Figure 2.

PKCδ inhibition attenuates Aβ25-35-mediated CCR and caspase-3 cleavage in post-mitotic neurons. (A–C) Primary cortical cultures were transfected with the control LacZ-shRNA or Luc-shRNA, as indicated in the figure panels above, or PKCδ-shRNA for 24 h, followed by exposure to 10 μM Aβ25-35 for an additional 8 h, before detection of cyclin D1 by Western blotting in (A) or immunostaining with the antibodies against cyclin D1 (green) and MAP-2 (red) in (B,C); Hoechst 33258 (blue) served as counterstaining. The arrowheads denote the representative cyclin D1⁺ neurons. Scale bars = 10 μm. Representative micrographs from 3 independent experiments with similar results are shown in (B); quantitative analyses of the cyclin D1⁺/MAP-2⁺ cells are shown in (C). Mean ± S.E.M. from N = 3 in (A) and (C). *, #, and + all denote p < 0.05. (D–F) The experimental conditions were the same as described in (A–C) except cells were treated with Aβ25-35 for an additional 24 h before detection of PCNA (D), p-Histone H3 (E), as well as pro- and cleaved caspase 3 (F) by Western blotting. α-Tubulin served as the internal control for equal loading of proteins in each lane. Mean ± S.E.M. from N = 5 in (D) and N = 3 in (E,F). *, #, and + all denote p < 0.05. (G,H) The experimental conditions were the same as described in (D-F) except that cells were subjected to immunostaining with the antibodies against BrdU (green) and NeuN (red); Hoechst 33258 (blue) served as counterstaining. The arrowheads denote the representative BrdU⁺ neurons. Scale bars = 20 μm. Representative micrographs from 3 independent experiments with similar results are shown in (G); quantitative analyses of the BrdU⁺/NeuN⁺ cells are shown in (H). Mean ± S.E.M. from N = 3 in (H). * and # denote p < 0.05.