Kimberly Woodward

Clinical and endoscopic characteristics do not reliably differentiate PPI-responsive esophageal eosinophilia and eosinophilic esophagitis in patients undergoing upper endoscopy: A Prospective Cohort Study

OBJECTIVES:Proton-pump inhibitor-responsive esophageal eosinophilia (PPI-REE) is a newly recognized entity that must be differentiated from eosinophilic esophagitis (EoE). Little is known about this condition. We aimed to determine the prevalence of PPI-REE and EoE in patients undergoing upper endoscopy and determine features that distinguish the two groups.METHODS:This prospective study conducted at the University of North Carolina from 2009 to 2011 enrolled consecutive adult patients undergoing outpatient upper endoscopy. Subjects had esophageal biopsies to quantify the maximum eosinophil count per high-power field (eos/hpf; hpf=0.24 mm2). If biopsies revealed ≥15 eos/hpf, subjects were treated with twice daily PPI for 8 weeks and endoscopy was repeated. If ≥15 eos/hpf persisted despite PPI therapy, EoE was diagnosed. If there were <15 eos/hpf, PPI-REE was diagnosed. The proportion of patients in each group was calculated, and patients with EoE and PPI-REE were compared.RESULTS:Of the 223 subjects enrolled, 173 had dysphagia and 50 did not. Of those with dysphagia, 66 (38%) had ≥15 eos/hpf. After the PPI trial, 40 (23%) were confirmed to have EoE, and 24 (14%) had PPI-REE. Of those without dysphagia, 2 (4%) had ≥15 eos/hpf, and after the PPI trial, 1 (2%) had EoE. Compared with EoE, PPI-REE patients were more likely to be older and male and less likely to have typical endoscopic findings of EoE. However, none of the individual factors was independently predictive of PPI-REE status on multivariable analysis. Similarly, although some endoscopic findings were differentially distributed between PPI-REE and EoE, none were significantly associated with disease status on multivariable analysis.CONCLUSIONS:Esophageal eosinophilia is common among patients undergoing esophagogastroduodenoscopy for dysphagia. Although EoE was seen in nearly a quarter of patients with dysphagia, PPI-REE was almost as common, and accounted for over one-third of those with ≥15 eos/hpf. No clinical or endoscopic features independently distinguished PPI-REE from EoE before the PPI trial.

Viscous topical is more effective than nebulized steroid therapy for patients with eosinophilic esophagitis

We performed a randomized trial to compare nebulized and viscous topical corticosteroid treatments for eosinophilic esophagitis (EoE). Subjects with incident EoE (n = 25) received budesonide 1 mg twice daily, either nebulized and then swallowed (NEB) or as an oral viscous slurry (OVB), for 8 weeks. Baseline eosinophil counts for the NEB and OVB groups were 101 and 83 (P = .62). Posttreatment counts were 89 and 11 (P = .02). The mucosal medication contact time, measured by scintigraphy, was higher for the OVB group than the NEB group (P < .005) and was inversely correlated with eosinophil count (R = -0.67; P = .001). OVB was more effective than NEB in reducing numbers of esophageal eosinophils in patients with EoE. OVB provided a significantly higher level of esophageal exposure to the therapeutic agent, which correlated with lower eosinophil counts.

Markers of Eosinophilic Inflammation for Diagnosis of Eosinophilic Esophagitis and Proton Pump Inhibitor–Responsive Esophageal Eosinophilia: A Prospective Study

