Activation of the BMP4 Pathway and Early Expression of CDx2 Characterize the Development of Nonspecialized Columnar Metaplasia in a Human Model of Barrett Esophagus
Manuel R. Pera Roman*1, Daniel Castillo1, Sonia Puig1, Carme De BolóS5, Mar Iglesias2, Agustin Seoane3, Laura Comerma2, Vicente Munitiz4, Pascual Parrilla4, Richard Poulsom6, Luis Grande1
1Section of Gastrointestinal Surgery, Hospital Universitario del Mar, Barcelona, Spain; 2Department of Pathology, Hospital Universitario del Mar, Barcelona, Spain; 3Service of Gastroenterology, Hospital Universitario del Mar, Barcelona, Spain; 4Service of General and Digestive Surgery, Hospital Virgen de la Arrixaca, Murcia, Spain; 5Programa de Recerca en Cancer, Institut de Recerca Hospital del Mar (IMIM), Barcelona, Spain; 6Histopathology Unit, London Research Institute, Cancer Research UK, London, United Kingdom
BACKGROUND: The cardia type epithelium, a nonspecialized columnar type of metaplasia (NSCM), has been proposed as an intermediate stage in the transdifferentiation process from normal squamous epithelium into the specialized intestinal type of differentiation present in Barrett esophagus. Recent studies suggest that the BMP4 pathway is involved in the transition of squamous epithelium into nonspecialized epithelium, and that CDX2 and CDX1 transcription factors seem critical for acquisition of the intestinal phenotype. Using an in vivo human model of pathologic reflux disease, we assessed prospectively the development of NSCM and the involvement of the BMP4 pathway and CDX2 expression in this intermediate phenotypic change.METHODS: Biopsy samples from the remnant esophagus of patients having an esophagectomy with gastric preservation were taken at different time periods (6, 12, 18, 24 and 36 months) after their operation and examined for the activation of the BMP pathway (BMP4 / pSMAD 1/5/8) and expression of CDX2 and CDX1. Samples were prepared from squamous esophageal mucosa just above the esophagogastric anastomosis, from columnar-appearing mucosa and from squamous esophageal mucosa 2 cm below the upper esophageal sphincter level and were assessed by immunohistochemistry, quantitative real-time PCR (qRT-PCR) and Western blot (WB). RESULTS: Since June 2006, 18 patients (16 male) were included in the study. Thirteen had an intrathoracic anastomosis. A short segment (mean length: 15.6 mm; longest 30 mm) of NSCM was detected in 10 (56%) patients, with an increasing prevalence along the time periods (17%, 25%, 38%, 42% and 71% at 6, 12, 18, 24 and 36 months, respectively). All cases except one were detected in patients having an intrathoracic anastomosis. Seven of 10 NSCM segments were first detected at 12-18 months postoperatively. Immunohistochemistry detected nuclear expression of pSMAD 1/5/8 in the squamous epithelium close to the anastomosis with strong expression in all epithelial cells of the NSCM areas. These results were confirmed by WB analysis. Scattered nuclear expression of CDX2 was observed in 44/59 biopsies (75%) with NSCM. Two cases showed isolated glands at 18 and 36 months that fully expressed CDX2 and co-expressed CDX1. BMP4 mRNA and CDX2 mRNA expression were significantly greater in NSCM compared with squamous epithelium.CONCLUSIONS: In this human model of columnar metaplasia, we identified early activation of BMP4 in squamous epithelium and NSCM and observed that CDX2 but not CDX1 expression was detected in NSCM before the appearance of the intestinal phenotype.
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