SARS-CoV-2 infection and replication in human gastric organoids.

COVID-19 typically manifests as a respiratory illness, but several clinical reports have described gastrointestinal symptoms. This is particularly true in children in whom gastrointestinal symptoms are frequent and viral shedding outlasts viral clearance from the respiratory system. These observations raise the question of whether the virus can replicate within the stomach. Here we generate gastric organoids from fetal, pediatric, and adult biopsies as in vitro models of SARS-CoV-2 infection. To facilitate infection, we induce reverse polarity in the gastric organoids. We find that the pediatric and late fetal gastric organoids are susceptible to infection with SARS-CoV-2, while viral replication is significantly lower in undifferentiated organoids of early fetal and adult origin. We demonstrate that adult gastric organoids are more susceptible to infection following differentiation. We perform transcriptomic analysis to reveal a moderate innate antiviral response and a lack of differentially expressed genes belonging to the interferon family. Collectively, we show that the virus can efficiently infect the gastric epithelium, suggesting that the stomach might have an active role in fecal-oral SARS-CoV-2 transmission.

Perspective: the nose and the stomach play a critical role in the NZACE2-Patari* (modified ACE2) drug treatment project of SARS-CoV-2 infection.

Background: COVID-19 has caused calamitous health, economic and societal consequences globally. Currently, there is no effective treatment for the infection. Areas covered: We have recently described the NZACE2-Patari project, which seeks to administer modified Angiotensin Converting Enzyme 2 (ACE2) molecules early in the infection to intercept and block SARS-CoV-2 binding to the pulmonary epithelium. Expert opinion: Since the nasopharyngeal mucosa is infected in the first asymptomatic phase of the infection, treatment of the nose is likely to be safe and potentially effective. The intercepted virus will be swallowed and destroyed in the stomach. There is however a limited window of opportunity to alter the trajectory of the infection in an individual patient, which requires access to rapid testing for SARS-CoV-2. The proposed strategy is analogous to passive immunization of viral infections such as measles and may be of particular benefit to immunodeficient and unvaccinated individuals.

Ephrin receptor A2, the epithelial receptor for Epstein-Barr virus entry, is not available for efficient infection in human gastric organoids.

Epstein-Barr virus (EBV) is best known for infection of B cells, in which it usually establishes an asymptomatic lifelong infection, but is also associated with the development of multiple B cell lymphomas. EBV also infects epithelial cells and is associated with all cases of undifferentiated nasopharyngeal carcinoma (NPC). EBV is etiologically linked with at least 8% of gastric cancer (EBVaGC) that comprises a genetically and epigenetically distinct subset of GC. Although we have a very good understanding of B cell entry and lymphomagenesis, the sequence of events leading to EBVaGC remains poorly understood. Recently, ephrin receptor A2 (EPHA2) was proposed as the epithelial cell receptor on human cancer cell lines. Although we confirm some of these results, we demonstrate that EBV does not infect healthy adult stem cell-derived gastric organoids. In matched pairs of normal and cancer-derived organoids from the same patient, EBV only reproducibly infected the cancer organoids. While there was no clear pattern of differential expression between normal and cancer organoids for EPHA2 at the RNA and protein level, the subcellular location of the protein differed markedly. Confocal microscopy showed EPHA2 localization at the cell-cell junctions in primary cells, but not in cancer cell lines. Furthermore, histologic analysis of patient tissue revealed the absence of EBV in healthy epithelium and presence of EBV in epithelial cells from inflamed tissue. These data suggest that the EPHA2 receptor is not accessible to EBV on healthy gastric epithelial cells with intact cell-cell contacts, but either this or another, yet to be identified receptor may become accessible following cellular changes induced by inflammation or transformation, rendering changes in the cellular architecture an essential prerequisite to EBV infection.

ARID1A deficiency in EBV-positive gastric cancer is partially regulated by EBV-encoded miRNAs, but not by DNA promotor hypermethylation.

