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(GFAP, a specific marker for astrocytes) and Akt1 are all expressed by both F3 and
F3.Akt1 cells. However, GFAP band at F3.Akt1 lane detected was lighter than F3
parental cells. In addition transcriptional level of Akt1 gene is higher in F3.Akt1
cells as compared to parental F3 cells.
Figure 1.
Characterization of human NSC lines.
A: The retroviral vector encoding Akt1 (pLHCX.Akt1) used
in the present study for the generation of HB1.F3.Akt1 (F3.Akt1) human neural stem cell (NSC) line. B and
C: Phase contrast microscopy of F3 and F3.Akt1 human NSCs. Bar inciates 20 µm. D: Gene expression of cell
type-specific markers as studied by RT-PCR in F3 and F3.Akt1 human NSCs. Both of F3 and F3.Akt1 NSC
lines express cell type-specific markers Nestin (for neural stem cells), NF-L, NF-M and NF-H (for neurons),
GFAP (for astrocytes) and Akt1.
Hydrogen Peroxide (H
2
o
2
)-Induced Cell Death
To determine the protective effects of Akt1 on the H
2
O
2
-induced cell death in
hNSCs, F3 and F3.Akt1 hNSCs were exposed to varying concentration of H
2
O
2
.
F3.Akt1 cells showed a higher survival rate to H
2
O
2
-induced cell death as com-
pared to the F3 controls (Figures 2A-C). Whether H
2
O
2
induces changes in
phosphorylation status of Akt1 and caspase-3 cleavage in F3 and F3.Akt1 cells
was investigated. When F3 and F3.Akt1 cells were exposed to 0.5 mM H
2
O
2
for
6 hr, phosphorylated form of Akt1 and inactivated form of caspase-3 were found
in F3.AKt1 cells (Figure 2D), while F3 cells showed the opposite expression pat-
tern (Figure 2D). An increase in active fragment of caspase 3 (20 kDa) was found
in F3 cells following H
2
O
2
treatment. These results indicate that the enhanced
cell survival of F3. Akt1 cells as compared to parental F3 cells following H
2
O
2
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