FAS APO-1 (CD95) IS NOT TRANSLOCATED TO THE CELL-MEMBRANE IN ESOPHAGEAL ADENOCARCINOMA/

This study describes Fas (CD95) expression in Barrett's esophagus, ade nocarcinomas of the esophagus, and three esophageal adenocarcinoma cel l tines. Immunohistochemical analysis of Barrett's esophagus demonstra ted cell surface expression of Fas protein, In contrast, 30.5% of esop hageal adenocarcinomas examined by immunohistochemical analysis demons trated faint cytoplasmic staining, and 69.5% were negative for Fas, Si milar levels of Fas mRNA were identified in tumors compared to mRNA le vels in esophageal squamous mucosa or Barrett's esophagus, rin approxi mately M-r 48,000 Fas protein was identified by Western blot analysis in tumors that were negative for Fas expression by immunohistochemical analysis, The esophageal adenocarcinoma cell line Seg-l was negative for Fas expression by immunohistochemical analysis, but Western blot a nalysis demonstrated abundant, appropriately sized Fas protein, In agr eement with the immunohistochemical analysis, now cytometry of Seg-1 s howed minimal amounts of Fas on the cell surface, which correlated wit h resistance to Fas-mediated apoptosis, No mutations in the Seg-1 Fas coding sequence or exon 1 were identified by sequence analysis, This w as confirmed by transient transfection of COS cells with expression ve ctors generated from the Seg-1 Fas cDNA, which resulted in cell surfac e expression of the Fas protein, Stable transfection of Seg-1 with a F as expression vector did not result in efficient Fas expression on the cell surface, Seg-1 cells, transiently transfected with a Fas-FLAG ex pression vector and examined for protein expression using confocal mic roscopy and an anti-FLAG antibody, showed that the Fas-FLAG protein wa s not present on the cell surface but was present in the cytoplasm, Ta ken together, these results indicate that expression of Fas on the cel l surface by esophageal adenocarcinoma is reduced, In an esophageal ad enocarcinoma cell line, wild-type Fas protein is retained in the cytop lasm, and this correlates with resistance to Fas-mediated apoptosis, T he retention of wild-type Fas protein within the cytoplasm may represe nt a mechanism by which malignant tells evade Fas-mediated apoptosis.