The homeostasis of normal breast depends on interactions between epithelial cells and their associated stroma. Previous studies indicated that in breast cancer carcinoma, tumor associated stromal cells with different functions appear to be emerged. These stromal elements include fibroblasts which modulate tumor behavior providing various growth factors and extracellular matrix components. Our aim was to evaluate the differential gene expression between fibroblasts derived from malignant (n=4, ECII) and benign (n=4) breast lesions. Other aim was to analyze the influence of normal epithelial cells (MCF10A) on gene expression profile of fibroblasts obtained from breast cancer. Fibroblast primary cultures were established and expression of vimentin and smooth cell actin was positive. Co-culture of these cell types separated by inserts, which allow the passage of soluble factors, was done and total RNA was extracted. After mRNA amplification using a template-switching prime, cDNA probes were synthesized, labeled with fluorochrome conjugated deoxynucleotide, a competitive hybridization was undertaken onto cDNA microarray glass slides in which 4,608 ORESTES (open reading frame expressed sequence tags) from Instituto Ludwig de Pesquisa sobre o Câncer/FAPESP bank were spotted and fluorescent signals were quantified. After normalization, the differentially expressed genes, at a False Discovery Ratio (FDR) less then 0.05, were selected for further analysis. We found 284 differentially expressed genes in tumoral fibroblasts when compared with benign disease derived fibroblasts. Among these genes, 187 were quantitatively down regulated (fold ranging from 1.05 to 4.14) against 96 up regulated (fold ranging from 1.17 to 7.73). The majority of alterations were related to membrane transport, signaling transduction and biosynthesis. Overall these results could suggest a reduced gene expression along transformation process. After coculture with MCF10A cells, we found 566 differentially expressed genes in tumoral fibroblasts, 323 were down regulated (fold ranging from 1.09 to 10.62) and 243 up regulated (fold ranging from 1.03 to 16.62). MCF10A influence in tumoral fibroblasts gene expression could be seen trough the expression of some genes possibly related with a reversion in the malignant phenotype of these cells. Supported by Fapesp 01/13515-1, CNPQ and Fundação Faculdade de Medicina.
[Proc Amer Assoc Cancer Res, Volume 46, 2005]