Figure 4: Everolimus Silences E2F Target Gene Expression

hSMC were treated with 10 μmol/l everolimus and stimulated with growth medium. (A) Minichromosome maintenance protein 7 (MCM7), Cyclin A, and proliferating cell nuclear antigen (PCNA) mRNA expression levels were analyzed by real-time PCR and normalized to transcript levels of the housekeeping gene TBP. Data are presented as mean ± SEM (n = 9 samples/group) relative to quiescent hSMC. Significance levels versus quiescent cells: MCM7, *p < 0.001; Cyclin A, *p < 0.001; PCNA, *p = 0.008. (B) Western blotting for MCM7, Cyclin A, and PCNA expression in hSMC treated with everolimus. Densitometric quantification from 3 independent experiments was normalized to glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and expressed as mean fold increase ± SEM (n = 6 samples/group) relative to quiescent hSMC. Significance levels versus quiescent cells: MCM7, *p < 0.001; Cyclin A, *p < 0.001; PCNA, *p = 0.006. Significance levels versus growth medium + vehicle: MCM7, #p < 0.001; Cyclin A, #p < 0.001; PCNA, #p = 0.032.

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