AT-rich interactive domain 1A (ARID1A), which is a tumor suppressor gene, is frequently mutated in Epstein-Barr virus-positive gastric cancer [EBV (+) GC]. While most ARID1A mutations in GC are truncating mutations, leading to loss of ARID1A protein expression, epigenetic modifications appear to contribute to ARID1A deficiency in EBV (+) GC harboring wild-type ARID1A. Based on the significant role of epigenetic modifications in EBV (+) GC that contributes to ARID1A deficiency, the methylation status of ARID1A was evaluated in EBV-infected cells and GC patients using a publicly available microarray and the Cancer Genome Atlas (TCGA) database. EBV-encoded miRNAs that potentially target ARID1A were identified as an additional epigenetic modulator by computational prediction. In vitro experiments were conducted to evaluate how EBV-encoded miRNAs affected ARID1A mRNA and protein levels. In clinical GC samples, the expression of predicted miRNAs and ARID1A and the mutation status of ARID1A was evaluated. As results, ARID1A was not hypermethylated in EBV (+) GC samples or EBV-infected GC cells. EBV infection did not alter ARID1A mRNA levels, suggesting that ARID1A protein deficiency was caused by post-transcriptional gene silencing in ARID1A-WT EBV (+) GC. Overexpression of miR-BART11-3p and miR-BART12, which were identified as miRNAs that potentially bind ARID1A, suppressed ARID1A protein expression in MKN7 and NCI-N87 cells. Highly expressed miR-BART11-3p and miR-BART12 were correlated with decreased ARID1A levels in GC tumors which did not harbor ARID1A mutations. The present findings revealed that ARID1A expression was epigenetically regulated by miR-BART11-3p and miR-BART12 in EBV (+) GC.

Association between Epstein-Barr virus infection and gastric cancer: a systematic review and meta-analysis.

BACKGROUND: Numerous studies conducted over the past 30 years have pointed to the presence of Epstein-Barr virus (EBV) in gastric cancer samples. This study was aimed to provide a meta-analytic review of the prevalence of EBV in gastric cancer patients, and to clarify the relationship between EBV infection and gastric cancer. METHODS: A literature search was performed electronically using online databases for English language publications until July 1, 2019. The pooled EBV prevalence and 95% confidence intervals (CIs) were estimated using a random-effects model. To determine the association between EBV and gastric cancer, pooled odds ratio (OR) and its 95% CI were computed for case-control studies. Two separate analyses were performed on data from case-control studies with matched and non-match pairs designs to calculate the pooled estimates of ORs. RESULTS: The pooled prevalence of EBV in 20,361 gastric cancer patients was 8.77% (95% CI: 7.73-9.92%; I2 = 83.2%). There were 20 studies with matched pairs design, including tumor and tumor-adjacent normal tissue pairs from 4116 gastric cancer patients. The pooled ORs were 18.56 (95% CI: 15.68-21.97; I2 = 55.4%) for studies with matched pairs design and 3.31 (95% CI: 0.95-11.54; I2 = 55.0%) for studies with non-matched pairs design. The proportion of EBV-associated gastric cancer among male cases was significantly higher than among female cases (10.83%, vs. 5.72%) (P < 0.0001). However, the pooled OR estimate for EBV-associated gastric cancer was significantly higher among females (21.47; 95% CI: 15.55-29.63; I2 = 0%) than in males (14.07; 95% CI: 10.46-18.93; I2 = 49.0%) (P = 0.06). EBV was more prevalent in the cardia (12.47%) and the body (11.68%) compared to the antrum (6.29%) (P = 0.0002). CONCLUSIONS: EBV infection is associated with more than 18 times increase the risk of gastric cancer. Although the prevalence of EBV was higher in male patients than in female patients with gastric cancer, women are more likely than men to develop EBV-associated gastric cancer. Our findings showed that using tumor-adjacent normal tissues as the control group provides more robust and accurate results regarding the relationship between EBV infection and gastric cancer.

A Mouse Model of SARS-CoV-2 Infection and Pathogenesis.

