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SARS-CoV-2 infection spawns from an asymptomatic condition to a fatal disease. Age, comorbidities, and several blood biomarkers are associated with infection outcome. We searched for biomarkers by untargeted and targeted proteomic analysis of saliva, a source of viral particles and host proteins.Saliva samples from 19 asymptomatic and 16 symptomatic SARS-CoV-2 infected subjects, and 20 controls were analyzed by LC-MS/MS for untargeted peptidomic (flow through of 10 kDa filter) and proteomic (trypsin digestion of filter retained proteins) profiling.Peptides from 53 salivary proteins were identified. ADF was detected only in controls, while IL1RA only in infected subjects. PRPs, DSC2, FABP5, his-1, IL1RA, PRH1, STATH, SMR3B, ANXA1, MUC7, ACTN4, IGKV1-33 and TGM3 were significantly different between asymptomatic and symptomatic subjects. Retained proteins were 117, being 11 highly different between asymptomatic and symptomatic (fold change ≥2 or ≤-2). After validation by LC-MS/MS-SRM (selected reaction monitoring analysis), the most significant discriminant proteins at PCA were IL1RA, CYSTB, S100A8, S100A9, CA6, and FABP5.The differentially abundant proteins involved in innate immunity (S100 proteins), taste (CA6 and cystatins), and viral binding to the host (FABP5), appear to be of interest for use as potential biomarkers and drugs targets.

OBJECTIVE Cardiovascular disease (CVD) accounts for most deaths in patients with type 1 diabetes (T1D); however, the determinants of plaque composition are unknown. MicroRNAs (miRNAs) regulate gene expression, participate in the development of atherosclerosis and represent promising CVD biomarkers. This study analyzed the circulating miRNA expression profile in T1D with carotid either calcified (CCP) or fibrous (CFP) plaque. RESEARCH DESIGN AND METHODS Circulating small non-coding RNA were sequenced and quantified using NGS and bioinformatic analysis in an exploratory set of 26 T1D with CCP and in 25 with CFP. Then, in a validation set of 40 CCP, 40 CFP and 24 controls T1D, selected miRNAs expression was measured by digital droplet PCR. Putative gene targets enriched for pathways implicated in atherosclerosis/vascular calcification/diabetes were analyzed. Patient’s main clinical characteristics were also recorded. RESULTS miR-503-5p, let-7d-5p, miR-106b-3p, miR-93-5p were significantly up-regulated, while miR-10a-5p downregulated in CCP patients compared to CFP (all FC>±1.5, p CONCLUSIONS These findings characterize miRNAs and their signature in the regulatory network of carotid plaque phenotype, in T1D, providing new insights into plaque pathophysiology, and possibly novel biomarkers of plaque composition.

The CYP4F2 gene is known to influence mean coumarin dose. The aim of the present study was to undertake a meta-analysis at individual patients' level to capture the possible effect of ethnicity, gene-gene interaction or other drugs on the association and to verify if inclusion of CYP4F2*3 variant into dosing algorithms improves the prediction of mean coumarin dose. We asked the authors of our previous meta-analysis (30 articles) and of 38 new articles retrieved by a systematic review to send us individual patients' data. The final collection consists 15,754 patients split into a derivation and validation cohort. The CYP4F2*3 polymorphism was consistently associated with an increase in mean coumarin dose (+9% (95%CI 7-10%), with a higher effect in females, in patients taking acenocoumarol and in Whites. The inclusion of the CYP4F2*3 in dosing algorithms slightly improved the prediction of stable coumarin dose. New pharmacogenetic equations potentially useful for clinical practice were derived. This article is protected by copyright. All rights reserved.

BackgroundSARS-CoV-2 quick testing and reporting are now considered relevant for the containment of new pandemic waves. Antigen testing in self-collected saliva might be useful. We compared the diagnostic performance of salivary and naso-pharyngeal swab (NPS) SARS-CoV-2 antigen detection by a rapid chemiluminescent assay (CLEIA) and two different point-of-care (POC) immunochromatographic assays, with that of molecular testing.Methods234 patients were prospectively enrolled. Paired self-collected saliva (Salivette) and NPS were obtained to perform rRT-PCR, chemiluminescent (Lumipulse G) and POC (NPS: Fujirebio and Abbott; saliva: Fujirebio) for SARS-CoV-2 antigen detection.ResultsThe overall agreement between NPS and saliva rRT-PCR was 78.7%, reaching 91.7% at the first week from symptoms onset. SARS-CoV-2 CLEIA antigen was highly accurate in distinguishing between positive and negative NPS (ROC-AUC=0.939, 95%CI:0.903-0.977), with 81.6% sensitivity and 93.8% specificity. This assay on saliva had an overall good accuracy (ROC-AUC=0.805, 95%CI:0.740-0.870), reaching the optimal value within 7 days from symptom onset (Sensitivity: 72%; Specificity: 97%). POC antigen in saliva had a very limited sensitivity (13%), performing better in NPS (Sensitivity: 48% and 66%; Specificity: 100% and 99% for Espline and Abbott respectively), depending on viral loads.ConclusionsSelf-collected saliva is a valid alternative to NPS for SARS-CoV-2 detection not only by molecular, but also by CLEIA antigen testing, for which the highest diagnostic accuracy was achieved in the first week from symptom onset. Saliva is not suitable for POC, although the accuracy of these tests appears satisfactory for NPS with high viral load.

Objectives The direct identification of SARS-CoV-2 RNA in nasopharyngeal swabs is recommended for diagnosing the novel COVID-19 disease. Pre-analytical determinants, such as sampling procedures, time and temperature storage conditions, might impact on the end result. Our aim was to evaluate the effects of sampling procedures, time and temperature of the primary nasopharyngeal swabs storage on real-time reverse-transcription polymerase chain reaction (rRT-PCR) results. Methods Each nasopharyngeal swab obtained from 10 hospitalized patients for COVID-19 was subdivided in 15 aliquots: five were kept at room temperature; five were refrigerated (+4 °C); five were immediately mixed with the extraction buffer and refrigerated at +4 °C. Every day and for 5 days, one aliquot per condition was analyzed (rRT-PCR) for SARS-CoV-2 gene E and RNaseP and threshold cycles (Ct) compared. To evaluate manual sampling, 70 nasopharyngeal swabs were sampled twice by two different operators and analyzed separately one from the other. Results A total of 6/10 swabs were SARS-CoV-2 positive. No significant time or storage-dependent variations were observed in SARS-CoV-2 Ct. Re-sampling of swabs with SARS-CoV-2 Ct lower than 33 resulted in highly reproducible results (CV=2.9%), while a high variability was observed when Ct values were higher than 33 (CV=10.3%). Conclusions This study demonstrates that time and temperature of nasopharyngeal swabs storage do not significantly impact on results reproducibility. However, swabs sampling is a critical step, and especially in case of low viral load, might be a potential source of diagnostic errors.

