Gastroenterology and Hepatology from Bed to Bench

Non-alcoholic fatty liver disease (NAFLD) represents a broad spectrum of liver damage, ranging from simple steatosis to steatohepatitis and fibrosis; as well, there is a close association between NAFLD, obesity, metabolic syndrome and type 2 diabetes mellitus. There is a certain degree of uncertainty regarding the natural history and prognosis of NAFLD; however, several methods are currently used for its diagnostic approach. In the first instance, non-invasive tests could be used to identify patients at low risk of developing fibrosis and to establish more easily the need for a liver biopsy, whose accuracy in the evaluation of fibrosis has been questioned, mainly due to errors of intra and interobserver sampling, technical problems and cost, which limits its use. Therefore, it is essential to determine the diagnostic strategy for patients with NAFLD.

Keywords: Fibrosis, Liver, Steatohepatitis, Steatosis.

(Please cite as Salva-Pastor N, Chávez-Tapia NC, Uribe M, Nuño-Lámbarri N. Diagnostic and initial approach of the patient with non-alcoholic fatty liver disease: role of the primary care provider. Gastroenterol Hepatol Bed Bench 2019;12(4):267-277).

Aim: To analyze the development of gliadin-specific immune responses in children with a genetic risk for CD and to determine whether these could be detected before the clinical onset of the disease by using immunological tests.

Background: Clinical manifestations of celiac disease (CD) in the first year of life is uncommon, which is due to the suboptimal sensitivity of tissue transglutaminase IgA antibodies (tTG-IgA) at this age and other possible causes of malabsorption in infants. The development of Deamidate gliadin peptide-specific antibodies (in particular DGP-IgG) in young children was poorly considered in the CD diagnosis.

Methods: We conducted a retrospective cross-sectional study on children between one month and forty-eight months of life, which performed in our health center from 2016 to 2018. Three hundred and fifty children were selected according to strict inclusion criteria:  positive for HLA-DQA1 and DQB1 alleles, positive anti tTG-IgA/IgG and/or positive DGP-IgG/IgA. Eighty-two children were selected and divided into two different groups of patients: Group one (forty newborns under twenty-four months of life) and Group two (children from twenty-five months to 48 months of life).

Results: Anti-DGP-IgG antibodies precede anti tTG-IgA seroconversion in children under two years in 80% of cases. Anti-DGP-IgG positive patients had milder symptomatic forms of CD than anti tTG-IgA positive children, characterized by gastrointestinal symptoms in the presence of normal growth, normal serum iron, and low MCH level. At tTG-IgA seroconversion, children present gastrointestinal clinical forms associated with impaired growth. The combined use of tTG-IgA and DGP-IgG antibodies upgrade the diagnostic sensitivity from 50% to 92%.

Conclusion: Anti-DGP-IgG antibodies precede tTG-IgA seroconversion in newborns and identified two distinct clinical phenotypes. At this point, if you wanted to test your newborn patients for CD serology, how would you proceed?

Keywords: Diagnostic tests, Transglutaminase antibodies, Deamidated gliadin peptides.

(Please cite as Assandri R, Montanelli A. Diagnosis of gluten-related enteropathy in a newborn: how and when? Gastroenterol Hepatol Bed Bench 2019;12(4):278-286).

Aim: This research aimed to evaluate the effect of gastroesophageal reflux disease (GERD) on pulmonary volumes, airflows, and airway resistance in the patients without respiratory symptoms and compare them with the healthy subjects.

Background: GERD is the return of gastric content into the esophagus and beyond. GERD may play an essential role in the extraesophageal diseases, including chest pain, asthma, laryngitis, chronic cough, and sinusitis. The relation between GERD and airway involvement in asthma and also bronchoconstrictor effects of GERD are well recognized, but its impact on lung parameters in the patients with GERD without respiratory symptoms is unclear.

Methods: In a case-control study, 78 GERD patients without pulmonary symptoms and 93 healthy subjects as control group were enrolled. The impulse oscillometry examined airway resistance. The body plethysmograph measured the pulmonary volumes and airflows.

