Biomedical Engineering Reference
In-Depth Information
2003
; Dressler
2006
). In
2000
, Meng et al. reported that spermatogenesis is
disrupted in mice carrying a single
Gdnf
null allele, while an accumulation of A
pr
and A
al
spermatogonia was observed in the testes of mice over-expressing GDNF.
In serum-free culture conditions, supplementation with GDNF is essential for
SSC self-renewal
in vitro
for extended periods of time (Kubota et al.
2004a
).
Additionally, studies by Kanatsu-Shinohara et al. (
2003
) showed a similar impor-
tance of GDNF in supporting survival and proliferation of mouse gonocytes
in vitro
. Moreover, the self-renewal of rat and hamster SSCs
in vitro
is also
supported by exposure to GDNF (Hamra et al.
2005
; Ryu et al.
2005
; Kanatsu-
Shinohara et al.
2008
). Maintenance of proliferating spermatogonia in defined
culture conditions with GDNF supplementation supports the formation of germ
cell clumps. These clumps can be maintained and expanded for extended periods
of time and upon transplantation, some of the cells can reestablish spermatogen-
esis in a recipient testis demonstrating the presence of SSCs within the germ cell
clump (Kanatsu-Shinohara et al.
2003
; Kubota et al.
2004a
; Ryu et al.
2005
).
However, the germ cell clumps are not composed purely of SSCs and also contain
other non stem cell spermatogonia, which are likely A
pr
-like and/or A
al
-like sper-
matogonia produced upon SSC differentiation (Kubota et al.
2004a
; Kanatsu-
Shinohara et al.
2005
; Dann et al.
2008
). The concentration of SSCs within germ
cell clumps can vary widely throughout a given culture period (Kubota et al.
2004a
; Kanatsu-Shinohara et al.
2005
), estimated to be as low as 0.02% of the
cell population in one study (Kanatsu-Shinohara et al.
2005
). Regardless, the
truest measure of an SSC is the ability of a germ cell to reestablish spermatogen-
esis following transplantation and cells capable of fulfilling this criterion are
present in cultured spermatogonial populations. These observations indicate that
both self-renewal and differentiation of SSCs is supported in clump-forming
spermatogonial cultures, which provides an excellent
in vitro
model system to
study SSC fate decisions.
Self-renewal of stem cells can be defined by the ability of a cell to undergo
mitosis producing two new daughter cells of which one is identical to the parent
cell and does not enter a differentiation pathway, maintaining the capability to
undergo future self-renewing divisions. Response to specific growth factors is one
major regulatory mechanism of this fate decision and requires the binding and
stimulation of specific receptor complexes expressed by stem cells. Although
GDNF is an essential regulator of SSC proliferation, its receptor complex
consisting of GDNF family receptor a 1 (GFRa1) and c-RET are expressed by
A
s
, A
pr
, and A
al
spermatogonia (Dettin et al.
2003
; Naughton et al.
2006
). The
GFRa1+ cell fraction is lesser than twofold enriched for SSCs in the pre-pubertal
mouse testis and depleted of SSCs in the adult mouse testis (Buageaw et al.
2005
;
Ebata et al.
2005
; Grisanti et al.
2009
). Additionally, fewer SSCs are found in the
c-Ret+ cell population of both pre-pubertal and adult mouse testes compared to
the total testis cell population (Ebata et al.
2005
). Because GDNF action is not
specific to SSCs, functional transplantation assays must be conducted when exam-
ining GDNF actions on spermatogonia to make unequivocal conclusions about the
biology of SSCs.
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