Abstract 3363: Identification and characterization of stem-like cells in human esophageal epithelial and adenocarcinoma cells

Abstract

Abstract The aim of this study was to identify and characterize stem-like cells in human normal esophageal epithelial and adenocarcinoma cell lines. The immortalized esophageal epithelial cell line, Het-1A, and the primary esophageal adenocarcinoma (EADC) cell line, OE33, were first studied by flow cytometry and immunofluorescence using a panel of antibodies to putative stem cell biomarkers which included CD34, CD44, CD44/CD24, CD117/integrin α6, Musashi I, EpiCam, and Oct 4. Fluorescence activated cell sorting (FACS) was then used to isolate 15% of cells with the highest CD44 staining (CD44high) or reactive oxygen species (ROS) (ROShigh) and 15% of cells with the lowest (including negative) CD44 staining (CD44low) or ROS (ROSlow). Each cell population was further characterized using sphere-forming assays, soft agar clonogenic assays, flow cytometry for ROS, and synchrotron mid-infrared (IR) spectromicroscopy. CD44 positivity was found in 90% of Het-1A and 75% of OE33 cells overall. For both Het-1A and OE33 cell lines, spheroid formation (mean±SD/5000 cells) was significantly higher in cells expressing high levels of CD44 (Het-1A:138.33±2.08 vs. 97.67±1.53, p<0.01; OE33: 150.00±31.19 vs. 100.67±7.02, p=0.056) and low levels of ROS (Het-1A: 149.33±20.43 vs. 92.00±8.89, p=0.011; OE33: 199.67±31.63 vs. 119.33±14.47, p=0.016). Colony formation (mean±SD/1000 cells) was also found to be significantly higher in CD44high (Het-1A: 15.33±4.51 vs. 8.00±1.00, p=0.051; OE33: 92.67±11.02 vs. 18.00±1.73, p<0.01) and ROSlow (Het-1A: 45.00±1.00 vs. 31.67±3.79, p<0.01; OE33: 33.00±5.29 vs. 14.67±5.03, p=0.012) cell populations. Whereas OE33 CD44high cells were found to express consistently lower levels of ROS compared with OE33 CD44low cells (p<0.01), no differences in ROS expression were found in Het-1A cells sorted by CD44 expression. Spectromicroscopic analysis revealed only small non-significant differences between cell populations sorted by CD44 and ROS expression. We conclude that the higher rates of spheroid formation and clonogenicity of Het-1A and OE33 cells expressing high levels of CD44 (CD44high) and low levels of ROS (ROSlow) reflect an enrichment of stem-like cell populations in esophageal cell lines. However, the significant differences in ROS levels between CD44high OE33 cells and CD44high Het-1A cells suggests unique and biologically distinct pathways in stem-like cells derived from normal and malignant esophageal epithelia. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 3363.