Cancer Epidemiology Unit-CeRMS, Department of Medical Sciences, University of Turin and CPO Piemonte, Via Santena 7, 10126, Turin, Italy.

Background: Men often undergo repeat prostate biopsies because of suspicion of missed cancer. We assessed if (i) methylation of selected genes in prostate tissue vary with aging and (ii) methylation alterations in repeat biopsies predict missed prostate cancer.

Methods: We conducted a case-control study among men who underwent at least two negative prostate biopsies followed by a sampling either positive (cases n = 111) or negative (controls n = 129) for prostate cancer between 1995 and 2014 at the University Hospital (Turin, Italy). Two pathology wards were included for replication purposes. We analyzed methylation of GSTP1, APC, PITX2, C1orf114, GABRE, and LINE-1 in the first two negative biopsies. Conditional logistic regression was used to estimate odds ratios (ORs) and 95% confidence intervals (CIs) of the association between genes methylation and prostate cancer.

Results: Age at biopsy and time interval between the two negative biopsies were not associated with methylation levels of the selected genes in neither cases nor controls. GSTP1 methylation in the first and in the second negative biopsy was associated with prostate cancer detection [OR per 1% increase: 1.14 (95% CI 1.01-1.29) for the second biopsy and 1.21 (95% CI 1.07-1.37) for the highest methylation level (first or second biopsy)]. A threshold > 10% for GSTP1 methylation corresponded to a specificity of 0.98 (positive likelihood ratio 7.87). No clear association was found for the other genes. Results were consistent between wards.

Conclusions: Our results suggest that GSTP1 methylation in negative prostate biopsies is stable over time and can predict missed cancer with high specificity.
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http://dx.doi.org/10.1186/s13148-019-0746-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6820908PMC
October 2019

DNA methylation, colon cancer and Mediterranean diet: results from the EPIC-Italy cohort.

Epigenetics 2019 10 14;14(10):977-988. Epub 2019 Jun 14.

h Unit of Cancer Epidemiology, Città della Salute e della Scienza University-Hospital and Center for Cancer Prevention (CPO) , Turin , Italy.

The biological mechanisms through which adherence to Mediterranean Diet (MD) protects against colon cancer (CC) are poorly understood. Evidence suggests that chronic inflammation may be implicated in the pathway. Both diet and CC are related to epigenetic regulation. We performed a nested case-control study on 161 pairs from the Italian component of the European Prospective Investigation into Cancer and Nutrition (EPIC) cohort, in which we looked for the methylation signals in DNA extracted from leucocytes associated with both CC and MD in 995 CpGs located in 48 inflammation genes. The DNA methylation signals detected in this analysis were validated in a subgroup of 47 case-control pairs and further replicated (where validated) in 95 new pairs by means of pyrosequencing. Among the CpG sites selected in inflammation-related genes, seven CpG sites were found to be associated with CC status and with MD, in line with its protective effect. Only two CpG sites (cg17968347- and cg20674490-) were validated using bisulphite pyrosequencing and, after replication, we found that DNA methylation of cg20674490- may be a potential molecular mediator explaining the protective effect of MD on CC onset. The use of a 'meet-in-the-middle' approach to identify the overlap between exposure and predictive markers of disease is innovative in studies on the relationship between diet and cancer, in which exposure assessment is difficult and the mechanisms through which the nutrients exert their protective effect is largely unknown.
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http://dx.doi.org/10.1080/15592294.2019.1629230DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6691992PMC
October 2019

Differentially methylated DNA regions in early childhood wheezing: An epigenome-wide study using saliva.

Pediatr Allergy Immunol 2019 05 17;30(3):305-314. Epub 2019 Feb 17.

Cancer Epidemiology Unit, Department of Medical Sciences, University of Turin, Turin, Italy.

Background: Epigenetics may play a role in wheezing and asthma development. We aimed to examine infant saliva DNA methylation in association with early childhood wheezing.

Methods: A case-control study was nested within the NINFEA birth cohort with 68 cases matched to 68 controls by sex, age (between 6 and 18 months, median: 10.3 months) and season at saliva sampling. Using a bumphunting region-based approach, we examined associations between saliva methylome measured using Illumina Infinium HumanMethylation450k array and wheezing between 6 and 18 months of age. We tested our main findings in independent publicly available data sets of childhood respiratory allergy and atopic asthma, with DNA methylation measured in different tissues and at different ages.

Results: We identified one wheezing-associated differentially methylated region (DMR) spanning ten sequential CpG sites in the promoter-regulatory region of PM20D1 gene (family-wise error rate < 0.05). The observed associations were enhanced in children born to atopic mothers. In the publicly available data sets, hypermethylation in the same region of PM20D1 was consistently found at different ages and in all analysed tissues (cord blood, blood, saliva and nasal epithelia) of children with respiratory allergy/atopic asthma compared with controls.

Conclusion: This study suggests that PM20D1 hypermethylation is associated with early childhood wheezing. Directionally consistent epigenetic alteration observed in cord blood and other tissues at older ages in children with respiratory allergy and atopic asthma provides suggestive evidence that a long-term epigenetic modification, likely operating from birth, may be involved in childhood atopic phenotypes.
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http://dx.doi.org/10.1111/pai.13023DOI Listing
May 2019

Single-nucleotide polymorphisms in DNMT3B gene and DNMT3B mRNA expression in association with prostate cancer mortality.

Prostate Cancer Prostatic Dis 2019 05 19;22(2):284-291. Epub 2018 Oct 19.

Cancer Epidemiology Unit-CERMS, Department of Medical Sciences, University of Turin, and CPO-Piemonte, Turin, Italy.

Background: Germline variants in DNA methyltransferase 3B (DNMT3B) may influence DNMT3B enzymatic activity, which, in turn, may affect cancer aggressiveness by altering DNA methylation.