Since December 2019, a novel coronavirus SARS-CoV-2 has emerged and rapidly spread throughout the world, resulting in a global public health emergency. The lack of vaccine and antivirals has brought an urgent need for an animal model. Human angiotensin-converting enzyme II (ACE2) has been identified as a functional receptor for SARS-CoV-2. In this study, we generated a mouse model expressing human ACE2 (hACE2) by using CRISPR/Cas9 knockin technology. In comparison with wild-type C57BL/6 mice, both young and aged hACE2 mice sustained high viral loads in lung, trachea, and brain upon intranasal infection. Although fatalities were not observed, interstitial pneumonia and elevated cytokines were seen in SARS-CoV-2 infected-aged hACE2 mice. Interestingly, intragastric inoculation of SARS-CoV-2 was seen to cause productive infection and lead to pulmonary pathological changes in hACE2 mice. Overall, this animal model described here provides a useful tool for studying SARS-CoV-2 transmission and pathogenesis and evaluating COVID-19 vaccines and therapeutics.

An improved quantitative real-time polymerase chain reaction technology for Helicobacter pylori detection in stomach tissue and its application value in clinical precision testing.

BACKGROUND: Helicobacter pylori (H. pylori) infection is a serious human health threat. The empiric H. pylori treatment paradigm guided by traditional testing technologies has led to antibiotic resistance. Here, we improved the qPCR method to provide technical support for precision H. pylori diagnosis and treatment. METHODS: Two pairs of primers and probes targeting the glmM gene were designed to detect H. pylori, and a multiplex qPCR method was established for virulence factor detection. Then, a rapid urease test (RUT), culturing and qPCR were performed on 141 specimens collected from Xinqiao Hospital of China in 2017 to evaluate the qPCR detection capability. Finally, the H. pylori infectious amount and virulence genes were detected by qPCR. RESULTS: 1. The improved qPCR method which used two pairs of primers had a higher detection rate (100%) and better accuracy (p = 0.000), compared with the qPCR using a pair of primers. It also had better consistency with the bacterial culture than with RUT (Kappa =0.440, p < 0.001). 2. The H. pylori infectious amount was significantly positively associated with gastritis in corpus (p = 0.003) and gastric erosion (p = 0.043). The H. pylori infectious amount in gastric precancerous patients was significantly lower than that in H. pylori-positive patients (p < 0.05), and the infectious H. pylori-vacA s1+ amount was significantly greater than that of H. pylori-vacA s1- (p < 0.05). 3. The vacA s1 frequency was significantly higher than that of vacA m1/cagA+/babA2+ in chronic superficial gastritis (p = 0.000), peptic ulcer (p = 0.037) and gastric erosion (p = 0.009). The H. pylori-vacA+/cagA+/babA2+ frequency showed a significant positive correlation (p < 0.05). CONCLUSIONS: The H. pylori infectious amount and presence of H. pylori virulence factors showed complex correlations with gastric disease occurrence and development. The improved qPCR with good detection performance can be used for quantitative H. pylori detection and testing for the virulence genes vacA s1, vacA m1, cagA and babA2 simultaneously. These findings will provide valuable information for disease diagnosis and treatment.

Potential prognostic impact of EBV RNA-seq reads in gastric cancer: a reanalysis of The Cancer Genome Atlas cohort.

Epstein-Barr virus (EBV)-associated gastric cancer (EBVaGC), whose prognosis remains controversial, is diagnosed by in situ hybridization of EBV-derived EBER1/2 small RNAs. In The Cancer Genome Atlas (TCGA) Stomach Adenocarcinoma (STAD) project, the EBV molecular subtype was determined through a combination of multiple next-generation sequencing methods, but not by the gold standard in situ hybridization method. This leaves unanswered questions regarding the discordance of EBV positivity detected by different approaches and the threshold of sequencing reads. Therefore, we reanalyzed the TCGA-STAD RNA sequencing (RNA-seq) dataset including 375 tumor and 32 normal samples, using our analysis pipeline. We defined a reliable threshold for EBV-derived next-generation sequencing reads by mapping them to the EBV genome with three different random arbitrary alignments. We analyzed the prognostic impact of EBV status on the histopathological subtypes of gastric cancer. EBV-positive cases identified by reanalysis comprised nearly half of the cases (49.6%) independent from infiltrating lymphocyte signatures, and showed significantly longer overall survival for adenocarcinomas of the 'not-otherwise-specified' type [P = 0.016 (log-rank test); hazard ratios (HR): 0.476; 95% CI: 0.260-0.870, P = 0.016 (Cox univariate analysis)], but shorter overall survival for the tubular adenocarcinoma type [P = 0.005 (log-rank test); HR: 3.329; 95% CI: 1.406-7.885, P = 0.006 (Cox univariate analysis)]. These results demonstrate that the EBV positivity rates were higher when determined by RNA-seq than when determined by EBER1/2 in situ hybridization. The RNA-seq-based EBV positivity demonstrated distinct results for gastric cancer prognosis depending on the histopathological subtype, suggesting its potential to be used in clinical prognoses.

