Physiol Rev 85: 1255–1270, 2005;
doi:10.1152/physrev.00025.2004.
Regulation of Pancreatic Beta-Cell Mass
LUC BOUWENS AND ILSE ROOMAN
Cell Differentiation Unit, Vrije Universiteit Brussel-Free University of Brussels, Brussels, Belgium
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I. Introduction
II. Prenatal Development of the Beta-Cell Mass
A. Embryogenesis
B. Fetal growth
III. Postnatal Growth of the Beta-Cell Mass
A. Neonatal growth
B. Compensation for body mass
C. Compensation in pregnancy
D. Regulation by growth factors
IV. Regeneration
A. Destruction of beta cells
B. Surgical injury
C. Transgenic mice and inflammation
V. Stem Cells and Phenotypic Plasticity in the Pancreas
A. Stem cells
B. Transdifferentiation
VI. Diabetes in Humans
VII. Summary
Bouwens, Luc, and Ilse Rooman. Regulation of Pancreatic Beta-Cell Mass. Physiol Rev 85: 1255–1270, 2005;
doi:10.1152/physrev.00025.2004.—Beta-cell mass regulation represents a critical issue for understanding diabetes, a
disease characterized by a near-absolute (type 1) or relative (type 2) deficiency in the number of pancreatic beta
cells. The number of islet beta cells present at birth is mainly generated by the proliferation and differentiation of
pancreatic progenitor cells, a process called neogenesis. Shortly after birth, beta-cell neogenesis stops and a small
proportion of cycling beta cells can still expand the cell number to compensate for increased insulin demands, albeit
at a slow rate. The low capacity for self-replication in the adult is too limited to result in a significant regeneration
following extensive tissue injury. Likewise, chronically increased metabolic demands can lead to beta-cell failure to
compensate. Neogenesis from progenitor cells inside or outside islets represents a more potent mechanism leading
to robust expansion of the beta-cell mass, but it may require external stimuli. For therapeutic purposes, advantage
could be taken from the surprising differentiation plasticity of adult pancreatic cells and possibly also from stem
cells. Recent studies have demonstrated that it is feasible to regenerate and expand the beta-cell mass by the
application of hormones and growth factors like glucagon-like peptide-1, gastrin, epidermal growth factor, and
others. Treatment with these external stimuli can restore a functional beta-cell mass in diabetic animals, but further
studies are required before it can be applied to humans.
I. INTRODUCTION
Blood glucose homeostasis is controlled by the endocrine cells of the pancreas, which are located in the
islets of Langerhans. The islet cells monitor the concentration of glucose in the blood and secrete hormones with
opposite effects. When after a meal the blood glucose
concentration is increasing, the beta cells, which are the
most numerous islet cells, secrete the hormone insulin to
reduce blood glucose. Insulin stimulates the uptake of
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glucose by cells of the body and stimulates the conversion
of glucose to glycogen in the liver. If the glucose level falls
too far, islet alpha cells secrete the hormone glucagon,
which stimulates the breakdown of glycogen to glucose in
the liver and therefore increases blood glucose between
meals. Optimal control of blood glucose levels depends
on delicate changes in insulin production and secretion
by the pancreatic beta cells and on their capacity for a
large increase of secretion after meals, requiring large
stores of insulin (68). Very important is the need for the
0031-9333/05 $18.00 Copyright © 2005 the American Physiological Society
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II. PRENATAL DEVELOPMENT OF THE
BETA-CELL MASS
During the gastrulation process of embryonic development, three germ layers are formed: ectoderm,
endoderm, and mesoderm. Although old text books may
still proclaim that pancreatic endocrine cells are derivatives of the neural crest (ectoderm) and exocrine cells of
the endoderm, it is now generally accepted that they both
derive from the endoderm. The first formal proof for this
was provided by coculturing genetically tagged pancreatic epithelium with unlabeled mesenchyme. Endocrine
and exocrine cells arise from the epithelium (106).
A. Embryogenesis
The embryonic endoderm develops into the primitive
gut from which the digestive system originates. The pancreas derives from two parts of epithelium that bud dorsally and ventrally from the foregut/midgut around embryonic day 9 in mouse and which fuse later on (reviewed
in Ref. 139). There has been much progress in recent
years in our understanding of the major steps leading to
the embryonic development of the endocrine pancreas. In
general, the induction and patterning of organs are regulated by extracellular signals from neighboring cells. This
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leads to the expression of tissue- and cell type-specific
patterns of transcription factors. In particular, the repertoire of transcription factors has been identified that
drives the differentiation of endodermal cells to beta cells
(reviewed in Refs. 38, 39, 66, 98, 165). Pancreatic development occurs in an endodermal region where the transcription factor Pdx1 (pancreas-duodenum homeobox) is
expressed and where the extracellular signaling molecule
sonic hedgehog is repressed. Ptf1a (pancreas transcription factor, a basic helix-loop-helix transcription factor,
or bHLH) is required for specification of ventral pancreas
development, whereas the homeobox transcription factor
Hlxb9 is required for dorsal specification. It has been well
established by genetic tracing experiments that both exocrine and endocrine cells of the pancreas derive from a
pool of endodermal cells that express the transcription
factors Pdx1 (55) and Ptf1a (72). The “protodifferentiated” pancreatic progenitor cells also express the bHLH
transcription factor Hes1 (hairy and enhancer of split
homolog), which is a transcriptional repressor keeping
cells undifferentiated (4). Hes1 is a downstream target of
the Delta-Notch signaling system that inhibits the endocrine cell fate, by a process called lateral inhibition.
Knocking out Hes1 or other mediators of the Notch signaling pathway in transgenic mice leads to accelerated
formation of endocrine cells, but abnormally few cells are
formed as there is insufficient expansion of progenitors
(64, 65). Proliferation of the progenitor cells is stimulated
by fibroblast growth factors (FGFs) (40). Other factors
produced by the surrounding mesenchyme are crucial for
proper pancreatic development (reviewed in Ref. 130).
The homeoprotein Isl1 (Islet-1) is required for development of the mesenchyme of the dorsal pancreatic bud and
also for differentiation of the dorsal pancreatic epithelium
to endocrine cells (1). Between E14 and E17 in embryonic
mice, endocrine cells are derived from a subset of HNF1bexpressing duct cells (88), which transiently express the
transcription factor Ngn-3 (55). Among the Ngn-3 expressing cells, transient Pax4 (paired homeobox gene) expression specifies the beta-cell phenotype, and differentiated
beta cells eventually express high levels of Pdx-1, Nkx6.1,
Nkx2.2, and Pax6 transcription factors (reviewed in Ref.
66). When the first hormone-expressing cells appear, they
stop dividing and are considered “postmitotic” cells (64).
However, later in fetal growth, endocrine cells appear to
be able to reenter the cell cycle (see below).
