Expression microarray analysis identifies novel epithelial-derived protein markers in eosinophilic esophagitis

Abstract

Gene expression studies in eosinophilic esophagitis support an immune-mediated etiology associated with differential regulation of inflammatory and epithelial-derived genes. We aimed to further characterize epithelial gene expression alterations in eosinophilic esophagitis and to explore the use of immunohistochemistry to identify these alterations. Esophageal biopsies from pediatric patients with eosinophilic esophagitis before and after therapy with topical steroids (N ¼ 7) were screened by gene expression microarray and results were validated by RT-PCR. A larger group of eosinophilic esophagitis patients (N ¼ 42) was then used to evaluate protein expression by immunohistochemistry compared with reflux patients (N ¼ 15) and normal controls (N ¼ 17). Microarray and RT-PCR studies identified overexpression of ALOX15 and tumor necrosis factor alphainduced factor 6 (TNFAIP6) and underexpression of filaggrin (FLG), SLURP1 and cysteine-rich secretory protein 3 (CRISP3) in eosinophilic esophagitis. Immunohistochemistry for ALOX15 was positive in 95% of eosinophilic esophagitis and negative in all controls, all eosinophilic esophagitis after therapy and all reflux biopsies (Po0.001). TNFAIP6 was positive in 88% of eosinophilic esophagitis samples versus 47% of controls, 29% of eosinophilic esophagitis after therapy and 40% of reflux samples (P ¼ 0.002). Overexpression of both ALOX15 and TNFAIP6 directly correlated with the degree of eosinophilic infiltration. FLG was positive in 88% of controls and 100% of reflux biopsies, but negative in all eosinophilic esophagitis samples, and its expression was regained in 86% of eosinophilic esophagitis after therapy patients (Po0.001). SLURP1 expression was positive in all controls and reflux samples, but only positive in 5% of eosinophilic esophagitis and was re-expressed to 100% positivity in eosinophilic esophagitis after therapy patients (Po0.001). The majority of controls (89%) and reflux biopsies (100%) were positive for CRISP3 while eosinophilic esophagitis before therapy were positive in 14% of samples (Po0.001) with partial recovery after treatment (43%, P ¼ 0.105). This study identified five epithelial-derived markers differentially expressed in eosinophilic esophagitis easily detectable by immunohistochemistry with potential diagnostic utility.

Pediatric Eosinophilic Esophagitis Symptom Scores (PEESS v2.0) identify histologic and molecular correlates of the key clinical features of disease

“…Previous studies of EoE have identified sets of genes (~500) whose esophageal expression can discriminate EoE from non-EoE. (6, 10, 32, 33) This work led to the development of a smaller set of genes that provides strong discriminatory value (the EDP) for individuals who have ≥ 15 eosinophils/hpf in the esophagus compared to those with ≤2 eosinophils/hpf in the esophagus. (11) Thus, the PEESS ® v2.0 dysphagia domain captures information associated with alterations in esophageal gene expression.…”

Filaggrin Dysfunction and Its Association with Inflammatory Conditions of the GI Tract

“…In the same study, 6.1 % of patients with EoE had a FLG loss-of-function mutation versus only 1.3 % of normal controls ( p = .0172) [ 21 ]. A more recent study on a cohort of 47 EoE patients found that fi laggrin was greatly underexpressed compared to controls [ 22 ]. Specifi cally, fi laggrin expression was positive in 88 % of controls, in 100 % of GERD patients, and in 0 % of patients with EoE ( p < .001) [ 22 ].…”

“…A more recent study on a cohort of 47 EoE patients found that fi laggrin was greatly underexpressed compared to controls [ 22 ]. Specifi cally, fi laggrin expression was positive in 88 % of controls, in 100 % of GERD patients, and in 0 % of patients with EoE ( p < .001) [ 22 ]. While FLG mutation likely does not explain the entire disease process of EoE, it shows how GI pathology is indeed linked to this barrier protein and its dysfunction.…”

Eosinophils in Gastrointestinal Disorders

“…A recent twin study revealed that if a sibling has EoE, there exists a risk of 2.4% of subsequent children developing EoE. 7 A variety of EoE genes provide clues to dysfunction of the epithelial barrier (filaggrin), 8 immune system (thymic stromal lymphopoietin, eotaxin-3) 9,10 11 and other yet to be identified areas (calpain-14). 12,13 …”