Project description:The gut and local esophageal microbiome progressively shift from healthy commensal bacteria to inflammatory-linked pathogenic bacteria in patients with gastroesophageal reflux disease, Barrett’s esophagus and esophageal adenocarcinoma (EAC). However, mechanisms by which microbial communities contribute to reflux-driven EAC remain incompletely understood and challenging to target. Herein, we utilized a rat reflux-induced EAC model to investigate targeting the gut microbiome-esophageal metabolome axis with cranberry proanthocyanidins (C-PAC) to inhibit EAC progression. Sprague Dawley rats, with or without reflux-induction received water or C-PAC ad libitum (700 µg/rat/day) for 25 or 40 weeks. C-PAC exerted prebiotic activity abrogating reflux-induced dysbiosis, and mitigating bile acid metabolism and transport, culminating in significant inhibition of EAC through TLR/NF-κB/TP53 signaling cascades. At the species level, C-PAC mitigated reflux-induced pathogenic bacteria (Streptococcus parasanguinis, Escherichia coli, and Proteus mirabilis). C-PAC specifically reversed reflux-induced bacterial, inflammatory and immune-implicated proteins and genes including Ccl4, Cd14, Crp, Cxcl1, Il6, Il1β, Lbp, Lcn2, Myd88, Nfkb1, Tlr2 and Tlr4 aligning with changes in human EAC progression, as confirmed through public databases. C-PAC is a safe promising dietary constituent that may be utilized alone or potentially as an adjuvant to current therapies to prevent EAC progression through ameliorating reflux-induced dysbiosis, inflammation and cellular damage.
Project description:Transcriptional profiling of adult esophageal epithelium comparing wild-type mice with Nrf2-/- mice with or without gastroesophageal reflux for 4 weeks. Goal was to determine the role of Nrf2 on the barrier function of mouse esophageal epithelium. Two-class comparisons. Wild-type/without reflux vs. Nrf2-/-/without reflux; Wild-type/gastric reflux vs. Nrf2-/-/gastric reflux; Wild-type/duodenal reflux vs. Nrf2-/-/duodenal reflux; Wild-type/mixed reflux vs. Nrf2-/-/mixed reflux. Biological replicates: 3 replicates for each group.
Project description:Transcriptional profiling of adult esophageal epithelium comparing wild-type mice with Nrf2-/- mice with or without gastroesophageal reflux for 4 weeks. Goal was to determine the role of Nrf2 on the barrier function of mouse esophageal epithelium.
Project description:Non-Erosive Reflux Disease (NERD) is one of the most prominent and common forms of gastroesophageal reflux disease (GERD). We performed transcriptomic analysis (RNA sequencing) of esophageal biopsies from patients with NERD and healthy controls to increase understanding of complex cellular and molecular pathways in NERD.
Project description:Background and Purpose: The current standard of care for locally advanced esophageal and gastroesophageal junctional cancer is neoadjuvant chemoradiation (NCRT) followed by surgery. The genomic and proteomic pathways responsible for response to neoadjuvant chemoradiation are sparsely described, and thus response to treatment cannot be reliably predicted. In this study, we performed an in-depth proteomic analysis of esophageal and gastroesophageal tumors, to describe differences in pathway activation between patients with good and poor prognosis following neoadjuvant chemoradiation. Materials and Methods: This study included locally advanced esophageal and gastroesophageal cancer patients treated with NCRT. The study cohort was dichotomized into two groups of patients- good prognosis (GP) and bad prognosis (BP) according to the post-operative disease-free interval. We performed a mass spectrometry analysis of proteins extracted from the malignant regions of surgical specimens and analyzed data from electronic medical records. Clinical data was correlated with differences in protein expression between GP and BP using validated gene expression pathways. Results: The study included thirty-five patients with adenocarcinoma. GP and BP had statistically significant differences in protein expression patterns. GP exhibited differential enrichment of pathways related to cellular respiration, oxidative phosphorylation and proteins of the RAS oncogene family. Conclusion: In this study we identify enrichment of pathways related to oxidative phosphorylation and RAS oncogene pathway in esophageal cancer patients with a favorable response to NCRT. Larger transcriptomic studies are warranted to portray potential surrogate signature of biomarkers based upon these potential pathways.