Telomere deregulation is a hallmark of cancer. Telomere length measured in lymphocytes (LTL) has been shown to be a risk marker for several cancers. For pancreatic ductal adenocarcinoma (PDAC) consensus is lacking whether risk is associated with long or short telomeres. Mendelian randomization approaches have shown that a score built from SNPs associated with LTL could be used as a robust risk marker. We explored this approach in a large scale study within the PANcreatic Disease ReseArch (PANDoRA) consortium. We analyzed 10 SNPs (ZNF676-rs409627, TERT-rs2736100, CTC1-rs3027234, DHX35-rs6028466, PXK-rs6772228, NAF1-rs7675998, ZNF208-rs8105767, OBFC1-rs9420907, ACYP2-rs11125529 and TERC-rs10936599) alone and combined in a LTL genetic score ("teloscore", which explains 2.2% of the telomere variability) in relation to PDAC risk in 2,374 cases and 4,326 controls. We identified several associations with PDAC risk, among which the strongest were with the TERT-rs2736100 SNP (OR = 1.54; 95%CI 1.35-1.76; p = 1.54 × 10-10 ) and a novel one with the NAF1-rs7675998 SNP (OR = 0.80; 95%CI 0.73-0.88; p = 1.87 × 10-6 , ptrend = 3.27 × 10-7 ). The association of short LTL, measured by the teloscore, with PDAC risk reached genome-wide significance (p = 2.98 × 10-9 for highest vs. lowest quintile; p = 1.82 × 10-10 as a continuous variable). In conclusion, we present a novel genome-wide candidate SNP for PDAC risk (TERT-rs2736100), a completely new signal (NAF1-rs7675998) approaching genome-wide significance and we report a strong association between the teloscore and risk of pancreatic cancer, suggesting that telomeres are a potential risk factor for pancreatic cancer.

A previous trial failed to demonstrate the superiority of a demographic-genetic algorithm in predicting warfarin (W) dose over a standard clinical approach. The purpose of the present study is to re-analyse the results in subgroups of patients with differing baseline sensitivity to W, integrated with additional pharmacokinetic data. The original trial allocated 180 treatment-naive patients with non-valvular atrial fibrillation to a control arm (CTL, n = 92) or a genetic-guided arm (GEN, n = 88). Before starting anticoagulation treatment, all patients were genotyped for CYP2C9, VKORC1 and CYP4F2 variants and classified into four quartiles (Q1, Q2, Q3, Q4) according to the algorithm-predicted W maintenance dose. International normalised ratios (INR) and plasma concentrations of S-warfarin [S-W]s and R-warfarin [R-W]s were measured at baseline and on days 5, 7, 9, 12, 15 and 19 of therapy. In the lowest dose quartile (Q1), the number of INRs > 3 and mean INR values on days 5 and 7 were significantly higher in CTL than in GEN. In Q3 and Q4, the mean INR values reached therapeutic level (> 2) 2 days later in CTL than in GEN. During follow-up, the mean time courses of INRs and [S-W]s in GEN were remarkably stable in all dose quartiles. Thus, mean changes from starting to final doses were significantly smaller in GEN than in CTL. Plasma concentrations of R-W (a partially active enantiomer) steadily increased from day 5 to day 19 in all Qs in both CTL and GEN, except in the Q1 CTL group, due to the marked dose reduction required. This analysis showed that the demographic-genetic algorithm used to predict the W dose can identify patients with differing degrees of sensitivity to W and to ‘normalise’ their average anticoagulant responses. The progressive rise in [R-W]s throughout the 19-day follow-up indicates that the (partial) contribution of R-W to the W anticoagulant effect changes continually during the early phase of treatment.

Pancreatic ductal adenocarcinoma (PDAC) is a very aggressive tumor with a five-year survival of less than 6%. Chronic pancreatitis (CP), an inflammatory process in of the pancreas, is a strong risk factor for PDAC. Several genetic polymorphisms have been discovered as susceptibility loci for both CP and PDAC. Since CP and PDAC share a consistent number of epidemiologic risk factors, the aim of this study was to investigate whether specific CP risk loci also contribute to PDAC susceptibility. We selected five common SNPs (rs11988997, rs379742, rs10273639, rs2995271 and rs12688220) that were identified as susceptibility markers for CP and analyzed them in 2,914 PDAC cases, 356 CP cases and 5,596 controls retrospectively collected in the context of the international PANDoRA consortium. We found a weak association between the minor allele of the PRSS1-PRSS2-rs10273639 and an increased risk of developing PDAC (ORhomozygous = 1.19, 95% CI 1.02-1.38, p = 0.023). Additionally all the SNPs confirmed statistically significant associations with risk of developing CP, the strongest being PRSS1-PRSS2-rs10273639 (ORheterozygous = 0.51, 95% CI 0.39-0.67, p = 1.10 × 10-6 ) and MORC4-rs 12837024 (ORhomozygous = 2.07 (1.55-2.77, ptrend = 0.7 × 10-11 ). Taken together, the results from our study do not support variants rs11988997, rs379742, rs10273639, rs2995271 and rs12688220 as strong predictors of PDAC risk, but further support the role of these SNPs in CP susceptibility. Our study suggests that CP and PDAC probably do not share genetic susceptibility, at least in terms of high frequency variants.

// Daniela Basso 1 , Elisa Gnatta 1 , Andrea Padoan 1 , Paola Fogar 1 , Sara Furlanello 1 , Ada Aita 1 , Dania Bozzato 1 , Carlo-Federico Zambon 2 , Giorgio Arrigoni 2,3 , Chiara Frasson 4 , Cinzia Franchin 2,3 , Stefania Moz 1 , Thomas Brefort 5,6 , Thomas Laufer 6 , Filippo Navaglia 1 , Sergio Pedrazzoli 7 , Giuseppe Basso 4 and Mario Plebani 1 1 Department of Medicine – DIMED, University of Padova, Padova, Italy 2 Department of Biomedical Sciences, University of Padova, Padova, Italy 3 Proteomic Center, University of Padova, Padova, Italy 4 Department of Woman and Child Health, Oncohematology Laboratory, University of Padova, Padova, Italy 5 Eurofins Medigenomix GmbH, Ebersberg, Germany 6 Comprehensive Biomarker Center GmbH (Recently re-named to Hummingbird Diagnostics GmbH), Heidelberg, Germany 7 Association Wirsung Onlus, Padova, Italy Correspondence to: Daniela Basso, email: // Keywords : pancreatic cancer, exosomes, myeloid derived suppressor cells, calcium, SMAD4 Received : March 03, 2017 Accepted : August 04, 2017 Published : September 13, 2017 Abstract Tumor genetics and escape from immune surveillance concur in the poor prognosis of PDAC. In this study an experimental model was set up to verify whether SMAD4 , deleted in about 55% PDAC and associated with poor prognosis, is involved in determining immunosuppression through Exosomes (Exo). Potential mechanisms and mediators underlying SMAD4 -dependent immunosuppression were evaluated by studying intracellular calcium (Fluo-4), Exo-miRNAs (microarray) and Exo-proteins (SILAC). Two PDAC cell lines expressing (BxPC3- SMAD4 +) or not-expressing (BxPC3) SMAD4 were used to prepare Exo-enriched conditioned media, employed in experiments with blood donors PBMCs. Exo expanded myeloid derived suppressor cells (gMDSC and mMDSC, flow cytometry) and altered intracellular calcium fluxes in an SMAD4 dependent manner. BxPC3- SMAD4 +, but mainly BxPC3 Exo, increased calcium fluxes of PBMCs ( p = 0.007) and this increased intracellular calcium trafficking characterized mMDSCs. The analysis of de-regulated Exo-miRNAs and transfection experiments revealed hsa-miR-494-3p and has-miR-1260a as potential mediators of SMAD4- associated de-regulated calcium fluxes. Eleven main biological processes were identified by the analysis of SMAD4 -associated de-regulated Exo-proteins, including translation, cell adhesion, cell signaling and glycolysis. A reverse Warburg effect was observed by treating PBMCs with PDAC-derived Exo: BxPC3 Exo induced a higher glucose consumption and lactate production than BxPC3- SMAD4 + Exo. Conclusion: PDAC-derived Exo from cells with , but mainly from those without SMAD4 expression, create an immunosuppressive myeloid cell background by increasing calcium fluxes and glycolysis through the transfer of SMAD4 -related differentially expressed miRNAs and proteins.