Methods: The study involves two Italian cohorts (NTAT cohort, n = 157, and 1980s biopsy cohort, n = 182) and two U.S. cohorts (Health Professionals Follow-Up Study, n = 214, and Physicians' Health Study, n = 298) of prostate cancer (PCa) patients, and a case-control study of lethal (n = 113) vs indolent (n = 290) PCa with DNMT3B mRNA expression data nested in the U.S. cohorts. We evaluated the association between: three selected DNMT3B variants and global DNA methylation using linear regression in the NTAT cohort, the three DNMT3B variants and PCa mortality using Cox proportional hazards regression in all cohorts, and DNMT3B expression and lethal PCa using logistic regression, with replication in publicly available databases (TCGA, n = 492 and MSKCC, n = 140).

Results: The TT genotype of rs1569686 was associated with LINE-1 hypomethylation in tumor tissue (β = -2.71, 95% CI: -5.41, -0.05). There was no evidence of association between DNMT3B variants and PCa mortality. DNMT3B expression was consistently associated with lethal PCa in the two U.S. cohorts (3rd vs 1st tertile, combined cohorts: OR = 2.04, 95% CI: 1.13, 3.76); the association was replicated in TCGA and MSKCC data (3rd vs 1st tertile, TCGA: HR = 3.00, 95% CI: 1.78, 5.06; MSKCC: HR = 2.22, 95% CI: 1.01, 4.86).

Conclusions: Although there was no consistent evidence of an association between DNMT3B variants and PCa mortality, the TT genotype of rs1569686 was associated with LINE-1 hypomethylation in tumor tissue and DNMT3B mRNA expression was associated with an increased risk of lethal PCa.
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http://dx.doi.org/10.1038/s41391-018-0102-5DOI Listing
May 2019

Methylation in host and viral genes as marker of aggressiveness in cervical lesions: Analysis in 543 unscreened women.

Gynecol Oncol 2018 11 29;151(2):319-326. Epub 2018 Aug 29.

Unit of Cancer Epidemiology-CeRMS, Department of Medical Sciences, University of Turin, Via Santena 7, 10126 Turin, Italy. Electronic address:

Objective: The present study aimed to evaluate the association between altered methylation and histologically confirmed high grade cervical intraepithelial neoplasia (hgCIN).

Methods: Methylation levels in selected host (CADM1, MAL, DAPK1) and HPV (L1_I, L1_II, L2) genes were measured by pyrosequencing in DNA samples obtained from 543 women recruited in Curitiba (Brazil), 249 with hgCIN and 294 without cervical lesions. Association of methylation status with hgCIN was estimated by Odds Ratio (OR) with 95% confidence interval (CI).

Results: The mean methylation level increased with severity of the lesion in the host and viral genes (p-trend < 0.05), with the exception of L1_II region (p-trend = 0.075). Positive association was found between methylation levels for host genes and CIN2 and CIN3 lesions respectively [CADM1: OR 4.17 (95%CI 2.03-8.56) and OR 9.54 (95%CI 4.80-18.97); MAL: OR 5.98 (95%CI 2.26-15.78) and OR 22.66 (95%CI 9.21-55.76); DAPK1: OR 3.37 (95%CI 0.93-12.13) and OR 6.74 (95%CI 1.92-23.64)]. Stronger risk estimates were found for viral genes [L1_I: OR 10.74 (95%CI 2.66-43.31) and OR 15.00 (95%CI 3.00-74.98); L1_II: OR 73.18 (95%CI 4.07-1315.94) and OR 32.50 (95%CI 3.86-273.65); L2: OR 4.73 (95%CI 1.55-14.44) and OR 10.62 (95%CI 2.60-43.39)]. The cumulative effect of the increasing number of host and viral methylated genes was associated with the risk of CIN2 and CIN3 lesions (p-trend < 0.001).

Conclusions: Our results, empowered by a wide cervical sample series with a large number of hgCIN, supported the role of methylation as marker of aggressiveness.
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http://dx.doi.org/10.1016/j.ygyno.2018.08.031DOI Listing
November 2018

The role of maternal anorexia nervosa and bulimia nervosa before and during pregnancy in early childhood wheezing: Findings from the NINFEA birth cohort study.

Int J Eat Disord 2018 08 2;51(8):842-851. Epub 2018 May 2.

Department of Medical Sciences, University of Turin, Turin, Italy.

Objective: This study evaluates associations of maternal eating disorders (bulimia nervosa, anorexia nervosa, and purging behaviors) with infant wheezing and examines the effects of eating disorders on several wheezing determinants.

Method: We studied 5,150 singletons from the NINFEA birth cohort. Maternal bulimia nervosa and anorexia nervosa diagnoses were ascertained from the questionnaires completed in pregnancy and 6 months after delivery, and were analyzed as: ever diagnosis, only before pregnancy, and during pregnancy. Purging behaviors were assessed for 12 months before or during pregnancy. The associations with wheezing between 6 and 18 months of age were assessed in models adjusted for a priori selected confounders.

Results: Children born to mothers with lifetime eating disorders were at an increased risk of developing wheezing (adjusted OR 1.68; [95% CI: 1.08, 2.60]), and this risk further increased when the disorders were active during pregnancy (2.52 [1.23, 5.19]). Increased risk of offspring wheezing was observed also for purging behaviors without history of eating disorder diagnosis (1.50 [1.10, 2.04]). The observed associations were not explained by comorbid depression and/or anxiety. Bulimia nervosa and/or anorexia nervosa during pregnancy were also associated with several risk factors for wheezing, including maternal smoking, adverse pregnancy outcomes, shorter breastfeeding duration, and day-care attendance.

Discussion: The associations of maternal eating disorders with offspring wheezing suggest long-term adverse respiratory outcomes in children of mothers with eating disorders. A better understanding of mechanisms implicated is necessary to help reduce the respiratory disease burden in these children.
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http://dx.doi.org/10.1002/eat.22870DOI Listing
August 2018

Epigenome-wide association study of adiposity and future risk of obesity-related diseases.

