Biology Reference
In-Depth Information
Serum and feeder-based strategies for culturing PGCs of other mammals
have been attempted. Limited survival of pig PGCs on a STO feeder layer, in
serum-based medium, has been observed (
Takagi et al., 1997
), and similar
results have been reported in rabbits (
Kakegawa et al., 2008
), cow (
Cherny
et al., 1994
), goats (
K
¨
hholzer et al., 2000
), and sheep (
Ledda et al., 2010
).
A number of growth factors have been reported to increase the survival
and/or proliferation of PGCs or the derivation efficiency of EG cells in
feeder cultures.
5.2. Growth factors
In 1991, three papers reported a positive effect of SF on PGC cultures (
Dolci
et al., 1991; Godin et al., 1991; Matsui et al., 1991
). Dolci et al. and Godin
et al. concluded that SF improves PGC survival but not proliferation. While
they provide good evidence for the former claim, the latter is complicated by
the fact that both studies performed BrdU incorporation assays on a STO
feeder layer which itself expresses high levels of SF (
Dolci et al., 1991
).
Matsui et al. performed the same assay on a SF null feeder line. They show
that addition of SF increases BrdU incorporation to a level comparable to
that measured on STO cells, providing a clear demonstration that SF can
act both as a survival factor and a mitogen (
Matsui et al., 1991
). Furthermore,
they demonstrate that membrane-bound SF increases PGC number further,
even when saturating concentrations of soluble SF are added. These findings
are consistent with the phenotypes of the various Steel alleles described in
mice (see earlier).
Leukemia inhibitory factor (LIF) can also act as both a survival factor and
a mitogen for PGCs and is capable of acting synergistically with SF (
Matsui
et al., 1991
). A subsequent report concluded that LIF acts as a “putative sur-
vival factor for proliferating PGCs” (
de Felici & Dolci, 1991
). Indeed, both
SF and LIF can act on PGCs to block apoptosis (
Pesce, Farrace, Piacentini,
Dolci, & de Felici, 1993
), although this effect does not preclude other roles
for either factor. When basic fibroblast growth factor (bFGF) is combined
with LIF and soluble SF, there is a dramatic increase in the number of PGCs
(
Matsui et al., 1992
) but reprogramming to EG cells only occurs in the pres-
ence of membrane-bound SF (
Matsui et al., 1992; Resnick et al., 1992
).
bFGF can bind to, and act directly on, germ cells to increase their numbers
(
Resnick, Ortiz, Keller, & Donovan, 1998
) and is widely believed to act as
a mitogen (
de Felici, Dolci, & Pesce, 1993; Matsui et al., 1992; Resnick
et al., 1992; Takeuchi, Molyneaux, Runyan, Schaible, & Wylie, 2005
)