Disease-independent sources of biomarker variability include pre-analytical, analytical and biological variance. The aim of the present study was to evaluate whether the pre-analytical phase has any impact on the emerging heart disease TWEAK and HMGB1 protein markers and miRNA biomarkers, and whether peptidome profiling allows the identification of pre-analytical quality markers.An assessment was made of sample type (serum, EDTA-Plasma, Citrate-Plasma, ACD-plasma, Heparin-plasma), temperature of sample storage (room temperature or refrigerated), time of sample storage (0.5, 3, 6 and 9h) and centrifugation (one or two-step). Aliquots of all processed samples were immediately frozen (-80°C) before analysis. Proteins were assayed by ELISAs, miRNA expression profile by microarray and peptidome profiling by MALDI-TOF/MS.Temperature, time and centrifugation had no impact on TWEAK and HMGB1 results, which were significantly influenced by matrix type, TWEAK levels being significantly higher (F=194.7, p0.0001), and HMGB1 levels significantly lower (F=36.32, p0.0001) in serum than in any other plasma type. Unsuitable miRNA results were obtained using Heparin-plasma. Serum miRNA expression profiles depended mainly on temperature, while EDTA-plasma miRNA expression profiles were strongly affected by the centrifugation method used. MALDI-TOF/MS allowed the identification of seven features as indices of pre-analytical serum (m/z at 1206, 1350, 1865 and 2021) or EDTA-plasma (m/z 1897, 2740 and 2917) degradation.Serum and EDTA-plasma allow the analysis of both proteins and miRNA emerging biomarkers of heart diseases. Refrigerated storage prevents an altered miRNA expression profile also in cases of a prolonged time-interval between blood drawing and processing.

Background: Pancreatic neuroendocrine tumors (PNETs) are rare neoplasms for which very little is known about either environmental or genetic risk factors. Only a handful of association studies have been performed so far, suggesting a small number of risk loci. Methods: To replicate the best findings, we have selected 16 SNPs suggested in previous studies to be relevant in PNET etiogenesis. We genotyped the selected SNPs (rs16944, rs1052536, rs1059293, rs1136410, rs1143634, rs2069762, rs2236302, rs2387632, rs3212961, rs3734299, rs3803258, rs4962081, rs7234941, rs7243091, rs12957119, and rs1800629) in 344 PNET sporadic cases and 2,721 controls in the context of the PANcreatic Disease ReseArch (PANDoRA) consortium. Results: After correction for multiple testing, we did not observe any statistically significant association between the SNPs and PNET risk. We also used three online bioinformatic tools (HaploReg, RegulomeDB, and GTEx) to predict a possible functional role of the SNPs, but we did not observe any clear indication. Conclusions: None of the selected SNPs were convincingly associated with PNET risk in the PANDoRA consortium. Impact: We can exclude a major role of the selected polymorphisms in PNET etiology, and this highlights the need for replication of epidemiologic findings in independent populations, especially in rare diseases such as PNETs. Cancer Epidemiol Biomarkers Prev; 26(8); 1349–51. ©2017 AACR.

Telomere deregulation is a hallmark of cancer. Telomere length measured in lymphocytes (LTL) has been shown to be a risk marker for several cancers. For pancreatic ductal adenocarcinoma (PDAC) consensus is lacking whether risk is associated with long or short telomeres. Mendelian randomization approaches have shown that a score built from SNPs associated with LTL could be used as a robust risk marker. We explored this approach in a large scale study within the PANcreatic Disease ReseArch (PANDoRA) consortium. We analyzed 10 SNPs (ZNF676-rs409627, TERT-rs2736100, CTC1-rs3027234, DHX35-rs6028466, PXK-rs6772228, NAF1-rs7675998, ZNF208-rs8105767, OBFC1-rs9420907, ACYP2-rs11125529 and TERC-rs10936599) alone and combined in a LTL genetic score ("teloscore", which explains 2.2% of the telomere variability) in relation to PDAC risk in 2,374 cases and 4,326 controls. We identified several associations with PDAC risk, among which the strongest were with the TERT-rs2736100 SNP (OR = 1.54; 95%CI 1.35-1.76; p = 1.54x10-10 ) and a novel one with the NAF1-rs7675998 SNP (OR = 0.80; 95%CI 0.73-0.88; p = 1.87x10-6 , ptrend = 3.27x10-7 ). The association of short LTL, measured by the teloscore, with PDAC risk reached genome-wide significance (p = 2.98x10-9 for highest vs. lowest quintile; p = 1.82x10-10 as a continuous variable). In conclusion, we present a novel genome-wide candidate SNP for PDAC risk (TERT-rs2736100), a completely new signal (NAF1-rs7675998) approaching genome-wide significance and we report a strong association between the teloscore and risk of pancreatic cancer, suggesting that telomeres are a potential risk factor for pancreatic cancer. This article is protected by copyright. All rights reserved.

