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astrocytes, and promoted functional recovery in these animals [9]-[12], [15]-
[17]. However, low survival rate of grafted F3 NSCs in ischemia and ICH rats in
the previous studies is a grave concern; less than 50% of grafted NSCs survived in
ICH mice at 2-weeks post transplantation and 30% at 8-weeks [15], [16].
One possible way to promote extended survival of transplanted NSCs in ani-
mal brain is to modulate properties of the NSCs, and this might be accomplished
by over-expressing Akt1 protein which is known as a general mediator of cell sur-
vival signals in the NSCs. Akt, a serine/threonine kinase, plays a critical role in the
modulation of cell proliferation, growth, and survival. The PI3K-Akt signal path-
way is well-known for the cell survival and it exhibits anti-apoptotic effects against
a variety of apoptotic paradigms including withdrawal of extracellular signaling
factors, oxidative and osmotic stress, irradiation and ischemic shock [23]-[26].
Previous studies have demonstrated that overexpression of Akt prevents cerebellar
granule cells from apoptotic cell death during growth factor withdrawal [23], and
promotes cell survival during free radical exposure to free radical or hypoxia in
hippocampal neurons [27]-[29].
Considering evidence of functional recovery in stroke animals following brain
transplantation of human NSCs and Akt1 protein as a general mediator of sur-
vival signals, the present study is designed to investigate whether human NSCs
overexpressing Akt1 can lead to the prolonged cell survival of grafted human
NSCs and functional recovery in the mouse ICH stroke model.
Materials and Methods
cell culture
Primary dissociated cell cultures of fetal human brain tissues of 14 weeks gesta-
tion were prepared as described previously [30], [31]. The cells were grown in
T25 flasks in Dulbecco's modified Eagle medium with high glucose (DMEM;
HyClone, Logan, UT), 5% fetal bovine serum (FBS) and 20 µg/ml gentamicin
(Sigma, St Louis, MO) (Sigma). The medium was changed twice a week. The
permission to use the fetal tissues was granted by the Clinical Research Screening
Committee involving Human Subjects of the University of British Columbia,
and the fetal tissues were obtained from the Anatomical Pathology Department
of Vancouver General Hospital.
Stable, immortal human neural stem cell line, HB1.F3 (F3), was generated
from the primary fetal human brain cell culture as described previously [15],
[32]-[34]. PA317 amphotropic packaging cell line was infected with the recom-
binant replication-incompetent retroviral vector pLNX.v-myc, and the superna-
tants from the packaging cells were used to infect NSCs in human telencephalon
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