Kelley E. Capocelli

The oesophageal string test: a novel, minimally invasive method measures mucosal inflammation in eosinophilic oesophagitis

Objective Eosinophil predominant inflammation characterises histological features of eosinophilic oesophagitis (EoE). Endoscopy with biopsy is currently the only method to assess oesophageal mucosal inflammation in EoE. We hypothesised that measurements of luminal eosinophil-derived proteins would correlate with oesophageal mucosal inflammation in children with EoE. Design The Enterotest diagnostic device was used to develop an oesophageal string test (EST) as a minimally invasive clinical device. EST samples and oesophageal mucosal biopsies were obtained from children undergoing upper endoscopy for clinically defined indications. Eosinophil-derived proteins including eosinophil secondary granule proteins (major basic protein-1, eosinophil-derived neurotoxin, eosinophil cationic protein, eosinophil peroxidase) and Charcot–Leyden crystal protein/galectin-10 were measured by ELISA in luminal effluents eluted from ESTs and extracts of mucosal biopsies. Results ESTs were performed in 41 children with active EoE (n=14), EoE in remission (n=8), gastro-oesophageal reflux disease (n=4) and controls with normal oesophagus (n=15). EST measurement of eosinophil-derived protein biomarkers significantly distinguished between children with active EoE, treated EoE in remission, gastro-oesophageal reflux disease and normal oesophagus. Levels of luminal eosinophil-derived proteins in EST samples significantly correlated with peak and mean oesophageal eosinophils/high power field (HPF), eosinophil peroxidase indices and levels of the same eosinophil-derived proteins in extracts of oesophageal biopsies. Conclusions The presence of eosinophil-derived proteins in luminal secretions is reflective of mucosal inflammation in children with EoE. The EST is a novel, minimally invasive device for measuring oesophageal eosinophilic inflammation in children with EoE.

Eosinophilic Esophagitis: Epithelial Mesenchymal Transition Contributes to Esophageal Remodeling and Reverses with Treatment

BACKGROUND Mechanisms underlying esophageal remodeling with subepithelial fibrosis in subjects with eosinophilic esophagitis (EoE) have not been delineated. OBJECTIVES We sought to explore a role for epithelial mesenchymal transition (EMT) in subjects with EoE and determine whether EMT resolves with treatment. METHODS Esophageal biopsy specimens from 60 children were immunostained for epithelial (cytokeratin) and mesenchymal (vimentin) EMT biomarkers, and EMT was quantified. Subjects studied had EoE (n = 17), indeterminate EoE (n = 15), gastroesophageal reflux disease (n = 7), or normal esophagus (n = 21). EMT was analyzed for relationships to diagnosis, eosinophil counts, and indices of subepithelial fibrosis, eosinophil peroxidase, and TGF-β immunostaining. EMT was assessed in pretreatment and posttreatment biopsy specimens from 18 subjects with EoE treated with an elemental diet, 6-food elimination diet, or topical corticosteroids (n = 6 per group). RESULTS TGF-β1 treatment of esophageal epithelial cells in vitro for 24 hours induced upregulation of mesenchymal genes characteristic of EMT, including N-cadherin (3.3-fold), vimentin (2.1-fold), and fibronectin (7.5-fold). EMT in esophageal biopsy specimens was associated with EoE (or indeterminate EoE) but not gastroesophageal reflux disease or normal esophagus and was correlated to eosinophil counts (r = 0.691), eosinophil peroxidase (r = 0.738), and TGF-β (r = 0.520) immunostaining and fibrosis (r = 0.644) indices. EMT resolved with EoE treatments that induced clinicopathologic remission with reduced eosinophil counts. EMT decreased significantly after treatment by 74.1% overall in the 18 treated subjects with EoE; pretreatment versus posttreatment EMT scores were 3.17 ± 0.82 versus 0.82 ± 0.39 (P < .001), with similar decreases within treatment groups. Pretreatment/posttreatment EMT was strongly correlated with eosinophil counts for combined (r = 0.804, P < .001) and individual treatment groups. CONCLUSIONS EMT likely contributes to subepithelial fibrosis in subjects with EoE and resolves with treatments that decrease esophageal inflammation, and its resolution correlates with decreased numbers of esophageal eosinophils.