Pancreatic neuroendocrine neoplasms (pNEN) account for less than 5% of all pancreatic neoplasms and genetic association studies on susceptibility to the disease are limited. We sought to identify possible overlap of genetic susceptibility loci between pancreatic ductal adenocarcinoma (PDAC) and pNEN; therefore, PDAC susceptibility variants (n = 23) from Caucasian genome-wide association studies (GWAS) were genotyped in 369 pNEN cases and 3277 controls from the PANcreatic Disease ReseArch (PANDoRA) consortium to evaluate the odds associated with pNEN risk, disease onset and tumor characteristics. Main effect analyses showed four PDAC susceptibility variants-rs9854771, rs1561927, rs9543325 and rs10919791 to be associated with pNEN risk. Subsequently, only associations with rs9543325, rs10919791 and rs1561927 were noteworthy with false positive report probability (FPRP) tests. Stratified analyses considering age at onset (50-year threshold), showed rs2736098, rs16986825 and rs9854771 to be associated with risk of developing pNEN at a younger age. Stratified analyses also showed some single nucleotide polymorphisms to be associated with different degrees of tumor grade, metastatic potential and functionality. Our results identify known GWAS PDAC susceptibility loci, which may also be involved in sporadic pNEN etiology and suggest that some genetic mechanisms governing pathogenesis of these two entities may be similar, with few of these loci being more influential in younger cases or tumor subtypes.

Additional file 2: Intermediate data results. Intermediate results generated step-by-step, following the manuscript details.

Background: The appropriate clinical use of fecal calprotectin (fCal) might be compromised by incomplete harmonization between assays and within- and between-subjects variability. Our aim was to investigate the analytical and biological variability of fCal in order to provide tools for interpreting fCal in the clinical setting. Methods: Experiments were conducted to investigate the effects of temperature and storage time on fCal. Thirty-nine controls were enrolled to verify biological variability, and a case-control study was conducted on 134 controls and 110 IBD patients to compare the clinical effectiveness of three different fCal assays: ELISA, CLIA and turbidimetry. Results: A 12% decline in fCal levels was observed within 24 h following stool collection irrespective of storage temperature. Samples were unstable following a longer storage time interval at room temperature. Within- and between-subjects fCal biological variability, at 31% and 72% respectively, resulted in a reference change value (RCV) in the region of 100%. fCal sensitivity in distinguishing between controls and IBD patients is satisfactory (68%), and the specificity high (93%) among young ( Conclusions: We recommend a standardized preanalytical protocol for fCal, avoiding storage at room temperature for more than 24 h. Different cutoffs are recommended for different fCal assays. In monitoring, the difference between two consecutive measurements appears clinically significant when higher than 100%, the fCal biological variability-derived RCV.

A small number of common susceptibility loci have been identified for pancreatic cancer, one of which is marked by rs401681 in the TERT-CLPTM1L gene region on chromosome 5p15.33. Because this region is characterized by low linkage disequilibrium, we sought to identify whether additional single nucleotide polymorphisms (SNPs) could be related to pancreatic cancer risk, independently of rs401681. We performed an in-depth analysis of genetic variability of the telomerase reverse transcriptase (TERT) and the telomerase RNA component (TERC) genes, in 5,550 subjects with pancreatic cancer and 7,585 controls from the PANcreatic Disease ReseArch (PANDoRA) and the PanScan consortia. We identified a significant association between a variant in TERT and pancreatic cancer risk (rs2853677, odds ratio=0.85; 95% confidence interval=0.80-0.90, p=8.3 × 10-8). Additional analysis adjusting rs2853677 for rs401681 indicated that the two SNPs are independently associated with pancreatic cancer risk, as suggested by the low linkage disequilibrium between them (r2=0.07, D′=0.28). Three additional SNPs in TERT reached statistical significance after correction for multiple testing: rs2736100 (p=3.0 × 10-5), rs4583925 (p=4.0 × 10-5) and rs2735948 (p=5.0 × 10-5). In conclusion, we confirmed that the TERT locus is associated with pancreatic cancer risk, possibly through several independent variants. What's new? Most pancreatic cancer patients do not survive long after diagnosis, and, so far, there are not many genetic markers to help screen for the disease. In search of genetic predictors of pancreatic cancer, the authors zoomed in on a region linked to susceptibility to the disease. They measured the frequency of different variants of two genes, telomerase reverse transcriptase and telomerase RNA component, among thousands of pancreatic cancer patients and controls. They identified several variants of the TERT gene that indicate a boosted pancreatic cancer risk, and which may develop into useful prognostic tools.

Expression of the solute carrier (SLC) transporter SLC22A3 gene is associated with overall survival of pancreatic cancer patients. This study tested whether genetic variability in SLC22A3 associates with pancreatic cancer risk and prognosis. Twenty four single nucleotide polymorphisms (SNPs) tagging the SLC22A3 gene sequence and regulatory elements were selected for analysis. Of these, 22 were successfully evaluated in the discovery phase while six significant or suggestive variants entered the validation phase, comprising a total study number of 1,518 cases and 3,908 controls. In the discovery phase, rs2504938, rs9364554, and rs2457571 SNPs were significantly associated with pancreatic cancer risk. Moreover, rs7758229 associated with the presence of distant metastases, while rs512077 and rs2504956 correlated with overall survival of patients. Although replicated, the association for rs9364554 did not pass multiple testing corrections in the validation phase. Contrary to the discovery stage, rs2504938 associated with survival in the validation cohort, which was more pronounced in stage IV patients. In conclusion, common variation in the SLC22A3 gene is unlikely to significantly contribute to pancreatic cancer risk. The rs2504938 SNP in SLC22A3 significantly associates with an unfavorable prognosis of pancreatic cancer patients. Further investigation of this SNP effect on the molecular and clinical phenotype is warranted. National Institute of Public Health Ministry of Education Youth and Sports of the Czech Republic Italian Cancer Genome Project Ministero Salute Pancobank Heidelberger Stiftung Chirurgie EU ERA-Net TRANSCAN Italian Ministry of Health Division of Gastroenterology, IRCCS “Casa Sollievo della Sofferenza” Hospital, San Giovanni Rotondo, Italy

Anti-transglutaminase (tTG) or anti-deamidated gliadin peptides (DGP) serum determination is the first step in diagnosing celiac disease (CD). Our aims were to: compare the performance of novel chemiluminescent tool in the detection of tTG and DGP (Q-Flash®, Inova) with that of the established ELISA (Q-Lite®, Inova) methods; identify the more reliable index for making a sound diagnosis and monitoring therapy.Using Q-Flash® and Q-Lite®, IgA and IgG class tTG and DGP were measured in the sera of 155 CD pediatric patients and 166 healthy age-matched controls. Forty-two of the patients had a follow-up one year after starting gluten free diet (GFD).Q-Flash® IgA tTG, the more accurate (intra-assay CV for low, intermediate and high values: 2.2%, 1.6%, and 1.1%; inter-assay CV: 2.8%, 4%, and 3%), sensitive (96.1%) and specific (97%) test for diagnosing CD, was the only variable to be significantly correlated with CD at binary logistic regression analysis (r=0.263, p0.0001, Exp(B)=1.0506, 95% CI=1.0286-1.0731). Q-Flash® IgA tTG or DGP screen were more accurate than Q-Lite® IgA tTG in monitoring CD patients on GFD (p=0.003).Q- Flash® IgA tTG measurement is an extremely precise, sensitive and specific index for not only diagnosing CD, but also monitoring therapy.

