Endocrine cells of the adult pancreas are organized into the
islets of Langerhans, which are scattered throughout the
exocrine tissue. There are four major islet cell types: α, β, ∂
and PP cells that synthesize glucagon (GLU), insulin (IN),
somatostatin (SOM) and pancreatic polypeptide (PP), as their
principal differentiated hormone products. In addition to the
four hormones, islet cells synthesize several neuronal-specific
markers such as tyrosine hydroxylase (TH) and neuronal
specific enolase (Alpert et al., 1988 and references therein).
The mammalian pancreas develops by fusion of dorsal and
ventral primordia that appear as evaginations of the gut. The
first differentiated pancreatic cells appear in the endodermal
layer of the gut in embryos of about 20 somites, correspond-
ing to day 9.5 of development (E9.5) in mouse and E10.5 in
the rat (Wessels and Evans, 1968; Pictet and Rutter, 1972;
Kaufman, 1992). The two primitive glands grow indepen-
dently, forming both endocrine and exocrine tissues, and
finally merge at E10.5 in mouse and E11.5 in rat (Pictet and
Rutter, 1972). Although it is generally agreed that precursor
cells in the pancreatic duct give rise to the endocrine and
exocrine compartment of the pancreas, the origin of these two
cell types from common endodermal precursors has been con-
troversial and it was hypothesized that islet cells were neural
crest derivatives (Pearse, 1977).
Previous studies suggest that all four islet cell types arise
from common multipotent precursors that coexpress several
hormones and neural markers when they first differentiate
(Alpert et al., 1988; De Krieger et al., 1992; Lukinius et al.,
1992). As these stem cells mature, their antigenic repertoire
becomes restricted to a single hormone (Alpert et al., 1988).
Cells coexpressing GLU and insulin C-peptide (IN C-P) first
appear in primordial pancreas at E9.5. The number of IN+
GLU+cells decreases during gestation and, by E14.5, a sig-
nificant number of cells express only one hormone. Cells con-
taining SOM and PP first differentiate at E14.5 and postnatal
day 1 (P1) respectively, Each of these cell types also co-
express IN and GLU when they first appear (Alpert et al.,
1988). Co-expression of several polypeptide hormones is also
Development 121,11-18 (1995)
Printed in Great Britain © The Company of Biologists Limited 1995
The XlHbox 8 homeodomain protein of Xenopus and STF-
1, its mammalian homolog, are selectively expressed by β
cells of adult mouse pancreatic islets, where they are likely
to regulate insulin expression. We sought to determine
whether the expression of the homeobox protein/s during
mouse embryonic development was specific to β cells or,
alternatively, whether XlHbox 8/STF-1 protein/s were
initially expressed by multipotential precursors and only
later became restricted to the insulin-containing cells. With
two antibodies, we studied the localization of STF-1 during
murine pancreatic development. In embryos, as in adults,
STF-1 was expressed by most β cells, by subsets of the other
islet cell types and by mucosal epithelial cells of the
duodenum. In addition, most epithelial cells of the pancre-
atic duct and exocrine cells of the pancreas transiently
contained STF-1. We conclude that in mouse, STF-1 not
only labels a domain of intestinal epithelial cells but also
provides a spatial and temporal marker of endodermal
commitment to a pancreatic and subsequently, to an
endocrine β cell fate. We propose a model of pancreatic cell
development that suggests that exocrine and endocrine (α,
β, ∂ and PP) cells arise from a common precursor pool of
STF-1+cells and that progression towards a defined mono-
specific non-β cell type is correlated with loss of STF-1
Key words: pancreatic islets, insulin gene, ontogeny, homeoprotein,
duodenum, pancreatic cell lineage, murine pancreas
Expression of murine STF-1, a putative insulin gene transcription factor, in β
cells of pancreas, duodenal epithelium and pancreatic exocrine and
endocrine progenitors during ontogeny
Y. Guz1, M. R. Montminy2, R. Stein3, J. Leonard2,†, L. W. Gamer4, C. V. E. Wright4and G. Teitelman1,*