B. Fetal Growth
The fastest expansion of the beta-cell mass occurs in
late fetal gestation, with an approximate doubling of the
beta-cell population each day starting from the 16th day
postconception in rats (93). In fetal rats, mitotic activity is
restricted to a limited number of beta cells (ⱕ10%) that
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beta-cell mass to be closely regulated by glucose and
hormonal effects on beta-cell replication, size, apoptotic
elimination and, under certain conditions, neogenesis
from progenitor cells. Failure to adapt to changes in body
mass, pregnancy, insulin sensitivity of peripheral tissues,
or tissue injury may lead to the development of chronically elevated blood glucose, or diabetes (68, 92, 126). The
increasing global prevalence of diabetes (164) has stimulated efforts to develop new therapeutic strategies like
beta-cell replacement or regenerative medicine. The existing therapies with exogenous insulin or hypoglycemic
agents for type 1 and type 2 diabetes are unsatisfactory,
since they do not offer a cure and are mostly insufficient
for preventing the secondary complications associated
with diabetes (99). Transplantation of a sufficient number
of pancreatic beta-cells can normalize blood glucose levels and may prevent the devastating complications of
diabetes (73, 143). However, beta-cells from cadaver pancreases are in such short supply that transplants can be
provided only to a limited number of patients. This problem could be overcome by devising ways of generating
more beta-cells from the available donors. An alternative
approach to cell transplantation would be to induce an
increase or regeneration of the endogenous beta-cell
mass in the pancreas by growth factors. Both approaches
require understanding of the mechanisms that regulate
the pancreatic beta-cell mass.
REGULATION OF PANCREATIC BETA-CELL MASS
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mass remain decreased to 32 and 47% of control values,
respectively. Although neonates until 14 days of age display normal basal plasma glucose, the adult diabetic GK
rats exhibit higher basal plasma glucose and become
glucose intolerant. Although fetal development is impaired in the GK pancreas, cultures of explanted pancreatic rudiments did not reveal deficiencies in growth or
differentiation compared with Wistar rats (95). This suggests that extrinsic factors regulating fetal growth, differentiation, and survival of beta cells and their precursors
are deficient in this strain.
III. POSTNATAL GROWTH OF THE
BETA-CELL MASS
It is sometimes believed that we are born with a fixed
number of beta cells that will not increase further during
life. However, the available evidence shows that this is
not true but that the beta-cell population still grows at
least until adolescence, as most other tissues and organs.
Furthermore, most tissues in our body are subjected to
continuous turnover, since the life span of most cell types
is shorter than that of the organism. The average life span
of cells can differ greatly dependent on cell type. In the
case of human beta cells, however, it remains unknown.
A. Neonatal Growth
In neonatal rodents, growth of the beta-cell mass still
occurs but at a reduced rate compared with late fetal
growth (71, 93, 94, 158). Evidence has been reported for
the existence of both beta-cell replication and neogenesis,
the latter from ductlike cells that surround neonatal islets
and exhibit a much higher replicative activity than the
differentiated islet cells (20). These islet precursor cells
disappear after the first week of life, and no further morphological evidence of neogenesis from such precursors
is found thereafter. With a recent transgenic approach, it
was confirmed that the beta-cell mass expands by selfreplication and not by neogenesis after the first week of
life. In knockout mice lacking the cyclin D2 positive regulator of the cell cycle, neonatal beta cells failed to replicate, whereas replicative activity of acinar and ductal
cells remained unaffected (52). Although born with a
normal beta-cell mass, these mice failed to further expand
their beta-cell mass in the neonatal period between 7 and
14 days.
In a rat model of food-restricted mothers it was found
that from birth to 3 mo of age, the offspring showed a
reduced beta-cell mass while beta-cell replication was
similar to or higher than in controls. Apparently, even
increased beta-cell replication is insufficient to restore a
functional beta-cell mass when neogenesis and beta-cell
mass are already affected at birth (49).
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can engage into the cell cycle (58, 146). A daily cell birth
rate of 10% was calculated from the observed beta-cell
mitotic indexes. Thus, in the rat fetus where the beta-cell
number increases at a rate of ⬃100% per day, beta-cell
division can account for not more than 10 –20% of the
total growth (147). The remaining 80% or more has been
attributed to the process of neogenesis from rapidly proliferating “undifferentiated” precursor cells. A much
higher frequency of DNA-synthesizing cells has been observed in the immediate vicinity of the rapidly growing
fetal islets than in the islets themselves. These proliferating islet precursors express the same cytokeratin proteins
that are characteristic for the cells lining exocrine ducts
in the adult pancreas (21). “Transitional” forms of differentiation, represented by cells coexpressing insulin and
duct-type cytokeratin, have been taken as evidence for
the conversion of ductlike precursors into beta cells (21).
Similar conclusions were drawn from studies of human
fetal pancreas, where it was shown that immature beta
cells express cytokeratin-19, an intermediate filament protein characteristic for adult duct cells (23). In human fetal
pancreas, rapid expansion of the beta-cell mass has been
noted from ⬃20 wk (141). The fraction of endocrine cells
undergoing replication in human fetal pancreas is extremely low (23, 70, 111), which is in favor of massive
differentiation from nonendocrine progenitor cells as the
major mechanism of beta-cell mass expansion during this
phase.
It appears that fetal life represents a critical “window” in which the appropriate number of beta cells is set
in place. If this goes wrong due to for example intrauterine growth retardation (IUGR), initially the number of
beta cells may be sufficient but from a certain age on the
beta-cell mass becomes unable to sufficiently compensate
for the increased needs of the body. In a model of IUGR
induced by reducing the blood flow to the fetus, no significant differences in beta-cell mass were found at 1 and
7 wk of age, but in 15-wk-old IUGR rats, the relative
beta-cell mass was 50% that of controls, and by 26 wk of
age, beta-cell mass was less than one-third that of controls (137). These animals developed glucose intolerance
as seen in type 2 diabetes. Fetal malnutrition resulting
from maternal food restriction in rats leads to a reduced
beta-cell mass in the offspring (48). Beta-cell replication
was not affected in this model, indicating that a deficient
neogenesis of beta cells during fetal life caused the effects. Perinatal malnutrition also leads to the development of type 2 diabetes later in life, apparently by the
inability of the beta-cell mass to adapt to additional demands placed by ageing (50). Similar observations were
made in the Goto-Kakisaki (GK) rat, which is a genetic
model of type 2 diabetes obtained by selective inbreeding
of mildly glucose-intolerant Wistar rats (97). GK fetuses
show a reduction of the beta-cell mass to 23% of control
values. The adult pancreatic insulin content and beta-cell
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A transient “wave” of beta-cell apoptosis has been
reported around the time of weaning and is associated
with a transiently decreased growth rate of islets in rodents (129), pigs (13), and humans (70). It may be that in
this period, important changes take place at the level of
beta-cell maturation.
There is no direct evidence of beta-cell neogenesis
from stem/progenitor cells in the undisturbed adult rodent pancreas (42).