The CDKN2A(p16) gene plays a key role in pancreatic cancer etiology. It is one of the most commonly somatically mutated genes in pancreatic cancer, rare germline mutations have been found to be associated with increased risk of developing familiar pancreatic cancer and CDKN2A promoter hyper-methylation has been suggested to play a critical role both in pancreatic cancer onset and prognosis. In addition several unrelated SNPs in the 9p21.3 region, that includes the CDNK2A, CDNK2B and the CDNK2B-AS1 genes, are associated with the development of cancer in various organs. However, association between the common genetic variability in this region and pancreatic cancer risk is not clearly understood. We sought to fill this gap in a case-control study genotyping 13 single nucleotide polymorphisms (SNPs) in 2,857 pancreatic ductal adenocarcinoma (PDAC) patients and 6,111 controls in the context of the Pancreatic Disease Research (PANDoRA) consortium. We found that the A allele of the rs3217992 SNP was associated with an increased pancreatic cancer risk (OR het=1.14, 95% CI 1.01-1.27, p=0.026, ORhom =1.30, 95% CI 1.12-1.51, p=0.00049). This pleiotropic variant is reported to be a mir-SNP that, by changing the binding site of one or more miRNAs, could influence the normal cell cycle progression and in turn increase PDAC risk. In conclusion, we observed a novel association in a pleiotropic region that has been found to be of key relevance in the susceptibility to various types of cancer and diabetes suggesting that the CDKN2A/Blocus could represent a genetic link between diabetes and pancreatic cancer risk.

Based on the knowledge that matrix metalloproteinases (MMPs) and S100A8/A9 synergistically work in causing PDAC-associated type 2 diabetes mellitus (T2DM), we verified whether tissue and blood MMP8, MMP9, S100A8 and S100A9 expression might help in distinguishing PDAC among diabetics.Relative quantification of MMP8, MMP9, S100A8 and S100A9 mRNA was performed in tissues obtained from 8 PDAC, 4 chronic pancreatitis (ChrPa), 4 non-PDAC tumors and in PBMCs obtained from 30 controls, 43 T2DM, 41 ChrPa, 91 PDAC and 33 pancreatic-biliary tract tumors.T2DM was observed in PDAC (66%), in pancreatic-biliary tract tumors (64%) and in ChrPa (70%). In diabetics, with or without PDAC, MMP9 tissue expression was increased (p0.05). Both MMPs increased in PDAC and MMP9 increased also in pancreatic-biliary tract tumors PBMCs. In diabetics, MMP9 was independently associated with PDAC (p=0.025), but failed to enhance CA 19-9 discriminant efficacy. A highly reduced S100A9 expression, found in 7 PDAC, was significantly correlated with a reduced overall survival (p=0.015).An increased expression of tissue and blood MMP9 reflects the presence of PDAC-associated diabetes mellitus. This finding fits with the hypothesized role of MMPs as part of the complex network linking cancer to diabetes.

// Stefania Moz 1 , Daniela Basso 1 , Dania Bozzato 1 , Paola Galozzi 2 , Filippo Navaglia 1 , Ola H. Negm 3, 4 , Giorgio Arrigoni 5, 6 , Carlo-Federico Zambon 1 , Andrea Padoan 1 , Paddy Tighe 3 , Ian Todd 3 , Cinzia Franchin 5, 6 , Sergio Pedrazzoli 7 , Leonardo Punzi 2 , Mario Plebani 1 1 University of Padova, Laboratory Medicine, Department of Medicine - DIMED, Padova, Italy 2 University of Padova, Rheumatology Unit, Department of Medicine - DIMED, Padova, Italy 3 University of Nottingham, School of Life Sciences, Queen’s Medical Centre, Nottingham, UK 4 Mansoura University, Medical Microbiology and Immunology Department, Faculty of Medicine, Mansoura City, Egypt 5 University of Padova, Department of Biomedical Sciences, Padova, Italy 6 Proteomics Center, University of Padova and Azienda Ospedaliera di Padova, Padova, Italy 7 Associazione Wirsung-onlus, Padova, Italy Correspondence to: Daniela Basso, email: daniela.basso@sanita.padova.it Keywords: pancreatic cancer, SMAD4, signalling, epidermal growth factor, transforming growth factor β1 Received: June 30, 2016 Accepted: September 05, 2016 Published: September 16, 2016 ABSTRACT Epidermal Growth Factor (EGF) receptor overexpression, KRAS, TP53, CDKN2A and SMAD4 mutations characterize pancreatic ductal adenocarcinoma. This mutational landscape might influence cancer cells response to EGF, Transforming Growth Factor β1 (TGFβ1) and stromal inflammatory calcium binding proteins S100A8/A9. We investigated whether chronic exposure to EGF modifies in a SMAD4-dependent manner pancreatic cancer cell signalling, proliferation and invasion in response to EGF, TGFβ1 and S100A8/A9. BxPC3, homozigously deleted (HD) for SMAD4, and BxPC3-SMAD4+ cells were or not stimulated with EGF (100 ng/mL) for three days. EGF pre-treated and non pretreated cells were stimulated with a single dose of EGF (100 ng/mL), TGFβ1 (0,02 ng/mL), S100A8/A9 (10 nM). Signalling pathways (Reverse Phase Protein Array and western blot), cell migration (Matrigel) and cell proliferation (XTT) were evaluated. SMAD4 HD constitutively activated ERK and Wnt/β-catenin, while inhibiting PI3K/AKT pathways. These effects were antagonized by chronic EGF, which increased p-BAD (anti-apoptotic) in response to combined TGFβ1 and S100A8/A9 stimulation. SMAD4 HD underlied the inhibition of NF-κB and PI3K/AKT in response to TGFβ1 and S100A8/A9, which also induced cell migration. Chronic EGF exposure enhanced cell migration of both BxPC3 and BxPC3-SMAD4+, rendering the cells less sensitive to the other inflammatory stimuli. In conclusion, SMAD4 HD is associated with the constitutive activation of the ERK and Wnt/β-catenin signalling pathways, and favors the EGF-induced activation of multiple signalling pathways critical to cancer proliferation and invasion. TGFβ1 and S100A8/A9 mainly inhibit NF-κB and PI3K/AKT pathways and, when combined, sinergize with EGF in enhancing anti-apoptotic p-BAD in a SMAD4-dependent manner.