1Department of Anatomy and Cell Biology, SUNY Health Science Center at Brooklyn, Brooklyn, New York 11203, USA
2The Clayton Foundation Laboratories for Peptide Biology, The Salk Institute, La Jolla, California, 9203, USA
3Department of Molecular Physiology and Biophysics, Vanderbilt University Medical Center, Nashville, Tennessee 37232, USA
4Department of Cell Biology, Vanderbilt University Medical Center, Nashville, Tennessee 37232-2175, USA
†Present address: Strang-Cornell Cancer Research Laboratory, 510 East 73 Street, New York, NY 10021, USA
*Author for correspondence
reported in individual cells of embryonic human and porcine
pancreas (De Krieger et al., 1992; Lukinius et al., 1992). These
findings suggested that common factors are involved in islet
cell determination, whereas a distinct set of signals restrict
expression of differentiated islet hormone genes in α, β, ∂ and
The mechanisms that govern islet-specific gene expression
have been intensely investigated in recent years. Because the
promoter regions required for islet expression of insulin,
somatostatin and glucagon include critical AT-rich elements,
it has been presumed that homeodomain-containing factors
are important activators of these genes. The homeodomain is
a sequence-specific DNA-binding motif present in numerous
developmentally regulated transcription factors, some of
which are thought to be required for the expression of lineage
specific genes (Bodner et al., 1988; Ingraham et al., 1988). It
was initially suggested that the homeodomain-containing
factor XlHbox 8 might function in pancreatic development
because its expression in Xenopus embryos corresponded to
endodermal cells of the duodenum and developing pancreas
(Wright et al., 1988). Subsequently, the putative murine and
rat homologues of XlHbox 8 have been isolated and variously
termed as STF-1 (Leonard et al., 1993), IPF-1 (Ohlsson et al.,
1993) and IDX-1 (Miller et al., 1994). Characterization of all
three mammalian proteins revealed their close similarity to
XlHbox 8 in the N-terminal region and 100% identity in the
homeodomain. For clarity, we will refer to this protein as
STF-1. DNA-binding and transactivation assays suggest that
STF-1 may be an important regulator of insulin (and somato-
statin) gene expression in the islet (Ohlsson et al., 1993;
Leonard et al., 1993; Peshavaria et al., 1994; Miller et al.,
Immunohistochemical studies reported the localization of
STF-1 to β cells of embryos and adults (Ohlsson et al., 1993)
suggesting that in mammals, in contrast to amphibians, STF-
1 was a specific β cell marker. We have also reported that
STF-1 is primarily restricted to β cells (91%) of adult mice
pancreata (Peshavaria et al., 1994). However, we also detected
STF-1 expression in approximately 3% of α and 15% of ∂
cells (Peshavaria et al., 1994), indicating a less stringent dis-
tribution of the homeoprotein. In light of the differences
between our observations and those reported by Ohlsson et al.
(1993), we have examined SFT-1 expression in pancreas
during embryonic development and in gut of embryos and
Here we find that STF-1 is expressed during islet develop-
ment in β cells, in other precursor cells destined to become α,
∂ and PP cells as well as in the epithelial layer of the duodenal
mucosa. Moreover,we also find that exocrine cells of the
pancreas and most epithelial cells of the pancreatic duct tran-
siently express STF-1 during development. These observa-
tions suggest that STF-1 expression during development iden-
tifies an endodermal domain in the gut with two
compartments, one that will generate the epithelial layer of the
duodenum and a second subset that is endowed with the
potential to form pancreas. This latter compartment of STF-
1+precursor cells subsequently gives rise to pancreatic
exocrine and endocrine cells. With further development, STF-
1 expression in the pancreas becomes highly confined to β
cells where it plays a critical role in the regulation of insulin
MATERIALS AND METHODS
Animals and tissue processing
Pregnant CD-1 mice were purchased from Charles River. The appear-
ance of the vaginal plug was considered day 0.5 of gestation (E0.5).