Epstein-Barr virus is absent in gastric superficial neoplastic lesions.

Epstein-Barr virus (EBV) has been associated with about 9% of all gastric carcinomas, but its role in gastric carcinogenesis remains unclear since there is lack of evidence of EBV presence in pre-neoplastic lesions of gastric mucosa. This study intends to determine the prevalence of EBV in gastric dysplasia and superficial neoplasia to clarify whether EBV infection is an early or late event in gastric cancer development. This retrospective study included a total of 242 gastric lesions from 199 consecutive patients who were referred for endoscopic resection. The histological classification of lesions includes 137 low- and high-grade dysplasia and 105 superficial carcinomas. EBV infection was investigated by EBER-ISH. Results showed that EBV was not detected in any epithelial cells of any case with dysplasia or superficial carcinomas, although we observed the presence of a small number of EBV-infected lymphocytes in 2.1% of all lesions. These results showed that EBV is not present in gastric dysplasia neither in superficial carcinomas suggesting that EBV carcinogenesis is a late event in well/moderately differentiated gastric carcinogenesis.

Association between Gastric Pathology and Hepatitis B Virus Infection in Patients with or without Helicobacter Pylori.

Background: In the recent years, hepatitis B virus (HBV) infection has been considered as a risk factor for gastric cancer, but further studies are required to confirm these claim. The present study was aimed to evaluate the correlation between gastric pathology (precancerous and cancerous conditions) with HBV infection in Helicobacter pylori (H. pylori) positive or negative patients. Methods: In this cross-sectional study, 728 patients under endoscopy examination in Yazd Shaheed Sadoughi Hospital between 2017 and 2018 were participated. Histopathological analysis was performed on gastric specimens. Hepatitis B surface antigen (HBsAg) in sera was detected by the enzyme-linked immunosorbent assay (ELISA). The relationship between gastric pathology and HBV infection were explored by logistic regression analysis. Results: Of 728 patients, HBsAg and H. pylori infection were detected in 83 and 408 patients, respectively. Sixty nine patients were co-infected with H. pylori/HBV. H. pylori infection detected in patients with HbsAg positive significantly more than those were negative for HbsAg (p=0.029). None of HBV/H. pylori co-infected patients did not have normal gastric tissue. A significant difference was seen in histopathology of gastric tissue between HBsAg positive patients with and without H. pylori infection (p<0.0001). The HBsAg was associated with histopathology of gastric (OR=21.56, 95℅CI=7.070-65.741, p<0.001) and as a risk factor for gastritis (OR=12.457, 95℅CI= 3.007-51.614, P=0.001) but no cancer (OR=2.127, 95℅CI=0.242-18.704, P=0.496). Conclusion: The HBV infection alone is associated with some precancerous lesions but is not correlated with gastric cancer. It can increase development of premalignant conditions and carcinoma significantly in H. pylori positive patients.

PARP1: A potential biomarker for gastric cancer.