A systematic review and a meta-analysis were performed to quantify the accumulated information from genetic association studies investigating the impact of the CYP4F2 rs2108622 (p.V433M) polymorphism on coumarin dose requirement. An additional aim was to explore the contribution of the CYP4F2 variant in comparison with, as well as after stratification for, the VKORC1 and CYP2C9 variants. Thirty studies involving 9,470 participants met prespecified inclusion criteria. As compared with CC-homozygotes, T-allele carriers required an 8.3% (95% confidence interval (CI): 5.6�11.1%; P < 0.0001) higher mean daily coumarin dose than CC homozygotes to reach a stable international normalized ratio (INR). There was no evidence of publication bias. Heterogeneity among studies was present (I2 = 43%). Our results show that the CYP4F2 p.V433M polymorphism is associated with interindividual variability in response to coumarin drugs, but with a low effect size that is confirmed to be lower than those contributed by VKORC1 and CYP2C9 polymorphisms. Wiley Online Library

Background The identification of specific serological algorithms allowing the diagnosis of celiac disease (CD) is a new challenge for both the clinic and the laboratory. We compared the diagnostic accuracy of three new tests proposed for CD screening with that of the well established IgA tTG, and ascertained whether any combination of these tools might enhance accuracy in diagnosing CD. Methods In sera from 329 CD and 374 control children, the following were assayed: IgA tTG; IgA/IgG, which identify tTG-gliadin complexes (Aeskulisa Celi Check and CeliCheck IgGA); IgA/IgG, which identify deamidated gliadin peptides and tTG (QUANTA Lite TM h-tTG/DGP Screen). Results When specificity was set at 100%, the most sensitive index of CD was IgA tTG (75.7%, cut-off = 100 U), followed by QUANTA Lite TM h-tTG/DGP Screen (65.3%, cut-off 145 U), Aeskulisa Celi Check (62.6%, cut-off 909 U/mL) and CeliCheck IgGA (59.6%, cut-off 977 U/mL). Three algorithms were obtained by combining IgA tTG with each of the new tests. The algorithm obtained by measuring IgA tTG and QUANTA Lite TM h-tTG/DGP Screen allowed the correct identification of CD in 78.7% of cases (negative predictive value = 97.3%). Conclusions The two-test based strategy could be used for the cost effective diagnosis of CD.

The present article provides an overview on the rapidly evolving scenario in clinical molecular diagnostics. The concept of clinical validity and utility underpins the use of molecular diagnostic tests in the clinical laboratory, and continuous sharing of expertise between clinical and laboratory partners will enhance appropriateness of molecular testing and interpretation, thus improving personalized medicine. Thanks to the technological advances made in molecular diagnostics, above all in next generation sequencing (NGS) and digital polymerase chain reaction (dPCR) platforms, clinical laboratories benefit from rapid translation of established and emerging molecular tests. The examples of their application which will be discussed encompass cystic fibrosis (CF) (qualitative molecular) and BCR/ABL1 (quantitative molecular) tests. A final consideration on total quality assurance in molecular testing is also made and the role of accreditation is underlined as one of the main factors in guaranteeing the overall quality of molecular test results.

The role of pharmacogenomics and tamoxifen was investigated by analyzing several polymorphisms of cytochrome P450 and SULT1A1 gene in a nested case control study from the Italian Tamoxifen Prevention Trial. This study included 182 Caucasian subjects, 47 breast cancer (BC) cases and 135 matched controls. We used the AmpliChip CYP450 Test to screen 33 alleles of CYP2D6 and 3 of CYP2C19. One more variant for CYP2C19*17 and two single-nucleotide polymorphisms for the gene SULT1A1 were also performed. By using the AmpliChip CYP450 Test, out of 182 subjects, we identified 8 poor metabolizer (PM), 17 intermediate metabolizer (IM), 151 extensive metabolizer (EM) and 3 ultrarapid metabolizer (UM). PM women allocated to the tamoxifen arm showed a higher risk of developing BC compared to the remaining phenotypes (P=0.035). In an exploratory analysis, among 58 women with a CYP2D6*2A allele, 9 BCs were diagnosed in the placebo arm and only 1 in the tamoxifen arm (P=0.0001). CYP2C19 and SULT1A1 polymorphisms did not show any correlation with tamoxifen efficacy. Tamoxifen showed reduced efficacy in CYP2D6 PMs in the chemoprevention setting. Conversely, the CYP2D6*2A allele may be associated with increased efficacy of tamoxifen. These findings support the relevance of pharmaco-genomics in tailoring tamoxifen treatment.

The Th2 cytokine IL-4 might limit H. pylori associated gastric inflammation and favour H. pylori clearance. The aim of the study was to verify whether IL-4 -588CT SNP, or two SNPs of the gene coding the alpha chain of IL-4 receptor (IL-4RA Ex5+14AG, IL-4RA Ex11+828AG) considered singly or as haplotypes, are correlated with H. pylori virulence genes or H. pylori associated diseases.We studied 144 patients with non-cardia gastric cancer (NCGC)(41/50 with present or past H. pylori infection), 75 with duodenal ulcer (DU)(66 H. pylori infected) and 171 with gastritis (CG)(107 H. pylori infected). cagA gene was present in 24/28 NCGC, 45/59 DU and 56/107 CG.All SNPs were in Hardy-Weinberg equilibrium. IL-4RA haplotypes frequencies were estimated using Arlequin software. Neither the SNPs nor the IL-4RA haplotype correlated with disease diagnosis, H. pylori infection, degree of mucosal inflammation or intestinal metaplasia. IL-4 -588T allele (OR=3.69, 95% CI:1.34-10.16) and IL-4RA GA haplotype (p0.05) enhanced the risk for cagA positive infections. IL-4RA GA haplotype correlated with IL-4 protein levels in H. pylori infected gastric mucosa.IL-4 and IL-4RA gene polymorphisms concur in selecting the H. pylori infecting strain, probably influencing the IL-4 signalling pathway.

UNLABELLED:Genotype-guided warfarin dosing have been proposed to improve patient’s management. This study is aimed to determine whether a CYP2C9- VKORC1- CYP4F2-based pharmacogenetic algorithm is superior to a standard, clinically adopted, pharmacodynamic method. Two-hundred naïve patients with non-valvular atrial fibrillation were randomized to trial arms and 180 completed the study. No significant differences were found in the number of out-of-range INRs (INR3.0) (p = 0.79) and in the mean percentage of time spent in the therapeutic range (TTR) after 19 days in the pharmacogenetic (51.9%) and in the control arm (53.2%, p = 0.71). The percentage of time spent at INR>4.0 was significantly lower in the pharmacogenetic (0.7%) than in the control arm (1.8%) (p = 0.02). Genotype-guided warfarin dosing is not superior in overall anticoagulation control when compared to accurate clinical standard of care. TRIAL REGISTRATION:ClinicalTrials.gov NCT01178034.