B. Compensation for Body Mass
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Whereas in some rapidly renewing gastrointestinal
tissues like the gut mucosa, there is a constant replacement of cells that are derived from a stem/progenitor
pool, this has not been shown to be the case in the
pancreas. However, the beta-cell population remains a
dynamic mass, probably throughout life. After the growth
of the beta-cell mass has considerably slowed down by
the time of weaning in rats, it continues to expand at a
relatively slow pace during adult life (96, 159). Islet mass
appears to increase mainly by an increased size of individual islets rather than by the formation of new islets
(138). Between 1 and 7 mo of age in rats, the beta-cell
mass increases six- to sevenfold in females and males,
respectively (94). Comparable figures were obtained by
other investigators using tissue morphometry (96, 159).
On average, the postnatal beta-cell population grows at a
rate between 2 and 3% per day in male rats (42, 96, 159).
Growth is significantly less in females, which obtain a
lower body weight than males (159). After 7 mo of age,
growth of the beta-cell population slows down considerably but still continues until 20 mo, which is old age for
rats (96). A linear correlation was found between beta-cell
mass and body weight (96), and such a relationship was
also found in larger mammals like pigs (13). However,
with increasing age, the beta cell number per kilogram
body weight decreases significantly in rats (159). A critical number seems to be ⬃10 million beta cells per kilogram body weight. When beta cell number decreases
below this value, e.g., in streptozotocin-treated rats where
part of the beta cells have been destroyed, animals become glucose intolerant (159).
Excessive increase in body weight leads to obesity.
Obesity is correlated with insulin resistance and is associated with a compensatory physiological response at the
level of beta-cell mass increase. In nondiabetic animal
models of obesity, for example, the Zucker fa/fa rat, beta
cell mass is increased fourfold compared with lean controls (109). Animals eventually become diabetic when
they age and become more obese and glucose intolerant.
There is also evidence for an increased beta-cell mass in
nondiabetic obese humans (74). In hyperglycemic obese
ob/ob mice, insulin resistance progressively develops up
to 6 mo of age and is accompanied by marked islet
hyperplasia in response to the sustained hyperglycemia
(14, 151). Beta-cell hyperplasia is also found in db/db
mutant mice (46, 162). The sand rat, Psammomys obesus,
is an animal that is adapted to a low-calorie diet and
shows insulin resistance. When the animal is fed a highcalorie diet, it is unable to compensate for the hyperglycemia. A transient increase in beta-cell replication is insufficient to cope with the observed increase in apoptotic
cell death, and hence, the animals become diabetic (36).
Histologically, dividing beta cells can be recognized
in adult pancreas, but they are very scarce. Using immunohistochemical demonstration of bromodeoxyuridine incorporation (a thymidine analog), the quantitative importance of beta-cell replication can be evaluated. Beta-cell
replication is progressively reduced during postnatal life,
with ⬍0.25% of beta cells being in S-phase of the cell cycle
after 3 mo of age (96, 159). Yet, when the beta-cell birth
rate is calculated based on the observed frequency of cells
in S-phase and an S-phase duration of 6.4 h (146), it was
concluded that beta-cell replication can theoretically account for the observed net increase in cell number (3–20
wk of age) (159). A recent genetic tracing experiment in
mice has demonstrated that this is indeed the case (37). In
this study, total beta cell numbers were counted from
enzymatically dissociated pancreata. A 6.5-fold increase
was noted between 3 and 12 mo of age, which corresponds to an average daily growth rate in the range of
2–3%, as in rats. With the use of a tamoxifen-induced
Cre/Lox system, a pulse of reporter gene expression in
insulin-transcribing cells could be followed over time.
This allowed the discrimination between beta cells generated from preexisting beta cells and beta cells derived
from another source of cells which did not previously
express insulin. The results demonstrated that new beta
cells in the adult animals were formed exclusively by
replication of preexisting beta cells (37). Thus these findings confirm the conclusion drawn from cytokinetic studies, i.e., that self-replication is sufficient for the slow
expansion of the beta-cell mass as a compensation for
normal growth in rats and mice.
The study of Dor et al. (37) also suggested that no new
islets are formed during adult life but rather that islets
present in old mice are all derived from islets that were
present at ⬃2 mo of age. It is possible, however, that growing islets can generate smaller ones by fission as indicated
by the study of Seymour et al. (134). In this study, the cell
composition of pancreatic islets was analyzed in mosaic
mice with a lacZ insertion on the X-chromosome. In female
heterozygous mice, due to random silencing of one of the
two X-chromosomes during development, tissues become a
mosaic of cells, some of which express the lacZ-reporter
(␤-galactosidase) and others not. The authors of this study
noted that islets are initially of mixed composition and
hence must be formed by aggregation of several cells. Later
REGULATION OF PANCREATIC BETA-CELL MASS
C. Compensation in Pregnancy
Besides adaptation to increased body weight and
thus increased demand for insulin, the beta cell mass can
also compensate for the increased demand during pregnancy. Failure to compensate is thought to lead to maternal or gestational diabetes. With an incidence of 3–5%, this
represents one of the top health concerns related to pregnancy. Although gestational diabetes usually goes away
after delivery, mothers are at risk for developing type 2
diabetes later in life. At the end of pregnancy in rats, the
beta-cell mass is ⬃2.5-fold increased compared with nonpregnant females (10, 156). This results from both an
increased beta cell number and cellular hypertrophy. Islet
enlargement and beta-cell hyperplasia have also been observed in autopsies from pregnant humans (155). During
experimental diabetes induced by streptozotocin in rats,
however, the endocrine pancreas has an impaired capacity to compensate during pregnancy (156). After pregnancy, a rapid decrease of the beta-cell mass occurs in
postpartum rats (89). This is accompanied by decreased
beta-cell replication and beta-cell size, and by an increased frequency of apoptosis (128).
These studies demonstrate the capability of beta
cells to up- and downregulate their mass using the mechanisms of beta-cell replication and apoptotic cell death,
and changes in beta cell size, to achieve homeostasis in
conditions of changing insulin demand. It is not clear
whether neogenesis from precursor cells is contributing
to beta-cell mass increase during pregnancy.
D. Regulation by Growth Factors
Which molecules regulate the beta cell mass? Insulin,
several other hormones, and glucose have been reported
to play an important role.
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1. Insulin
Transgenic mice with a beta cell-specific knockout of
the insulin receptor exhibit a decreased beta cell mass in
adults and develop diabetes (102). Insulin resistance,
leading to hyperinsulinemia, stimulates an increase in
beta-cell mass. This was shown in mice that were made
double heterozygous for null alleles in the insulin receptor
and insulin receptor substrate-1 genes (IR/IRS mice). The
animals showed an expected 50% reduction in expression
of these two proteins and exhibited severe insulin resistance with 5- to 50-fold elevated plasma insulin levels. A
compensatory beta-cell hyperplasia was observed with a
significant increase in beta-cell mass (27). In insulin-resistant IR/IRS mice that were crossed with Pdx1-heterozygous mice, there was a high number of apoptotic beta
cells, and the compensatory beta-cell growth response
was completely abrogated (75). These data indicate that
the Pdx1 transcription factor is a crucial regulator of the
adaptative response of the beta-cell mass in insulin-resistant mice. This homeodomain protein seems to be important for beta-cell proliferation and apoptosis (75) as well
as for neogenesis (135).