Int J Obes (Lond) 2018 12 1;42(12):2022-2035. Epub 2018 May 1.

Department of Medical Sciences, Unit of Cancer Epidemiology-CERMS, University of Turin, Turin, Italy.

Background: Obesity is an established risk factor for several common chronic diseases such as breast and colorectal cancer, metabolic and cardiovascular diseases; however, the biological basis for these relationships is not fully understood. To explore the association of obesity with these conditions, we investigated peripheral blood leucocyte (PBL) DNA methylation markers for adiposity and their contribution to risk of incident breast and colorectal cancer and myocardial infarction.

Methods: DNA methylation profiles (Illumina Infinium HumanMethylation450 BeadChip) from 1941 individuals from four population-based European cohorts were analysed in relation to body mass index, waist circumference, waist-hip and waist-height ratio within a meta-analytical framework. In a subset of these individuals, data on genome-wide gene expression level, biomarkers of glucose and lipid metabolism were also available. Validation of methylation markers associated with all adiposity measures was performed in 358 individuals. Finally, we investigated the association of obesity-related methylation marks with breast, colorectal cancer and myocardial infarction within relevant subsets of the discovery population.

Results: We identified 40 CpG loci with methylation levels associated with at least one adiposity measure. Of these, one CpG locus (cg06500161) in ABCG1 was associated with all four adiposity measures (P = 9.07×10 to 3.27×10) and lower transcriptional activity of the full-length isoform of ABCG1 (P = 6.00×10), higher triglyceride levels (P = 5.37×10) and higher triglycerides-to-HDL cholesterol ratio (P = 1.03×10). Of the 40 informative and obesity-related CpG loci, two (in IL2RB and FGF18) were significantly associated with colorectal cancer (inversely, P < 1.6×10) and one intergenic locus on chromosome 1 was inversely associated with myocardial infarction (P < 1.25×10), independently of obesity and established risk factors.

Conclusion: Our results suggest that epigenetic changes, in particular altered DNA methylation patterns, may be an intermediate biomarker at the intersection of obesity and obesity-related diseases, and could offer clues as to underlying biological mechanisms.
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http://dx.doi.org/10.1038/s41366-018-0064-7DOI Listing
December 2018

Assessment of viral methylation levels for high risk HPV types by newly designed consensus primers PCR and pyrosequencing.

PLoS One 2018 26;13(3):e0194619. Epub 2018 Mar 26.

Centre for Oncologic Prevention, Turin, Italy.

Background: Measuring viral DNA methylation in human papillomavirus (HPV) infected women showed promise for accurate detection of high-grade cervical lesions and cancer. Methylation status has been widely investigated for HPV16, sporadically for other HPV types.

Methods: Objective of this methodological study was to set up molecular methods to test the methylation levels in the twelve oncogenic HPV types by pyrosequencing, minimizing the number of HPV type-specific PCR protocols. Target CpGs were selected on the HPV L1 (two regions, L1 I and L1 II) and L2 genes. Study samples included DNA stored at Turin, Italy, purified by cervical cells collected in Standard Transport Medium or PreservCyt from women who participated in two studies (N = 126 and 140) nested within the regional organized screening programme. PCR consensus primers were designed by PyroMark Assay Design software to be suitable for amplification of many different oncogenic HPV types.

Results: Generation of consensus primers was successful for L1 I and II regions, unsuccessful for L2 region, for which HPV type-specific primers remained necessary. The difference between replicated tests on the same sample was ≤4% in 88%, 77% and 91% of cases when targeting the L1 I, L1 II and L2 regions, respectively. The corresponding intra-class correlation coefficients (ICC) were 0.94, 0.87 and 0.97 respectively. When comparing methylation measures based on consensus and type-specific primers, ICC was 0.97 for the L1 I region and 0.99 the for L1 II region.

Conclusions: The proposed protocols, applying consensus primers suitable to amplify the oncogenic HPV types and minimize the number of PCR reactions, represent a promising tool to quantify viral methylation in women positive for any high risk HPV type.

Impact: Potential application of these methylation protocols in screening settings can be explored to identify women with high probability of progression to high grade lesions.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0194619PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5868804PMC
July 2018

Human papilloma virus genotyping for the cross-sectional and longitudinal probability of developing cervical intraepithelial neoplasia grade 2 or more.

Int J Cancer 2018 07 9;143(2):333-342. Epub 2018 Mar 9.

Center for Cancer Epidemiology and Prevention (CPO), Turin, Italy.

Human papilloma virus (HPV) testing is more sensitive but less specific than cytology. We evaluated stand-alone genotyping as a possible triage method. During a multicentre randomised controlled trial comparing HPV testing to conventional cytology, HPV-positive women were referred to colposcopy and followed up if no high-grade lesion was detected. HPV-positive samples were genotyped by GP5+/GP6+ primed polymerase chain reaction followed by reverse line blot. Genotypes were hierarchically ordered by positive predictive value (PPV) for CIN grade 2 or more (CIN2+), and grouped by cluster analysis into three groups (A, B and C in decreasing order). Receiver operating characteristic curves were computed. Among 2,255 HPV-positive women with genotyping, 239 CIN2+ (including 113 CIN3+) were detected at baseline or during a 3-year follow-up. HPV33 had the highest PPV with CIN2+ and CIN3+ as the endpoint and when considering lesions detected at baseline or also during follow-up. HPV16 and HPV35 were the second and third, respectively. Cross-sectional sensitivity for CIN2+ at baseline was 67.3% (95% CI 59.7-74.2), 91.8% (95% CI 86.6-95.5) and 94.7% (95% CI 90.2-97.6), respectively, when considering as "positive" any of the HPV types in group A (33, 16 and 35), A or B (31, 52, 18, 59 and 58) and A or B or C (39, 51, 56, 45 and 68). The corresponding cross-sectional PPVs for CIN2+ were 15.8% 95% (CI 13.2-18.7), 12.0% (95% CI 10.3-13.9) and 9.6% (95% CI 8.2-11.1), respectively. HPV33, 16 and 35 confer a high probability of CIN2+ but this rapidly decreases when adding other genotypes.
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http://dx.doi.org/10.1002/ijc.31326DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6099271PMC
July 2018

Pre-diagnostic metabolite concentrations and prostate cancer risk in 1077 cases and 1077 matched controls in the European Prospective Investigation into Cancer and Nutrition.

