1 Department of Developmental Biology, Stanford University, Stanford, CA
94305-5329, USA
2 Department of Medicine (Oncology Division), Stanford University, Stanford, CA
94305-5329, USA
Author for correspondence (e-mail:
seungkim{at}cmgm.stanford.edu)
Accepted 21 September 2004
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SUMMARY |
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Key words: Gdf11, TGF-ß, Pancreas, Islet of Langerhans, Insulin, Stem cell
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Introduction |
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Both positive and negative regulators control the transient amplification
of NGN3+ islet progenitor cells in the developing pancreas.
Loss-of-function studies in mice, and in vitro analyses suggest that HNF6
(ONECUT1 Mouse Genome Informatics) is a direct activator of
Ngn3 transcription (Jacquemin et
al., 2000; Lee et al.,
2001
). By contrast, the product of hairy and enhancer of split 1
(HES1) probably functions as a direct repressor of Ngn3 transcription
(Jensen et al.,
2000a
,b
;
Lee et al., 2001
), and prior
studies provide evidence that pancreatic HES1 expression is controlled by
Notch signaling in the pancreas (Apelqvist
et al., 1999
; Schwitzgebel et
al., 2000
; Jensen et al.,
2000b
). However, it is not known if other intercellular signaling
pathways regulate development of NGN3+ islet progenitor cells.
During islet cell maturation, the progeny of hormone-negative,
proliferating NGN3+ cells exit the cell cycle, begin hormone
production and aggregate to form polyclonal islets with stereotyped
architecture (Gradwohl et al.,
2000; Schwitzgebel et al.,
2000
; Deltour et al.,
1991
). In neonatal mice, islets represent 2-3% of total pancreatic
mass, and insulin-producing ß-cells constitute about 80-90% of the islet.
Experiments in mice have identified a sequence of transcription factors,
including Ngn3, Pax6, Neurod1, Nkx2.2, Nkx6.1 and Isl1,
essential for maturation of ß-cells (reviewed by
Wilson et al., 2003
). Thus
far, experiments fail to demonstrate that a single transcription factor is
sufficient to direct pancreas development. Rather, classical and modern
studies reveal that islet cell maturation, apportioning of
endocrine-to-exocrine cell ratio, and islet morphogenesis are orchestrated by
multiple intercellular signals (Wessels and Cohen, 1967) (reviewed by
Kim and Hebrok, 2001
;
Edlund, 2002
;
Ober et al., 2003
;
Wilson et al., 2003
).
Prior studies of embryonic chicks and mice have implicated the transforming
growth factor ß (TGF-ß) signaling pathways as regulators of
pancreatic differentiation and islet development
(Hebrok et al., 1998;
Yamaoka et al., 1998
;
Shiozaki et al., 1999
;
Kim et al., 2000
;
Dichmann et al., 2003
;
Sanvito et al., 1995
;
Lee et al., 1995
).
Additionally, in vitro studies (Sanvito et
al., 1994
; Miralles et al.,
1998
; Miralles et al.,
1999
) have shown that TGF-ß ligand activity may regulate the
ratio of endocrine to exocrine cells during pancreas development. However, an
endogenous pancreatic TGF-ß ligand that regulates development of islet
progenitor cells has not yet been identified. Here we provide evidence that
growth differentiation factor 11 (GDF11; also known as BMP11) is a pancreatic
TGF-ß ligand that negatively regulates the production of NGN3+
pancreatic islet precursor cells, and is required for regulating islet size,
ß-cell maturation and ß-cell mass.