Feeding Dysfunction in Children With Eosinophilic Gastrointestinal Diseases

OBJECTIVES: Feeding dysfunction (FD) seen in younger children with eosinophilic gastrointestinal disease (EGID) has not been well described. Thus, our aim was to further characterize FD in children with EGIDs. METHODS: A retrospective medical record analysis of 200 children seen over 12 months in a multidisciplinary Gastrointestinal Eosinophilic Diseases Program was performed. The clinical data of 33 children identified as also having FD were examined, including information obtained by history, physical examination, feeding evaluation, review of nutritional data, allergy testing and histologic assessment of mucosal biopsies. RESULTS: Of 200 children with EGIDs, 16.5% had significant FD. The median age of this group was 34 months (range: 14–113 months). A variety of learned maladaptive feeding behaviors were reported in 93.9%. Frequent gagging or vomiting occurred in 84.8%. Food sensitivity was documented in 88% while 52% had other allergic disease. Twenty one percent were diagnosed with failure to thrive and 69.7% required individual or group feeding therapy. Forty-two percent had residual eosinophilia of >15 per HPF on esophageal biopsies performed at the time of symptoms. CONCLUSIONS: FD is prevalent in children with EGIDs often presenting as maladaptive learned feeding behaviors with altered mealtime dynamics and physical difficulties in eating mechanics. FD can persist even after eosinophilic inflammation is successfully treated. Awareness of the increased prevalence of FD in children with EGIDs with enable earlier recognition of this problem, resulting in a comprehensive, individualized treatment plan with the desired outcome of improving the development, feeding, and nutrition of these children.

Effect of proton pump inhibitor on esophageal eosinophilia.

Objective: Differentiation between the common etiologies of dense esophageal eosinophilia such as gastroesophageal reflux disease (GERD) and eosinophilic esophagitis can be difficult. We hypothesized that histologic features may provide diagnostic clues concerning the etiology of esophageal eosinophilia. Methods: We performed a retrospective chart review of 204 children with the diagnosis of esophagitis characterized by ≥15 eosinophils (eos) per high-power field (HPF) in at least 1 biopsy. We then restricted our analysis to subjects who had received at least 8 weeks of only proton pump inhibitors (PPIs) followed by endoscopy and who had a clinicopathologic response to this treatment. Symptoms, endoscopic findings, and pathologic descriptions were reviewed and an eosinophil peroxidase (EPX) index was determined to assess for degranulation/eosinophil activation. Results: Of the 204 identified charts, 7 subjects identified met the inclusion criteria. Five of these 7 patients showed a clinicopathologic response to PPIs after their follow-up endoscopy, (mean peak eosinophil count: 92 vs 5 eos/HPF, and EPX index: 39.2 vs 14.6, pre- and posttreatment, respectively). Two patients experienced initial resolution of symptoms and esophageal eosinophilia with PPI therapy; however, within 17–23 months they redeveloped symptoms and esophageal eosinophilia while receiving PPI therapy at the time of a third endoscopy (mean peak eosinophil count: 40 vs 11 vs 36 eos/HPF, and EPX index: 44 vs 21 vs 36.5, pre-, post- and posttreatment, respectively). No clinicopathologic features or degranulation patterns differentiated subjects with GERD/PPI responsive esophageal eosinophilia from those who had transient response to PPI treatment. Conclusions: No clinicopathologic features differentiated subjects who responded to PPI treatment. PPI treatment can be helpful to exclude GERD and PPI responsive esophageal eosinophilia but long-term follow-up is critical in the management of esophagitis.

