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. 2013 Oct 2;8(10):e77639. doi: 10.1371/journal.pone.0077639

Figure 2. Effects of PD-0332991 on BSG cell lines derived from genetically engineered mouse models.

Figure 2

Cells were treated with PD-0332991 for 48 hours before being harvested for MTT or BrdU assays to assess cell survival and proliferation, respectively. (A) MTT assays (left panel) show minimal cytotoxic activity at doses up to 5µM in cell lines derived from PDGF-B; Ink4a-ARF deficient BSGs. BrdU assays (right panel) showed inhibition of proliferation with an IC50 of 1.8µM. Assays performed in two independent cell lines. (B) Apoptosis assays showed a very small but significant increase in apoptosis at 2µM and 5µM (p=0.0006 and p=0.0002, respectively). (C) Western blot analysis shows inhibition of Rb phosphorylation at the protein level, with inhibition observed at 2µM in the PDGF-B; Ink4a-ARF deficient line after only 24 hours of treatment. (D) MTT assays (left panel) show the minimal cytotoxic activity only at doses of 5µM in cell lines derived from PDGF-B; p53 deficient BSGs. BrdU assays (right panel) showed significant inhibition of proliferation at 5µM but IC50 was not reached. (E) Apoptosis assays did not show any difference in apoptosis between vehicle and up to 5µM PD. Assays performed in two independent cell lines. Error bars represent SEM from three independent experiments. Statistical significance was determined using One-way ANOVA; paired student’s t-test was used to compare within groups. (F) PDGF-B; p53 deficient BSG cells show no decrease in pRb at doses up to 2µM even after 48 hours of treatment with PD.