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Materials and methods |
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Histological analysis
Digoxigenin-labeled RNA probes of Gdf11
(McPherron et al., 1999),
Ngn3 (Dr D. Anderson, California Institute of Technology), and
Hes1 (Dr T. Vogt, Princeton University) were prepared and hybridized
as previously described (Kim et al.,
2000
). To detect hybridization of digoxigenin-UTP (dig-UTP)
labeled riboprobes to tissue sections, we used antibodies that detect dig-UTP
and NBT/BCIP (blue) or INT/BCIP (rust brown, Roche, Indianapolis, IN) as
previously described (Kim et al.,
2000
). Immunohistochemical analyses were performed as described
(Kim et al., 2000
;
Kim et al., 2002
). The primary
antisera used were: rabbit anti-caboxypeptidase A (1:200, Biogenesis,
Kingston, NH), guinea pig anti-glucagon (1:100, Linco, St Charles, Missouri),
rabbit anti-ghrelin (1:400, Dr Lori Sussel, University of Colorado Health
Sciences Center), rabbit anti-HES1 (1:500, Dr T. Sudo, Toray Industries Inc.,
Kamakura, Japan), rabbit anti-HNF6 (1:100, Dr G. Rousseau, Université
Catholique de Louvain, Brussels), guinea pig anti-insulin (1:100, Linco),
rabbit anti-insulin (1:100, ICN, Aurora, OH), hamster anti-mucin (1:200,
NeoMarkers, Fremont, CA), mouse anti-NKX2.2 (both 1:25, Developmental Studies
Hybridoma Bank, University of Iowa), rabbit anti-NKX6.1 (1:500, Dr M. German,
University of California, San Francisco), rabbit anti-PDX1 (1:200, Dr C.
Wright, Vanderbilt University, Nashville, TN), rabbit anti-PP and rabbit
anti-somatostatin (both 1:200, Dako, Carpinteria, CA), and mouse
anti-synaptophysin (1:50, BioGenex, San Ramon, CA). Secondary antibodies
(Jackson ImmunoResearch, West Grove, PA) for immunofluorescent or peroxidase
detection of primary antibodies were respective animal Ig conjuguated to Cy3
(1:800), FITC (1:200) or biotin (1:100). Ghrelin was detected using a
biotinylated secondary antibody followed by streptavidin-Cy3 (1:1000, Jackson
ImmunoResearch). To stain ductal cells, biotinylated lectin Dolichos
biflorus agglutinin (DBA) was used at 1:200 (Vector Laboratories, San
Bruno, CA) and visualized with streptavidin-FITC (1:200, Jackson
ImmunoResearch). Immunoperoxidase detection was performed using Vectastain
Elite ABC and DAB kits (Vector Laboratories). TUNEL assays were performed
using the Fluorescein In Situ Cell Death Detection Kit (Roche).
Immunofluorescent stains were imaged using a BioRad confocal microscope.
Immunoperoxidase pictures were captured using a Zeiss Axiophot microscope.
Morphometric quantification of pancreas tissue and cell numbers
Cell counting, point-counting morphometry, and ß-cell and -cell
mass determination were performed using standard morphometric techniques
(Kim et al., 2002
). To obtain
representative results, all quantification of immunostaining was performed by
counting numbers of positive-stained cells and dividing by the area of total
pancreatic tissues using a standard 10x10 microscope grid. For
quantification of cells expressing Ngn3, Nkx6.1, Nkx2.2, insulin,
glucagon, Hnf6, Hes1, carboxypeptidase A, mucin, DBA and
synaptophysin, pancreata were fixed, and sectioned to generate seven
micron-thick tissue sections. Appropriately stained cells were counted in a
minimum of 30 random microscopic fields obtained from at least three animals
per genotype (if greater, then number indicated in the appropriate figure
legend). All data represent the average of the indicated number of animals
± the standard error of the mean (s.e.m.). Two-tailed t-tests
were conducted to determine statistical significance.
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Results |
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|
Gdf11 deficiency leads to malformations of the stomach, spleen and pancreas
We analyzed Gdf11-deficient mice
(McPherron et al., 1999) to
elucidate the role of Gdf11 in development of the pancreas and
adjacent organs. In Gdf11-/- embryos, inspection by
stereomicroscopy revealed defects in stomach, spleen and pancreas morphology.