BACKGROUND & AIMS Distinguishing between eosinophilic esophagitis (EoE), gastroesophageal reflux disease, and proton pump inhibitor-responsive esophageal eosinophilia (PPI-REE) is challenging. We assessed whether immunohistochemical analysis of esophageal tissues for major basic protein (MBP), eotaxin-3, and tryptase can be used for diagnosis of EoE and to differentiate EoE from PPI-REE. METHODS We conducted a prospective study of 196 consecutive adults who underwent outpatient endoscopy at the University of North Carolina from 2009 through 2012. Incident cases of EoE were diagnosed per consensus guidelines. Patients with gastroesophageal reflux disease or dysphagia served as controls. PPI-REE was defined as a symptomatic and histologic response to a PPI. Immunohistochemistry was performed to quantify MBP, eotaxin-3, and tryptase. The maximum density of epithelial staining was determined for each assay; levels were compared between EoE and control groups and then EoE and PPI-REE groups, and receiver operating characteristic curves were constructed. RESULTS Esophageal tissues from patients with EoE (n = 50) had a median 951 MBP-positive cells/mm(2), whereas those from controls (n = 123) had a median 2 MBP-positive cells/mm(2) (P < .001). Samples from patients with EoE had a median 155 eotaxin-3-positive cells/mm(2), and those from controls (n = 123) had 18 eotaxin-3-positive cells/mm(2) (P < .001). Samples from patients with EoE had a median 249 tryptase-positive cells/mm(2), and those from controls (n = 123) had 11 tryptase-positive cells/mm(2) (P < .001). Levels of MBP, eotaxin-3, tryptase, and the combination of all 3 identified patients with EoE with area under the receiver operating characteristic curve values of 0.99, 0.94, 0.99, and 1.00. Analyses of only samples with eosinophil counts of 10-100 eosinophils per high-power field produced similar results. No marker distinguished EoE from PPI-REE. Esophageal tissues from patients with PPI-REE (n = 23) had 987 MBP-positive cells/mm(2) (P = .18, compared with EoE), 160 eotaxin-3-positive cells/mm(2) (P = .33), and 243 tryptase-positive cells/mm(2) (P = .28). CONCLUSIONS Esophageal tissues from patients with EoE have substantially higher levels of MBP, eotaxin-3, and tryptase than controls on the basis of immunohistochemical analysis. Assays for the 3 markers identify patients with EoE with 100% accuracy but cannot distinguish EoE from PPI-REE.

Utility of a Noninvasive Serum Biomarker Panel for Diagnosis and Monitoring of Eosinophilic Esophagitis: A Prospective Study.

OBJECTIVES:Noninvasive biomarkers would be valuable for diagnosis and monitoring of eosinophilic esophagitis (EoE). The aim of this study was to determine the utility of a panel of serum biomarkers for the diagnosis and management of EoE.METHODS:We conducted a prospective cohort study of consecutive adults undergoing outpatient esophagogastroduodenoscopy. Incident cases of EoE were diagnosed per consensus guidelines; controls had gastroesophageal reflux disease (GERD) or dysphagia and did not meet the EoE criteria. EoE cases were treated with topical steroids and had repeat endoscopy. Pre- and post-treatment serum samples were analyzed in a blinded manner for interleukin (IL)-4, IL-5, IL-6, IL-9, IL-13, transforming growth factor (TGF)-α, TGF-β, tumor necrosis factor-α, eotaxin-1, -2, and -3, thymic stromal lymphopoietin (TSLP), major basic protein, and eosinophil-derived neurotoxin. Cases and controls were compared at baseline, and pre- and post-treatment assays were compared in cases.RESULTS:A total of 61 incident EoE cases and 87 controls were enrolled; 51 EoE cases had post-treatment serum analyzed. There were no significant differences in any of the biomarkers between EoE cases and controls at baseline. IL-13 and eotaxin-3 for cases and controls were 85±160 vs. 43±161 pg/ml (P=0.12) and 41±159 vs. 21±73 (P=0.30). There were no significant differences in assay values among cases before and after treatment. There were also no differences after stratification by atopic status or treatment response.CONCLUSIONS:A panel of inflammatory factors known to be associated with EoE pathogenesis were not increased in the serum, nor were they responsive to therapy. None of these biomarkers are likely candidates for a serum test for EoE. Histologic analysis for diagnosis and management of EoE continues to be necessary, and novel, less invasive, biomarkers are needed.

