Identification of the putative intestinal stem cell marker doublecortin and CaM kinase-like-1 in Barrett's esophagus and esophageal adenocarcinoma

Abstract

Background and aim: In Barrett's esophagus (BE), the normal esophageal squamous epithelium is replaced with a specialized metaplastic columnar epithelium. BE is a premalignant lesion that can progress to esophageal adenocarcinoma (EAC). Currently, there are no early molecular indicators that would predict progression from BE to EAC. As the only permanent residents of the epithelium, stem cells have been implicated in this metaplastic progression. The aim of the present study was to determine the expression of doublecortin and CaM kinase-like-1 (DCAMKL-1) and other putative gastrointestinal stem cell markers in normal esophageal mucosa (NEM), BE, and EAC.

Methods: Human NEM, BE, EAC, and multitissue microarrays were analyzed for DCAMKL-1, and immunohistochemically scored based on staining intensity and tissue involvement, with epithelia and stroma scored separately. Total RNA isolated from BE and paired NEM was subjected to real-time reverse-transcription-polymerase chain reaction analysis for DCAMKL-1, leucine-rich repeat-containing G-protein-coupled receptor (LGR5), and Musashi-1 (Msi-1) mRNA expression.

Results: DCAMKL-1 was minimally expressed in squamous NEM, but increased in BE (with and without dysplasia) and EAC tissues. In EAC, we found increased stromal DCAMKL-1 staining compared to adjacent epithelia. Within the submucosa of dysplastic BE tissues, an increase in the endothelial cell expression of DCAMKL-1 was observed. Finally, an upregulation of DCAMKL-1, LGR5, and Msi-1 mRNA was seen in BE compared to squamous NEM.

Conclusions: In the present study, we report the progressive increase of DCAMKL-1 expression in BE from dysplasia to EAC. Furthermore, there was an increase in putative stem cell markers DCAMKL-1, LGR5, and Msi-1 mRNA. Taken together, these data suggest that the regulation of resident stem cells might play an important role in the progression of BE to EAC.

Conflict of interest statement

Authors have no conflict of interest

© 2011 Journal of Gastroenterology and Hepatology Foundation and Blackwell Publishing Asia Pty Ltd.

Figures

Figure 1

Immunohistochemical expression of DCAMKL-1 in Normal, BE without dysplasia, BE with dysplasia and Adenocarcinoma/EAC. (A) Minimal DCAMKL-1 epithelial staining in normal squamous epithelium. (B–D) Increased expression of DCAMKL-1 in stroma of biopsies of BE with no dysplasia (B) and BE with dysplasia (C) as well as Adenocarcinoma/EAC in situ (D). Brown indicates cells positive for DCAMKL-1. (E–F) Immunohistochemical scoring of DCAMKL-1 in epithelium (E) and stroma (F) of various tissues as indicated. Values in the bar graphs are given as average ± SEM, and asterisks denote statistically significant differences (* p<0.05 = BE with dysplasia compared to Normal; ** p<0.05 = Adenocarcinoma compared to Normal).

Figure 2

Immunohistochemical expression of DCAMKL-1 in endoscopically obtained, histologically confirmed squamous esophageal mucosa. (A) Minimal DCAMKL-1 expression in normal squamous epithelium. (B) Increased DCAMKL-1 expression in the squamous cells of BE without dysplasia; Progressive increase of perinuclear DCAMKL-1 expression in reactive, irregular cells of BE with dysplasia (C) and in dysplastic cells of Adenocarcinoma/EAC (D). Brown indicates cells positive for DCAMKL-1.

Figure 3

Immunohistochemical expression of DCAMKL-1 within endoscopically obtained, histologically confirmed glandular epithelium, BE without dysplasia, BE with dysplasia and Adenocarcinoma/EAC. (A) Scattered individual epithelial cells were positive for DCAMKL-1 in normal patients without BE. (B–D) The appearance of DCAMKL-1 expression in columnar epithelial cells in BE without dysplasia (B), increased expression of DCAMKL-1 in the columnar epithelia and the appearance of stromal cells positive for DCAMKL-1 in BE with dysplasia (C) overall increase in the number and intensity of DCAMKL-1 expression in both columnar and stromal compartments in Adenocarcinoma/EAC. (D). Minimal DCAMKL-1 immunostaining is observed in endothelial cells in patients with BE without dysplasia (arrows indicates endothelial cells) (E). Increased DCAMKL-1 staining is observed endothelial cells in BE with dysplasia (F). Brown indicates cells positive for DCAMKL-1 (arrows).

Figure 4

DCAMKL-1 is overexpressed in BE. (A) Increased DCAMKL-1 mRNA expression in BE compared to normal. (B–C) Increased Msi-1 (B) and LGR5 (C) mRNA expression in BE compared to normal. Values in the bar graphs are given as average ± SEM, and asterisks denote statistically significant differences (* p<0.01) compared to normal.