Gastric cancer (GC) is the third leading cause of cancer mortality worldwide, with an overall 5-y survival rate of 25%. The majority of GCs are caused by infectious agents, including the bacterium Helicobacter pylori (H. pylori) and Epstein-Barr virus (EBV). Furthermore, inappropriate repair of DNA damage can also result in genomic instability, which has shown to be a key factor in carcinogenesis of different regions including gastric region. Present study was designed to explore the association between base excision repair pathway genes, PARP1 and APEX1 and gastric pathology and H. pylori infection. Two hundred gastric cancer tissue samples (114 H. pylori positive and 86 H. pylori negative) and adjacent uninvolved area taken as controls was used for expression analysis of BER pathway genes at mRNA level and protein levels using quantitative PCR (qPCR) and immunohistochemistry (IHC) respectively. Oxidative stress and DNA damage was also determined by measuring the level of antioxidant enzymes and comet assay respectively. Significant upregulation in PARP1 (p < 0.001) and APEX1 (p < 0.02) was observed in GC tissue samples compared to controls and this upregulation was more pronounced in H. pylori positive cases (HPGC) (PARP1, p < 0.02: APEX1, p < 0.04) than H. pylori negative cases (HNGC). Upregulation of BER pathway genes in HPGC was found correlated with smoking status (p < 0.0001), T stage (p < 0.01) and lymph node metastasis (p < 0.03). Moreover, immunohistochemical staining of BER pathway genes was found correlated with a number of clinicopathological characteristics such as tumor type (p < 0.03), tumor size (p < 0.01) and lymph node metastasis (p < 0.01). Expression levels of APEX1 and PARP1 gene also correlated with increased oxidative burden (p < 0.0001) and DNA damage (p < 0.001) in GC patients. Survival analysis showed that upregulation of PARP1 gene was associated with poor overall survival outcome of gastric cancer patients (HR = 2.04 (95% CI = 1.10-3.76; p < 0.02). Univariate and multivariate cox regression analysis showed the upregulated PARP1 gene (HR = 5.03; 95%CI (2.22-11.35); p = 0.0001), positive smoking status (HR = 3.58; 95%CI (1.67-7.65); p = 0.001), positive status for H pylori infection (HR = 4.38; 95%CI (1.82-10.56); p = 0.001) and advance N-stage (HR = 5.29; 95%CI (2.28-12.24); p = 0.0001) were independent prognostic factors for gastric cancer and may serve as a valuable biomarker for the diagnosis and progression of GC and can be helpful in developing individualized treatment strategies for treating GC.

Simulated human digestion of N1-aryl-2-arylthioacetamidobenzimidazoles and their activity against Herpes-simplex virus 1 in vitro.

Drug performance in the gastrointestinal tract (GIT) plays a crucial role in determining release and absorption. In the present work, we assessed the in vitro digestion of two synthetic N1-aryl-2-arylthioacetamidobenzimidazoles (NAABs), NAAB-496 and NAAB-503, using bio-relevant models of the human stomach and small intestine. The activity of NAAB-496 and NAAB-503 against herpes simplex virus (HSV-1) replication was also investigated. NAAB-496 was resistant to pepsin in the gastric environment, with a virtual 100% recovery, which decreased to 43.2% in the small intestine. NAAB-503 was sensitive to pepsin, with 65.7% degradation after 120 min gastric phase. 1H Nuclear magnetic resonance (NMR) post in vitro digestion highlighted an alteration of NAAB-496 after the gastric phase, whereas NAAB-503 appeared comparable to the original spectral data. Both NAAB-496 and NAAB-503 revealed some antiviral activity anti-HSV-1. The 50% effective concentration (EC50) of the compounds was 0.058 mg/mL for NAAB-496 and 0.066 for NAAB-503. Future studies will evaluate the behavior of NAAB-496 within pharmaceutical formulations.

CT findings and clinical features of Epstein-Barr virus-associated lymphoepithelioma-like gastric carcinoma.