BMC Med 2017 07 5;15(1):122. Epub 2017 Jul 5.

Cancer Epidemiology Unit, Nuffield Department of Population Health, University of Oxford, Oxford, OX3 7LF, UK.

Background: Little is known about how pre-diagnostic metabolites in blood relate to risk of prostate cancer. We aimed to investigate the prospective association between plasma metabolite concentrations and risk of prostate cancer overall, and by time to diagnosis and tumour characteristics, and risk of death from prostate cancer.

Methods: In a case-control study nested in the European Prospective Investigation into Cancer and Nutrition, pre-diagnostic plasma concentrations of 122 metabolites (including acylcarnitines, amino acids, biogenic amines, glycerophospholipids, hexose and sphingolipids) were measured using targeted mass spectrometry (AbsoluteIDQ p180 Kit) and compared between 1077 prostate cancer cases and 1077 matched controls. Risk of prostate cancer associated with metabolite concentrations was estimated by multi-variable conditional logistic regression, and multiple testing was accounted for by using a false discovery rate controlling procedure.

Results: Seven metabolite concentrations, i.e. acylcarnitine C18:1, amino acids citrulline and trans-4-hydroxyproline, glycerophospholipids PC aa C28:1, PC ae C30:0 and PC ae C30:2, and sphingolipid SM (OH) C14:1, were associated with prostate cancer (p < 0.05), but none of the associations were statistically significant after controlling for multiple testing. Citrulline was associated with a decreased risk of prostate cancer (odds ratio (OR) = 0.73; 95% confidence interval (CI) 0.62-0.86; p  = 0.0002) in the first 5 years of follow-up after taking multiple testing into account, but not after longer follow-up; results for other metabolites did not vary by time to diagnosis. After controlling for multiple testing, 12 glycerophospholipids were inversely associated with advanced stage disease, with risk reduction up to 46% per standard deviation increase in concentration (OR = 0.54; 95% CI 0.40-0.72; p  = 0.00004 for PC aa C40:3). Death from prostate cancer was associated with higher concentrations of acylcarnitine C3, amino acids methionine and trans-4-hydroxyproline, biogenic amine ADMA, hexose and sphingolipid SM (OH) C14:1 and lower concentration of glycerophospholipid PC aa C42:4.

Conclusions: Several metabolites, i.e. C18:1, citrulline, trans-4-hydroxyproline, three glycerophospholipids and SM (OH) C14:1, might be related to prostate cancer. Analyses by time to diagnosis indicated that citrulline may be a marker of subclinical prostate cancer, while other metabolites might be related to aetiology. Several glycerophospholipids were inversely related to advanced stage disease. More prospective data are needed to confirm these associations.
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http://dx.doi.org/10.1186/s12916-017-0885-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5497352PMC
July 2017

LINE-1 methylation status in prostate cancer and non-neoplastic tissue adjacent to tumor in association with mortality.

Epigenetics 2017 01 28;12(1):11-18. Epub 2016 Nov 28.

a Cancer Epidemiology Unit-CERMS , Department of Medical Sciences , University of Turin and CPO-Piemonte , Turin , Italy.

Aberrant DNA methylation seems to be associated with prostate cancer behavior. We investigated LINE-1 methylation in prostate cancer and non-neoplastic tissue adjacent to tumor (NTAT) in association with mortality from prostate cancer. We selected 157 prostate cancer patients with available NTAT from 2 cohorts of patients diagnosed between 1982-1988 and 1993-1996, followed up until 2010. An association between LINE-1 hypomethylation and prostate cancer mortality in tumor was suggested [hazard ratio per 5% decrease in LINE-1 methylation levels: 1.40, 95% confidence interval (CI): 0.95-2.01]. After stratification of the patients for Gleason score, the association was present only for those with a Gleason score of at least 8. Among these, low (<75%) vs. high (>80%) LINE-1 methylation was associated with a hazard ratio of 4.68 (95% CI: 1.03-21.34). LINE-1 methylation in the NTAT was not associated with prostate cancer mortality. Our results are consistent with the hypothesis that tumor tissue global hypomethylation may be a late event in prostate cancerogenesis and is associated with tumor progression.
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http://dx.doi.org/10.1080/15592294.2016.1261786DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5270633PMC
January 2017

Prevalence of Primary HPV in Djibouti: Feasibility of Screening for Early Diagnosis of Squamous Intraepithelial Lesions.

J Low Genit Tract Dis 2016 Oct;20(4):321-6

1National Institute for Health, Migration and Poverty (INMP), Rome, Italy; 2Epidemiology Unit, Local Health Authority-IRCCS Arcispedale Santa Maria Nuova, Reggio Emilia, Italy; 3Centre National de Référence en Santé de la Reproduction Housseina, Djibouti; 4Centro Unico Screening Cervico Vaginale-AO Città della Salute e della Scienza, Torino, Italy; and 5Molecular Epidemiology Lab-CERMS-University of Turin, Torino, Italy.

Objective: In many African Sub-Saharan countries, human papilloma virus (HPV) prevalence data are not available. The current study estimated the prevalence of HPV virus in the female population of Djibouti.

Methods: Approximately 1000 asymptomatic women 16 to 64 years old were enrolled from 3 of the main health structures of Djibouti in 2014 and 2015; 998 cervical samples were tested for HPV-DNA of high risk types, 499 during the first year, and 499 during the second. Positive samples were typed with an HPV genotyping kit.