Distribution and variability of esophageal eosinophilia in patients undergoing upper endoscopy

The variability of eosinophilic infiltrates in eosinophilic esophagitis is not well described. This study aimed to determine the distribution of esophageal eosinophilia and the utility of histologic cut-points for eosinophilic esophagitis diagnosis in subjects undergoing endoscopy. We performed a prospective study of adults undergoing outpatient endoscopy. Research protocol esophageal biopsies were obtained from all subjects. Incident cases of eosinophilic esophagitis were diagnosed per consensus guidelines. Biopsies were interpreted following a validated protocol, and maximum eosinophil counts (eosinophils per high-power field; eos/hpf) were determined. Histologic analyses were performed on a per-patient, per-biopsy, and per-hpf basis. There were 213 patients, yielding 923 esophageal biopsies with 4588 hpfs. Overall, 48 patients (23%), 165 biopsy fragments (18%), and 449 hpfs (10%) had ≥15 eos/hpf; most subjects had no or low levels of eosinophils. In the eosinophilic esophagitis cases, 119 biopsy fragments (63%) and 332 hpfs (36%) had ≥15 eos/hpf. There was a mean 104-fold difference between the lowest and highest hpf eosinophil count for the eosinophilic esophagitis patients; 85% of the biopsies from eosinophilic esophagitis cases also had at least one hpf with <15 eos/hpf. The cut-point of 15 eos/hpf had a sensitivity of 100% and a specificity of 96% for diagnosis of eosinophilic esophagitis. In conclusion, most patients have little to no esophageal eosinophilia. In patients with eosinophilic esophagitis, there was marked variability in the eosinophil counts by biopsy and by hpf within a given biopsy. Additionally, the 15 eos/hpf cut-point was highly sensitive and specific for eosinophilic esophagitis. Multiple esophageal biopsies from different locations should be obtained to optimize eosinophilic esophagitis diagnosis.

Epstein-barr virus infected gastric adenocarcinoma expresses latent and lytic viral transcripts and has a distinct human gene expression profile

BackgroundEBV DNA is found within the malignant cells of 10% of gastric cancers. Modern molecular technology facilitates identification of virus-related biochemical effects that could assist in early diagnosis and disease management.MethodsIn this study, RNA expression profiling was performed on 326 macrodissected paraffin-embedded tissues including 204 cancers and, when available, adjacent non-malignant mucosa. Nanostring nCounter probes targeted 96 RNAs (20 viral, 73 human, and 3 spiked RNAs).ResultsIn 182 tissues with adequate housekeeper RNAs, distinct profiles were found in infected versus uninfected cancers, and in malignant versus adjacent benign mucosa. EBV-infected gastric cancers expressed nearly all of the 18 latent and lytic EBV RNAs in the test panel. Levels of EBER1 and EBER2 RNA were highest and were proportional to the quantity of EBV genomes as measured by Q-PCR. Among protein coding EBV RNAs, EBNA1 from the Q promoter and BRLF1 were highly expressed while EBNA2 levels were low positive in only 6/14 infected cancers. Concomitant upregulation of cellular factors implies that virus is not an innocent bystander but rather is linked to NFKB signaling (FCER2, TRAF1) and immune response (TNFSF9, CXCL11, IFITM1, FCRL3, MS4A1 and PLUNC), with PPARG expression implicating altered cellular metabolism. Compared to adjacent non-malignant mucosa, gastric cancers consistently expressed INHBA, SPP1, THY1, SERPINH1, CXCL1, FSCN1, PTGS2 (COX2), BBC3, ICAM1, TNFSF9, SULF1, SLC2A1, TYMS, three collagens, the cell proliferation markers MYC and PCNA, and EBV BLLF1 while they lacked CDH1 (E-cadherin), CLDN18, PTEN, SDC1 (CD138), GAST (gastrin) and its downstream effector CHGA (chromogranin). Compared to lymphoepithelioma-like carcinoma of the uterine cervix, gastric cancers expressed CLDN18, EPCAM, REG4, BBC3, OLFM4, PPARG, and CDH17 while they had diminished levels of IFITM1 and HIF1A. The druggable targets ERBB2 (Her2), MET, and the HIF pathway, as well as several other potential pharmacogenetic indicators (including EBV infection itself, as well as SPARC, TYMS, FCGR2B and REG4) were identified in some tumor specimens.ConclusionThis study shows how modern molecular technology applied to archival fixed tissues yields novel insights into viral oncogenesis that could be useful in managing affected patients.