Epstein-Barr virus (EBV)-associated lymphoepithelioma-like gastric carcinoma (LELGC) is a rare primary stomach tumor, which has overlapping imaging features with mass forming gastric carcinoma (GC). The aim of our study was to present the computed tomography (CT) findings and clinical features of EBV-associated LELGC to increase awareness of this entity.The CT findings and clinical features of 4 patients with pathologically documented EBV-associated LELGC were retrospectively analyzed.Among the 4 patients, 3 were male, and 1 was female. The medium age was 51 years old. All cases were single lesions including 1 was in the gastric cardia, 1 was in the gastric body, and 2 were in the gastric antrum. A focal thickening of the gastric wall was conducted, with a large thickness-to-length ratio. The low-density stripe of the normal gastric wall abruptly terminated at the edge of the lesion. The CT images of 4 cases showed inhomogeneous density with the radiodensity values ranging from 25 to 48 HU. In addition, an ulcer was demonstrated with an irregular base and slightly raised borders in all cases. Enhancement after injection of contrast material was heterogeneous enhancement (n = 3) or homogeneous (n = 1). After enhancement, obvious enhancement was seen in 1 case, moderate enhancement was seen in 3 cases, with the peak value of the tumor in the portal phase. No evidence of lymph node involvement and distant invasion was observed.Although LELGC is quite rare, it should be considered in differential diagnosis of early GC, advanced GC, and lymphoma. The relatively typical CT appearance, combined the age and sex of patients, can suggest the diagnosis of LELGC.

Distribution of the Epstein-Barr virus in the normal stomach and gastric lesions in Thai population.

The Epstein-Barr virus (EBV) is one of the infectious agents found in stomach tissue. Recently, EBV-associated gastric carcinoma (EBVaGC) was classified as a new subtype of gastric carcinoma. To date, there is a lack of knowledge about the distribution and prevalence of EBV infection in both the normal stomach and various gastric lesions, including EBVaGC, in the Thai population. In this study, we detected EBV in the normal stomach (NS; n = 19), chronic gastritis (CG; n = 36), intestinal metaplasia (IM; n = 40), gastric dysplasia (GD; n = 15), and gastric adenocarcinoma (GC; n = 33) by polymerase chain reaction (PCR) amplification of the latent membrane protein (LMP1) gene of EBV. EBV-PCR amplification was positive in 42.1%, 36.1%, 22.5%, 13.3%, and 33.3% of NS, CG, IM, GD, and GC, respectively. For further clarification in EBVaGC, we performed EBV-encoded small RNA in situ hybridization (EBER-ISH) in PCR-positive cases of GD and GC. Four GC cases were EBER-ISH positive (12.1%), while both GD cases were EBER-ISH negative. In addition, we determined the distribution of the EBV strain (type A or B) based on EBNA3C sequence and EBV variants based on LMP1 variation (wild-type and 30-bp deletion variants; wt-LMP1 or del-LMP1). The results showed that type A and wt-LMP1 were the most prevalent in all lesions. In conclusion, EBV is common in both the NS and gastric lesions, and the frequency of EBVaGC was 12.1% in Thai patients.

Development of a Predictive Model Using Endoscopic Features for Gastric Cytomegalovirus Infection in Renal Transplant Patients.

BACKGROUND: Cytomegalovirus (CMV) is a common viral pathogen in transplant patients which often targets the stomach. However, the endoscopic characteristics of gastric CMV infection are not well established. We aimed to develop a predictive model using endoscopic findings for gastric CMV infection in renal transplant patients. METHODS: A retrospective study of 287 kidney transplant recipients who underwent endoscopy with biopsy for suspected CMV infection from January 2006 to November 2015 at a tertiary referral hospital was performed. CMV infection was defined based on inclusion bodies in hematoxylin and eosin and immunohistochemical staining. Endoscopic and clinical parameters related to gastric CMV infection were selected by univariate analyses. Multivariate logistic regression was used to create a predictive model from ß-coefficients. RESULTS: CMV was present in 107 (37.7%) of the 287 patients. Multivariate analysis found age (odds ratio [OR], 0.964; 95% confidence interval [CI], 0.938-0.99; P = 0.008), erosions with surface exudate (OR, 5.34; 95% CI, 2.687-10.612; P < 0.001), raised shape of erosions (OR, 3.957; 95% CI, 1.937-8.083; P < 0.001), and antral location of ulcers (OR, 15.018; 95% CI, 5.728-39.371; P < 0.001) as independent predictive factors for gastric CMV infection. Using the predictive model created from this analysis, the sensitivity, specificity, positive predictive value, negative predictive value, and accuracy were 71.03%, 85.56%, 74.51%, 83.24%, and 80.14%, respectively. The area under the receiver operating characteristic curve of this model for detecting CMV infection was 0.850 (95% CI, 0.803-0.889; P < 0.001). CONCLUSIONS: The predictive model with typical endoscopic findings may be useful for detecting gastric CMV infection in renal transplant patients.