Results: The mean age of GERD patients and the healthy subjects were 37.30±9.76 and 34.74±11.10, respectively. A total of 53.8% of patients and 67.7% of healthy subjects were male. The lung volumes measured by the body plethysmography were normal in both patients and healthy subjects. However, there was a significant difference between the groups in forced expiratory volume in 1 second (FEV1)/forced vital capacity (FVC) (P=0.01) and maximal mid expiratory flow (MMEF) (P=0.008). Airway resistance at R5Hz was significantly higher in the case group than the control group (P=0.001).

Conclusion: The results of the current study demonstrated that GERD patients have small airway disease even in the absence of respiratory symptoms.

Keywords: Gastroesophageal reflux, Lung function, Plethysmography, Airway resistance, Oscillometry.

(Please cite as Nazemiyeh M, Nouri-Vaskeh M, Somi MH, Saeedi E, Sharifi A. Lung function parameters in patients with gastroesophageal reflux without respiratory symptoms: a case-control study. Gastroenterol Hepatol Bed Bench 2019;12(4):287-291).

Aim: In the present study, a new formulation of HBsAg vaccine was developed and compared with a commercial peer.

Background: Vaccination of hepatitis B infection has been an unavoidable affair since the 1980s, though it has numerous limitations such as inefficacy in the induction of cellular immune responses. To address these limitations, research on novel formulations is necessary to develop a superior formulation with the potency of induction of both cellular and humoral immune responses.

Methods: HBsAg was formulated in oil-in-water adjuvant Montanide ISA-266 (5 µg/dose) using homogenizer. Balb/C mice were then immunized three times at days 0, 14, and 28 with HBsAg/Montanide ISA-266 or HBsAg/alum with proper control groups. Two weeks after the last immunization, immunological parameters including IL-2, IL-4, TNF-?, IFN-?, total IgG and IgG1/IgG2a isotypes were assessed by ELISA.

Results: The results demonstrated that the formulation of HBsAg with Montanide ISA-266 enhanced humoral immune responses versus the commercial vaccine and control groups. No significant difference in terms of Th1 pattern was found between HBsAg/Montanide ISA-266 and the commercial vaccine.

Conclusion: Formulation of HBsAg with an oil-based adjuvant may be useful for the induction of a more potent humoral immune response compared to the commercially available HBV vaccine.

Keywords: Adjuvant, Hepatitis B vaccine, Montanide ISA 266, Immune response.

(Please cite as Savoji MA, Haghighat S, Mirzaee M, Golkaran B, Mirzaee R, Esfandiari B, et al. Formulation of HBs antigen in Montanide ISA266 shows superiority to commercial HBsAg vaccine in the induction of humoral immune responses. Gastroenterol Hepatol Bed Bench 2019;12(4):292-300).

Aim: This paper aimed to assess and follow up the course of resolved HBV (hepatitis B virus) during and after treatment with direct-acting antiviral drugs (DAAs).

Background: Co-infection with hepatitis B and hepatitis C is increasingly recognized in patients with chronic hepatitis. Resolved HBV in patients with chronic HCV (hepatitis C virus) infection has been investigated during interferon therapy, and the investigators suggest a possible correlation with a lower response to anti-viral treatment, higher grades of liver histological changes, and development of hepatocellular carcinoma.

Methods: Three hundred and thirteen patients were included in our observational and prospective study; two hundred and fifty-three patients had chronic hepatitis C (CHC) (group I), and sixty patients had both CHC and resolved HBV-infection (group II). They all were eligible for treatment with DAAs therapy for chronic HCV in our hepatology unit, Internal Medicine Department, Zagazig University Hospitals from December 2017 to September 2018. They were subjected to thorough history taking, full clinical examination, routine laboratory investigations, HCV antibody, HCV RNA, HBV surface antigen (HBsAg), HBV surface antibody (anti-HBs) HBV core antibody (anti-HBc), and HBV-DNA quantitative levels. All patients were followed up at baseline, at the end of week 4 of anti-viral therapy, at the end of treatment and 12 weeks after treatment.

