Western blotting

Hippocampal tissues were homogenized and centrifuged at 13,000 g at 4 °C for 10 min. Proteins in the supernatant were separated by sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS-PAGE). Proteins were then transferred onto a nitrocellulose membrane (Hybond-ECL, Amersham Biosciences, Little Chalfont, UK). Membranes were blocked and incubated overnight at 4 °C with primary antibodies against octamer-binding transcription factor 4 (OCT4; 1:200), doublecortin (DCX; 1:1000), or β-actin (1:200) (all from Santa Cruz Biotechnology, Santa Cruz, CA, USA); against SOX2 (1:200), Ki67 (1:1000), nestin (1:1000), or brain lipid-binding protein (BLBP; 1:800) (all from Abcam, Cambridge, MA, USA); or against glial fibrillary acidic protein (GFAP; 1:1000; EMD Millipore, Billerica, MA, USA). After thorough washing, blots were incubated with secondary antibodies (Sigma-Aldrich, St. Louis, MO, USA) conjugated with horseradish peroxidase. Protein bands were detected using enhanced chemiluminescence (Amersham Biosciences) and quantitated using the Quantity One software (Bio-Rad Laboratories, Hercules, CA, USA). Band intensities were normalized to β-actin.