Eosinophilic esophagitis: the newest esophageal inflammatory disease

Eosinophilic esophagitis (EoE) is a chronic esophageal inflammatory disease of undetermined pathophysiology that results in dense mucosal eosinophilia and esophageal dysfunction. In childhood, vague symptoms associated with GERD and feeding difficulties are the first manifestations of EoE. Adults typically present with dysphagia and food impaction. No pathognomonic features have been identified for EoE and, therefore, its diagnosis must be made on both clinical and histological grounds. Effective treatments rely on steroids and dietary exclusions.

Local hypersensitivity reaction in transgenic mice with squamous epithelial IL-5 overexpression provides a novel model of eosinophilic oesophagitis

Objective Eosinophilic oesophagitis (EoE) is a chronic inflammatory condition of the oesophagus with limited treatment options. No previous transgenic model has specifically targeted the oesophageal mucosa to induce oesophageal eosinophilia. Design We developed a mouse model that closely resembles EoE by utilising oxazolone haptenation in mice with transgenic overexpression of an eosinophil poietic and survival factor (interleukin (IL)-5) in resident squamous oesophageal epithelia. Results Overexpression of IL-5 in the healthy oesophagus was achieved in transgenic mice (L2-IL5) using the squamous epithelial promoter Epstein–Barr virus ED-L2. Oxazolone-challenged L2-IL5 mice developed dose-dependent pan-oesophageal eosinophilia, including eosinophil microabscess formation and degranulation as well as basal cell hyperplasia. Moreover, oesophagi expressed increased IL-13 and the eosinophil agonist chemokine eotaxin-1. Treatment of these mice with corticosteroids significantly reduced eosinophilia and epithelial inflammation. Conclusions L2-IL5 mice provide a novel experimental model that can potentially be used in preclinical testing of EoE-related therapeutics and mechanistic studies identifying pathogenetic features associated with mucosal eosinophilia.

Thymoma in Children: Report of 2 Cases and Review of the Literature

Thymoma is an uncommon and slow-growing neoplasm. It is derived from thymic epithelial cells and comprises about 20% to 30% of mediastinal masses in adults, but only about 1% in pediatric patients. Patients usually present with mass-associated respiratory symptoms, superior vena cava syndrome, or paraneoplastic syndrome including myasthenia gravis, pure red cell aplasia, or acquired hypogammaglobulinemia, and connective tissue disorders. Due to the limited number of cases, knowledge, and experience with thymoma in pediatric patients, the diagnosis and treatment are very challenging for this age group. In this article, we report 2 cases of thymoma in childhood and provide a comprehensive review and analysis of the reported pediatric cases in the past 30 years (total of 32 cases). We found that patients younger than age 10 years were predominantly male (M:F = 6:1) and had advanced tumor stage more frequent than patients older than age 10 (P = .03). There were also significant associations of male sex with more advanced tumor stage and less favorable outcome (P = .03). These findings suggest that age and sex may be additional potential prognostic contributors in pediatric patients with thymoma. The clinicopathologic features, differential diagnosis, and current therapeutic recommendations of this uncommon tumor in pediatric patients are also addressed.

Eosinophils and IL-33 Perpetuate Chronic Inflammation and Fibrosis in a Pediatric Population with Stricturing Crohn's Ileitis