Results: The women were an average age of 38.8 years (SD, 10.2); 54 women tested positive for HPV (prevalence rate, 5.4% [95% confidence interval, 4.0-6.8]). The highest prevalence was observed among the women younger than 35 years. HPV66 was the most prevalent (15.4% of the infections), followed by HPV31 and HPV52 (10.8% both) and HPV16 (9.2%). All 54 women who tested HPV-positive underwent a Pap test, which was positive in 8 cases (14.8%): 2 high-grade squamous intraepithelial lesion (HSIL) and 6 low-grade (LSIL).

Conclusions: The HPV prevalence shows a curve by age similar to that of other African countries. The proportion of HPV16 is among the lowest ever seen in similar studies. The findings suggest to Djibouti the choice of a strategy of screening that includes forms of cytological triage, thus limiting recourse to colposcopy.
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http://dx.doi.org/10.1097/LGT.0000000000000240DOI Listing
October 2016

Ki-67 proliferation index but not mitotic thresholds integrates the molecular prognostic stratification of lower grade gliomas.

Oncotarget 2016 Apr;7(16):21190-8

Department of Medical Sciences, University of Turin, Turin, Italy.

Despite several molecular signatures for "lower grade diffuse gliomas" (LGG) have been identified, WHO grade still remains a cornerstone of treatment guidelines. Mitotic count bears a crucial role in its definition, although limited by the poor reproducibility of standard Hematoxylin & Eosin (H&E) evaluation. Phospho-histone-H3 (PHH3) and Ki-67 have been proposed as alternative assays of cellular proliferation. Therefore in the present series of 141 LGG, the molecular characterization (namely IDH status, 1p/19q co-deletion and MGMT promoter methylation) was integrated with the tumor "proliferative trait" (conventional H&E or PHH3-guided mitotic count and Ki-67 index) in term of prognosis definition. Exclusively high PHH3 and Ki-67 values were predictor of poor prognosis (log rank test, P = 0.0281 for PHH3 and P = 0.032 for Ki-67), unlike standard mitotic count. Based on Cox proportional hazard regression analyses, among all clinical (age), pathological (PHH3 and Ki-67) and molecular variables (IDH, 1p/19q codeletion and MGMT methylation) with a prognostic relevance at univariate survival analysis, only IDH expression (P = 0.001) and Ki-67 proliferation index (P = 0.027) proved to be independent prognostic factors. In addition, stratifying by IDH expression status, high Ki-67 retained its prognostic relevance uniquely in the IDH negative patient (P = 0.029) doubling their risk of death (hazard ratio = 2.27). Overall, PHH3 immunostaining is the sole reliable method with a prognostic value to highlight mitotic figures in LGG. Ki-67 proliferation index exceeds PHH3 mitotic count as a predictor of patient's prognosis, and should be integrated with molecular markers in a comprehensive grading system for LGG.
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http://dx.doi.org/10.18632/oncotarget.8498DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5008278PMC
April 2016

Performance of Different Analytical Software Packages in Quantification of DNA Methylation by Pyrosequencing.

PLoS One 2016 2;11(3):e0150483. Epub 2016 Mar 2.

Cancer Epidemiology Unit - C.E.R.M.S, Department of Medical Sciences, University of Turin, Turin, Italy.

Background: Pyrosequencing has emerged as an alternative method of nucleic acid sequencing, well suited for many applications which aim to characterize single nucleotide polymorphisms, mutations, microbial types and CpG methylation in the target DNA. The commercially available pyrosequencing systems can harbor two different types of software which allow analysis in AQ or CpG mode, respectively, both widely employed for DNA methylation analysis.

Objective: Aim of the study was to assess the performance for DNA methylation analysis at CpG sites of the two pyrosequencing software which allow analysis in AQ or CpG mode, respectively. Despite CpG mode having been specifically generated for CpG methylation quantification, many investigations on this topic have been carried out with AQ mode. As proof of equivalent performance of the two software for this type of analysis is not available, the focus of this paper was to evaluate if the two modes currently used for CpG methylation assessment by pyrosequencing may give overlapping results.

Methods: We compared the performance of the two software in quantifying DNA methylation in the promoter of selected genes (GSTP1, MGMT, LINE-1) by testing two case series which include DNA from paraffin embedded prostate cancer tissues (PC study, N = 36) and DNA from blood fractions of healthy people (DD study, N = 28), respectively.

Results: We found discrepancy in the two pyrosequencing software-based quality assignment of DNA methylation assays. Compared to the software for analysis in the AQ mode, less permissive criteria are supported by the Pyro Q-CpG software, which enables analysis in CpG mode. CpG mode warns the operators about potential unsatisfactory performance of the assay and ensures a more accurate quantitative evaluation of DNA methylation at CpG sites.

Conclusion: The implementation of CpG mode is strongly advisable in order to improve the reliability of the methylation analysis results achievable by pyrosequencing.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0150483PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4775062PMC
July 2016

Prenatal exposure to antibiotics and wheezing in infancy: a birth cohort study.

Eur Respir J 2016 Mar 2;47(3):810-7. Epub 2015 Dec 2.

Dept of Medical Sciences, University of Turin, Turin, Italy CPO Piemonte, Turin, Italy AOU Città della Salute e della Scienza, Turin, Italy.

