FIGURE 6.

Restoration of normal proliferation of Atm^-/- astrocytes by inactivation of ERK1/2 signaling or knockdown of p16^Ink4a. A, growth curves of Atm^+/+ and Atm^-/- astrocytes, cultured either in the presence or absence of 50 μm PD98059. PD98059 partially rescued Atm^-/- astrocytes from defective proliferation. The means ± S.D. of three independent countings are shown. ^*, p < 0.05 when PD98059-treated Atm^-/- astrocytes were compared with untreated Atm^-/- astrocytes. B, photomicrographs of Atm^+/+ and Atm^-/- astrocytes culture in the presence or absence of PD98059. Scale bars, 50 μm. C, Atm^+/+ astrocytes were left untreated. Atm^-/- astrocytes were either untreated left or treated with 1 mm NAC or 50 μm PD98059 for 2 days. The levels of phospho-ERK1/2, ERK1/2, Bmi-1, p16^Ink4a, and phospho-Rb were determined by direct Western blotting analysis. D, Atm^+/+ and Atm^-/-astrocytes were transfected with Ink4a siRNA. p16^Ink4a levels were determined by direct Western blotting analysis (left panel). Proportions of senescent cells were determined by SA β-galactosidase expression 2 days after cultivation (right panel). The means ± S.D. of three independent experiments are shown. ^*, p < 0.05 when untransfected Atm^-/-astrocytes were compared with untransfected Atm^+/+ culture or when Ink4a siRNA-transfected Atm^-/-astrocytes were compared with untransfected Atm^-/- astrocytes.