Gene expression profiles of epithelial cells microscopically isolated from a breast-invasive ductal carcinoma and a nodal metastasis.

Expression profiles of breast carcinomas are difficult to interpret when they are obtained from tissue in toto, which may contain a large proportion of non-cancer cells. To avoid this problem, we microscopically isolated cells from a primary invasive ductal carcinoma of the breast and from an axillary node harboring a metastatic breast carcinoma, to obtain pure populations of carcinoma cells ( approximately 500) and used them for serial analysis of gene expression. The expression profiles generated from both populations of cells were compared with the profile of a disease-free mammary epithelium. We showed that the expression profiles obtained are exclusive of carcinoma cells with no contribution of non-epithelial cells. From a total of 16,939 unique tags analyzed, we detected 559 statistically significant changes in gene expression; some of these genes have not been previously associated with breast cancer. We observed that many of the down-regulated genes are the same in both cancers, whereas the up-regulated genes are completely different, suggesting that the down-regulation of a set of genes may be the basic mechanism of cancer formation, while the up-regulation may characterize and possibly control the state of evolution of individual cancers. The results obtained may help in characterizing the neoplastic process of breast cancer.

Association of rat8 with Fyn protein kinase via lipid rafts is required for rat mammary cell differentiation in vitro.

We previously identified rat8 in the pathway involved in epithelial cell differentiation that occurs in the rat mammary gland at pregnancy when tubules and alveoli are formed. rat8, which encodes an IFN-inducible membrane protein, is the rat homologue of the mouse gene fragilis. By differential detergent extraction and isopycnic sucrose density gradients, we show that rat8 protein is associated to lipid membrane domains together with Lyn and Fyn, members of the Src tyrosine kinase family. We also show that recruitment of rat8 to lipid membrane domains is a necessary step in mammary epithelial cell differentiation. Immunoprecipitation analysis, performed with an anti-Fyn protein antibody, shows that rat8 was present in the Fyn immunoprecipitate. Antisense oligonucleotides, used to inhibit Fyn protein expression, block mammary cell differentiation. Taken together, these results suggest that the functional interaction, via lipid membrane domains, of rat8 and Fyn proteins is required for mammary cell differentiation. Therefore, rat8, like fragilis, may be involved in developmental decisions and the demarcation of a subset of cells in the mammary gland that cause epithelial cells to develop into a network of tubuloalveolar structures involved in secretion.