The role of prenatal antibiotic exposure in the development of childhood wheezing is debated. We evaluated whether this association could potentially be explained by confounding factors.Antibiotic use in the first and third trimester of pregnancy, wheezing in children aged ≤18 months and confounding factors were assessed in singletons participating in the NINFEA (Nascita e Infanzia: gli Effetti dell'Ambiente) birth cohort (n=3530 for first-trimester exposure and n=3985 for third-trimester exposure).There was no evidence of an association between antibiotic exposure in the first trimester of pregnancy and ever-wheezing (adjusted risk ratio (RR) 1.02, 95% CI 0.80-1.30) or recurrent wheezing (RR 0.99, 95% CI 0.54-1.82). For the third-trimester exposure, the crude RRs (95% CI) of ever-wheezing and recurrent wheezing were 1.34 (1.10-1.64) and 2.72 (1.80-4.11), respectively, which decreased to 1.12 (0.90-1.39) and 2.09 (1.32-3.29) after adjustment. The RRs of wheezing after genitourinary infections during pregnancy were increased independently of antibiotic treatment.In conclusion, the association between prenatal antibiotic exposure and infant wheezing could be largely explained by confounding factors, in particular respiratory infections during pregnancy. An excess risk of wheezing after antibiotic exposure during the third trimester of pregnancy remains after adjustment.
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http://dx.doi.org/10.1183/13993003.00315-2015DOI Listing
March 2016

Temozolomide as salvage treatment for recurrent intracranial ependymomas of the adult: a retrospective study.

Neuro Oncol 2016 Feb 30;18(2):261-8. Epub 2015 Aug 30.

Department of Neuro-Oncology, University and City of Health and Science Hospital, Torino, Italy (R.R., C.B., M.M., F.F., A.P., R.S.); Department of Clinical Epidemiology, University and City of Health and Science University Hospital, Torino, Italy (V.F., M.T.); Department of Biomedical Sciences and Oncology, University and City of Health and Science Hospital, Torino 10126, Italy (P.C.).

Background: Few data are available on temozolomide (TMZ) in ependymomas.We investigated the response, survival, and correlation with MGMT promoter methylation in a cohort of patients with adult intracranial ependymoma receiving TMZ as salvage therapy after failure of surgery and radiotherapy.

Patients And Methods: We retrieved clinical information from the institutional database and follow-up visits, and response to TMZ on MRI was evaluated according to the MacDonald criteria.

Results: Eighteen patients (median age, 42 y), with either WHO grade III (10) or grade II (8) ependymoma were evaluable. Tumor location at diagnosis was supratentorial in 11 patients and infratentorial in 7. Progression before TMZ was local in 11 patients, local and spinal in 6 patients, and spinal only in one patient. A median of 8 cycles of TMZ (1-24) was administered. Response to TMZ consisted of complete response (CR) in one (5%) patient, partial response (PR) in 3 (17%) patients, stable disease (SD) in 7 (39%) patients, and progressive disease (PD) in 7 (39%) patients. Maximum response occurred after 3, 10, 14, and 15 cycles, respectively, with neurological improvement in 2 patients. All 4 responding patients were chemotherapy naïve. Both anaplastic (2) and grade II (2) tumors responded. Median progression-free survival and overall survival were 9.69 months (95% CI, 3.22-30.98) and 30.55 months (95% CI, 12.85-52.17), respectively. MGMT methylation was available in 11 patients and was not correlated with response or outcome.

Conclusion: TMZ has a role in recurrent chemo-naïve adult patients with intracranial ependymoma, regardless of tumor grade and MGMT methylation. We suggest that, after failure of surgery and radiotherapy, TMZ should be considered as a possible first-line treatment for recurrent ependymoma.
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http://dx.doi.org/10.1093/neuonc/nov167DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4724181PMC
February 2016

Prenatal Paracetamol Exposure and Wheezing in Childhood: Causation or Confounding?

PLoS One 2015 25;10(8):e0135775. Epub 2015 Aug 25.

Cancer Epidemiology Unit, Department of Medical Sciences, AOU Città della Salute e della Scienza di Torino, CPO Piemonte, CERMS, and University of Turin, Turin, Italy.

Background: Several studies have reported an increased risk of wheezing in the children of mothers who used paracetamol during pregnancy. We evaluated to what extent this association is explained by confounding.

Methods: We investigated the association between maternal paracetamol use in the first and third trimester of pregnancy and ever wheezing or recurrent wheezing/asthma in infants in the NINFEA cohort study. Risks ratios (RR) and 95% confidence intervals (CI) were estimated after adjustment for confounders, including maternal infections and antibiotic use during pregnancy.

Results: The prevalence of maternal paracetamol use was 30.6% during the first and 36.7% during the third trimester of pregnancy. The prevalence of ever wheezing and recurrent wheezing/asthma was 16.9% and 5.6%, respectively. After full adjustment, the RR for ever wheezing decreased from 1.25 [1.07-1.47] to 1.10 [0.94-1.30] in the first, and from 1.26 [1.08-1.47] to 1.10 [0.93-1.29] in the third trimester. A similar pattern was observed for recurrent wheezing/asthma. Duration of maternal paracetamol use was not associated with either outcome. Further analyses on paracetamol use for three non-infectious disorders (sciatica, migraine, and headache) revealed no increased risk of wheezing in children.

Conclusion: The association between maternal paracetamol use during pregnancy and infant wheezing is mainly, if not completely explained by confounding.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0135775PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4549146PMC
May 2016

Seasonality modifies methylation profiles in healthy people.

PLoS One 2014 11;9(9):e106846. Epub 2014 Sep 11.

Unit of Cancer Epidemiology - CERMS, Department of Medical Sciences, University of Turin and Città della Salute e della Scienza Hospital, Turin, Italy.