Analysis of sectioned stomachs from Gdf11-/- neonates and
their control littermates confirmed that there was a 2-fold reduction in the
thickness of the gastric wall, accompanied by a reduction in the number of
characteristic folds in the stomach, called gastric rugae
(Fig. 2A,B). The spleen in
Gdf11-/- mice was also invariably malformed
(Fig. 2A,B, insets), similar to
splenic defects seen in mice lacking the type IIB activin receptor (ACTRIIB;
ACVR2B Mouse Genome Informatics)
(Oh and Li, 1997
;
Oh et al., 2002
). The pancreas
in Gdf11-/- mice appeared grossly normal at E11.5 (data
not shown) but had an abnormally compact appearance by E12-13.5 and was
malformed by postnatal day (P) 1 (Fig.
2C,D). Thus, consistent with its expression in foregut epithelium,
GDF11 is required for morphogenesis of organs in the posterior foregut.
|
Disrupted pancreatic islet development in Gdf11-deficient mice
Immunohistochemical analysis identified defects of pancreatic islet
development in Gdf11-deficient mice. At birth, wild-type mouse
pancreatic islets contain clustered insulin-producing ß-cells surrounded
by other hormone-producing cells, including cells producing glucagon,
somatostatin or pancreatic polypeptide
(Fig. 3A,E,I,L,O). In
heterozygous Gdf11+/- mice at E17.5 and P1, the number of
islets was slightly increased, but the islets were clearly smaller than in
wild type (Fig. 3O,P). Total
ß-cell mass was not reduced (Fig.
3A,B,D,) but -cell mass in Gdf11+/-
mice was increased approximately 3-fold
(Fig. 3E,F,H). After weaning,
we did not detect alterations of glucose regulation in random-fed or
glucose-challenged Gdf11+/- mice (data not shown). In
Gdf11-/- mice, the mass of insulin+ cells was
reduced approximately 2-fold (Fig.
3A,C,D), and the mass of glucagon+ cells was increased
(Fig. 3E,G,H), resulting in
islets of reduced size (Fig.
3Q). We did not observe a significant change in the numbers of
cells expressing somatostatin or pancreatic polypeptide in
Gdf11+/- or Gdf11-/- mice
(Fig. 3I-K,L-N). Together,
these data provide evidence that GDF11 regulates islet cell fate and
morphogenesis.
|
|
To explore further the basis for accumulation of NGN3+ cells in
mice lacking GDF11, we examined expression of HNF6, an established positive
regulator of pancreatic Ngn3 transcription
(Jacquemin et al., 2000;
Lee et al., 2001
). The
expression of HNF6 in ductal cells normally peaks around E15.5, coinciding
with the peak in NGN3+ cell number
(Jacquemin et al., 2000
;
Gannon et al., 2000
;
Gradwohl et al., 2000
). By
E17.5 the number of these HNF6+ cells declines, paralleling a
decrease in NGN3+ cell number. Using anti-HNF6 antisera
(Jacquemin et al., 2000
), we
observed two levels of HNF6 protein accumulation in the embryonic pancreas at
E17.5. In acinar cells, low concentrations of HNF6 were detected in nuclei. By
contrast, we observed dark nuclear staining of cells associated with, and
comprising, ductal epithelia (Fig.
4M-O). Consistent with previous reports
(Gannon et al., 2000
), we
found few HNF6+ cells associated with ducts in wild-type mice at
E17.5 (Fig. 4M), corresponding
with a pronounced decrease in numbers of NGN3+ cells in wild-type
mice at this stage (Fig. 4G;
production of both HNF6- and NGN3-antisera in rabbits precluded
immunohistochemical co-localization studies). By contrast, we observed that
the number of these HNF6+ cells was increased in
Gdf11+/- and Gdf11-/- mice
(Fig. 4N-P). These increases
paralleled the accumulation of NGN3+ cells observed in these mice
(Fig. 4H,I). Together, these
data suggest that GDF11 may modulate the expression of HNF6 during
NGN3+ cell development, although further quantitative studies of
Hnf6 mRNA levels are required to confirm this possibility.