Project description:The major aetiological risk factor for Barrett's oesophagus and oesophageal adenocarcinoma is gastroesophageal reflux. This study's aim was to identify genes involved in the celular response to reflux in vitro. The Barrettâ??s oesophagus cell line, CP-A hTERT, was exposed to media with acid, deoxycholic acid or a primary bile salt mixture. RNA expression was compared with controls on Affymetrix U133 Plus 2.0 arrays. In CP-A hTERT, the greatest number of changes in gene expression was observed after treatment with deoxycholic acid, pH 4.5; 152 genes were up-regulated at 2 hours (91 at 6 hours) and 10 down-regulated at 2 hours (34 at 6 hours). 12 genes were identified and were subsequently assessed in patients with non-erosive reflux disease, oesophagitis, Barrett's oesophagus and oesophageal adenocarcinoma; Background and Aims: The major etiological risk factor for Barrettâ??s esophagus and esophageal adenocarcinoma is gastro-esophageal reflux. This studyâ??s aim was to identify genes involved in the cellular response to components of reflux both in vitro and in patients with reflux-related disease. Methods: The Barrettâ??s cell line, CP-A hTERT, was exposed to media with acid, deoxycholic acid or a primary bile salt mixture. RNA expression was compared with controls on Affymetrix U133 Plus 2.0 arrays. 12 genes of interest were analysed by Real Time PCR both in cell line and biopsies from 110 patients with non-erosive reflux disease, esophagitis, Barrettâ??s esophagus and esophageal adenocarcinoma. Results: In CP-A hTERT, the greatest number of changes in gene expression was observed after treatment with deoxycholic acid, pH 4.5. Of 12 genes analysed in biopsies, 10 were significantly different between the 4 groups with the largest change for anterior gradient homolog 2, which may modulate p53 function. This had highest expression in biopsies from Barrettâ??s esophagus (median gene fold change for Barrettâ??s esophagus versus non-erosive reflux disease, 411.2 (95% CI 290.5-682.7; p<0.01); esophageal adenocarcinoma versus non-erosive reflux disease 68.1 (20.5-161.4; p<0.01)). In addition 4 genes associated with development/differentiation were upregulated in Barrettâ??s biopsies compared to those from non-erosive reflux disease (SEL1L, MFNG, CRIP1 and EFNA1). Conclusions: Novel genes have been identified, whose expression is altered after acid and bile exposure in vitro and in biopsies from patients with reflux related diseases. These genes may have utility as biomarkers of response to reflux and should be assessed in prospective studies. Experiment Overall Design: The Barrett's oesophagus cell line CP-A hTERT was treated with a 15 minute exposure of acid (pH 4.5), a mixture of primary bile acids (pH 4.5) or deoxycholic acid (pH 4.5). RNA extraction occurred in treatment and non-treated cells at 2 hours and 6 hours. The treatments were performed in duplicate on 2 different days. RNA was compared in each treatment to each control at the relevant time points, in a 2 x 2 manner by using Affymetrex U133 Plus 2.0 arrays. Results of 12 genes were confirmed by Real Time PCR and were subsequently assessed in patients with non-erosive reflux disease, oesophagitis, Barrett's oesophagus and oesophageal adenocarcinoma.
Project description:Identification of Novel Immune Cell Signature in Gastroesophageal Reflux Disease: Altered Mucosal Mast Cells and Dendritic Cell Profile. The mechanisms underlying the most troublesome symptom of gastroesophageal reflux disease (GERD), heartburn, remain incompletely understood. The pathogenesis of heartburn in GERD is likely to involve not only central mechanisms of sensitization including hypervigilance, but also multiple mucosal factors including maintenance of epithelial barrier integrity via tight junction proteins, expression of acid-sensing ion channels on nerve endings, and mucosal inflammation . We hypothesized that immune cell components play a regulatory role in mucosal mechanisms of sensitization in GERD, and aimed to identify differences in the immune cell signature and sensory mucosal markers between reflux phenotypes and healthy asymptomatic subjects.