DNA methylation is a well-characterized epigenetic modification that plays an important role in the regulation of gene expression. There is growing evidence on the involvement of epigenetic mechanisms in disease onset, including cancer. Environmental factors seem to induce changes in DNA methylation affecting human health. However, little is known about basal methylation levels in healthy people and about the correlation between environmental factors and different methylation profiles. We investigated the effect of seasonality on basal methylation by testing methylation levels in the long interspersed nucleotide element-1 (LINE-1) and in two cancer-related genes (RASSF1A and MGMT) of 88 healthy male heavy smokers involved in an Italian randomized study; at enrolment the subjects donated a blood sample collected in different months. Methylation analyses were performed by pyrosequencing. Mean methylation percentage was higher in spring and summer for the LINE1, RASSF1A and MGMT genes (68.26%, 2.35%, and 9.52% respectively) compared with autumn and winter (67.43%, 2.17%, and 8.60% respectively). In particular, LINE-1 was significantly hypomethylated (p = 0.04 or 0.05 depending on the CpG island involved) in autumn and winter compared with spring and summer. Seasonality seems to be a modifier of methylation levels and this observation should be taken into account in future analyses.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0106846PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4161384PMC
May 2015

Age and geographic variability of human papillomavirus high-risk genotype distribution in a large unvaccinated population and of vaccination impact on HPV prevalence.

J Clin Virol 2014 Jul 18;60(3):257-63. Epub 2014 Apr 18.

Centre for Cancer Prevention, Via S. Francesco da Paola 31, 10123 Turin, Italy. Electronic address:

Background: The prevalence of infections with human papillomavirus (HPV) specific genotypes differs by age and areas. Knowledge of these differences will help predicting how prophylactic HPV vaccination and screening program could best be integrated.

Objectives: To investigate variations in the HPV distribution between areas and ages in Italy and the impact of vaccination on HPV prevalence.

Study Design: 37,367 women aged 25-60 years who attended cervical screening in eight different areas in Northern and Central Italy were tested for HPV infection with the high-risk hybrid capture (hr-HC2) assay. hr-HC2 positive samples were genotyped by an intensive integrated strategy.

Results: hr-HPV types were detected in 79.1% of HC2 positive women. HPV16 was the most frequent type, followed by HPV31, HPV18 and HPV56. A statistically significant variability in HPV type distribution between centres (overall χ84df(2)=195.86p<0.001) was observed. No significant overall difference in the HPV type distribution was observed in the age groups 25-34, 35-44 and 45-60 years. Considering cross-protection, overall 57.6% (95%CI 56.0-59.3) of all infections by hr-HPV types was preventable by vaccination with the bivalent vaccine and 49% (95%CI 46.9-51.1) with the quadrivalent vaccine. The variability between centres was statistically significant with both bivalent (χ7df(2)=43.8, p<0.0001) and quadrivalent vaccine (χ7df(2)=32.9, p<0.0001).

Conclusions: We observed differences in HPV genotype distribution according to centres but not to age. Results suggest that the higher proportion of HPV16/18 related high grade CIN in younger women could be the result of faster progression and not of earlier infection by these types.
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http://dx.doi.org/10.1016/j.jcv.2014.04.009DOI Listing
July 2014

MGMT promoter methylation in plasma of glioma patients receiving temozolomide.

J Neurooncol 2014 Apr 12;117(2):347-57. Epub 2014 Feb 12.

Unit of Cancer Epidemiology - CERMS, Department of Medical Sciences, University of Turin, Città della Salute e della Scienza Hospital, Via Santena 7, 10126, Turin, Italy,

Promoter methylation of the O6-methylguanine-DNA methyltransferase (MGMT) gene plays a role in cellular response to alkylating agents. In the present study aimed to: (i) evaluate the concordance between MGMT promoter methylation status in tumor tissue and plasma; (ii) monitor MGMT promoter methylation status in plasma taken before and during temozolomide treatment; (iii) explore the value of MGMT promoter methylation status in plasma as a prognostic/predictive biomarker in glioma patients. We enrolled 58 patients with histologically confirmed glioma at different grades of malignancy. All patients underwent surgical resection and temozolomide treatment. Paraffin-embedded tumor tissue was available for 48 patients. Blood samples were collected from all patients before temozolomide treatment (baseline) and at each MRI examination for a 12-month period. MGMT promoter methylation status was assessed in both sample types by real time PCR with a specific probe. The frequency of MGMT promoter methylation was 60.4 % in tumor tissue and 41.38 % in plasma. MGMT promoter methylation status was concordant in the two sample types (Kappa = 0.75, 95 % confidence interval (CI) 0.57-0.93; p value <0.001). Overall and progression-free survival were longer in patients with methylated MGMT promoter. Mortality was higher in patients with unmethylated MGMT promoter, whether in tumor tissue [hazard ratio (HR) 2.21; 95 % CI 0.99-4.95] or plasma (HR 2.19; 95 % CI 1.02-4.68). Progression-free survival was shorter in patients with unmethylated MGMT promoter, whether in tissue (HR 2.30; 95 % CI 1.19-4.45) or plasma (HR 1.77; 95 % CI 0.95-3.30). The cumulative incidence of unmethylated MGMT promoter in plasma at baseline was 58 %, and reached virtually 100 % at 12 months. In conclusion MGMT promoter methylation status in tumor tissue and plasma was highly concordant, and both were associated with longer survival, supporting the role of the detection of methylated MGMT promoter in predicting treatment response. However we suggest caution in using plasma as a surrogate of tumor tissue due to possible false-negative results.
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http://dx.doi.org/10.1007/s11060-014-1395-4DOI Listing
April 2014

Piccolipiù, a multicenter birth cohort in Italy: protocol of the study.

BMC Pediatr 2014 Feb 7;14:36. Epub 2014 Feb 7.

Background: The fetal and infant life are periods of rapid development, characterized by high susceptibility to exposures. Birth cohorts provide unique opportunities to study early-life exposures in association with child development and health, as well as, with longer follow-up, the early life origin of adult diseases. Piccolipiù is an Italian birth cohort recently set up to investigate the effects of environmental exposures, parental conditions and social factors acting during pre-natal and early post-natal life on infant and child health and development. We describe here its main characteristics.