Results: Assessment at 28 days showed significant decreases in ALT and AST levels in both groups, with stabilization of these levels on follow-up at 12 and 24 weeks. The efficacy of treatment was comparable in both groups. No case of ALT flare was observed in either group. Similar outcomes regarding AST and ALT levels were found in patients with diseases associated with immune derangement.

Conclusion: The risk of resolved HBV reactivation during or after treatment with DAAs is low.

Keywords: HCV infection, Resolved HBV, HBV flare, Direct-acting antivirals.

(Please cite as Mohamed SY, Gaballah BA, Mohamed Elsadek H, Hassan Emara M, Hamed EF. Resolved HBV behavior during the treatment of chronic HCV infection with direct-acting antivirals. Gastroenterol Hepatol Bed Bench 2019;12(4):301-308).

Aim: This study aimed to evaluate rs1179251 single nucleotide polymorphism in the IL-22 gene as a host factor and its effect on chronic hepatitis B infection.

Background: Interleukin 22 (IL-22) belongs to a group of recently discovered cytokines, and it is produced and secreted by innate lymphoid cells (ILCs) and T helper 22 (Th22) cells. This cytokine plays dual roles as pro-inflammatory and anti-inflammatory effects in various conditions and different tissues of the body.

Methods: This study was performed based on a case-control format to assess IL-22 rs1179251 single nucleotide polymorphism genotypic and allelic frequencies among 227 hepatitis B chronic patients and 227 healthy controls. The polymerase chain reaction and restriction fragment length polymorphism techniques were employed to determine the polymorphism’s genotypes.

Results: Genotypes Frequencies in patients’ group were determined CC 59.91%, CG 37.89%, and GG 2.20% respectively in comparison to CC 63.44%, CG 31.72% and GG 4.85% in control group. The findings revealed that there was no statistically significant difference in the genotypes (P=0.156) frequencies of IL-22 gene polymorphism (rs1179251) between patients and control groups.

Conclusion: No association was found between rs1179251 single nucleotide polymorphism in IL-22 gene and chronic hepatitis B infection. So, in spite of the importance of IL-22 gene in immune responses, the studied polymorphism does not serve a decisive role in susceptibility to hepatitis B virus chronic infection.

Keywords: Interleukin 22, Single nucleotide polymorphism, Hepatitis B virus, Immune response, Inflammation.

(Please cite as Asadi P, Mohebbi SR, Hosseini SM, Zali MR. Evaluation of single nucleotide polymorphism in interleukin 22 (IL-22) gene and its association with chronic hepatitis B infection Gastroenterol Hepatol Bed Bench 2019;12(4):309-314).

Aim: Survey of the survival levels of gastric cancer and its effective causes.

Background: The survival of gastric cancer because of the advances in this type of cancer cures has been increased during the last decades.

Methods: 643 patients evolved by gastric cancer referred to Imam Khomeini hospital of Sari (2007- 2013) were studied. According to this method, the numbers of 74 patients were neglected because of defective data, and the number of 569 patients went under study. The level of survival was determined by use of Kaplan Meier, so to determine the causes affecting on the patients' survival, the univariate analysis of Log-rank test was used.

Results: Regarding the follow up of these patients during 2013 Nov-Dec the one, 2, 3, 4 and 5 years of survival of these patients were estimated equal to 0.77, 0.65, 0.52, 0.44, 0.27 percent and the survival median equal to 19 months, so survival means equal to 24.49 months. Based on the ranked logarithm test and FDR method some variables like stage (p<0.001) Radiotherapy (p<0.005) and undergo Surgery before Adjuvant chemotherapy (p<0.001) were determined as the effective factors on the survival probability.

Conclusion: The life length of the patients under this article in comparison with developed countries is shallow that might be because of late reference or delayed diagnosis and the shortness of cure facilities. In this way, some materials like soon diagnosis and screen methods could be effective on the increase in patients' survival.

Keywords: Gastric cancer, Survival analysis, Kaplan Meier estimate.

(Please cite as Nikpour A, Khalilian A, Maleki I, Mohsenipouya H, Yazdani Charati J. Survival of gastric cancer patients based on pathologic and demographic characteristics in Mazandaran between 2007 and 2013. Gastroenterol Hepatol Bed Bench 2019;12(4):315-321).

