Figure 5

A, Immunohistochemical staining of proliferating cells in SVZ of brain slices cultured for 14 days. Brain slices from 21-day-old BALB/c mice were cultured for 0 days (a, e, and i) or 14 days (c, d, g, h, k, l, and m to p), infected with mutant MCMV (RM461) for 3 days and then stained with X-Gal. Some brain slices were not infected after 14 days of culturing (b, f, and j). Additional immunohistochemical staining was performed with antibodies to GFAP (a, b, c, and d), nestin (e, f, g, and h), and Musashi-1 (i, j, k, and l). Most of the proliferating X-Gal-positive cells in the SVZ after incubation for 2 weeks were stained with PCNA antibody (m and n) and specifically labeled with BrdU when it was added to the medium for 14 days (o and p). The proliferating periventricular cells after incubation for 2 weeks were positive for GFAP (c), nestin (g), and Musashi-1 (k) and susceptible to MCMV infection. Most of the infected cells were double-stained with the antibodies to GFAP (d), nestin (h), and Musashi-1 (l). Original magnification, × 10 (a to c, e to g, and i to k), × 100 (d, h, and l), × 30 (m and o), × 150 (n), and × 120 (p). B, Comparison of infection ratios and numbers of neural progenitor cells in the brain sections of 0-day-cultures versus 14-day-cultures. Brain slices from 21-day-old mice were cultured for 0 or 14 days before infection. Infection ratios (a) and the number of cells immunostained for nestin (b) and Musashi-1 (c) were determined as described in Figure 3. The number of cells immunostained for neural immature/progenitor cell markers significantly increased in the brain sections of 14-day cultures under both serum-containing and serum-free culture conditions, but the number of cells stained for neural immature/progenitor cell markers was significantly lower in slices cultured in serum-free medium than in those cultured in serum-containing medium. Data are expressed as the mean ± sd of at least three independent experiments. ★ p < 0.05 and ★★ p < 0.01.