Pregnant females were killed by cervical dislocation, the uterus was
removed and placed in 4% paraformaldehyde buffered to pH 7.4 with
0.1 M sodium phosphate buffer (PBS). The embryos were dissected
in the fixative solution and were postfixed for 1 hour in the same
solution. Embryos were examined at E9.0, 9.5, 11.5, 13.5, 15.5, and
16.5. Postnatal and adult CD-1 mice were perfused through the heart
with the fixative solution; the pancreas was then removed and
postfixed for 1 hour. The fixed tissues were infiltrated overnight in
30% sucrose, mounted in embedding matrix (Lipshaw Co., Pittsburg,
Pa) and 15-20 µm cryostat sections were collected onto gelatin-coated
Source of antibodies and purified peptides
The following antisera were used to stain cryostat sections: guinea
pig antibodies to bovine insulin and rat C-peptide were purchased
from Linco Research Inc (Eureka, MO); rabbit antisera to human
glucagon was purchased from Calbiochem (San Diego, CA); rabbit
antisera to human PP and somatostatin were supplied by Peninsula
Labs (Belmont CA); rabbit antiserum to human α amylase was
purchased from Accurate Chem. Sci. Corp. (Westbury, N.Y.)
Biotinylated goat anti-rabbit IgG, goat anti-guinea pig IgG and
avidin-labelled peroxidase were purchased from Vector Laboratories
(Burlingame, CA). Antiserum to the N-terminal domain of XIHbox
8 was raised against the first seventy five amino acids of XIHbox 8
as a GST/XIHbox 8 fusion protein (Wright et al., 1988; Peshavaria
et al., 1994). STF-1 antiserum was raised in rabbits using a synthetic
STF-1 peptide extending from amino acids 196-214 (Leonard et al.,
Characterization of homeoprotein antibodies
Anti-HlXbox 8 serum was purified with a GSE-E. coli extract
depletion matrix and then affinity purified on a column containing
immobilized N-terminal fusion protein. The affinity purified antibody
reacts with N-terminal fusion protein on western blots of bacterial
proteins. Preincubation of the antibody with the fusion protein blocks
all staining in western blots (results not shown). Western blot analysis
of XIHbox 8 and STF-1 antisera revealed that both interact with a
47×10−3Mrprotein on HIT and β TC cells (Peshavaria et al., 1994).
Occasionally, immunoblots of β TC cells also revealed a 39×10−3
band, which could be a degradation product; this band was never
observed in HIT or α cell extracts.
Immunolabeling of cryostat sections using peroxidase
Sections on slides were transferred to Tris-saline solution (TS; 0.9%
NaCl in 0.1 M Tris, pH 7.4) and were immunostained using the
avidin-biotin-HRP method. In brief, the sections were incubated
sequentially in: (a) 0.3% Triton X-100 in a 1% solution of goat serum
in TS for 15 minutes; (b) a 1:30 dilution of goat serum (Gibco) in TS
for 30 minutes; (c) an empirically derived optimal dilution of control
serum or primary antibody raised in species ‘X’ containing 1% goat
serum in TS for 18 hours; (d) a 1:50 dilution of anti-(species x)
biotinylated IgG solution in 1% goat serum in TS for 30 minutes and
(e) a 1:100 dilution of peroxidase-avidin complex for 30 minutes.
Following these incubations, the bound peroxidase was visualized by
reaction for 6 minutes in a solution containing 22 mg of 3,3′-
diaminobenzidine (DAB) and 10 µl of 30% H2O2in 100 ml of 0.1 M
TS. All incubations were carried out at room temperature. After the
DAB step, sections were dehydrated and mounted with Permount.