Background:Fibrostenosis and stricture are well-recognized endpoints in Crohns disease (CD). We hypothesized that stricturing CD is characterized by eosinophilia and epithelial IL-33. We proposed that eosinophil exposure to IL-33 would perpetuate inflammatory chronicity and subsequent fibrostenosis. Methods:We performed a retrospective study of 74 children with inflammatory and stricturing ileal CD comparing clinicopathological features to immunohistochemical measures of eosinophilia and IL-33. To scrutinize eosinophil patterns, we developed a novel eosinophil peroxidase score encompassing number, distribution, and degranulation. Human eosinophils and intestinal fibroblasts were cultured with IL-33 and IL-13, and inflammatory and remodeling parameters were assessed. Antieosinophil therapy was also administered to the Crohns-like ileitis model (SAMP1/SkuSlc). Results:Our novel eosinophil peroxidase score was more sensitive than H&E staining, revealing significant differences in eosinophil patterns, comparing inflammatory and stricturing pediatric CD. A significant relationship between ileal eosinophilia and complicated clinical/histopathological phenotype including fibrosis was determined. IL-33 induced significant eosinophil peroxidase secretion and IL-13 production. Exposure to eosinophils in the presence of IL-33, “primed” fibroblasts to increase proinflammatory cytokines (TNF-&agr;, IL-1&bgr;, and IL-6), eosinophil-associated chemokines (CCL24 and CCL26), and IL-13R&agr;2 production. Production of fibrogenic molecules (collagen 1A2, fibronectin, and periostin) increased after exposure of “primed” fibroblasts to IL-13. Epithelial-IL-33 was increased in pediatric Crohns ileitis and strongly associated with clinical and histopathological activity, ileal eosinophilia, and complicated fibrostenotic disease. SAMP1/SkuSlc eosinophil-targeted treatment resulted in significant improvements in inflammation and remodeling. Conclusions:Our study of specimens from pediatric patients with ileal CD linked eosinophil patterns and IL-33 to fibrosis and suggested that these may contribute to the perpetuation of inflammation and subsequent stricture in pediatric CD.

Ultrastructural features of eosinophilic oesophagitis: impact of treatment on desmosomes

Aims A growing body of evidence suggests a role for altered epithelial barrier function in the pathophysiology of eosinophilic oesophagitis (EoE), but few have described the epithelial structure during inflammation. The purpose of this study was to define ultrastructural features of active, inactive EoE and control subjects oesophageal epithelia. Methods We prospectively enrolled patients undergoing diagnostic upper endoscopy for evaluation of EoE. Mucosal pinch biopsies were obtained from the distal oesophagus and processed for routine histology and electron microscopic assessment. Clinical features of enrolled subjects were analysed and subjects were divided into four groups: normal, gastroesophageal reflux disease (GERD), inactive EoE and active EoE. Representative photomicrographs of the basal and superficial epithelia were reviewed for abnormalities. Desmosomes were quantified on the surface of epithelia three to four prickle-cell layers above the basal layer. Results Twenty-nine paediatric cases (ages 2–18 years) were enrolled in the study. We observed a significant decrease in the number of desmosomes per cell (DPC) of subjects with active EoE compared with inactive EoE, GERD and normal epithelia. With respect to DPC, no significant differences were found between inactive EoE compared with GERD or normal subjects. Additional ultrastructural features observed included epithelial microplicae and evidence of eosinophil transmigration, degranulation, and sombrero formation. Conclusions Consistent with clinical and molecular findings, our ultrastructural data provide support for an altered oesophageal barrier in paediatric cases with active EoE, which may improve following treatment.

Novel platelet storage conditions: additive solutions, gas, and cold.

Purpose of reviewPlatelets are a frequently requested blood product today and are often in limited supply because of a shelf life of 5–7 days, depending on the country. Room temperature storage is associated with an increased risk of transfusion-transmitted infection. Plasma used for platelet storage is unavailable for other uses, and allogeneic plasma carries with it risks for adverse transfusion reactions. This review looks at recent activities evaluating alternative conditions for the storage of platelets. Recent findingsNew-generation platelet additive solutions are being evaluated and applied as a strategy to reduce the volume of allogeneic plasma transfused and to support storage following pathogen reduction treatments. There is a renewed interest in refrigerator temperature and frozen storage of platelets to improve availability, to reduce septic transfusion risk, and to enhance hemostatic efficacy in the bleeding patient. SummaryUse of platelet additive solutions has been shown to reduce the incidence of allergic and nonhemolytic febrile transfusion reactions in two large studies. Results of ongoing research and new clinical trials in cold storage methods will be forthcoming and may present solutions for platelet availability problems and new choices for therapeutic transfusion of the bleeding patient.