Aim: To evaluate the expression of the growth arrest-specific 8 (GAS8) and its antisense (GAS8-AS1) in gastric cancer.

Background: GAS8 exists in a genomic region that is recurrently deleted in breast and prostate cancer. This gene contains a long non-coding RNA, namely GAS8-AS1 whose roles in the regulation of GAS8 has been reported in hepatocytes. GAS8-AS1 has also been regarded as a putative tumor suppressor gene in papillary thyroid cancer and hepatocellular carcinoma.

Methods: In the present study, we evaluated expression levels of GAS8 and GAS8-AS1 in 30 gastric cancer tissues and their corresponding adjacent non-cancerous tissues (ANCTs).

Results: GAS8 was significantly down-regulated in tumor tissues compared to ANCTs (Expression ratio=0.29, p<0.001). Although the expression of GAS8-AS1 was higher in tumor tissues compared to ANCTs (Expression ratio=2.15), it did not reach the level of significance (p=0.12). GAS8 expression was associated with the site of the primary tumor (p=0.01). GAS8-AS1 expression was significantly higher in tumors with lymphatic/ vascular invasion compared with those without lymphatic/ vascular invasion (p=0.03). Significant pairwise correlations were detected between expression levels of GAS8 and GAS8-AS1 in tumor tissues and ANCTs. Based on the results of the ROC curve, the diagnostic power of transcript levels of GAS8 in gastric tissues was estimated to be 76%.

Conclusion: The current study underscores the roles of GAS8 and GAS8-AS1 in gastric carcinogenesis and warrants future functional studies to unravel the underlying mechanism of such contribution.

Keywords: GAS8, RNA, Long noncoding, Gastric cancer.

(Please cite as Esfandi F, Mohammad Rezaei F, Taheri M, Naby Gol M, Kholghi Oskooei V, Namvar A, et al. GAS8 and GAS8-AS1 expression in gastric cancer. Gastroenterol Hepatol Bed Bench 2019;12(4):322-327).

Aim: This paper aimed to identify new candidate biomarkers in blood for early diagnosis of CRC.                                                            

Background: Colorectal cancer (CRC) is the third most widespread malignancies increasing globally. The high mortality rate associated with colorectal cancer is due to the delayed diagnosis in an advanced stage while the metastasis has occurred. For better clinical management and subsequently to reduce mortality of CRC, early detection biomarkers are in high demand.

Methods: A 2D-PAGE separation of proteins was performed followed by tandem mass Spectrometry (MALDI-TOF-TOF) to discover potential plasma protein markers for CRC and AA (advanced adenomas). Furthermore, western blot method was used to confirm a part of the results in colorectal tissue samples.

Results: The significantly altered proteins including HPR, HP, ALB, KRT1, APOA1, FGB, IGJ and C4A were down-regulated in polyp relative to normal, and CRC compare to polyp surprisingly, and inversely, ORM2 was up-regulated with the fold change ? 2 and p-value ? 0.05. We also surveyed APOA1, FGB, and C4A for further confirmation of their expression changes by western blotting. All three of them showed a decreasing trend from normal toward CRC tissue samples as it mentioned before, but just changes of FGB and C4A were significant.

Conclusion: The results demonstrated that plasma proteins can be less invasive markers for the detection of CRC. FGB and C4A can be considered as plasma potential biomarkers to early diagnosis of CRC patients and understanding the underlying procedures in tumorigenesis. Undoubtedly, the additional study must be conducted on large scale cohorts to verify the results.

Keywords: Colorectal cancer, Advanced adenomatous polyp, Early detection, Plasma biomarker, proteomics.

(Please cite as Fayazfar S, Zali H, Arefi Oskouie A, Asadzadeh Aghdaei H, Rezaei Tavirani M, Nazemalhosseini Mojarad E. Early diagnosis of colorectal cancer via plasma proteomic analysis of CRC and advanced adenomatous polyp. Gastroenterol Hepatol Bed Bench 2019;(4):328-339).