Sirt1 Gene Expression and Gastric Epithelial Cells Tumor Stage in Patients with Helicobacter pylori Infection

Introduction: The World Health Organization has categorized Helicobacter pylori as a carcinogen for gastric cancer, which causes human mortality worldwide. A number of studies have shown that H. pylori affects cell signaling in gastric epithelial cells and changes the expression of some proteins such as proinflammatory cytokines. Bacterial infections may alter sirt1 and sirt2 genes expression in inflammatory tissues and cancer cells. In this study, sirt1 and sirt2 genes expression in gastric cancers was surveyed with reference to H. pylori status. Methods: Stomach biopsies were collected from 50 gastric cancer patients, 25 H. pylori-positive and 25 H. pylori-negative as determined by the urea rapid test. Tumor grade was determined by a pathologist. After total RNA extraction from gastric cancer biopsy samples and cDNA synthesis, sirt1 and sirt2 genes expression levels were determined by Real Time PCR and ΔΔCT methods. Results: There was no statistically significant link between H. pylori infection and sirt1 (P<0.899) and sirt2 (P<0.169) genes expression in gastric epithelial cells. However, pathologic findings showed that there is a statistically significant relationship between sirt1 gene expression and the tumor grade (P<0.024). Discussion: A statistically significant association was found between sirt1 gene expression and tumor grade of gastric cancers that could be due to effects on progression of cancer cells infected with H. pylori.

Clinical and pathological characterization of Epstein-Barr virus-associated gastric carcinomas in Portugal.

AIM: To determine the prevalence of Epstein-Barr virus (EBV)-associated gastric carcinomas in the North Region of Portugal and to study its clinicopathological characteristics. METHODS: We have performed a retrospective study including a total of 179 consecutive patients with gastric cancer (GC) submitted to gastrectomy during 2011 at the Portuguese Oncology Institute of Porto. Clinical and pathological data was collected from individual clinical records and inserted on a database with unique codification. Tumour tissues were collected from the institutional tumour bank. EBV was detected by in situ hybridization for the detection of EBV-encoded small RNAs (EBERs) and EBV latent proteins (LMP1 and LMP2A) were detected by immunohistochemistry. RESULTS: The analysis showed that EBV-associated gastric carcinomas (EBVaGC) represents 8.4% (15/179) of all GC cases, with a significant differential distribution among histological types (P < 0.001): 100% (3/3) of medullary carcinomas, 100% (1/1) of adenosquamous carcinoma, 8.7% (8/92) of tubular adenocarcinomas, 8.0% (2/25) of mixed carcinomas and 2% (1/51) in poorly cohesive carcinomas. The analysis revealed a higher predominance of EBVaGC in the upper third and middle (cardia, fundus and body) of the stomach (P = 0.041), a significant lower number of regional lymph nodes invasion (P = 0.025) and a tendency for better prognosis (P = 0.222). EBV latent protein expression revealed that all EBVaGC cases were LMP1-negative, nevertheless 6 cases (40%) expressed LPM2A, which reveals that these cases show a distinct EBV-Latency profile (latency II-like). CONCLUSION: EBVaGC represents 8.4% of all GC in the North Region of Portugal. The EBV-infected patients have specific clinic-pathological features that should be further explored to develop new strategies of management and treatment.

Gastrosplenic fistula occurring in lymphoma patients: Systematic review with a new case of extranodal NK/T-cell lymphoma.