An increased beta-cell number was also observed in
acutely euglycemic-hyperinsulinemic infused rats (103).
On the other hand, hyperinsulinemia caused by ectopic
transplantation of a rat insulinoma has been shown to
lead to a significant reduction of the beta-cell mass in the
pancreas (11). This reduction in beta-cell mass was nonimmune-mediated and resulted from apoptosis of beta
cells. These observations indicate that insulin can control
beta cell population dynamics, although the mechanism is
still unclear.
2. Glucose
Glucose itself has an effect on beta-cell population
dynamics. With fetal islets, it is possible to increase the
proportion of cycling beta cells in vitro by increasing the
glucose concentration of the culture medium (146). Glucose infusion in rats during a period of only 24 h increases
the beta cell number by ⬃50% (8). Seven days after stopping glucose infusion, the beta-cell mass returned to basal
values, and this was accompanied by increased beta-cell
apoptosis. By combining simultaneous infusion of either
glucose and/or insulin or glucose and diazoxide, hyperglycemic-hyperinsulinemic rats, hyperglycemic-euinsulinemic rats, and euglycemic-hyperinsulinemic rats could
be studied (103). In all groups, beta cell number was
significantly increased, namely, with 50 –70% compared
with controls, within 48 h. The rate of beta-cell proliferation decreased, which indicates that neogenesis from precursor cells was responsible for the increased beta-cell
mass (103). A longer glucose infusion time was reported
to result in increased beta-cell replication and hypertrophy besides neogenesis, and leads to sustained effects on
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on in postnatal life, the heterogeneity of islets diminished.
This suggests that following cell turnover, older islets contain descendants of only a small number of the preexisting
islet cells that have replicated.
Species-related differences cannot be excluded, and it
remains to be found whether the same applies to human
pancreas. Differences do exist between human and rodent
pancreas, for example, with respect to cellular composition,
tissue organization, and beta-cell replicative activity. For
instance, in human pancreas there exists a population of
extra-islet beta cells that occur scattered over the exocrine
tissue and represent 15% of all beta cells (24). Such cells are
quite rare in normal rodent pancreas. Also, replicative activity of human beta cells is lower compared with rodents and
decreases with age (154). In mice, an age-dependent decrease of beta-cell replicative capacity has been noted, and
this capacity depends on the genetic background (148).
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vitro. These include insulin-like growth factors (IGF-I and
-II), platelet-derived growth factor (PDGF), epidermal
growth factor (EGF), and others (reviewed in Refs. 47,
60). IGF-II exerts an islet protective role in late fetal
gestation and immediately after birth and thereby is important in defining the adult beta-cell mass (61, 108). In
general however, unless transgenic approaches are used
to induce overexpression of these factors in islets, the
mitogenic effect on beta-cell mass is rather modest, and
their role under normal physiological conditions is not
clear. When these growth factors are added to primary
cultured adult beta cells, no significant expansion of cell
number has so far been reported. An exception may be
the stimulation of human islet cell expansion in vitro by
hepatocyte growth factor (HGF) (57). However, it was
subsequently reported that HGF stimulates replication of
contaminating ductal cells but not of the insulin-positive
beta cells (81). Ductal cells are known to contaminate
human islet preparations (73, 143).
3. Placental hormones
During pregnancy, placental hormones, especially
placental lactogens, are mainly responsible for the
changes in beta-cell mass (reviewed in Refs. 100, 140).
These hormones stimulate beta-cell proliferation in isolated islets, and beta-cell mass is reduced 26 – 42% in
receptor-deficient mice (44).
4. Glucagon-like peptide-1
The gastrointestinal incretin hormone glucagon-like
peptide-1 (Glp-1) has been repeatedly reported to affect
beta-cell function, replication, apoptosis, and neogenesis
(reviewed in Refs. 25, 28). In obese hyperglycemic ob/ob
mice, a Glp-1 analog ameliorates glycemia and increases
beta-cell mass and beta-cell replication (116). Glp-1 delays
the onset of diabetes in db/db mice (162), and in diabetic
mice it lowers glycemia and increases beta-cell mass
through neogenesis from Pdx1-expressing ductal precursor cells (142). Similar results were obtained with glucose-intolerant aged Wistar rats (107). With the use of rat
and human duct cell lines in vitro, it was found that Glp-1
can induce differentiation to a functional beta-cell phenotype in cells expressing Pdx1 (63, 170). Glp-1 therefore
seems to be a differentiation factor that can play an
important role in neogenesis, besides having stimulatory
effects on already differentiated beta cells via increased
replication and decreased apoptosis (84). Surprisingly,
pancreatic beta-cell mass is normal in Glp-1-receptor
knockout mice (85), and it can normally adapt to obesity
(132). It is possible that other gastrointestinal/incretin
hormones can compensate when Glp-1 is knocked out.
5. Other growth factors
Several growth factors have been reported to exert a
stimulatory effect on beta-cell replication in vivo or in
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6. Cell cycle regulators
It seems that mature beta cells are highly refractory
to mitotic stimulation. Cell cycling is regulated intracellularly by protooncoproteins like cyclins and cyclin-dependent kinases (cdks), which are positive regulators,
and by tumor suppressors or cdk inhibitors as negative
regulators. Little is known on the molecular mechanisms
that regulate, and mainly suppress, beta-cell replication.
Using cyclin D2 ⫺/⫺ mice, it was shown that this particular D-cyclin is dispensable during the embryonic-fetal
development of the beta-cell mass but that it is indispensable for neonatal replication of beta cells (52). Neonatal
beta cells apparently fail to efficiently upregulate other
D-cyclins, in contrast to acinar and ductal cells which
retained replicative activity in these mice. Thus the cyclin
D2 positive regulator of the cell cycle plays an important
role in the control of beta-cell replication. Growth hormone and prolactin increase the expression of cyclin D2
via the STAT5 pathway in beta cells (45). Overexpression
of cdk-4, or certain mutated form of this kinase, leads to
insensitivity to cell cycle suppressors of the INK4 type.
