SummaryThe protein expression patterns of normal, metaplastic and malignant oesophageal tissues were analysed by two-dimensional polyacrylamide gel electrophoresis (2D-PAGE) to identify changes associated with Barrett's metaplasia and transformation to oesophageal adenocarcinoma. Heat-shock protein 27 (Hsp27), a small heat-shock protein which is protective against cytotoxic stresses, was abundant in normal oesophagus. However, Hsp27 expression was markedly lower in Barrett's metaplasia and oesophageal adenocarcinomas. This was confirmed by immunohistochemical analysis. Hsp27 protein was most highly expressed in the upper layers of squamous epithelium and exhibited a pattern of expression that corresponded with the degree of squamous maturation. Northern and Southern analysis demonstrated Hsp27 to be regulated at the level of mRNA transcription or abundance. Normal oesophageal tissues were examined for gender differences in Hsp27 expression. Women expressed fourfold higher levels of Hsp27 mRNA, however, this difference was not appreciable in protein expression. Hsp27 protein was inducible by heat shock in Barrett's adenocarcinoma cell lines and an immortalized oesophageal epithelial cell line (HET-1A), but not by oestradiol. These results demonstrate abundant constitutive expression of the stress-response protein Hsp27 in the normal oesophagus, and suggest that low-level expression in Barrett's metaplasia may be one factor which may influence susceptibility to oesophageal adenocarcinoma development.

We have developed a data base of lymphoid proteins detectable by two-dinensional polyacrylamide gel electrophoresis. The data base contains two-dimensional patterns and derived information pertaining to polypeptide constituents of unstimulated and stimulated mature T cells and immature thymocytes, single-cell-derived T-and B-cel While the coordinated acquisition or loss of cell-surface differentiation antigens during lymphoid development is a well-characterized process, little is known regarding the regulated expression of proliferation-related proteins during lymphoid differentiation and maturation (1). Recently, we have used two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) to analyze the expression patterns of two cell cycle-regulated proteins, proliferating cell nuclear antigen (PCNA) and oncoprotein 18 (Opl8), during lymphoid development. PCNA and Opl8 are two proliferation-related polypeptides that in mature T cells are induced in late G1 after mitotic stimulation (2-4). Rather surprisingly, we found that these two proteins were strongly expressed in noncycling (as well as cycling) immature thymocytes, suggesting that the molecular pathways leading to proliferation differ in important aspects in mature versus immature T cells (5, 6). During our studies it became readily apparent that many other proteins were also differentially regulated during lymphoid maturation. Since many of these proteins had not yet been identified, it was clear that detailed comparative studies of protein expression patterns in this manner would be substantially facilitated by the construction of a data base.We now report the development of a lymphoid protein data base derived from the analysis of T-cell and thymocyte polypeptides by 2-D PAGE. Using this data base, we have found that although a large proportion of polypeptides induced with proliferation in mature T cells are similarly induced in immature thymocytes, substantial differences also exist. A subset of polypeptides that are induced in mature T cells after mitotic stimulation are constitutively expressed in immature thymocytes. We also describe the identification through amino acid microsequencing of several constitutively expressed or mitotically induced proteins in the different lymphoid populations analyzed. This data base provides a powerful tool for the continued analysis of the cell biology of lymphoid differentiation and activation. These studies also indicate the feasibility of this approach for the examination of other cell types. METHODSIsolation, Stimulation, and Metabolic Labeling of Lymphoid Cells. Thymocyte subpopulations were isolated as described (7) from thymic tissue obtained from children under age 6 undergoing cardiac surgery. Immature CD4+ CD8+ (double positive) and mature CD4+ or CD8+ (single positive) thymocytes were obtained by negative selection for CD28 or CD1, respectively, as CD28 is expressed at high levels only in single-positive thymocytes and CD1 is expressed at high levels only in double-positive thymocytes (7,8). Alte...