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=7) were screened by gene expression microarray and results were validated by RT-PCR. A larger group of eosinophilic esophagitis patients (N=42) was then used to evaluate protein expression by immunohistochemistry compared with reflux patients (N=15) and normal controls (N=17). Microarray and RT-PCR studies identified overexpression of ALOX15 and tumor necrosis factor alpha-induced 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 (P<0.001). 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=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 (P<0.001). 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 (P<0.001). The majority of controls (89%) and reflux biopsies (100%) were positive for CRISP3 while eosinophilic esophagitis before therapy were positive in 14% of samples (P<0.001) with partial recovery after treatment (43%, P=0.105). This study identified five epithelial-derived markers differentially expressed in eosinophilic esophagitis easily detectable by immunohistochemistry with potential diagnostic utility.