Defective ß-cell maturation in mice lacking GDF11
Despite increased numbers of NGN3+ islet precursor cells, the
numbers of insulin-producing cells in Gdf11-/- mice was
reduced (Fig. 3D). As we did
not detect evidence of increased pancreatic cell death or reduced cell
proliferation in Gdf11-/- mice, we postulated that
maturation of ß-cell precursors into insulin-producing cells was
disrupted by the absence of GDF11 in these mice. To test this possibility, we
examined expression of gene products in Gdf11-/- mice that
regulate or define stages of ß-cell maturation. In the mid- and
late-gestational pancreas, NK-class homeodomain transcription factors NKX2.2
and NKX6.1 are expressed in subsets of islet precursor cells and in more
mature hormone-expressing islet cells, including ß-cells (Schwitzgebel et
al., 2001; Sander et al.,
2000; Sussel et al.,
1998
). In Gdf11-/- mice at E15.5, we observed
significantly increased numbers of cells expressing NKX2.2 and NKX6.1
(Fig. 5A-F). Further analysis
showed that all cells expressing NKX6.1, which is required for ß-cell
maturation (Sander et al.,
2000
), co-expressed synaptophysin, consistent with the view that
these cells have begun to differentiate toward an endocrine fate
(Fig. 5G,H)
(Tomita, 2002
). As mature
ß-cells express both NKX6.1 and insulin
(Sander et al., 2000
), and
insulin+ cell numbers were reduced in Gdf11-/-
mice, these results raised the possibility that immature ß-cells
expressing NKX6.1 accumulate in Gdf11-/- mice.
|
Smad2 mutation leads to pancreatic defects similar to those in Gdf11-deficient mice
SMAD2 and SMAD3, also known as R-SMADs, are intracellular signal
transduction proteins activated by ligand-bound activin receptors (reviewed by
Massague and Chen, 2000), and
both are expressed in the developing mouse pancreas
(Brorson et al., 2001
) (data
not shown). Recent biochemical studies demonstrate that binding of GDF11 to
type II activin receptors can activate SMAD2
(Oh et al., 2002
). However,
TGF-ß signals may stimulate other signaling pathways, including the
mitogen-activated protein kinase (MAPK) cascade (reviewed by
Massague, 1998
). Moreover,
Ogihara and co-workers recently provided evidence from in vitro studies that
activin signaling regulates Ngn3 expression in the tumor-derived
AR42J cell line, and that SMAD-dependent signaling may not be required for
this effect (Ogihara et al.,
2003
). Thus, it was unclear if perturbation of SMAD signaling in
vivo would perturb pancreatic islet development.
We postulated that if GDF11 activates SMAD2 to regulate pancreas
development, then loss of Smad2 functions might phenocopy pancreatic
defects observed in Gdf11-deficient mice. As
Smad2/ mice fail to form endoderm
(Nomura and Li, 1998;
Tremblay et al., 2000
), we
examined pancreas development in Smad2+/ mice. We
observed increased numbers of Ngn3+ cells and reduced
ß-cell mass in Smad2+/ mice
(Fig. 6A,D,E,H). We also found
increased numbers of NKX2.2+ and NKX6.1+ cells in the
pancreas of Smad2+/ mice at E17.5
(Fig. 6B,C,F,G). Spleen and
stomach development in Smad2+/ mice, by contrast,
appeared normal (data not shown), suggesting that pancreas defects in
Smad2+/ mice occurred in the absence of generalized
foregut defects. Thus, our results show that Smad2 functions are
essential for islet development, and suggest that Smad3 functions are
unable to compensate fully for loss of Smad2 function in the
embryonic pancreas. The similarity of pancreatic defects in mice deficient for
Smad2 or Gdf11 also provides indirect evidence that GDF11
and SMAD2 function in the same signaling pathway to control accumulation and
maturation of Ngn3-expressing cells in pancreas development. The
phenotypes observed in mice haploinsufficient for Smad2 or
Gdf11 suggest that pancreas cell fates are highly sensitive to
loss-of-function mutations affecting this signaling pathway.