This results in increased beta-cell proliferation in vitro
and in transgenic mice, apparently without loss of betacell function (34, 90, 114). In contrast, forcing beta cells to
reenter the cell cycle by overexpression of the protooncogene c-Myc leads to beta-cell apoptosis (79). Thus the
growth-promoting and oncogenic potential of c-Myc in
beta cells is normally strongly counterbalanced by apoptosis. However, upon transgenic coexpression of c-Myc
and the antiapoptotic factor Bcl-xl, the beta cells in the
transgenic mice proliferate and form aggressive tumors
(105). A previous study had shown that overexpression of
Bcl-xl in beta cells prevents their cell death but at the
same time leads to mitochondrial defects leading to im-
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beta-cell mass even after glucose infusion is stopped (16,
152). Glucose has been shown to promote beta-cell survival by suppressing a constitutive apoptotic program in
vitro (62). However, high glucose concentrations have
also been reported to stimulate beta-cell apoptosis in vitro
and in the sand rat Psammomys obesus, an animal model
of type 2 diabetes which becomes hyperglycemic upon
feeding a high-calorie diet (36). This “glucotoxicity” may
also involve cytokines like interleukin (IL)-1, which is secreted by human islets in the presence of high glucose and
leads to beta-cell apoptosis (87). Chronic hyperglycemia
may thus be detrimental to beta cells and plays a role in the
development of diabetes by decreasing the beta-cell mass
(reviewed in Ref. 69). Increased beta-cell apoptosis has indeed been observed in humans with type 2 diabetes (30).
Several studies based on tissue morphometry have reported
a diminished beta cell mass in type 2 diabetic patients compared with control subjects (30, 33, 74, 125, 168).
REGULATION OF PANCREATIC BETA-CELL MASS
IV. REGENERATION
In several organs of our body, tissue parts which
have been damaged can be regenerated. The regenerative
capacity of, for example, liver tissue is well-known.
It appears that in the absence of major external stimuli, the beta-cell population has only a very limited potential for regeneration. This is probably due to the limited
replication capacity of beta cells and to the fact that
neogenesis from precursor cells is not readily reactivated.
Yet, under certain conditions where major external stimuli are applied, there can be a quite vigorous regenerative
expansion of the beta-cell mass. Such regenerative
growth may result from activation of otherwise quiescent
precursor/progenitor or stem cells.
Regeneration of the endocrine pancreas has been
studied in different experimental models that differ in the
extent and selectivity of the tissue injury that is inflicted.
The two major types of injury are caused by toxic drugs
and surgery, respectively.
A. Destruction of Beta Cells
Selective destruction of beta cells can be obtained by
injecting streptozotocin or alloxan. Streptozotocin is a
DNA-alkylating agent and alloxan a generator of oxygen
free radicals, both causing extensive DNA damage (149).
Their selectivity is thought to be due to a better uptake by
beta cells, although at higher doses also liver and kidney
cells become affected. Experimental models can be quite
different depending on the dose and the route of administration; for example, intraperitoneal injection has less
severe effects than intravenous injection at a given dose.
In rats, intravenous administration of 100 mg/kg
streptozotocin on the day of birth reduces the total betacell mass by ⬃90% in 48 h. Twenty days later, ⬍40% of the
normal beta-cell mass is restored (158). Up to a certain
body weight the animals can maintain normoglycemia,
but at the age of ⬃6 wk they become glucose intolerant.
Physiol Rev • VOL
This neonatal-streptozotocin model is considered as an
experimental model of type 2 diabetes wherein beta-cell
mass compensation is deficient (15, 32). In this model,
replicative activity of beta cells is increased, but apparently insufficient to regenerate a functional mass (15).
The capacity to regenerate following a toxic insult
like streptozotocin rapidly declines during the first 5 days
of life in rats (159). This indicates that beta-cell regeneration beyond the critical perinatal time window is inefficient, possibly because there is no more neogenesis operating from precursor cells. Interestingly, when the hormone Glp-1 is administered to streptozotocin-treated
newborn rats, beta-cell neogenesis is stimulated, and this
results in improved glucose homeostasis persisting at
adult age (153).
When subtotal beta-cell destruction is accomplished
in adult rodents, there is no “spontaneous” regeneration,
and it is well known that the animals remain diabetic or
die (2, 83 121). The exception may be the selectively
perfused rat where alloxan is perfused in part of the
pancreas while another part is clamped off and spared
from the toxic effects (157). Preexisting beta cells from
the spared part proliferate, whereas neogenesis of beta
cells from duct cells leads to regeneration in the perfused
part.
After near-total destruction of the beta-cell mass,
application of a major external stimulus has been found to
induce regeneration. In a recently described experimental
model of beta-cell destruction by alloxan, the combination of two factors, gastrin and EGF, were found to restore glycemic control in mice (121). In this model of
beta-cell regeneration, a beta-cell growth rate of ⬎30% per
day was observed, leading to a beta-cell population doubling time of only 3 days. Although there was no complete
regeneration of the original beta-cell number after subtotal destruction, regenerative growth induced by the gastrin and EGF treatment led to the restoration of 30 – 40%
of the normal beta-cell mass within 7 days. The treatment
had no effect on beta-cell replication, cell size, or apoptosis, and therefore, the regenerative effect could be attributed to neogenesis from precursor cells. Indeed, a pulsechase labeling with the thymidine analog bromodeoxyuridine confirmed an influx of labeled cells from a replicating
insulin-negative pool into the regenerating islets. The proliferating precursor cells express ductal-type cytokeratin
markers. An “intermediate phenotype” characterized by
cells coexpressing the beta-cell marker insulin and the
ductal cytokeratin marker was taken as evidence for a
transition from exocrine duct cells to endocrine beta cells
(121). Gastrin hormone is expressed in fetal and in regenerating pancreas, and its high-affinity receptor cholecystokinin (CCK)-B is not expressed by beta cells themselves
(118, 124, 161). Gastrin can be considered an important
regulator of beta-cell neogenesis (see also sect. IVB).
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paired insulin secretion and hence hyperglycemia (171).
Interestingly, when the beta cell transcription factor
Pax-4 is overexpressed by adenoviral transduction in
vitro, an increase in beta cell proliferation was observed
with a concomitant increase in Bcl-xl and c-Myc. Thus
induction of Pax-4 activates c-myc and beta-cell proliferation, and at the same time activates Bcl-xl thereby suppressing beta-cell apoptosis (26). Bcl-xl activity, however,
alters mitochondrial calcium levels and ATP production,
which explains impaired insulin secretion.
Thus cell cycle regulators might represent therapeutic targets to manipulate the size of the beta-cell mass,
provided that the apoptotic program can be controlled
and that functional defects can be circumvented.
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LUC BOUWENS AND ILSE ROOMAN
Betacellulin, a member of the EGF family, has also
been reported to promote beta-cell neogenesis and to
improve glucose metabolism in adult animals after betacell destruction (83, 167).
In NOD mice, beta cells are destroyed by a spontaneous autoimmune reaction leading to a type 1-like diabetes condition. Immune suppression in combination
with Glp-1 (analog) treatment could restore normoglycemia and improve islet histology (101). The mechanism of
this remission was not clear, however, and it may have
involved regeneration as well as increased survival of beta
cells. It should be pointed out that Glp-1 has been shown
to protect beta cells from apoptosis (84).