Aim: Identification of the important processes and the related genes that are dis-regulated in the celiac disease (CD) was the aim of this study.

Background: Celiac disease is an autoimmune disorder which is characterized by immune reaction response mostly to wheat gluten. The gluten-free diet is the best-known treatment of the patients.

Methods: Significant differentially expressed proteins (DEPs) related to the CD are extracted from a published proteomics study and are included in protein-protein interaction PPI) network analysis by Cytoscape software and its applications. The central proteins and related processes are identified and discussed.

Results: Among 53 queried genes, 51 individuals were recognized by the database, and after network construction, 48 ones included in the network, and three genes remained as isolated nodes. Following 50 neighbors, the network was analyzed, and eight central genes were identified as dis-regulated elements. Related processes and the role of the central genes in celiac are discussed in detail.

Conclusion: CAT, ENO1, PCK2, ACO2, ALDOOB, GALM, ADA, and ACTBADA as critical genes and Antioxidant activity, carbohydrate metabolism, inflammation, cell growth processes are highlighted as the dis-regulated individuals in CD.

Keywords: Celiac disease, Anti-oxidant, Metabolism, Inflammation, Cell growth.

(Please cite as KhalKhal E, Rezaei-Tavirani M, Akbari Z, Rezaei-Tavirani S, Zali H, Rostamii-Nejad M. The critical role of dysregulation of antioxidant activity and carbohydrate metabolism in celiac disease. Gastroenterol Hepatol Bed Bench 2019;12(4):340-347).

Aim: This article aimed to analyze the diarrheagenic potential of E. coli isolated from urinary tract infection (UTI) and to recognize the presence of antibiotic resistance genes.

Background: The marked genome plasticity of Escherichia coli has allowed the emergence of resistant pathogenic strains displaying an unusual arrangement of genes.

Methods: In this cross-sectional study, 110 E. coli were isolated from patients with the symptoms of UTI in Sanandaj, west of Iran between July and September - 2015. The isolates were examined by the disk diffusion method for antibiotic susceptibility test and by polymerase chain reaction for the presence of genes characteristic of diarrheagenic E. coli (DEC), Uropathogenic E. coli (UPEC) virulence genes, extended-spectrum ?-lactamase bla_CTX-M and plasmid-mediated quinolone resistance determinants, qnrA, qnrB, and qnrS.

Results: The most and the least effective antibiotics were nitrofurantoin and cefotaxime (96.4% and 27.3% sensitivity, respectively). Of the 110 UTI isolates, 57.3% carried diarrheagenic genes. The bundle-forming pilus bfpA was the most prevalent diarrheagenic gene (39.1%). The most commonly detected DEC pathotype was enterotoxigenic E. coli (-ETEC, 12.7%). All the pathotypes carried the bla_CTX-M and qnr. The -UPEC hly hemolysin and  pap adhesin genes were mainly detected among ETEC isolates

Conclusion: Our results indicated the presence of resistant diarrheagenic pathotypes in UTI-associated E. coli. Such isolates may have the capacity of causing both extraintestinal and intestinal infections. Based on our knowledge, this is the first report of the presence of qnr in ETEC from urine.

Keywords: Diarrheagenic Escherichia coli, Resistance, Urinary tract infection, Virulence factors.

(Please cite as Derakhshan S, Farhadifar F, Roshani D, Ahmadi A, Haghi F. Study on the presence of resistant diarrheagenic pathotypes in Escherichia coli isolated from patients with urinary tract infection. Gastroenterol Hepatol Bed Bench 2019;12(4):348-357).

Aim: The aim of this study was to analyze the Clostridium difficile and their toxins in cancerous tissues in comparison to their adjacent healthy tissues in patients with colorectal cancer (CRC) in Iran.

Background: Intestinal infection or colonization by microbial pathogens and their released metabolites may have a role in the exacerbation of CRC.

Methods: A total of 60 biopsy samples from 30 cancerous and 30 adjacent healthy tissues were collected from patients with CRC. Biopsies were homogenized and cultured in cycloserine cefoxitin fructose agar-agar medium to investigate the presence of C. difficile. DNA was extracted, PCR was performed on pure colonies for bacteria detection, and toxin genes were evaluated in each bacterium positive cases. Real-time PCR was performed on extracted DNA for quantitative comparison of Clostridium difficile in healthy and tumor tissues in CRC patients.

