Neurogenesis by Progenitor Cells in the Ischemic Adult Rat Hippocampus

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Neurogenesis by Progenitor Cells in the Ischemic Adult Rat Hippocampus by Yoshiki Yagita, Kazuo Kitagawa, Toshiho Ohtsuki, Ken-ichiro Takasawa, Takaki Miyata, Hideyuki Okano, Masatsugu Hori, and Masayasu Matsumoto Stroke Volume 32(8):1890-1896 August 1, 2001 Copyright © American Heart Association, Inc. All rights reserved.

Figure 1. Postproliferation cells in the SGZ Figure 1. Postproliferation cells in the SGZ. All rats were administered BrdU (50 mg/kg IP) 3 times 1 day before they were killed. Figure 1. Postproliferation cells in the SGZ. All rats were administered BrdU (50 mg/kg IP) 3 times 1 day before they were killed. Note the marked increase in the BrdU-positive cells in the SGZ and GCL 8 days after ischemia (B), compared with the control (A). At this time point, few BrdU-positive cells were observed in the hilus and the molecular layer. BrdU-positive cells formed clusters in the SGZ (C). It seemed that these cells were divided from an original progenitor cell. Quantitative analysis showed that BrdU-positive cells increased ≈5.7-fold 8 days after ischemia compared with the control (D). In our experiments, the activity of cell proliferation was maximal at this time point. Five days after ischemia, BrdU-positive cells also increased significantly. Because many glial cells were still proliferating at this time point in the dentate gyrus, these BrdU-positive cells may have included not only proliferative neuronal progenitor cells but also glial cells. The significance of differences was determined using ANOVA followed by Scheffé’s post hoc test. Error bars indicate SD. *P<0.05 compared with the control group. D, C indicates control animal; 5, 8,11,15, and 29, 5, 8, 11, 15, and 29 days after ischemia. Bar (A and for B) 0.5 mm, bar (C) 30 μm. Yoshiki Yagita et al. Stroke. 2001;32:1890-1896 Copyright © American Heart Association, Inc. All rights reserved.

Figure 2. Proliferation and cluster formation of Msi1-positive cells in the SGZ. To label newborn cells, we administered to the rats BrdU (50 mg/kg IP) 3 times 7 days after ischemia and were killed the next day after BrdU injection. Figure 2. Proliferation and cluster formation of Msi1-positive cells in the SGZ. To label newborn cells, we administered to the rats BrdU (50 mg/kg IP) 3 times 7 days after ischemia and were killed the next day after BrdU injection. In the SGZ (BrdU [A], Msi1 [B], BrdU/Msi1 [C]), BrdU-positive cells (green in A), which formed clusters, colocalized with Msi1 (red in B), as indicated with arrows (yellow in C). In contrast, in the CA1 region (BrdU [D], Msi1 [E], BrdU/Msi1 [F]), BrdU (arrows in D and F) and Msi1-positive cells (arrowheads in E and F) did not colocalize. Bar (for A to F) 30 μm. Yoshiki Yagita et al. Stroke. 2001;32:1890-1896 Copyright © American Heart Association, Inc. All rights reserved.

Figure 3. Cluster-formed Msi1-positive cells did not colocalize with GFAP in the SGZ 8 days after ischemia. Figure 3. Cluster-formed Msi1-positive cells did not colocalize with GFAP in the SGZ 8 days after ischemia. To confirm Msi1-positive cells were not astrocytes, we performed double-immunolabeling with Msi1 and GFAP. In the SGZ (Msi1 [A], GFAP [B], Msi1/GFAP [C]), Msi1-positive cells (green in A and C, an arrow), which formed clusters, did not colocalize with GFAP (red in B and C, arrowhead). In contrast, in the CA1 region (Msi1 [D], GFAP [E], Msi1/GFAP [F]), all Msi1-positive cells (green in D) colocalized with GFAP (red in E), as indicated with arrows in F. Bar (for A to F) 30 μm. Yoshiki Yagita et al. Stroke. 2001;32:1890-1896 Copyright © American Heart Association, Inc. All rights reserved.

Figure 4. Neuronal differentiation of newborn cells after ischemia. Figure 4. Neuronal differentiation of newborn cells after ischemia. We administered to the rats BrdU (50 mg/kg IP) 3 times 7 days after ischemia and were killed 28 days after BrdU injection. In the SGZ and GCL (NeuN [A], BrdU [B], NeuN/BrdU [C]), arrows indicate newborn neurons, which show NeuN (green in A) and BrdU (red in B) double-positive findings (yellow in C). In contrast, BrdU (green) and GFAP (red) immunofluorescence did not colocalize in the SGZ (BrdU/GFAP [D]). Newborn neurons that migrated into the GCL were large and round, similar to mature granular cells (E and F). Bar (A, for B and C, and D and F) 30 μm, bar (E) 0.5 mm. Yoshiki Yagita et al. Stroke. 2001;32:1890-1896 Copyright © American Heart Association, Inc. All rights reserved.