Figure 4
SX13 cells expressing low-levels of IGF-1R are sensitive to the death-inducing effects of PD 98059. (a) Western blot showing low levels of IGF-1R, but comparable levels of ERα and cleaved PARP in SX13 and NEO cells undergoing the indicated hormonal treatments for 120 hours. (b, c) Cell counts showing that IGF-1 does not enhance E2-stimulated SX13 cell proliferation, but that PD 98059 can restore the growth-inhibitory effects of 4-OHT treatment. Cells (2 × 105) were seeded and, after 24 hours, treated with either 1% or 5% FBS-DCC serum in the absence (E2 ablation) or presence of E2 and/or IGF-1 for 168 hours (b) or 144 hours (c). Cell counts were performed with a Coulter counter (c). (d) Trypan blue exclusion assay shows that IGF-1 attenuates the death-inducing effects of 4-OHT and/or MIF treatments in an MEK1-dependent manner. At 144 hours after treatments, adherent and detached cells were collected and counted by using a hemacytometer. (e) Representative images show that PD 98059 effectively reduces cell number in the E2-treated cell population (compare a with b), and induces cell shrinkage and detachment, indicative of apoptosis, in the 4-OHT plus MIF-treated cell population (compare c with d). Data in (b through d) are expressed as mean ± SD (n = 3). The following show significant differences in the induction of apoptosis (number of detached cells) and cell proliferation for the hormonal therapies compared with: (a) E2; (b) 4-OHT + MIF; (c) E2+IGF-1; (d) E2+IGF-1+PD 98059. *P < 0.05.