|
|
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Discussion |
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GDF11 signaling regulates the number of pancreatic NGN3+ islet progenitor cells
One of the principal findings in this study is that GDF11 modulates the
number of NGN3+ cells, a pancreatic progenitor cell population
crucial for islet development. Our data support a model in which GDF11
negatively regulates the formation of NGN3+ cells in the developing
pancreas. Smad2 is also required for this process, suggesting that
Gdf11 and Smad2 function in the same pathway in pancreatic
development. While there is good evidence that Notch signaling regulates
production of these islet progenitors, to our knowledge this is the first
demonstration that TGF-ß signaling can impact the development of these
cells. Supporting this view, we observed that mice deficient for
Gdf11 harbor defects in islet and exocrine cell fates similar to
those observed in animals that mis-express Ngn3. For example,
following forced expression of Ngn3 in foregut endoderm, chick
embryos produce increased numbers of glucagon+ cells
(Grapin-Botton et al., 2001).
Likewise, in mice that prematurely express Ngn3, including mice
lacking Hes1 (Jensen et al.,
2000b
) or in transgenic mice expressing Ngn3 from
Pdx1 (also known as Ipf1) regulatory elements
(Apelqvist et al., 1999
;
Schwitzgebel et al., 2000
),
there is a relative increase in the number of glucagon+ cells,
accompanied by reductions in acinar and insulin+ cell numbers. We
suggest that increased production and accumulation of NGN3+ cells
in Gdf11-/- mice is also the basis for increased numbers
of glucagon+ cells, and reductions in insulin+ and
acinar cell numbers, observed in these mice.
While the cellular origin of NGN3+ cells in the developing
pancreas remains the focus of investigation, results from several studies
provide some evidence that embryonic NGN3+ cells derive from
pancreatic ductal cells (Gradwohl et al.,
2000; Jensen et al.,
2000b
; Schwitzgebel et al.,
2000
; Gu et al.,
2002
). Could the increased numbers of NGN3+ cells we
detected in Gdf11-deficient pancreata reflect a global increase in
this ductal cell compartment? To address this possibility, ductal cells were
marked and quantified using previously described immunohistochemical methods
(Reid and Harris, 1998
;
Kobayashi et al., 2002
;
Dor et al., 2004
). These
analyses demonstrated that ductal cell number was unchanged in
Gdf11-/- and heterozygous Gdf11+/-
mice. Thus, our results make it unlikely that a global expansion of ductal
cell mass is the basis for increased numbers of NGN3+ cells
observed in Gdf11-deficient mice.
Our observations further suggest that the increased number of
NGN3+ cells in Gdf11-deficient mice is not solely an
indirect consequence of defective ß-cell maturation. The majority of
islet cells are insulin+ ß-cells, which derive from
NGN3+ precursors (Gu et al.,
2002), and we found evidence for impaired ß-cell maturation
in Gdf11-/- mice. In Gdf11+/- and
Gdf11-/- mice, the increase in numbers of NGN3+
cells was similar. However, the ß-cell mass in heterozygous
Gdf11+/- and wild-type mice was indistinguishable. Thus,
in Gdf11+/- mice, the increase of NGN3+ cells
is genetically uncoupled from apparent defects in production of
insulin+ cells. These observations support our view that the
increased number of NGN3+ cells in mice lacking GDF11 or SMAD2
results from increased production of NGN3+ cells. We have tested
purified GDF proteins for activity when added to E10.5 dorsal pancreatic
rudiments grown in tissue culture. We have not yet detected significant
activity of these factors in regulating formation of Ngn3-expressing
cells (Å.A., E.H. and S.K., unpublished). Thus, our studies do not yet
establish whether GDF11 is sufficient to regulate the development of
endogenous pancreatic islet precursor cells.
