Transcriptional Characterization of Glioma Neural Stem Cells Diva ...

2.2 Neural Stem Cells Introduction
· neurospheres differentiate more promptly into astrocytes rather than
neurons both in vitro and when transplanted into mice;
· human NS cells cannot be properly screened for compounds affecting
them because quantification is made difficult by their slow growth in
suspension culture and their variable death rate;
· cellular responses cannot be monitored within aggregates and fusion of
neurospheres can confound results based on their number and size.
Therefore, a niche-independent environment is better suited for the growth of
stem cell cultures, in that differentiation towards a specific lineage can always
be traced back to the stem cells themselves [165]. Such an environment was
produced when the presence of FGF2 and EGF alone was discovered to be
sufficient for the continuous expansion of NS cells in adherent conditions [107].
Niche-independent NS cell derivation
The key to proper usage of ES cell-based technologies is the development of
robust, reproducible and reliable protocols for controlling propagation and dif-
ferentiation of cells, and an important goal in embryonic stem cell biology over
the past 10 years has been that of developing protocols to enable the conver-
sion of mouse and human ES cells to the neural lineage [400].
The default model of neural induction proposes that the key event is the re-
moval of BMP signaling with no positive induction required [352]. In mouse
ES cells, however, positive induction is necessary for differentiation to take
place, since these cells can be maintained in vitro through the addition of the
Leukemia inhibitory factor (LIF) and BMP extrinsic factors, as well as in-
trinsic determinants Sox2, Oct-4 and Nanog transcription factors, but require
replacement of LIF and BMP to specify the direction of differentiation. It
was initially thought that exposure to RA and serum 29 in suspension culture
was required, provided LIF retraction, to generate neurons [38,471], although
later reports showed RA was unnecessary and neural precursors could be en-
riched in a serum-free basal media [367]. During neural differentiation, ES
cells are believed to undergo progressive lineage restrictions similar to those
observed during normal fetal development, providing a means to isolate dis-
tinct neural precursor populations such as NEP cells and radial glia, as well as
29 Animal derived fluid most commonly drawn from a bovine fetus that contains hormones
and growth factors that allow cells in culture to proliferate.
43