MALDI-TOF profiling of low molecular weight peptides (peptidome) usage is limited due to the lack of reproducibility from the confounding inferences of sample preparation, data acquisition, and processing. We applied MALDI-TOF analysis to profile urine peptidome with the aims to: (i) compare centrifugal ultrafiltration and dialysis pretreatments, (ii) determine whether using signal LOD (sLOD), together with data normalization, may reduce MALDI-TOF variability. We also investigated the influence of peaks detection on reproducibility. Dialysis allowed to obtain better MALDI-TOF spectra than ultrafiltration. Within the 1000-4000 m/z range, we identified 120 and 129 peaks in intra- and interassay studies, respectively. To estimate the sLOD, serial dilution of pooled urines up to 1/256 were analyzed in triplicate. Six data normalization strategies were investigated-the mean, median, internal standard, relative intensity, TIC, and linear rescaling normalization. Normalization methods alone performed poorly in reducing features variability while when combined to sLOD adjustment showed an overall reduction in features CVs. Applying a feedback signal processing approach, after median normalization and sLOD adjustment, CVs were reduced from 103 to 26% and 113 to 25% for the intra- and interassay, respectively, and spectra became more comparable in terms of data dispersion.

OBJECTIVES Anti-tissue transglutaminase antibody (anti-tTG) determination using second-generation (human antigen) enzyme-linked immunoassays (ELISAs) is a very accurate test to diagnose celiac disease (CD). In this study, we compared 2 second-generation ELISAs (Celikey tTG; Pharmacia Diagnostics GmbH & Co, Freiburg, Germany, and QuantaLite; Inova Diagnostics, San Diego, CA) and antiendomysial antibodies (EMAs) with a new indirect chemiluminescence immunoassay (LIAISON tTG; DiaSorin S.p.A., Saluggia, Italy) in diagnosing and monitoring CD in children. PATIENTS AND METHODS Antiendomysial antibodies, anti-tTGs and total immunoglobulin A were measured in the sera of 103 control children, 101 children with histologically proven CD and 31 CD children on gluten-free diet (GFD). RESULTS Anti-tissue transglutaminase antibody mean levels were significantly higher in CD with respect to control or GFD children. The sensitivity value of EMAs, LIAISON tTG, Celikey tTG and QuantaLite in diagnosing CD was 97.7%, 97.0%, 94.1% and 98.0%, respectively, and the corresponding specificity values were 91.1%, 98.1%, 97.1% and 96.1%, respectively. The degree of mucosal destruction (Marsh criteria) was correlated with EMA semiquantification (P < 0.01) and with the circulating levels of anti-tTGs measured using LIAISON (P < 0.05) or QuantaLite (P < 0.01). Twenty-six CD children were followed up from 5 to 25 months after GFD. The circulating levels of anti-tTGs measured with any of the 3 assays significantly dropped after GFD. CONCLUSIONS Anti-tissue transglutaminase antibody determination with second-generation ELISAs is as effective as EMAs for CD diagnosis. The novel chemiluminescent method described in the present paper for the detection of anti-tTGs in the diagnosis of CD had the highest sensitivity and specificity values. The anti-tTG test correlates with the degree of mucosal destruction and is suitable for verifying patient compliance to dietary treatment.

Background Our aim was to identify the pancreatic cancer diabetogenic peptide. Methods Pancreatic tumor samples from patients with (n = 15) or without (n = 7) diabetes were compared with 6 non-neoplastic pancreas samples using SDS-PAGE. Results A band measuring approximately 1500 Da was detected in tumors from diabetics, but not in neoplastic samples from non-diabetics or samples from non-neoplastic subjects. Sequence analysis revealed a 14 amino acid peptide (1589.88 Da), corresponding to the N-terminal of the S100A8. At 50 nmol/L and 2 mmol/L, this peptide significantly reduced glucose consumption and lactate production by cultured C2C12 myoblasts. The 14 amino acid peptide caused a lack of myotubular differentiation, the presence of polynucleated cells and caspase-3 activation. Conclusions The 14 amino acid peptide from S100A8 impairs the catabolism of glucose by myoblasts in vitro and may cause hyperglycemia in vivo. Its identification in biological fluids might be helpful in diagnosing pancreatic cancer in patients with recent onset diabetes mellitus.

The aim of this study was to evaluate whether there was any correlation between Helicobacter pylori -associated diseases and (1) H. pylori virulence genes or (2) IL-1B , IL-1RN , IFN-G , TNF-A , IL-10 genetic polymorphisms. Patients with non-cardia gastric cancer (NCGC, n = 129) or benign gastroduodenal diseases ( n = 792) were studied. IL-1RN intron 2 VNTR polymorphism (PCR), IL-1B −31 C/T (RFLP), the SNPs of IFN-G (+874 A/T), TNF-A (−1031 C/T, −857 C/T, −376 A/G, −308 A/G, −238 A/G), IL-10 (−1082 A/G, −819 C/T, −592 A/C) (Taqman chemistry) were studied. cagA , s1 and m1 vacA , were PCR amplified. Duodenal ulcer was more frequent in TNF-A −857 TT and in IL-1RN 1,2 subjects. TNF-A −857 TT genotype was also correlated with gastric ulcer. IL-10 −819 TT genotype was associated with intestinal metaplasia and NCGC. Antral inflammation was associated with TNF-A −1031 TT, while corpus activity with IL-10 −819 CC. H. pylori infection was associated with TNF-A −308 AG genotype, while IFN-G +874 AA genotype was associated with cagA . In conclusion, among host genetic factors contributing to H. pylori disease outcome, IFN-G +874 AA genotype favors cagA positive infections, TNF-A −857 TT duodenal ulcer while IL-10 −819 TT intestinal metaplasia and NCGC.

Aim: To study in vivo whether pancreatic cancer tumour growth and metastasis can be modified by a gene construct with HSV-TK suicide gene and IL2 co-expression. Methods: Seventy-eight female SCID mice were i.p. inoculated with retrovirally transduced or control MIA PaCa 2, CAPAN-1 and PANC-1 cell lines. The animals were then randomly selected for saline or ganciclovir (GCV) treatment from the second week, for a total of two weeks. Results: Most inoculated mice developed tumour nodules and spleen metastases. The liver was colonized by control CAPAN-1 and MIA PaCa 2, but not by PANC-1. Tumours in transduced MIA PaCa 2 cell injected mice were smaller, and in transduced CAPAN-1 injected mice larger, than in control-inoculated mice. There were increased pancreatic and decreased spleen metastases from transduced CAPAN-1, and diminished liver involvement from transduced MIA PaCa 2. No differences were found between mice inoculated with transduced and control PANC-1 cell lines. GCV treatment had no effect on tumour's size or metastases. Conclusions: The HSV-TK suicide gene does not confer GCV sensitivity to pancreatic cancer in this in vivo model. Different pancreatic cancer cell lines cause different growth and metastasis patterns after inoculation in SCID mice, possibly because of variations in their inherent characteristics. The different effects of our vector on cell growth and metastasis may be attributable to the effects of the immunostimulatory cytokine IL2.