GDF11 and Notch signaling: parallel negative regulators of islet progenitor cell development?
Our data suggest that GDF11 and SMAD2 signaling activity regulate the
development of pancreatic NGN3+ pro-endocrine cells. Prior reports
suggested a role for the Notch signaling pathway in regulating the expansion
of the NGN3+ cell compartment in pancreas development
(Apelqvist et al., 1999;
Schwitzgebel et al., 2000
;
Jensen et al., 2001a; Jensen et al., 2001b). In certain contexts, others have
recently provided evidence for signaling interactions between the Notch and
TGF-ß pathways (Zavadil et al.,
2001
; Blokzijl et al.,
2003
). In our studies, the number of cells expressing Hes1, a
transcriptional repressor of Ngn3
(Jensen et al., 2000b
), was
unchanged in Gdf11 deficient mice. While further studies are needed
to exclude the possibility of other Notch signaling defects in
Gdf11-deficient mice, these data suggest that GDF11 activity may not
be required for pancreatic Notch signaling and production of HES1+
cells. Moreover, the number of pancreatic cells expressing Gdf11 was
similar in mice lacking Hes1 and control mice. Regulation of
TGF-ß ligand activity is thought to occur at several stages, including
transcription, post-translational processing, activation and inhibition
(Massague and Chen, 2000
;
Attisano and Wrana, 2002
).
Thus, it remains possible that Hes1-deficiency or other Notch
signaling defects may perturb Gdf11 signaling pathways. These caveats
notwithstanding, our results provide evidence for a model in which GDF11 and
Notch signaling function in parallel pathways to negatively regulate
production of pancreatic NGN3+ pro-endocrine cells.
Identification of unequivocal pancreatic endocrine defects arising from
Gdf11 or Smad2 haploinsufficiency suggests that developing
islet progenitor cells are particularly sensitive to perturbations in
TGF-ß signaling, and raises the possibility that subsets of familial
pancreatic endocrine disorders could result from heterozygous mutation in
Smad2 or Gdf11. In mice deficient for Gdf11, we
observed that some phenotypes, such as accumulation of excess
NGN3+, HNF6+ or glucagon+ cells, was
similarly severe in both Gdf11+/- and
Gdf11-/- mice, while others, such as reduction of
ß-cell mass, was observed only in Gdf11-/- mice.
While the molecular basis for these gene-dosage effects requires further
elucidation, we note that the correlation of phenotypic severity with
Gdf11 gene dosage is consistent with similar effects we previously
reported for mice harboring one, two or three null alleles of the type 2
activin receptors ActRIIA and ActRIIB
(Kim et al., 2000). Together,
these observations suggest that the strength or duration of TGF-ß
signaling may be a crucial determinant of cell fates in the developing
pancreas.
The preservation of some ß-cell development in mice deficient for
Gdf11 or Smad2 indicates that additional TGF-ß ligands
and intracellular SMADs may also regulate pancreatic islet development.
Consistent with this possibility, prior work has shown that TGF-ß ligands
like BMP7, activins, TGF-ß1, TGF-ß2, and TGF-ß3, and the R-SMAD
members SMAD1, SMAD3, and SMAD5 are expressed in the developing pancreas
(Lyons et al., 1995;
Miralles et al., 1999
;
Brorson et al., 2001
;
Dichmann et al., 2003
). Thus,
redundant activities from these factors in the pancreas might mitigate the
observed in vivo effects of Gdf11 or Smad2 loss of function.
Further experiments, including studies of genetic interactions between
Gdf11 and Smad2, are required to determine effects of more
severe disruption of TGF-ß signaling in islet development.