Antibodies were used at the following dilutions: guinea pig anti-
bovine insulin, 1:400; guinea pig anti-rat insulin C-peptide, 1:300;
Y. Guz and others
13 Homeoprotein expression in pancreas
rabbit anti-human glucagon, 1:12,000; rabbit anti-human somato-
statin, 1:8,000; rabbit anti-human pancreatic polypeptide, 1:20,000;
rabbit anti-human amylase, 1:1000; Rabbit anti-XIHbox 8 and rabbit
Sections were first incubated with antisera to STF-1 or XIHbox 8 and
the bound antibody was visualized by DAB (brown precipitate),
followed by incubation with antiserum to a hormone, which was visu-
Fig. 1. Gradual restriction of STF-1 expression to pancreatic islets during development. Photomicrographs illustrating immunohistochemical
localization of STF-1. (A) Cross section of E8.5 embryo. Note distribution of labelled nuclei (arrowheads) in the dorsal wall of the gut. Bar, 15
µm. N, neural tube; n, notochord; a, dorsal aorta; arrow indicates the communication of the foregut diverticulum with the yolk sac. (B) Cross
section of an E9.5 embryo illustrates the presence of labelled nuclei in the primordia of the dorsal pancreas (dp) and ventral (vp) pancreatic
primordia, bar, 50 µm. (C) E13.5 embryo. A large number of immunoreactive cells are distributed throughout the pancreatic duct (indicated
with arrowheads) and in small clusters; bar, 50 µm. (D) High magnification microphotograph of E13.5 pancreas illustrates the presence of
darkly and lightly stained nuclei; bar, 15 µm. (E) E17.5 pancreas. Stained cells are located in newly formed islets. Some cells expressing the
homeoprotein are seen in the pancreatic duct. Note that exocrine tissue is devoid of immunoreactive cells; bar, 15 µm. (F) In adults, the
homeoprotein is expressed exclusively by islet cells; bar, 15 µm. Darkly stained cells indicated with asterisks in B and D are nucleated red
blood cells that contain peroxidase. These cells are also seen in controls in which incubation with the first antibody has been omitted.
alized with the blue reaction product of the Vector SG substrate
Analysis of double label staining
Slides were examined with a Nikon Microphot SA microscope
equipped with Nomarski optics and using a 10× ocular and a 100× oil
immersion objective. Depth of focus was calculated according to man-
ufacture specifications and Klein and Furtak (1986) as follows:
D =n × λ/2(NA)2+ n/7 × NA × M
where n = refractive index on object side, λ = wavelength of light
(nm) and M = total magnification
D = 1.515 × 550/2(1,25)2+ 1.515/7 × 1.25 × 1250*
*Due to 1.25 differential interference contrast (DIC) magnification
factor in microscope.
With a depth of focus of approximately 0.4 µm only objects located
at ±0.4 µm are within the same plane of focus. Therefore, structures
from different cells can be easily distinguished since they are at
different plane of focus and cannot be focused simultaneously for
photography. The small depth of focus used allowed us to distinguish
clearly the presence (or absence) of staining in the nucleus and
cytoplasm of individual cells.
These immunohistochemical studies were conducted with two
different polyclonal antisera that recognized the STF-1 protein, one
raised to the N-terminal region of XlH box 8 (Peshavaria et al.,
1994) and the other to the C-terminal region of the rat STF-1
(Leonard et al., 1993). The same results were obtained with each.
In all stages examined, STF-1 immunoreactivity was localized to
the nucleus. The distribution of STF-1 was examined throughout
development beginning at day 8 (E8; 5 to 7 somites). Cells con-
taining STF-1 were first seen at E8.5 (Fig. 1A), which corre-
sponded to embryos with 11-13 somites (Kaufman, 1992). At E8.5,
STF-1+ cells were present in the endodermal layer of the dorsal
region of the gut (Fig. 1A) but not in the laterally located endo-
dermal cells. At this stage the midgut is still wide open into the
yolk sac and lacks a ventral wall. In 13- to 20-somite embryos (E9)
when the wide communication between the midgut and yolk sac
narrows and the gut has become tubular, cells containing STF-1
were seen in a transverse band of endoderm along the dorsal, lateral
and ventral regions of the presumptive duodenum (Fig. 2A) and in
cells forming the dorsal pancreatic primordium. The appearance of
STF-1 immunoreactivity at E8.5 preceded that of GLU or IN C-P,
which were first seen at E9.5 (20 somites; Teitelman et al., 1993).
