Growth of MCF10A-Ras cells in two- versus three-dimensions alters the expression levels of pStat3, IL-6 and E-Cadherin. (a) Tumors arising from immunocompromised mice injected with MCF10A-Ras cells were cultured and passaged four times on tissue culture plates. Extracts were isolated from an MCF10A-Ras primary tumor (Tu) and cultured cells following serial passage (P1, P2, P3 and P4) and analyzed for pStat3, Stat3, E-Cadherin and Tubulin by Western blot. Quantitative evaluation of pStat3/Stat3 (grey bars) and pStat3/Tubulin (black bars) immunoblots by densitometry. Results are representative of three individual experiments. Statistical significance is indicated with asterisks (*, P < 0.05; **, P < 0.01). (b) IL-6 levels by of supernatants from passaged cells were determined by ELISA. C. RNA from passaged cells (P1, P2, P3, P4) was analyzed for human IL-6 mRNA levels by real time RT-PCR (Q-PCR). Results shown are IL-6 expression normalized to HPRT from triplicates.