Total RNA was extracted from cultured cells as described²  and reverse transcribed with the MMuLV-RT and random hexaprimers, according to the supplier's instructions (Promega, Charbonnières, France). Real-time quantitative PCR was performed on an ABI PRISM 7700 Sequence Detector (PE Applied Biosystems, Foster City, CA) with the SYBR Green PCR master mix (PE Applied Biosystems). The relative quantitation of cyclin D1 expression was normalized to 18S rRNA as internal standard. The primers were designed as follows: cyclin D1 sense, 5′-ACA AAC AGA TCA TCC GCA AAC AC-3′; cyclin D1 antisense, 5′-TGT TGG GGC TCC TCA GGT TC-3′, 18S rRNA sense, 5′-GCT GGA ATT ACC GCG GCT-3′; 18S rRNA antisense, 5′-CGG CTA CCA CAT CCA AGG AA-3′. PCR was performed in 25 μL final volume containing 10 ng cDNA templates, 300 nM of each primer. The quantitation of cyclin D1 compared with 18S rRNA was evaluated using the comparative threshold (Ct) method with the ABI PRISM 7000 SDS software.⁵  Triplicate experiments were done for each sample; for each sample, the average Ct value for the internal standard (18S rRNA) was subtracted from the average Ct value for cyclin D1 to yield ΔCt. ΔCt obtained for the calibrator cell line GRANTA-519 was then subtracted from the ΔCt of each cell line to give ΔΔCt. The relative amount of cyclin D1 compared with the control was calculated by the formula N = 2^−ΔΔCt.