A Clinical Prediction Tool Identifies Cases of Eosinophilic Esophagitis Without Endoscopic Biopsy: A Prospective Study

OBJECTIVES:Eosinophilic esophagitis (EoE) is difficult to distinguish from gastroesophageal reflux (GERD) and other causes of dysphagia. We assessed the utility of a set of clinical and endoscopic features for predicting EoE without obtaining esophageal biopsies.METHODS:We prospectively enrolled consecutive adults undergoing outpatient upper endoscopy at the University of North Carolina from July 2011 through December 2013. Incident cases of EoE were diagnosed per consensus guidelines. Non-EoE controls had either GERD- or dysphagia-predominant symptoms. A predictive model containing clinical and endoscopic, but no histological, data was assessed. Receiver operator characteristic curves were constructed and the area under the curve (AUC) was calculated.RESULTS:A total of 81 EoE cases (mean age 38 years; 60% male; 93% white; 141 eosinophils per high-power field (eos/hpf)) and 144 controls (mean age 52, 38% male; 82% white; 3 eos/hpf) were enrolled. A combination of clinical (age, sex, dysphagia, food allergy) and endoscopic (rings, furrows, plaques, hiatal hernia) features was highly predictive of EoE. The AUC was 0.944, with sensitivity, specificity, and accuracy of 84, 97, and 92%. Similar values were seen after limiting controls to those with only reflux or dysphagia or to those with esophageal eosinophilia not due to EoE.CONCLUSIONS:We validated a set of clinical and endoscopic features to predict EoE with a high degree of accuracy and allow identification of those at very low risk of disease. Use of these predictors at the point-of-care will avoid the effort and expense of low-yield histological examinations for EoE.

Determination of esophageal eosinophil counts and other histologic features of eosinophilic esophagitis by pathology trainees is highly accurate

Many studies of eosinophilic esophagitis (EoE) use expert pathology review, but it is unknown whether less experienced pathologists can reliably assess EoE histology. We aimed to determine whether trainee pathologists can accurately quantify esophageal eosinophil counts and identify associated histologic features of EoE, as compared with expert pathologists. We used a set of 40 digitized slides from patients with varying degrees of esophageal eosinophilia. Each of 6 trainee pathologists underwent a teaching session and used our validated protocol to determine eosinophil counts and associated EoE findings. The same slides had previously been evaluated by expert pathologists, and these results comprised the criterion standard. Eosinophil counts were correlated, and agreement was calculated for the diagnostic threshold of 15 eosinophils per high-power field as well as for associated EoE findings. Peak eosinophil counts were highly correlated between the trainees and the criterion standard (ρ ranged from 0.87 to 0.92; P<.001 for all). Peak counts were also highly correlated between trainees (0.75-0.91; P<.001), and results were similar for mean counts. Agreement was excellent for determining if a count exceeded the diagnostic threshold (κ ranged from 0.83 to 0.89; P<.001). Agreement was very good for eosinophil degranulation (κ = 0.54-0.83; P<.01) and spongiosis (κ = 0.44-0.87; P<.01) but was lower for eosinophil microabscesses (κ = 0.37-0.64; P<.01). In conclusion, using a teaching session, digitized slide set, and validated protocol, the agreement between pathology trainees and expert pathologists for determining eosinophil counts was excellent. Agreement was very good for eosinophil degranulation and spongiosis but less so for microabscesses.

Su1112 A Set of Clinical and Endoscopic Features Predicts the Presence of Eosinophilic Esophagitis With High Accuracy, Allowing Omission of Low-Yield Biopsies: Results From a Prospective Study