At E9.5, STF-1+cells were also seen in the ventral pancreatic
primordia (Fig. 1B) and at E11.5, groups of cells adjacent to the
pancreatic duct were immunostained. Two days later, at E13.5, the
great majority of cells of the epithelium of the pancreatic duct
contained STF-1 immunoreactivity (Fig. 1C,D). In addition, the
pancreatic primordia contained numerous clusters of STF-1+cells,
some of which had darkly stained nuclei while other cell nuclei
contained less immunoreactive product (Fig. 1D). At E16.5, when
endocrine cells began to aggregate to form pancreatic islets, STF-
1+ cells were located in the newly formed islets, surrounded mainly
by unstained exocrine tissue. The number of immunostained pan-
creatic duct cells had also significantly decreased. One day later,
at E17.5, STF-1+cells in the pancreas were seen exclusively within
islets (Fig. 1E) and were nearly absent from the pancreatic duct.
As shown previously, STF-1 protein in adult pancreas is restricted
to islet cells (Fig. 1F), with no expression detected in exocrine cells
(Peshavaria et al., 1994).
In contrast to the non-β cells of the pancreas, expression of
STF-1 in the duodenal mucosa persisted throughout life. At
E9.5 STF-1 was expressed by most cells of the epithelium of
the mucosal layer but not by cells of the adjacent connective
tissue sheath. In duodenum of older embryos and adults, almost
all cells forming the simple columnar epithelium that line the
villi were STF-1+whereas the crypt cells did not contain the
homeoprotein. Cells of the other layers of the mucosa as well
as cells of the submucosa, muscularis and adventitial layers of
the wall of the duodenum also lacked STF-1 expression (Fig.
2B,C). Cells of the gut wall in other regions of the digestive
tract never contained STF-1 immunoreactivity (not shown).
To determine the identity of the islet cells expressing STF-1,
we performed double immunohistochemical visualization of the
homeoprotein with each of the four pancreatic hormones. At
E9.5, a time when glucagon and insulin first appeared, some STF-
Y. Guz and others
Fig. 2. Expression of STF-1 in the duodenum. (A) Photomicrographs
of a cross section of the duodenum at E9.5 illustrates that most cells
of the mucosa (arrowheads) express the homeoprotein; bar, 50 µm.
(C) Photomicrograph of an adult duodenum illustrates the presence
of STF-1+cells in the epithelium that lines the villi (arrowheads). In
contrast, nuclei of cells in the cript (arrows) lack STF-1
immunoreactivity. Dark cells, indicated with asterisks, are red blood
cells. Bar, 20 µm.
15Homeoprotein expression in pancreas
1+ cells coexpressed IN C-P or GLU (Fig. 3). While most IN+
cells expressed the homeoprotein, few GLU+ cells at E9.5 and at
later stages contained the antigen (Fig. 3A,B and Table 1). During
development, the number of IN+ cells increased and almost all of
them expressed STF-1 (Fig. 3C). At E14.5 and at P1, when ∂ and
PP cells first differentiate, STF-1 was detected in approximately
Fig. 3. Coexpression of STF-1 with hormones in the embryonic pancreas. These photomicrographs illustrate the immunohistochemical
localization of a hormone (visualized with a blue reaction product) and STF-1 (visualized with a brown reaction product) in the same tissue
section of embryonic pancreas. (A,B) Coexpression of STF-1 and glucagon at E9.5 (A), and E13.5 (B). A shows glucagon cells expressing the
homeoprotein (arrow), alpha cells with unlabeled nuclei (arrowheads) and cells in which only the nucleus is stained. Similar cells are also
visualized in B. Bar, 4 µm. (C) Coexpression of insulin and STF-1 at E14,5. Bar, 4 µm. (D) Cells indicated with arrows contain somatostatin
and STF-1. Bar, 4 µm. (E,F) Coexpression of STF-1 and PP at P1 (E) and in the adult (F). Also note that nuclei of exocrine cells at P1 are
unstained.Bar, 4 µm.