Results: Clostridium difficile was isolated from 18 of the cancerous tissue (60%) and 6 of their healthy adjacent tissue (20%) in the culture medium, but toxin genes were positive just in one sample in both groups. Real-time PCR showed the colonization in all samples.

Conclusion: This study showed a higher prevalence of Clostridium difficile in cancerous lesions in comparison to healthy tissues. We suggest that the investigation of the rate of CD of colorectal cancer patients before surgery is critical for patients. Further studies with more samples size to study the importance of this bacterium and its toxins in the investigation of colorectal cancer patients survey is recommended.

Keywords: CRC, Intestinal microbiota, Toxin, Clostridium difficile.

(Please cite as Jahani-Sherafat S, Azimirad M, Alebouyeh M, Ahmadi Amoli H, Hosseini P, Ghasemian-Safaei, Sharareh Moghim H. The rate and importance of Clostridium difficile in colorectal cancer patients. Gastroenterol Hepatol Bed Bench 2019;12(4):358-363).

Aim: This study aimed to investigate the presence of oocyst of Cryptosporidium sp., and egg of soil-transmitted helminths in market vegetables in the north of Iran.

Background: Fecal-oral transmission via consumption of contaminated food is the main route of transmission of intestinal parasites. Concerning the high risk of contamination of vegetable with intestinal parasites, raw consumption of crops can enhance the chance of transmission of intestinal parasites.

Methods: In this study, we collected 34 pre-washed vegetable samples including spinach, mint, parsley, oregano, chives, savory, radish, coriander, basil and tarragon from local markets in Tonekabon City, North of Iran. All vegetable samples were washed using sterile PBS. Parasitological examinations, including direct examination and staining with Lugol’s iodine and modified Ziehl–Neelsen were performed on the pellet resulted from the washing process.

Results: The findings showed that 14/34 (41.17%) of collected samples were contaminated with at least one parasite. Eggs of Toxocara sp., Ascaris sp., Fasciola sp., Toxoascaris leonine, Trichuris sp. and Enterobius together with free-living larvae, amoeba cyst, cyst of Entamoeba coli and oocyst of Cryptosporidium sp., were observed among the positive samples. Furthermore, statistical analysis indicated that there was no significant correlation between parasitic contamination of vegetables and seasonal changes.

Conclusion: This study signifies that some parasites due to their resistant cell wall usually remain in an environment with the harsh condition and thus, consumption of raw vegetables increases the risk of transmission of them.

Keywords: Iran, North of Iran, Vegetables, Parasitic contamination.

(Please cite as Taghipour A, Javanmard E, Haghighi A, Mirjalali H, Zali MR. The occurrence of Cryptosporidium sp., and eggs of soil-transmitted helminths in market vegetables in the north of Iran. Gastroenterol Hepatol Bed Bench 2019;12(4):364-369).

Malignancy can induce a hypercoagulable state and lead to an increased risk of thromboembolic events. The pathogenesis of the prothrombotic state in cancer is complicated but is thought to involve several mechanisms. Thrombosis predominantly affects the venous circulation and infrequently the arteries. Arterial occlusion as an initial manifestation of acute leukemia is unusual. This is a case of a 44-year-old male admitted with complete thrombotic occlusion of the superior mesenteric artery and treated with emergent thrombectomy. Hematologic work-up was consistent with a diagnosis of T-cell acute lymphoblastic leukemia. To our knowledge, this is the first case of complete occlusion of the superior mesenteric artery presenting as the initial manifestation of T-cell acute lymphoblastic leukemia.

Keywords:  Mesenteric ischemia, Leukemia, Arterial occlusion

(Please cite as Krill T, Baliss M, Zaibaq J, M. Abdulla H, Parupudi S. Acute lymphoblastic leukemia presenting with mesenteric ischemia. Gastroenterol Hepatol Bed Bench 2019;12(4):370-373).

Recent advances in endoscopy