Methods/design: Piccolipiù is a prospective cohort of expected 3000 newborns, who will be recruiting in six maternity units of five Italian cities (Florence, Rome, Trieste, Turin and Viareggio) since October 2011. Mothers are contacted during pregnancy or at delivery and are offered to participate in the study. Upon acceptance, their newborns are recruited at birth and followed up until at least 18 years of age. At recruitment, the mothers donate a blood sample and complete a baseline questionnaire. Umbilical cord blood, pieces of umbilical cord and heel blood spots are also collected. Postnatal follow-up currently occurs at 6, 12, and 24 months of age using on-line or postal self administered questionnaire; further questionnaires and medical examinations are envisaged. Questionnaires collect information on several factors, including mother's and/or child's environmental exposures, anthropometric measures, reproductive factors, diet, supplements, medical history, cognitive development, mental health and socioeconomic factors. Health promotion materials are also offered to parents.

Discussion: Piccolipiù will broaden our understanding of the contribution of early-life factors to infant and child health and development. Several hypotheses on the developmental origins of health can be tested or piloted using the data collected from the Piccolipiù cohort. By pooling these data with those collected by other existing birth cohorts it will be possible to validate previous findings and to study rare exposures and outcomes.
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http://dx.doi.org/10.1186/1471-2431-14-36DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3926689PMC
February 2014

Case-control study of HLA-G promoter methylation status, HPV infection and cervical neoplasia in Curitiba, Brazil: a pilot analysis.

BMC Cancer 2012 Dec 24;12:618. Epub 2012 Dec 24.

Department of Medical Sciences, Unit of Cancer Epidemiology - C,E,R,M,S, University of Turin, Turin, Italy.

Background: The causal association between persistent human papillomavirus (HPV) infection and cervical cancer has been established, but the mechanisms that favor HPV persistence in cervical cells are still unknown. The diminished capability of the immune system to control and resolve HPV infection is one of several hypotheses. The tolerogenic protein HLA-G has shown aberrant expression in a variety of cancers, which has been suggested as a mechanism for tumor escape from immunosurveillance. In the present study we evaluate the role of epigenetic modification (promoter de-methylation) of the HLA-G gene on susceptibility to HPV infection and development of high-grade cervical lesions.

Methods: A case-control study was carried out in Curitiba, Brazil, between February and June 2010. A total of 789 women aged 15-47 years were recruited: 510 controls with normal cervical cytology, and 279 cases with histologically confirmed cervical intraepithelial neoplasia grade 2 (CIN2, N = 150) or grade 3 (CIN3, N = 129). All women were administered a questionnaire by interview, which collected information on demographic and lifestyle factors, and a cervical sample was collected. HPV DNA detection was performed by GP5+/GP6+ primer-mediated PCR. HPV-positive samples were genotyped by multiplex PCR. A pilot analysis of HLA-G promoter methylation was carried out in a subset of the study population (96 cases and 76 controls) by pyrosequencing. HLA-G methylation and HPV infection status of cases and controls were compared, and confounding factors were computed by t Student and non-parametric Wilcoxon tests. Comparison of HLA-G methylation between cases and controls was assessed by the Bonferroni correction. The association of HLA-G methylation with CIN2/3 was evaluated by logistic regression.

Results: HPV prevalence was 19.6% in controls and 94.3% in CIN2/3 cases. HPV16, 31, 33, 35 and 18 were the most prevalent types. Methylation analysis of seven CpGs in the HLA-G promoter did not reveal any spontaneous de-methylation events in CIN2/3 cases (mean proportion of methylation: 75.8%) with respect to controls (mean 73.7%; odds ratio 1.01, 95% confidence interval 0.96, 1.07).

Conclusions: This study did not support the hypothesis that spontaneous de-methylation events in the HLA-G promoter play a primary role in promoting escape from immunosurveillance in the development of precancerous cervical lesions.
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http://dx.doi.org/10.1186/1471-2407-12-618DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3545901PMC
December 2012

Prevalence and follow-up of occult HCV infection in an Italian population free of clinically detectable infectious liver disease.

PLoS One 2012 22;7(8):e43541. Epub 2012 Aug 22.

Unit of Cancer Epidemiology, C.E.R.M.S., University of Turin, Turin, Italy.

Background: Occult hepatitis C virus infection (OCI) is a recently described phenomenon characterized by undetectable levels of HCV-RNA in serum/plasma by current laboratory assays, with identifiable levels in peripheral blood mononuclear cells (PBMCs) and/or liver tissue by molecular tests with enhanced sensitivity. Previous results from our group showed an OCI prevalence of 3.3% in a population unselected for hepatic disease. The present study aimed to evaluate OCI prevalence in a larger cohort of infectious liver disease-free (ILDF) subjects. Clinical follow-up of OCI subjects was performed to investigate the natural history of the infection.

Methods And Findings: 439 subjects referred to a Turin Blood Bank for phlebotomy therapy were recruited. They included 314 ILDF subjects, 40 HCV-positive subjects and 85 HBV-positive subjects, of whom 7 were active HBV carriers. Six subjects (4/314 ILDF subjects [1.27%] and 2/7 active HBV carriers [28%]) were positive for HCV-RNA in PBMCs, but negative for serological and virological markers of HCV, indicating OCI. HCV genotypes were determined in the PBMCs of 3/6 OCI subjects two had type 1b; the other had type 2a/2c. OCI subjects were followed up for at least 2 years. After 12 months only one OCI persisted, showing a low HCV viral load (3.73×10(1) UI/ml). By the end of follow-up all OCI subjects were negative for HCV. No seroconversion, alteration of liver enzyme levels, or reduction of liver synthesis occurred during follow-up.

Conclusions: This study demonstrated the existence of OCI in ILDF subjects, and suggested a high OCI prevalence among active HBV carriers. Follow-up suggested that OCI could be transient, with a trend toward the decrease of HCV viral load to levels undetectable by conventional methods after 12-18 months. Confirmation studies with a longer follow-up period are needed for identification of the OCI clearance or recurrence rates, and to characterize the viruses involved.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0043541PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3425488PMC
January 2013
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