40% of SOM+and PP+cells (Fig. 3D,E and Table 1). We found
that most β cells of adults contained STF-1 whereas this protein
was only present in a fraction of the α and ∂ cells (Table 1). In
addition we determined that some PP cells (8.6%) of adult islets
contain STF-1 immunoreactivity (Fig. 3F).
To ascertain whether STF-1 was expressed in precursors of the
exocrine pancreas, we co-localized amylase, an exocrine cell
marker, and STF-1 in embryonic and adult pancreas respectively.
While previous studies indicated that amylase immunoreactivity
was first detected at E14.5 (Teitelman et al.,1987), we now detect
cells expressing amylase at E13.5. The appearance of amy+cells
at an ealier stage is probably due to the improved sensitivity of
the avidin-biotin system. Examination of E13.5 pancreas revealed
that 41.3% of amylase+cells contained STF-1 (Fig. 4A,B and
Table 1). Coexpression of amylase and the homeoproteins
decreased dramatically at E17.5 (Fig. 4C
and Table 1) and, in adults, no amylase+
cells expressed STF-1 (Fig. 4D and
The studies reported here sought to
determine whether STF-1 expression
was restricted to the pancreatic
primordia, and, if so, whether a
specific cell type of the pancreas
contained the homeoprotein or, alter-
natively, whether the homeodomain
protein was initially expressed by mul-
tipotential precursors and only later
became restricted to a single cell type
during islet cell maturation.
We found that STF-1 expression first
appeared in a narrow transverse band
of endoderm in the duodenum at E8.5
and preceded that of the islet hormones.
At this stage the embryo has just
completed the process of turning, and
the tubular shape of the gut becomes
apparent (Kaufman, 1992). In addition,
we show that during development,
STF-1 was transiently expressed in
cells of the pancreatic duct and in
endocrine cells with each islet hormone
as well as in acinar cells with the
exocrine protein amylase. The distrib-
ution of STF-1+cells became gradually
restricted and, in the mature pancreas,
STF-1 immunoreactivity was localized
to β cells and to small subsets of the
other endocrine non-β cells of the islet.
suggest that in mouse, as in Xenopus,
STF-1 provides a spatial and temporal
marker of endodermal commitment to
a pancreatic and subsequently, to an
endocrine β cell fate.
The fact that a subset of GLU+cells
contained STF-1 from the time they first
Y. Guz and others
Table 1. Coexpression of STF-1 and pancreatic exocrine
and endocrine markers during development
Glu/STF-1 Som/STF-1PP/STF-1 Amy/STF-1
At least six embryonic and newborn pancreata were processed for each
antigen combination. Total number of cells scored was 234 cells at E9.5 and
over 500 cells at other developmental stages. The number of cells expressing
STF-1 and a pancreatic antigen is expressed as the mean percentage ±s.e.m.
of the cells immunoreactive to the antigen. Abbreviations: ND, not
determined; ne, not expressed (pancreatic exocrine or endocrine marker is not
expressed at that developmental stage). *Peshavaria et al. 1994.
Fig. 4. Exocrine cells transiently express STF-1 during development.(A,B) The colocalization of
the homeoprotein and amylase at E15.5. At this stage many amylase+cells express STF-1
(arrows). also shown are STF-1+Amy−cells (arrowheads); Bar, 4 µm. (C) Photomicrograph of
an E17.5 pancreas shows that, at this stage, most exocrine cells do not contain STF-1. Also
shown is a small cluster of STF-1+, amylase−cells. It is likely that these cells are islet cells. Bar,
40 µm. (D) In adult pancreas, homeoprotein expression is restricted to islet cells. Bar, 15 µm.