increasing in the population especially in pediatrics and reach 1:1000 in diseases such as EoE (Eosinophilic Esophagitis) (Lee 2013). CD24 is a small, mucin-like glycoprotein that functions both in signal transduction and as an adhesion molecule (Sagiv, 2008). CD24 is expressed on many hematopoietic cells, non-hematopoietic cells and malignancies (Fang, 2010). Previous studies in eosinophils have suggested CD24 as an IL-5-upregulated gene that is involved in eosinophil apoptosis and survival (Temple, 2011). The role evoked for CD24 in IL-5-induced survival of eosinophils remains to be investigated. We hypothesize that CD24 is a key molecular checkpoint, regulating eosinophil functions in chronic mucosal GI diseases, all of which are characterized by significant eosinophil infiltration. Aim: 1) To assess the expression of CD24 in eosinophils in the GI tract in wild-type (WT) mice; 2) To assess the level of eosinophils in the GI tract in CD24 knockout (-/-) and to compare it to WT mice.Methods: We have previously established a colony of CD24-/mice, on a C57BL/ 6J genetic background, and their WT littermates at our local animal facility. The levels of eosinophils were assessed in WT and CD24-/mice. Bone marrow (BM) cells were collected, and grown in medium containing 10 ng/mL recombinant mouse interleukin-5 (rmIL-5). On day 13 the cells were counted and levels of eosinophils were assessed by FACS. The levels of eosinophil and CD24 expression in different tissues were also evaluated. Results: Eosinophil counts in BM extracted from mice were significantly (p-value=0.0436) higher in CD24-/compared to WT mice, (Fig. 1). Fluorescense intensity was stronger in CD24-/compared to WT mice even though there were no differences in the eosinophil population of mature cells (Fig 2). Eosinophil count was higher in BM from CD24-/compared to WT mice, and lower in blood and addipose tissue (Fig. 3). CD24 expression was higher in peripheral tissues compared to the blood and BM (Fig 4). Conclusion: Based on our preliminary results, it is suggested that CD24 has an important role in negatively regulating eosinophil proliferation/maturation/chemotaxis. Understanding the role of eosinophil in GI disorders may have a large imapct on our understanding of the EGIDs. Targeting specific proteins that play an important role in these processes may lead to novel therapeutic approaches

Su1113 In Search of a Non-Invasive Blood Test for Diagnosis and Monitoring of EoE: A Prospective Study of a Serum Biomarker Panel

increasing in the population especially in pediatrics and reach 1:1000 in diseases such as EoE (Eosinophilic Esophagitis) (Lee 2013). CD24 is a small, mucin-like glycoprotein that functions both in signal transduction and as an adhesion molecule (Sagiv, 2008). CD24 is expressed on many hematopoietic cells, non-hematopoietic cells and malignancies (Fang, 2010). Previous studies in eosinophils have suggested CD24 as an IL-5-upregulated gene that is involved in eosinophil apoptosis and survival (Temple, 2011). The role evoked for CD24 in IL-5-induced survival of eosinophils remains to be investigated. We hypothesize that CD24 is a key molecular checkpoint, regulating eosinophil functions in chronic mucosal GI diseases, all of which are characterized by significant eosinophil infiltration. Aim: 1) To assess the expression of CD24 in eosinophils in the GI tract in wild-type (WT) mice; 2) To assess the level of eosinophils in the GI tract in CD24 knockout (-/-) and to compare it to WT mice.Methods: We have previously established a colony of CD24-/mice, on a C57BL/ 6J genetic background, and their WT littermates at our local animal facility. The levels of eosinophils were assessed in WT and CD24-/mice. Bone marrow (BM) cells were collected, and grown in medium containing 10 ng/mL recombinant mouse interleukin-5 (rmIL-5). On day 13 the cells were counted and levels of eosinophils were assessed by FACS. The levels of eosinophil and CD24 expression in different tissues were also evaluated. Results: Eosinophil counts in BM extracted from mice were significantly (p-value=0.0436) higher in CD24-/compared to WT mice, (Fig. 1). Fluorescense intensity was stronger in CD24-/compared to WT mice even though there were no differences in the eosinophil population of mature cells (Fig 2). Eosinophil count was higher in BM from CD24-/compared to WT mice, and lower in blood and addipose tissue (Fig. 3). CD24 expression was higher in peripheral tissues compared to the blood and BM (Fig 4). Conclusion: Based on our preliminary results, it is suggested that CD24 has an important role in negatively regulating eosinophil proliferation/maturation/chemotaxis. Understanding the role of eosinophil in GI disorders may have a large imapct on our understanding of the EGIDs. Targeting specific proteins that play an important role in these processes may lead to novel therapeutic approaches