Partial pancreatectomy represents another model of
tissue injury wherein regeneration has been studied in
rodents. In contrast to the robust liver regeneration following partial hepatectomy, surgical removal of part of
the pancreas is followed only by a limited regenerative
growth, and there is never a complete restoration of the
original pancreatic volume. Furthermore, the regenerative response is proportional to the amount of pancreas
removed. If half the pancreatic volume is left after
hemipancreatectomy, this residual pancreas will grow
only 20%. In the case of one-third of the volume being left
after two-thirds pancreatectomy, the residual pancreas
grows only 30%. When only one-tenth of the volume is left
after subtotal pancreatectomy, the residual pancreatic
volume grows 80% (104). Regenerative growth after partial pancreatectomy is thus not impressive and never
complete. Except maybe for subtotal (90 –95%) pancreatectomy, it cannot be considered as an important growth
stimulus. The same is true for the endocrine part of the
pancreas, with only a 30% expansion of the residual betacell mass being observed after two-thirds pancreatectomy
in mice, and this over a period of 5 wk (35). The beta-cell
growth rate following two-thirds pancreatectomy is thus
rather modest. Dor et al. (37) using a transgenic cell
labeling approach showed that after two-thirds pancreatectomy there is no evidence for the formation of new
beta cells by differentiation from insulin-negative progenitor or stem cells. This conclusion seems at stake with the
observations of others who reported neogenesis of beta
cells from proliferating ducts after subtotal pancreatectomy (17). This apparent discrepancy may be explained
by a difference in the extent of tissue damage and growthstimulating capacity between the two models that were
used, i.e., between 70 and 90% pancreatectomy. Another
important difference is that a hyperglycemic condition is
induced by 90% but not by 70% pancreatectomy. Thus it
remains possible that in the latter case only beta-cell
replication is activated, whereas in the case of 90% panPhysiol Rev • VOL
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B. Surgical Injury
createctomy, a more vigorous beta-cell growth is obtained
by neogenesis in addition to beta-cell replication. Unfortunately, these reports (17, 37) do not mention the betacell number before and after the pancreatectomy insult so
that it is impossible to compare the respective beta-cell
growth rates that were induced by the different treatments. However, in other studies of 90% pancreatectomized rats, a doubling of the remnant beta-cell mass was
obtained within 1 wk postsurgery (110), whereas only a
30 – 40% increase over 4-wk time was observed following
60% pancreatectomy (80). These data confirm that subtotal pancreatectomy is a much stronger growth stimulus
for the beta-cell population than two-thirds pancreatectomy. In 90% pancreatectomized rats, depletion of beta
cells by streptozotocin treatment before surgery does not
impair the regenerative capacity of the pancreas and demonstrates that partial beta-cell regeneration can be accomplished by neogenesis rather than replication of beta
cells (43).
Others have used 90 –95% pancreatectomy as a rodent model of type 2 diabetes and showed that daily
administration of the Glp-1 analog exendin-4 during 10
days postpancreatectomy attenuates the development of
diabetes. It was reported that exendin-4 stimulates the
regeneration of the pancreas and expansion of the betacell mass by the processes of neogenesis and replication
of beta cells (166). After 70% partial pancreatectomy,
Glp-1 was also found to act as a potent stimulator of
beta-cell regeneration (35). Glp-1 receptor knockout mice
showed worse glucose intolerance after partial pancreatectomy compared with wild-type mice, and this correlated with a significant defect in beta-cell mass regeneration (35). Another regeneration-promoting factor after
subtotal pancreatectomy is betacellulin, a growth factor
that belongs to the family of EGFs (82). Interestingly,
Glp-1 has been shown to transactivate the EGF receptor
and thereby stimulate beta-cell replication (29). On the
other hand, in the GK-rat genetic model which spontaneously develops “type 2” diabetes, regeneration of the betacell mass following 90% pancreatectomy is impaired due
to the decreased capacity for both replication and neogenesis (110).
An experimental model that leads to robust hyperplasia of beta cells is partial duct ligation. This consists in
the closure, or ligation, with surgical thread of part of the
main duct of the pancreas. As a consequence of this
obstruction, exocrine secretory products will leak into
the interstitial space and lead to tissue damage and inflammation. The part of the pancreas that lies downstream of the ligation (⬃50% of the pancreatic volume) is
not affected histologically and continues to function normally. During the first week postligation, a pronounced
beta-cell hyperplasia occurs in the ligated part, although
the animals remain normoglycemic (160). Beta-cell replication is only slightly elevated, which is strongly suggest-
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REGULATION OF PANCREATIC BETA-CELL MASS
Transgenic mice in which beta cells express insulin-like
growth factor (IGF)-I can recover from immune attack of
the islets following multiple low doses of streptozotocin
(51). IGF-I is thought to stimulate beta-cell proliferation and
neogenesis from ducts and has an antiapoptotic effect.
Transgenic mice that overexpress the inflammatory
cytokine interferon-␥ in pancreatic islets show features
indicative of rapid beta-cell renewal by the process of
neogenesis from ductal complexes (54). Beta-cell hyperplasia was also observed in transgenic mice with pancreatic overexpression of IL-6, another important inflammatory cytokine (31). These studies suggest that inflammatory cytokines may induce or regulate beta-cell
neogenesis. It can be assumed that these cytokines are
also increased in conditions like surgical or toxin-mediated tissue injury, where inflammation is obviously
present. Indeed, it has been reported that in the absence
of inflammation, subtotal pancreatectomy does not lead
to beta-cell regeneration (76). Recently, it was found that
the IL-6-related cytokine leukemia inhibitory factor (LIF)
acts synergistically with EGF to induce beta-cell neogenesis in vitro (5).
In summary, whereas self-replication of beta cells
leads to a slow expansion in physiological conditions,
neogenesis from progenitor cells can lead to vigorous
expansion and partial regeneration of the beta-cell mass
in short time (see also Table 1). This process can be
considerably enhanced by exogenous factors like the hormones gastrin and Glp-1, and possibly by inflammatory
mediators or cytokines. The precise molecular regulation
of neogenesis, however, remains to be unravelled.
V. STEM CELLS AND PHENOTYPIC PLASTICITY
IN THE PANCREAS
According to a growing number of reports, beta cells
or at least beta cell-like cells, can be generated ex vivo
1. List of some experimental models of beta-cell
regeneration wherein the increase in beta cell number
or mass was quantified
TABLE
C. Transgenic Mice and Inflammation
Interesting information concerning regenerative expansion of beta cells has been obtained from various
transgenic mouse models. TGF-␣ overexpressing transgenic mice develop metaplastic acinoductal complexes in
the pancreas and show signs of tissue inflammation (67,
127). When these mice are crossed with transgenic mice
overexpressing gastrin hormone under control of the insulin promoter, the double transgenic TGF-␣/gastrin mice
were found to contain significantly more beta cells (about
twice the number in control mice) (163). The role of
gastrin and EGF family members in the regulation of beta
cell neogenesis during regeneration has been confirmed
in other studies (119 –121).
Physiol Rev • VOL
Experimental Model
Mouse 70% pancreatectomy
Rat 60–70% pancreatectomy
Rat 90% pancreatectomy
Rat duct ligation
Rat duct ligation ⫹ gastrin
Mouse alloxan ⫹ gastrin/EGF
Beta-Cell Numerical
Increase/Time
Reference
Nos.