17Homeoprotein expression in pancreas
appeared supports our hypothesis that α and β cells arose from
a common pool of stem cells (Alpert et al., 1988; Teitelman et
al., 1993) and also indicates that most α cells extinguished STF-
1 expression rapidly. Conceivably, the GLU+/STF-1+ cells at
E9.5 also contained IN and represent the precursors of the other
islet cell types (Fig. 5). During midgestation, almost 50% of the
newly differentiated SOM+cells expressed STF-1, strongly sug-
gesting that ∂ cells were generated from precursors containing
this homeoprotein. The fact that, in embryos, ∂ cells coexpressed
IN when they first appeared (Alpert et al., 1988), is consistent
with the proposition that SOM precursor cells contained IN in
addition to the homeoprotein (Fig. 5). Similarly, PP cells also
probably arose from cells coexpressing STF-1 and IN (Fig. 5).
Since only a fraction of embryonic non-β cells expressed the
homeoprotein, STF-1 expression may be incompat-
ible with differentiation of mature α, ∂, PP and
exocrine cell types. Alternatively, STF-1 expression
may be lost because it plays no role in specific gene
transcription in non-β cells. In adults, we found that
STF-1 was expressed by most βcells and by a subset
of α,∂ and PP cells. It is tempting to speculate that
the non-β islet cells that contained STF-1 may
represent immature cells which possess a ‘precursor-
The observation that many exocrine cells
expressed STF-1 protein when they first differen-
tiate supports a common origin for pancreatic
exocrine and endocrine cells from STF+ endoder-
mal precursor cells present in the pancreatic duct.
The origin of acinar and islet cells from a common
stem cell population has been controversial. Since
pancreatic islet cells display a large number of
neural properties (Pearse, 1977; Alpert et al., 1988;
Le Douarin, 1988; Teitelman, 1990) it was
proposed that exocrine tissue derived from
endoderm, while endocrine cells were generated
by the neural crest (Pearse, 1977). However,
previous experiments performed in chick-quail
chimeras (Le Douarin, 1982) and mouse embryos
(Pictet et al., 1976; Teitelman, 1990) and the
present observations, support a common origin for
both endocrine and exocrine pancreatic cells as
indicated in our model (Fig. 5).
Since exocrine cells never expressed islet-
specific cell markers (Teitelman et al., 1987), the
subset of amylase+STF-1+precursor cells probably
diverged towards an exocrine pathway of differen-
tiation at the time they became specified. In
agreement with this possibility, recent studies by
Kruse et al. (1993) showed that an essential tran-
scription control element within exocrine specific
genes, including those of elastase I and amylase 2,
functions as an endocrine-specific control element
in transgenic animal. This element is also contained
within the transcription control region of the insulin
and somatostatin genes, and bears sequence simi-
larity to the STF-1 binding site. It was proposed
that this element may act during pancreatic deter-
mination prior to the divergence of the acinar and
islet cell lineages.
Our present study also documents that most
epithelial cells of the duodenal mucosa expressed STF-1 through-
out life. In contrast, Ohlsson et al. (1993) did not find homeo-
protein+cells in gut. In agreement with our finding, however,
Leonard et al. (1993) and Miller et al. (1994) reported STF-1
mRNA expression in duodenum of adults. In embryos, all cells
lining the lumen of the duodenum were STF-1+. The distribution
of STF-1+cells, however, changed during development due to
modifications in the cytoarchitecture of the gut. During matura-
tion, the duodenal mucosa forms numerous folds (villi), contain-
ing absorptive, secretory and enteroendocrine cells, and these
folds are continuous with invaginations or crypts, which are
formed mostly by stem cells (reviewed in Neutra, 1988). In
adults, STF-1 expression was restricted to epithelial cells lining
the villi but was absent from crypt cells. Since cells of the villi
Days of development
Commitment of STF-1 precursors
Fig. 5. Proposed model of pancreatic cell determination and differentiation. At
around E8.5, expression of STF-1 endows a dorsoventral portion of endodermal
cells with the positional status of caudal foregut-rostral midgut. Under the
influence of unknown factors, dorsal and ventral pancreatic anlagen bud from
this area, while the gut tube itself forms the the duodenum. STF-1 expression
throughout the dorsal, lateral and ventral duodenal epithelium lining the lumen
of the gut is maintained throughout life. The dorsal and ventral pancreatic
primordia arise at E9.5 and E10 respectively and initially all their cells are STF-
1+. The first glucagon and insulin cells are seen at E9.5. At this stage the vast
majority of the β cells are STF-1+, but only a subset of α cells contain the
homeoprotein. It is possible that the STF-1+GLU+cells also contain insulin and
represent the precursors of the β, ∂ and PP cells. In agreement with this
hypothesis, subsets of SOM+and PP+cells express insulin (Alpert et al., 1988)
and STF-1 (present work). STF-1 is also expressed by exocrine cell precursors.