AIM: To provide the overall spectrum of gastrosplenic fistula (GSF) occurring in lymphomas through a systematic review including a patient at our hospital. METHODS: A comprehensive literature search was performed in the MEDLINE database to identify studies of GSF occurring in lymphomas. A computerized search of our institutional database was also performed. In all cases, we analyzed the clinicopathologic/radiologic features, treatment and outcome of GSF occurring in lymphomas. RESULTS: A literature search identified 25 relevant studies with 26 patients. Our institutional data search added 1 patient. Systematic review of the total 27 cases revealed that GSF occurred mainly in diffuse, large B-cell lymphoma (n = 23), but also in diffuse, histiocytic lymphoma (n = 1), Hodgkin's lymphoma (n = 2), and NK/T-cell lymphoma (n = 1, our patient). The common clinical presentations are constitutional symptoms (n = 20) and abdominal pain (n = 17), although acute gastrointestinal bleeding (n = 6) and infection symptoms due to splenic abscess (n = 3) are also noted. In all patients, computed tomography scanning was very helpful for diagnosing GSF and for evaluating the lymphoma extent. GSF could occur either post-chemotherapy (n = 10) or spontaneously (n = 17). Surgical resection has been the most common treatment. Once patients have recovered from the acute illness status after undergoing surgery, their long-term outcome has been favorable. CONCLUSION: This systematic review provides an overview of GSF occurring in lymphomas, and will be helpful in making physicians aware of this rare disease entity.

Cytokine expression associated with Helicobacter pylori and Epstein-Barr virus infection in gastric carcinogenesis.

Helicobacter pylori and Epstein-Barr virus (EBV) infection, and associated cytokines are involved in gastric carcinogenesis. We investigated the expression of these cytokines and their relationship with clinicopathological characteristics. The study included specimens from 207 patients with gastric adenocarcinoma, 56 with chronic gastritis, 32 with metaplasia, and 30 with low-grade epithelial dysplasia. Tissue microarrays were constructed and immunohistochemical staining for IL-1ß, IL-6, IL-10, IL-17, p16, p21, TNF-α, and TNFR1 was performed. EBV and H. pylori infection status was determined. IL-1ß, IL-6, IL-17, p16, and p21 protein expression was significantly higher in adenocarcinoma cases than in the other cases (p < 0.05). EBV was only noted in adenocarcinoma (13 cases, 6.3%). The H. pylori infection rate in adenocarcinoma was significantly higher than that in the other cases (p < 0.005). IL-6 expression was associated with improved survival (p < 0.05), whereas IL-17 expression was associated with decreased survival (p < 0.05). IL-6 expression was inversely associated with angioinvasion, and disease stage (p < 0.05), whereas IL-17 expression was associated with disease stage (p < 0.05). IL-10 expression was correlated with IL-1ß and TNF-α expression, and p16 expression was correlated with IL-17 and EBV status. Our results indicate that IL-6 and IL-17 are associated with gastric carcinogenesis and may be considered prognostic factors.

Lymphoepithelioma-like gastric carcinoma: clinicopathological characteristics and infection status.

BACKGROUND: Lymphoepithelioma-like gastric carcinoma (LLGC) is a rare subtype of gastric carcinoma (GC) characterized by prominent lymphocytic infiltration. LLGC may be associated with latent Epstein-Barr virus (EBV) infection or microsatellite instability (MSI). This study aims to assess the clinicopathological characteristics, EBV infection, and MSI status in LLGC. METHODS: A retrospective analysis of GC patients submitted to potentially curative resection between 2009 and 2014 was performed. The LLGC subtype specimens were examined for EBV by in situ hybridization and MSI by immunohistochemical analysis. The LLGC profile was analyzed accordingly to clinicopathological parameters. RESULTS: From 255 patients, seven were identified on the pathological report as LLGC. Six cases were EBV-positive and one had MSI, showing loss of MLH1 and PMS2 expression. LLGC was more frequently seen in men, and the mean age was 69 years. When compared to non-LLGC, LLGC cases were larger (∼5.8 cm) poorly differentiated tumors and had lower incidence of lymph node metastasis (P = 0.045). Mean number of lymph nodes dissected in the LLGC group was 39.5, and only one patient had a single positive lymph node. In addition, two patients presented associated lesions. LLGC was not associated with HER-2, chromogranin and synaptophysin positivity or Helicobacter pylori infection. CONCLUSIONS: Distinct pathological aspects and clinical behavior of LLGC reinforce the need for proper recognition of this histological subtype to choose better therapeutic approaches.