GDF11 and SMAD2 activity regulate pancreatic ß-cell maturation and number
The second principal finding in our study is that GDF11 may regulate the
terminal differentiation of NKX6.1+ cells into insulin-producing
ß-cells. While GDF11 has been shown previously to regulate the number of
cellular progenitors in organs such as the olfactory primordium, there has
been no prior evidence, to our knowledge, for GDF11 roles in maturation of
cells toward a terminally differentiated fate. Thus, our study provides novel
insights into the range of developmental roles for GDF11 in neuroendocrine
cell differentiation.
Recent evidence suggests that GDF11 activity regulates neurogenesis in
embryonic olfactory epithelium (Wu et al.,
2003). In this study, Calof and colleagues described the number of
ngn1+ immediate neuronal precursors (INPs) in the
olfactory epithelium of Gdf11-/- mice as being increased
by 20%, with corresponding increases in the number of differentiated olfactory
neurons (Wu et al., 2003
).
Thus, GDF11 negatively regulates formation of ngn1+ INPs
but is not essential for neuronal cell maturation or survival in the
developing olfactory epithelium. In the pancreas of
Gdf11-/- mice we observed a 300% increase of
NGN3+ cells, accumulation of immature endocrine cells, and reduced
ß-cell mass. Together, our data show that GDF11 activity regulates
both the production of pancreatic islet progenitor cells and the
maturation of these progenitor cells into insulin-producing ß-cells.
Further support for this view also comes from a recent study by Sarvetnick and
colleagues, who show that TGF-ß signaling may regulate endocrine cell
maturation in the postnatal pancreas
(Zhang et al., 2004
). Future
studies should allow us to determine whether GDF11 and SMAD2 signaling also
serve to maintain the function and fate of fully differentiated endocrine
cells in the adult pancreas.
Cell cycle regulators also probably control maturation of ß-cells and
other islet cells, as the majority of islet cells are postmitotic, and
increased hormone production is associated with growth arrest
(Ptasznik et al., 1997).
TGF-ß signaling is an established regulator of the cell cycle, and in
specific contexts TGF-ß signals have been shown to control the expression
of cell cycle components, including p15, p18, p21 and p27kip1,
members of the family of cyclin-dependent kinase inhibitors
(Moustakas et al., 2002
). In
maturing neural progenitor cells, GDF11 activity increases expression of the
cyclin-dependent kinase inhibitor p27kip1
(Wu et al., 2003
), suggesting
that disrupted endocrine cell maturation in Gdf11-/- mice
might reflect an impairment of growth arrest. Indeed, we have also observed an
overall reduction of pancreatic p27kip1 expression in
Gdf11-/- mice (E. Harmon, PhD thesis, Stanford University,
2003), but it remains unclear from these and other studies whether
p27kip1 expression is reduced specifically in pancreatic endocrine
or ß-cell precursors (N.S., Å.A., E.H. and S.K., unpublished).
Thus, additional tests are required to clarify how GDF11 may promote
ß-cell maturation.
This study provides evidence that GDF11 is an endogenous inhibitor of
endocrine progenitor cell formation in the embryonic pancreas. Our studies
reveal that Gdf11 is expressed throughout the pancreatic epithelial
cell compartment when NGN3+ endocrine progenitor cells first
appear. Thus, we postulate that GDF11 is a crucial autocrine or paracrine
regulator of islet progenitor cell formation. Similar short-range signaling
roles have been proposed for regulation of myoblast development by GDF8, a
TGF-ß ligand highly homologous to GDF11, and for GDF11 in regulating
neurogenesis (Lee and McPherron,
1999; Thomas et al.,
2000
; Wu et al.,
2003
). Identification of TGF-ß signals that regulate
pancreatic islet development may prove useful for generating strategies to
regenerate or replace islets in patients with diabetes mellitus.
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ACKNOWLEDGMENTS |
---|
![]() |
Footnotes |
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* Present address: MRC Centre for Developmental Neurobiology, King's College
London, London SE1 1UL, UK
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