30% in 35 days
40% in 28 days
100% in 7 days
200% in 14 days
80% in 7 days
100% in 3 days
100% in 3 days
35
80
110
160
119
121
The regenerative growth rate of the beta-cell mass differs considerably between these models. Note that beta-cell replication was increased to some extent in the first four models but not in the latter two.
The highest growth rates are seen in the latter four experimental models,
wherein beta-cell neogenesis was shown to be active. EGF, epidermal
growth factor.
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ing that the increased beta-cell number results from neogenesis. This is also indirectly indicated by the observation of intermediate phenotypes between ductal and
endocrine cells (160) or exocrine acinar and endocrine
cells (9, 77). Whereas part of the affected acinar exocrine
cells are eliminated by apoptosis, the remaining ones
transdifferentiate into ductlike cells and thereby contribute to the formation of metaplastic ductal complexes.
These cells start to express transforming growth factor-␣
(TGF-␣), gastrin (161), and the high-affinity gastrin/CCK-B
receptor, which is not expressed on beta cells (118). Cells
in these ductal complexes proliferate and can differentiate into endocrine cells (160). The observed increase in
beta-cell number is 80% in only 7 days time. In this model
of stimulated expansion of the beta-cell mass, a further
increase in the beta-cell growth rate is achieved by infusing the rats with gastrin hormone (119). This leads to a
beta-cell number doubling in only 3 days time, or a growth
rate which is ⬃10 times faster than in control conditions.
Obviously, this robust hyperplasia cannot be accounted
for by beta cell self-replication, as the fraction of beta
cells being engaged in the cell cycle remains very low
(119). Administration of a selective CCK-B gastrin receptor antagonist completely prevents the duct ligation-induced increase in beta-cell mass (120). This means that
gastrin is a crucial factor in the regulation of beta-cell
neogenesis during regeneration.
A protein extract from partially obstructed pancreas
(by wrapping the organ with cellophane) has been reported to induce islet neogenesis from ducts, and it could
reverse streptozotocin-induced diabetes when administered to hamsters (122, 123). From this regenerating pancreas, a novel pancreatic gene called INGAP has been
identified whose protein product is capable of initiating
duct cell proliferation (112). INGAP belongs to the Reg
family of genes (150). However, no direct evidence has
been given to indicate that it can directly induce beta-cell
neogenesis.
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LUC BOUWENS AND ILSE ROOMAN
from either undifferentiated stem cells or from transdifferentiating mature pancreatic cells (Fig. 1). Even if the
physiological relevance of these findings remains unclear,
it is clear that they may find applications in cell therapy by
providing a source of cells to restore the beta-cell mass.
A. Stem Cells
Stem cells are defined as clonigenic, self-renewing,
multipotent cells and are known to reside in a number of
organs such as bone marrow, intestine, skin, brain, and
others. In the liver, oval cells are resident stem cells that
participate to tissue regeneration under certain conditions (3, 41). However, it is not yet clear whether such
cells also reside in the postnatal pancreas. It has been
claimed that exocrine ducts (18) and islets (56) would
harbor islet progenitor cells. Although this possibility has
already been raised for a long time (reviewed in Ref. 22),
only recently a few reports have given support to the
existence of true progenitor or stem cells in the adult
pancreas. First, it was reported that cells can be isolated
from normal rodent pancreas (ducts?), which can be subcultured over long periods of time. In differentiation conditions in vitro, these self-renewing cells generated insulin-producing cells that could reverse diabetes upon transplantation (113). Another report claimed that selfrenewing cells expressing the neural stem cell marker
nestin were able to generate cells with phenotypic characteristics of pancreatic endocrine and exocrine cells,
and of hepatocytes (172). More recently, clonigenic cells
Physiol Rev • VOL
were isolated from rodent pancreas that were able to give
rise to cells with neural or pancreatic features, including
expression and secretion of insulin, but these had only
limited self-renewal capacity (133). Others found clonigenic cells that were able to generate insulin-expressing
cells and that could be enriched by sorting for the c-met
receptor (receptor for HGF) (145). It is not clear whether
these progenitor or stem cells actually take part in renewal, expansion, or regeneration of beta cells or other
pancreatic cells in vivo. It is hoped, however, that if such
cells also reside in human pancreas, they could be used
for ex vivo generation of beta cells. Besides stem cells
that reside in the pancreas, extrapancreatic or embryonic
stem cells (ES cells) are candidates for ex vivo beta-cell
generation. There have been reports on the differentiation
of beta cell-like cells from mouse and human ES cells, rat
hepatic oval cells, mouse intestinal epithelium, and
mouse bone marrow (reviewed in Refs. 12, 144).
B. Transdifferentiation
Transdifferentiation has been originally defined as a
phenotypic switch from one differentiated state to another. In simpler vertebrates like the newt for example,
regeneration of major body parts like the limbs is accomplished by differentiated cells rather than by stem cells.
The differentiated cells at the site of tissue injury first
dedifferentiate whereby they reacquire embryonic plasticity, then proliferate and finally redifferentiate. This system
differs from stem cell-mediated regeneration in that the
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FIG. 1. Overview of the different hypotheses
concerning possible sources for beta cell neogenesis (see details in sect. V). Two major potential
sources of new beta cells can be discerned postnatally in experimental animals. One is represented by putative stem cells, which remain
poorly identified at present, and the other can
involve a range of proposed transdifferentiations
from previously mature cells like acinar, ductal,
mesenchymal, and islet cells. Solid lines represent possible transdifferentiations. Dotted line
represents stem cell derivation.
REGULATION OF PANCREATIC BETA-CELL MASS
VI. DIABETES IN HUMANS
A lot of information on beta-cell biology has been
obtained in animal studies. However, we are mainly interPhysiol Rev • VOL
ested in human beta cells and human diabetes. There is no
doubt that a critical reduction in beta-cell mass is responsible for the appearance of type 1 diabetes symptoms.
Hence, transplantation studies have demonstrated that
the diabetic metabolic defects can be restored in type 1
patients by restoring a functional beta-cell mass (73, 143).
Animal data (see sect. IIIB) strongly suggest that the impairment of insulin secretion in type 2 diabetes is also
partly related to reduction of beta-cell mass, at least
relative to prevailing insulin demand. In fact, this idea is
relatively new, and it is actually still a matter of debate
whether humans with type 2 diabetes have a reduced or
suboptimal beta-cell mass. In humans, such a defect in the
beta-cell mass may originate from genetic predisposition,
but the situation is likely worsened by environmental
factors such as hyperglycemic glucotoxicity and hyperlipidemic lipotoxicity. An increase in beta-cell apoptosis
may prevent the adaptive increase in beta-cell mass in
type 2 diabetes (30). For obvious reasons, it is difficult to
collect data on the size of the beta-cell mass in humans,
and there is probably a large variation in the beta-cell
volume density among individuals. Nevertheless, several
studies have clearly documented that the beta-cell mass in
type 2 diabetic patients is reduced at least 20% compared
with control subjects (30, 33, 74, 125, 168). We have also
to take into account that in insulin-resistant subjects,
there is need for a larger beta-cell mass than in normal
control subjects.