At E14.5, the first amylase+cells arise and many of these cells contain STF-1.
Homeopotein expression outside the islets, however, declines rapidly, so that at
E16.5, STF-1 signal is restricted to few ductal cells while the exocrine tissue is
almost uniformely unstained. These pathway lead to the adult pancreatic
expression pattern, in which STF-1 is found in 90% of β, and in small subsets of
∂ (15%), α (3%) and PP (9%) cells.
18 Download full-text
are constantly being renewed and cell replacement occurs by pro-
liferation, upward migration and differentiation of the crypt stem
cells, our findings indicate that cells of the duodenal epithelium of
adults initiated STF-1 expression after they migrated into the villi
and differentiated. As a consequence of this process of intestinal
cell neogenesis and maturation, the duodenum was lined by STF-
1+cells not only in embryos but also in adults. This suggests that
STF-1 controls the expression of, as yet, unidentified molecules
that are important for duodenal epithelial cell function.
The identity of the signals that initiate STF-1 expression in
the gut and pancreas are unknown. It has been suggested that the
notochord, which in early embryos is in close contact with
midgut endoderm, may play a role in the induction of SFT-1
expression (Ohlsson et al., 1993). This is an attractive hypothe-
sis, since the notochord is a known source of inductive signals
that may instruct surrounding tissues in a region specific manner
during development (Holtfreter and Hamburger, 1955;
Hemmati-Brivanlow et al., 1990). Our findings indicate that the
first inductive signal initiated STF-1 expression in stem cells of
both duodenum and pancreas. It is likely that the these two sets
of precursors diverged at around E8, when commitment of endo-
dermal cells to a pancreatic fate occurs (Wessels and Cohen,
1967). During maturation there were differences in the fate of
STF-1 expression in gut and pancreas. Thus, while epithelial
cells of the duodenum contained the homeoprotein throughout
life, STF-1 expression in pancreas was transient in non-β cells,
persisting only in the insulin-producing cells. This observation
suggest that STF-1 expression is down-regulated in all the cell
types of the pancreatic cell lineage that do not synthesize insulin,
and that other factor/s are required for its maintenance in β cells.
According to this proposition, these molecule/s are present in β
cells but not in ductal, exocrine or endocrine non-β cells.
The signal/s that instruct endodermal cells to follow
alternate pathways of differentiation remain to be determined.
Tissue co-culture experiments have revealed that, following
the initial commitment of the endoderm to a pancreatic fate,
further differentiation of the pancreatic primordium requires
interactions with mesoderm (Wessels and Cohen, 1967). It is
now well established that the mesoderm secretes a variety of
peptides implicated in growth and differentiation. Conceiv-
ably, factors secreted by mesodermal cells may be involved in
a paracrine fashion in different aspects of pancreatic differen-
tiation including the restriction of expression of STF-1 and
insulin to the β cells of the pancreas.
This work was supported by NIH grants, by the Juvenile Diabetes
Foundation International and in part by the Vanderbilt University
Diabetes Research and Training Center Molecular Biology Core Lab-
oratory and The Foundation for Medical Research. The authors thank
M. Ehrlich (New York University) for her helpful comments.
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(Accepted 22 September 1994)
Y. Guz and others