method have become a research trend. The ultimate biomarkers must be easily detectable with fine sensitivity and specificity and also must discriminate PC from other benign pancreatic diseases 9. Blood is a simply reachable and rather steady sample to find alerting biomarkers. Technological advances in the recent years have provided possibilities to detect circulating biomarkers based on "omics" research, relying on proteins, cell-free DNAs, non-coding RNAs, circulating tumor cells (CTCs), and exosomes molecular contents 10. MicroRNAs (miRs) are small (~22 nucleotides) non-coding RNAs that have gene regulatory roles via targeting the 3′-untranslated region (3′-UTR) of their target mRNA and finally cause either translational repression or mRNA degradation 11,12. Up to now, a number of biomarkers have been introduced as PC biomarkers such CEA, CA19-9, CA125 and CA72-4. Nonetheless, none of these tumor markers has shown efficient sensitivity or specificity for diagnosing PC at primary stages and have been used for post resection monitoring rather than earlier detection purposes. MiRNAs seem to be truly stable in blood and several authors reported that miRNAs show dysregulation in pancreatic diseases being able to differentiate PC from pancreatitis, pancreatic benign masses as well as normal subjects 13. Furthermore, owing to advanced technologies in high-throughput molecular methods the understanding of the pathophysiology of pancreatic cancer have been improved. Various genome-wide mRNA and miRNA expression profiling studies using microarray-based and NGS approaches have provided important insights into the phenotypic characteristics of pancreatic cancer 14. In this study, using multiple bioinformatics tools, we integrated various serum expression profiles of miRNAs to find the most significant potential miRNA signatures helpful in the diagnosis of PC and constructed a novel miRNA-mRNA regulatory network in PC using bioinformatics approaches. Next, we investigated the molecular mechanisms downstream of the captured miRNA signatures and their predicted target genes correlated to PC progression and analyzed them in a logistic model. Material and method Microarray datasets search. In order to find proper miRNA expression profiles in microarray datasets, we conducted a systematic search in Gene Expression Omnibus (GEO) database (https://www.ncbi.nlm.nih.gov/ geo/) 15. Using the keywords "Pancreatic cancer" and "Serum", at the first step we reached 900 datasets. Then, we limited the results using 'Homo sapiens' and 'Non-coding RNA profiling by array' filters, so we reached to 16 datasets. Finally, by setting the sample count on more than 200 samples, 4 final datasets were achieved. Differentially expressed miRNAs (DEMs) detection. The DEMs were obtained using the online tool GEO2R in the GEO database 15 , which makes evaluations using the GEOquery and limma R packages from the Bioconductor project to compare two or more groups of samples in a GEO dataset. Normalization has been carried out using the RMA algorithm. To ke...

Background and Aims:  Helicobacter pylori is a highly diverse pathogen, which encounters epithelial cells as the initial defense barrier during its lifelong infection. The structure of epithelial cells can be disrupted through cleavage of microfilaments. Cytokeratin 18 (CK18) is an intermediate filament, the cleavage of which is considered an early event during apoptosis following activation of effector caspases. Methods:  Helicobacter pylori strains were isolated from 76 dyspeptic patients. cagA 3’ variable region and CagA protein status were analyzed by PCR and western blotting, respectively. Eight hours post‐co‐culture of AGS cells with different H. pylori strains, flow cytometric analysis was performed using M30 monoclonal antibody specific to CK18 cleavage‐induced neo‐epitope. Results:  Higher rates of CK18 cleavage were detected during co‐culture of AGS cells with H. pylori strains bearing greater numbers of cagA EPIYA‐C and multimerization (CM) motifs. On the other hand, H. pylori strains with greater numbers of EPIYA‐B relative to EPIYA‐C demonstrated a decrease in CK18 cleavage rate. Thus, H. pylori‐mediated cleavage of CK18 appeared proportional to the number of CagA EPIYA‐C and CM motifs, which seemed to be downplayed in the presence of EPIYA‐B motifs. Conclusions:  Our observation associating the heterogeneity of cagA variants with the potential of H. pylori strains in the induction of CK18 cleavage as an early indication of apoptosis in gastric epithelial cells supports the fact that apoptosis may be a type‐specific trait. However, additional cagA‐targeted experiments are required to clearly identify the role of EPIYA and CM motifs in apoptosis and/or the responsible effector molecules.

Background: Multiple etiologic factors are suspected to cause gastric cancer, the most important of which is infection with virulent types of Helicobacter pylori. Materials and Methods: We have compared 102 gastric cancer patients with 122 non-ulcer, non-cancer dyspeptic patients. Gastric specimens were evaluated for H. pylori infection by tissue-based detection methods. Patient sera underwent antigen-specific ELISA and western blotting using a Helicoblot 2.1 kit and antibody responses to various H. pylori antigens were assessed. Results: The absolute majority (97-100%) of both groups were H. pylori seropositive. Multivariate regression analysis demonstrated serum antibodies to the low molecular weight 35kDa protein to be protective and reduce the risk of gastric cancer by 60% (OR:0.4; 95%CI:0.1-0.9). Conversely, seroreactivity to the 89kDa (VacA) protein was significantly higher in gastric cancer patients (OR:2.7; 95%CI:1.0-7.1). There was a highly significant association (p<0.001) between seroreactivity to the 116kDa (CagA) and 89kDa (VacA) proteins, and double positive subjects were found at nearly five fold (OR:4.9; 95%CI:1.0-24.4) enhanced risk of gastric cancer as compared to double negative subjects. Conclusions: Seroreactivity to H. pylori low (35kDa) and high (116kDa/89kDa) molecular weight antigens were respectively revealed as protective and risk indicators for gastric cancer.

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Helicobacter pylori, a microaerophilic fastidious bacterium, has been cultured on various plating and broth media since its discovery. Although the agar media can be sufficient for the identification, typing, and antibiotic resistance studies, no secretory antigen of H. pylori can be evaluated in such media. Thus, satisfactory growth of H. pylori in liquid culture which is needed for analysis of secretory proteins without the presence of interfering agents is in demand. We assessed the impact of beta-cyclodextrin, Fetal Bovine Serum (FBS), and charcoal as supplements for H. pylori growth. Furthermore, we aimed to identify the most favorable supplement that supports the secretion of the dominant secretory protein, vacuolating cytotoxin (VacA). Five clinical strains were cultured on broth media and the growth, viability, morphology, and protein content of each strain were determined. Our results revealed that beta-cyclodextrin supports the growth rate, viability, and cell lysate protein content to the extent similar to FBS. Application of beta-cyclodextrin is found to postpone spiral to coccoid conversion up to 72 h of incubation. Although FBS supports a higher VacA protein content, presence of interfering macromolecules in FBS questions its utility particularly for purposes of studying extra cellular proteins such as VacA. This study recommends further application of beta-cyclodextrin as a culture supplement with the potential capacity in neutralizing toxic compounds and flourishing the secretion of H. pylori proteins without addition of interfering proteins.

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Here, we show higher prevalence of mixed babA/B genotypes among BabA-low/Le(b)-low clinical strains. Recombination of babA and babB genes across their loci may yield lower BabA expression and lower Le(b) binding activity. We conclude that H. pylori strains with lower Le(b) binding activity are better adapted for colonization of the gastric metaplastic patches in the duodenum and enhance the risk of duodenal ulcers.

We, hereby, present a simple, consistent, and cost-efficient gastric cell culture system, which provides a suitable model for extended in vitro infection of H. pylori. This platform can be employed for a variety of gastric-related research.