Tumor cell-collagen interactions: Identification and semi-quantitative evaluation of selectively-expressed genes by combination of differential display- and multiplex-PCR.
It is widely acknowledged that the presence of extracellular matrix components as substrates can drastically modulate the phenotype and gene expression of cultured cells, including tumor cells. A number of published reports indicated that substrates made from two peculiar collagen species, i.e. type V and OF/LB, which are abnormally deposited in the stroma of primary ductal infiltrating carcinoma (d.i.c.) of the breast "in vivo," were able to exert marked and opposite effects on "in vitro" viability, growth and invasiveness of the 8701-BC cell line, isolated from d.i.c.-affected breast epithelium. To complement such functional data on the effect of cell-collagen interactions with information at molecular level, we have utilized a combination of differential display- and semi-quantitative multiplex-PCR techniques with the aim of detecting variations in the expression levels of selected genes by cells maintained in either culture condition. Here we report some prototypical data on the identification and semi-quantitation of three of the differentially-amplified PCR products found, i.e. HSP2A and MSF-B which are up-regulated in cells grown onto OF/LB collagen substrate, and SRCAP which is prominently down-regulated in the presence of type V collagen substrate. This protocol represents a powerful tool for evaluating changes in the levels and patterns of gene expression which can be theoretically adapted to any experimental model system.
Sirchia
Rosalia
R
Dipartimento di Biologia Cellulare e dello Sviluppo, Università di Palermo. Viale delle Scienze, 90128 Palermo. Italy.
Ciacciofera
Valentina
V
Luparello
Claudio
C