Project description:<p>The distal esophagus is an important anatomical area where gastric acid reflux can cause reflux esophagitis (RE), Barrett's esophagus (BE) (intestinal metaplasia), and esophageal adenocarcinoma (EA). The incidence of EA has increased 6-fold in the U.S. since the 1970s, parallel to a significant increase in the prevalence of gastroesophageal reflux diseases (GERD). Although specific host factors might predispose one to disease risk, such a rapid increase in incidence must be predominantly environmental. The cause remains unknown. Our hypothesis is that changes in the foregut microbiome are associated with EA and its precursors, RE and BE in the GERD sequence.</p> <p>We will conduct a case control study to characterize the microbiome in every stage of the GERD sequence as well as analyze the trend in changes in the microbiome along disease progression toward EA.</p> <p>Specific Aim 1. To conduct a comprehensive population survey of the foregut microbiome and demonstrate its association with GERD sequence, by a 16S rRNA gene survey. We will analyze samples of the foregut microbiome at three anatomic loci: mouth, distal esophagus, and gastric corpus. Changes of the microbiota in the distal esophagus will be correlated with the phenotypes. Spatial relationship between the esophageal microbiota and upstream (mouth) and downstream (stomach) foregut microbiotas as well as temporal stability of the microbiome-disease association will also be examined.</p> <p>Specific Aim 2. To define distal esophageal metagenome and demonstrate its association with GERD sequence, by shotgun metagenomic analysis. We will first classify samples of the metagenome into metagenotypes by between-sample k-mer distance and correlate the metagenotypes with the four phenotypes. Subsequent detailed analyses will include pathway-disease and gene-disease associations. DNA viruses and fungi, if identified, also will be correlated with the phenotypes.</p> <p>A significant association between the foregut microbiome composition and GERD sequence, if demonstrated, will be the first step for eventually testing the causal hypothesis that an abnormal microbiome is required for the development of the sequence of phenotypic changes toward EA. If EA and its precursors represent a microbial ecological disease, treating the cause of GERD might become possible, for example, by normalizing the microbiome through use of antibiotics, probiotics, or prebiotics. Causative therapy for GERD could prevent its progression and reverse the current trend of increasing incidence of EA.</p>
Project description:The major aetiological risk factor for Barrett's oesophagus and oesophageal adenocarcinoma is gastroesophageal reflux. This study's aim was to identify genes involved in the celular response to reflux in vitro. The Barrett’s oesophagus cell line, CP-A hTERT, was exposed to media with acid, deoxycholic acid or a primary bile salt mixture. RNA expression was compared with controls on Affymetrix U133 Plus 2.0 arrays. In CP-A hTERT, the greatest number of changes in gene expression was observed after treatment with deoxycholic acid, pH 4.5; 152 genes were up-regulated at 2 hours (91 at 6 hours) and 10 down-regulated at 2 hours (34 at 6 hours). 12 genes were identified and were subsequently assessed in patients with non-erosive reflux disease, oesophagitis, Barrett's oesophagus and oesophageal adenocarcinoma Background and Aims: The major etiological risk factor for Barrett’s esophagus and esophageal adenocarcinoma is gastro-esophageal reflux. This study’s aim was to identify genes involved in the cellular response to components of reflux both in vitro and in patients with reflux-related disease. Methods: The Barrett’s cell line, CP-A hTERT, was exposed to media with acid, deoxycholic acid or a primary bile salt mixture. RNA expression was compared with controls on Affymetrix U133 Plus 2.0 arrays. 12 genes of interest were analysed by Real Time PCR both in cell line and biopsies from 110 patients with non-erosive reflux disease, esophagitis, Barrett’s esophagus and esophageal adenocarcinoma. Results: In CP-A hTERT, the greatest number of changes in gene expression was observed after treatment with deoxycholic acid, pH 4.5. Of 12 genes analysed in biopsies, 10 were significantly different between the 4 groups with the largest change for anterior gradient homolog 2, which may modulate p53 function. This had highest expression in biopsies from Barrett’s esophagus (median gene fold change for Barrett’s esophagus versus non-erosive reflux disease, 411.2 (95% CI 290.5-682.7; p<0.01); esophageal adenocarcinoma versus non-erosive reflux disease 68.1 (20.5-161.4; p<0.01)). In addition 4 genes associated with development/differentiation were upregulated in Barrett’s biopsies compared to those from non-erosive reflux disease (SEL1L, MFNG, CRIP1 and EFNA1). Conclusions: Novel genes have been identified, whose expression is altered after acid and bile exposure in vitro and in biopsies from patients with reflux related diseases. These genes may have utility as biomarkers of response to reflux and should be assessed in prospective studies. Keywords: Acid (pH 4.5) and bile (mixture of primary bile salts or the secondary bile salt deoxycholic acid, both at pH 4.5) challenge to a Barrett's oesophagus cell line. RNA extraction at 2 and 6 hours. Comparison of treatment RNA to control (non-treatment) RNA,
Project description:The involvment of bile acids such as deoxycholic acid (DCA) in gastro-esophageal reflux disease and subsequent Barrettâ??s metaplsia has been postulated. This study examines gene expression induced by exposure to DCA in esophageal cells and may be utilised in cross-comparisions with data derived from gene expression studies of Barrettâ??s esophagus and associated adenocarcinoma. Additionally this study may be used to assess divergence in response to bile acids by comparisons with similar study performed in SKGT4 barrett''s assocaited adenocarcinoma cell line. HET-1A cells were exposed to 300um DCA over 24 hours in duplicate experiments including matched timepoint controls