Background. The aims of this retrospective study were to ascertain in large series of children and adults: the relationship of the infecting strain to gastric mucosal lesions; and the relationship of the infecting strain to its duodenal localization. Materials and Methods. We studied 307 and 604 consecutive children and adults. In gastric mucosal samples H. pylori was cultured, genotyped and histologically assessed, while inflammation, activity and intestinal metaplasia were graded. In a subset of 171 patients H. pylori ureaseA (ureA) and cagA genes were amplified (PCR) using mucosal biopsies from the duodenum. Results.H. pylori infection was diagnosed in 40 children and 308 adults. cagA was identified in 50% and 65.5% of infected children and adults. Antral activity was associated with the density of infecting bacteria (p

INTRODUCTION The transfer of drug-susceptible (suicide) genes to tumor cells by retroviral or adenoviral vectors is a novel approach to the treatment of human tumors. AIMS To ascertain the antitumor effect of retroviral transduction of the pancreatic cancer cell lines MIA PaCa 2, CAPAN-1, PANC1, and PSN1 with the herpes simplex virus thymidine kinase (HSV-TK) gene. METHODOLOGY The vector carried a neoselectable marker gene, the human interleukin-2 gene, an internal ribosome entry coding site, and the region coding HSV-TK. RESULTS Twenty micromoles or less of ganciclovir did not modify nontransduced TK- cell growth, whereas > or =100 micromol completely inhibited TK- cell growth, indicating that this dosage is cytotoxic per se. The 4 TK- and the 4 transduced cell lines were treated daily with 0.001, 0.01, 0.1, 1, 10, and 20 micromol of ganciclovir for 13 days. CAPAN-1 cell growth was completely inhibited by 0.1 micromol of ganciclovir; higher doses were required to kill PANC1 (10 micromol) and PSN1 (20 micromol). MIA PaCa 2 cell growth decreased following a 20-micromol ganciclovir dosing. The bystander effect was great in the CAPAN-1 cell line and moderate in PANC1; no bystander effect was recorded in MIA PaCa 2 and PSN1 cell lines. CONCLUSION Gene therapy with HSV-TK for pancreatic cancer seems effective in only a limited number of tumor-derived cell lines, and this limits its application in vivo.

Helicobacter pylori infection outcome might depend on genotypic polymorphisms of both the bacterium and the host. We ascertained: (1) the functionality of H. pylori oipA gene; (2) the polymorphism of the hostinterleukin (IL-1β) gene (−31 C/T) and of the IL-1RN gene (intron 2 VNTR); (3) the association between the above genes and the histological and pathological outcome of H. pylori infection. One hundred and sixty-five H. pylori positive and 137 H. pylori negative subjects (23 gastric adenocarcinoma, 58 peptic ulcer, 221 gastritis) were studied. oipA was sequenced, IL-1β was RFLP analysed. Antral and body mucosal biopsies were histologically evaluated. Functional oipA genes were correlated with cagA gene; both genes were significantly associated with gastritis activity, peptic ulcer and gastric adenocarcinoma. In these patients heterozygousIL-1RN 1/2 and IL-1β C/T genotypes were more frequent than in gastritis patients. Intestinal metaplasia was associated with cagA, functional oipA and IL-1RN 2 allele. In conclusion, peptic ulcer and the preneoplastic intestinal metaplasia are associated with H. pylori virulence genes and with IL-1RN 2 host allele. An interplay between bacterial virulence factors and cytokines genotypes, is probably the main route causing H. pylori infection to lead to benign mild disease, benign severe disease or preneoplastic lesions.

Leukocyte telomere length (LTL) has been observed to be hereditable and correlated with longevity. However, contrasting results have been reported in different populations on the value of LTL heritability and on how biology of telomeres influences longevity. We investigated whether the variability of genes correlated to telomere maintenance is associated with telomere length and affects longevity in a population from Southern Italy (20–106 years). For this purpose we analyzed thirty-one polymorphisms in eight telomerase-associated genes of which twelve in the genes coding for the core enzyme (TERT and TERC) and the remaining in genes coding for components of the telomerase complex (TERF1, TERF2, TERF2IP, TNKS, TNKS2 and TEP1). We did not observe (after correcting for multiple testing) statistically significant associations between SNPs and LTL, possibly suggesting a low genetic influence of the variability of these genes on LTL in the elderly. On the other hand, we found that the variability of genes encoding for TERF1 and TNKS2, not directly involved in LTL, but important for keeping the integrity of the structure, shows a significant association with longevity. This suggests that the maintenance of these chromosomal structures may be critically important for preventing, or delaying, senescence and aging. Such a correlation was not observed in a population from northern Italy that we used as an independent replication set. This discrepancy is in line with previous reports regarding both the population specificity of results on telomere biology and the differences of aging in northern and southern Italy.

Inflammatory bowel diseases (IBDs), which comprise the two major clinical subtypes, Crohn's disease and ulcerative colitis, incur high morbidity and potential mortality. The present study reviews data on the pathogenesis and diagnosis of IBDs. The pathogenesis depends on complex interactions between susceptibility genes, environmental factors, and innate and adaptive immunity, the understanding of which is crucial to discovering novel laboratory biomarkers. Traditional laboratory tests for the diagnosis, prognosis and assessment of disease activity of IBDs are reported on, and the biochemical properties, pre-analytical and analytical aspects and clinical utility of the fecal markers lactoferrin and calprotectin are described. DNA testing and established (ASCA and pANCA) and emerging (ACCA, ALCA, AMCA, OmpC) serum markers are described; a further aspect to be addressed is the clinical use of pharmacogenetics for the treatment of IBDs.

Background. The aims of our study were twofold. First, we sought to evaluate in symptomatic children the influence of the Helicobacter pylori genotype on gastritis, abdominal pain, and circulating anti–H. pylori IgG antibodies (anti–H. pylori IgG) or pepsinogen A (PGA) and C (PGC). Additionally, we sought to assess anti–H. pylori IgG, PGA, and PGC patterns in a large cohort (N = 921) of asymptomatic children. Materials and Methods. In 183 symptomatic children, H. pylori infection and the presence of gastritis were evaluated by histology. In a subgroup of 20 H. pylori–positive children, the H. pylori genotype was evaluated also by polymerase chain reaction. Nine hundred and twenty-one asymptomatic children, aged 11 to 14 years, were studied by anti–H. pylori IgG, PGA, and PGC serum determination. Results. The infection was found in 33 of 183 symptomatic children; among the 20 H. pylori–positive children for which the H. pylori genotype was available, cagA was present or absent in equal percentages. H. pylori infection was associated with more severe gastritis and higher serum levels of anti–H. pylori IgG and PGC but not with abdominal pain. In infected children, higher levels of anti–H. pylori IgG and the presence of abdominal pain were associated with infections caused by cagA-positive strains. In the cohort of 921 asymptomatic children, raised levels of anti–H. pylori IgG, PGA, and PGC were found in approximately 5% of the cases. Conclusions. Infection with cagA-positive H. pylori strains can be associated with increased frequency of reported abdominal pain and higher circulating levels of anti–H. pylori IgG. The serological assessment of H. pylori IgG using H. pylori antigens containing significant amounts of cagA protein may, therefore, underestimate the true prevalence of infection.