VII. SUMMARY
Based on the available evidence that we have discussed above, a general concept on beta-cell mass regulation can be proposed as follows. The number of beta
cells in the endocrine pancreas seems to be determined
during a critical window phase that is situated somewhere
in the last quarter of fetal gestation and in the first few
days after birth, at least in rodents. This critical beta-cell
population size is developed mainly by the process of
neogenesis. Neogenesis, which can be defined as the formation of new beta cells by the differentiation of previously insulin-negative precursor cells, is a process that
normally stops shortly after birth. Postnatal adaptation of
the beta-cell population size in response to changing metabolic demands is accomplished by an interplay of betacell replication and apoptosis. New beta cells are thus
added to the original number the individual is born with,
by replication of preexisting cells. It appears that if the
perinatal beta-cell number is suboptimal, for example, as
a result of deficient fetal nutrition, postnatal beta-cell
replication does not suffice to compensate later in life.
Nutritional overload, obesity, limited physical exercise,
and ageing will further aggravate the situation and lead to
the development of type 2 diabetes symptoms.
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precursor cells are not “undifferentiated” cells and are not
necessarily clonigenic. It is not clear whether this mechanism of regeneration may also be operational in mammals,
although there are examples of transdifferentiation. Most
notably, in the pancreas it is known that severe tissue injury
leads to acinoductal transdifferentiation or “metaplasia.” In
this case acinar exocrine cells switch to a ductlike phenotype (117, 118). Under other conditions, hepatocytes can
arise in the adult pancreas, for example, after feeding a
copper-deficient diet to rodents (115). It has been demonstrated that embryonic pancreatic cells, the acinar tumor
cell line AR42-J (136), and freshly isolated adult acinar cells
(78) can transdifferentiate to hepatocytes in vitro. AR42-J
cells can also transdifferentiate into endocrine cells (91,
170). Transdifferentiation of acinar exocrine cells to insulinproducing endocrine cells may also take place in vivo, as
indicated by the occurrence of transitional phenotypes coexpressing exocrine and endocrine cell markers in pancreatic duct-ligated rats (9, 77). Recently, in vitro transdifferentiation of exocrine cells into insulin-producing beta cells was
reported by the addition of EGF and LIF to the medium (5).
This treatment also induced reexpression of the embryonic
proendocrine transcription factor Ngn3 (unpublished observations). Forced expression of Ngn3 in adult human exocrine duct cells by adenoviral transfection turns on the
endocrine differentiation pathway (59). The mechanism of
acinar-to-islet transdifferentiation (via a ductlike intermediate) may also be involved in the gastrin-stimulated neogenesis process that has been observed in duct-ligated rats (119)
and in alloxan-treated mice (121). It has also been suggested
as the mechanism that causes beta-cell hyperplasia and
metabolic adaptation in the chronically glucose infused rat
(86, 152).
Others have suggested that human beta cells can
transdifferentiate into ductlike cells (131, 169). Such
trans- or dedifferentiated islet cells would lose insulin
expression and gain vigorous proliferative potential and
the ability to expand considerably by serial passages in
culture, but could not be induced to redifferentiate (6, 19).
Still others proposed that human beta cells can transdifferentiate in vitro to mesenchymal cells that have a vigorous proliferative potential. In the absence of serum in
the culture medium, these expanded cells could redifferentiate into functional beta cells (53). The transdifferentiation from mature beta cells, or other pancreatic cells, to
cells with a higher proliferative capacity and differentiation plasticity could represent an alternative explanation
to observations that were attributed by others to “progenitor” or “stem” cells with limited self-replicative activity
(e.g., Ref. 133).
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Physiol Rev • VOL
different, since this is not an autoimmune disease. Pharmacological regulation of the beta-cell mass therefore
represents an interesting target for the treatment of type
2 diabetes patients that benefit from insulin treatment or
that have become insulin dependent.
ACKNOWLEDGMENTS
Address for reprint requests and other correspondence: L.
Bouwens, Cell Differentiation Unit, Vrije Universiteit Brussel,
Laarbeeklaan 103, B-1090 Brussels, Belgium (E-mail: lucbo
@expa.vub.ac.be).
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Neogenesis by the process of transdifferentiation
and/or stem cells appears to be activated only after severe
injury to endocrine or exocrine pancreatic tissue, or when
strong additional stimuli like Glp-1 or gastrin are provided. Beta-cell self-replication can also be increased by
certain stimuli like glucose and other nutrients, insulin
and several growth factors, but it apparently cannot lead
to an efficient regeneration. This can be explained by the
small fraction of cells engaged in the cell cycle, by the
observation that increased beta-cell replication is associated with increased apoptotic death, and possibly by a
glucotoxic effect of chronic hyperglycemia.
Recently, some confusion has been raised as to the
relative importance of neogenesis, or even its mere existence, following the paper by Dor et al. (37). This paper
has shown that there is no neogenesis during normal
physiological renewal and compensatory growth of the
murine beta cell mass, confirming previous studies (see
sect. III). In addition, no evidence for neogenesis was
found following two-thirds pancreatectomy. Care should
be taken, however, to avoid over-interpretation of these
data, since this is an experimental model causing only
very limited regenerative growth (see sect. IV). In other
experimental models with a much more important betacell growth (Table 1), it is clear that beta-cell neogenesis
does operate. A comparable “duality” is known from liver
regeneration where there has been a long debate concerning the relative importance of hepatocyte self-replication
versus stem cell derivation. It is now known that under
some conditions, like after two-thirds hepatectomy, regeneration is caused by replication of residual hepatocytes, whereas under other conditions hepatic stem cells
are called upon, namely when hepatocyte replication is
compromised by certain drugs or when there is extensive
inflammation (3, 41). Nevertheless, it would be interesting
for future studies to apply a genetic tracing approach
similar to the one introduced by Dor et al., to confirm the
occurrence of neogenesis and to trace the beta cell precursors in the pancreas. It is to be expected that different
types of precursors may be involved, again much depending on the experimental conditions used to cause injury to
pancreatic tissue(s).
Combinations of agents that stimulate beta-cell replication, inhibit beta-cell apoptosis, and induce beta-cell
neogenesis, are probably the best option for pharmacological restoration of a functional beta-cell mass in diabetics. It is very likely, however, that a pharmacological
treatment resulting in regeneration of beta cells would be
inefficient in type 1 patients due to the expected recurrence of autoimmunity and inflammatory destruction of
the newly formed beta cells. So, whether the knowledge
on beta-cell mass control is to find application in cell
therapy (ex vivo generation) or in regenerative therapy,
controlling the immune system represents the major challenge. In the case of type 2 diabetes, the situation is
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