Islet Growth Research Articles

Incretins and their analogues as new antidiabetic drugs.

Glucagon-like peptide 1 (GLP-1) is a gut (incretin) hormone with multiple actions that could potentially contribute to an antidiabetic effect. This includes: (a) glucose-dependent insulinotropic actions; (b) glucagonostatic actions; (c) a reduction in appetite/promotion of satiety leading to reduced food intake and weight reduction; (d) the deceleration of gastric emptying; and (e) the stimulation of islet growth, differentiation and regeneration. Thus, multiple aspects of the type 2 diabetic phenotype can potentially be improved or even corrected by GLP-1. The native gut hormone, however, after intravenous injection or absorption from subcutaneous depots, is proteolytically degraded and eliminated from the circulation too quickly to be useful for the treatment of diabetes. GLP-1 derivatives (receptor agonists) with prolonged pharmacokinetics that promise a potential for clinical use in the near future are being developed.

Identification and characterization of small cells in the adult pancreas: potential progenitor cells?

In this report we describe the identification of a novel cell type in human and canine pancreas using tissue culture techniques. These cells, representing less than 1% of total islet cells, are of a small size (7-10 microm) and highly quiescent. They display a fairly immature morphology, which is characterized by a weakly developed protein synthesis machinery, a few mitochondria and a small number of neuroendocrine granules. These cells, which we have termed "small cells," are usually organized into small clusters, which can be identified within the islets of predominantly small size. They can also be collected as separate structures from preparations of freshly isolated islets. Immunohistochemically, small cells are positive for PDX-1, synaptophysin, insulin, glucagon, somatostatin, pancreatic polypeptide, alpha-fetaprotein and Bcl-2 and negative for cytokeratin 19 and nestin. Insulin secretion studies demonstrated that these cells secrete insulin in a glucose-responsive fashion, although do not respond to secretagogues such as IBMX and arginine as do mature beta cells. Although this study does not provide evidence of the proliferative and differentiation potential of small cells, their immature morphology, along with a small size and quiescence, let us hypothesize that these cells may serve as progenitors contributing to the islet growth.

Beta-cell-specific deletion of the Igf1 receptor leads to hyperinsulinemia and glucose intolerance but does not alter beta-cell mass.

Regulation of glucose homeostasis by insulin depends on the maintenance of normal beta-cell mass and function. Insulin-like growth factor 1 (Igf1) has been implicated in islet development and differentiated function, but the factors controlling this process are poorly understood. Pancreatic islets produce Igf1 and Igf2, which bind to specific receptors on beta-cells. Igf1 has been shown to influence beta-cell apoptosis, and both Igf1 and Igf2 increase islet growth; Igf2 does so in a manner additive with fibroblast growth factor 2 (ref. 10). When mice deficient for the Igf1 receptor (Igf1r(+/-)) are bred with mice lacking insulin receptor substrate 2 (Irs2(-/-)), the resulting compound knockout mice show a reduction in mass of beta-cells similar to that observed in pancreas of Igf1r(-/-) mice (ref. 11), suggesting a role for Igf1r in growth of beta-cells. It is possible, however, that the effects in these mice occur secondary to changes in vascular endothelium or in the pancreatic ductal cells, or because of a decrease in the effects of other hormones implicated in islet growth. To directly define the role of Igf1, we have created a mouse with a beta-cell-specific knockout of Igf1r (betaIgf1r(-/-)). These mice show normal growth and development of beta-cells, but have reduced expression of Slc2a2 (also known as Glut2) and Gck (encoding glucokinase) in beta-cells, which results in defective glucose-stimulated insulin secretion and impaired glucose tolerance. Thus, Igf1r is not crucial for islet beta-cell development, but participates in control of differentiated function.

Receptors for insulin and insulin-like growth factor-1 and insulin receptor substrate-1 mediate pathways that regulate islet function.

The insulin/insulin-like growth factor-1 (IGF-1) signalling pathways are present in most mammalian cells and play important roles in the growth and metabolism of tissues. Most proteins in these pathways have also been identified in the beta-cells of the pancreatic islets. Tissue-specific knockout of the insulin receptor (betaIRKO) or IGF-1 receptor (betaIGFRKO) in pancreatic beta-cells leads to altered glucose-sensing and glucose intolerance in adult mice, and betaIRKO mice show an age-dependent decrease in islet size and beta-cell mass. These data indicate that these receptors are important for differentiated function and are unlikely to play a major role in the early growth and/or development of the pancreatic islets. Conventional insulin receptor substrate-1 (IRS-1) knockouts manifest growth retardation and mild insulin resistance. The IRS-1 knockouts also display islet hyperplasia, defects in insulin secretory responses to multiple stimuli both in vivo and in vitro, reduced islet insulin content and an increased number of autophagic vacuoles in the beta-cells. Re-expression of IRS-1 in cultured beta-cells is able to partially restore the insulin content indicating that IRS-1 is involved in the regulation of insulin synthesis. Taken together, these data provide evidence that insulin and IGF-1 receptors and IRS-1, and potentially other proteins in the insulin/IGF-1 signalling pathway, contribute to the regulation of islet hormone secretion and synthesis and therefore in the maintenance of glucose homeostasis.

Drug-induced islet growth: a novel treatment for diabetes?

Drug-induced islet growth: a novel treatment for diabetes?

In Vitro Maturation of Neonatal Porcine Islets

The mechanisms involved in islet neogenesis have remained largely unexplored due to lack of an appropriate model. Furthermore, with the recent advances in islet transplantation, the need for alternative islet tissue sources is greater than ever. Therefore, the authors have refined a neonatal porcine islet (NPI) maturation model that offers an ideal tool to gain insight into islet growth as well as an alternative source of transplantable tissue. Recent knowledge in islet growth has resulted in endocrine tissue being derived from human pancreatic precursor tissue in vitro. The potential for large scale production of endocrine tissue in vitro has been indicated, however, more investigation must be done on the various signals and pathways involved in pancreatic development to optimize this technique. The authors believe that their NPI in vitro maturation model provides an ideal tool to study islet growth and maturation. Transduction of the NPI to overexpress genes of interest (i.e., PDX-1) or exposure of the NPI to various culture conditions will allow us to determine the effects on islet maturation. An understanding of NPI development gained will not only allow us to mature this unlimited tissue source for optimal xenotransplantation, but also elude to how human pancreatic endocrine precursor cells may be used to solve the current islet tissue supply problem.

Using beta-cell growth factors to enhance human pancreatic Islet transplantation.

This is a particularly exciting time in the field of pancreatic islet growth, development, and survival. The recent publication of a study demonstrating that human pancreatic islet transplantation is both technically and immunologically feasible has highlighted the need for large supplies of pancreatic islets or pancreatic beta cells for larger-scale islet transplantation in patients with diabetes. This, together with a rapid expansion in the past several years of the understanding of mechanisms of islet growth, development, and survival, has accelerated and invigorated efforts to therapeutically harness the cellular mechanisms responsible for pancreatic beta-cell proliferation, survival, and development and to take advantage of this new knowledge to enhance the availability, survival, and function of pancreatic beta cells in human islet transplantation for diabetes mellitus. Here, we briefly review the confluence of events that have provided optimism and energy to the islet transplant field, and we focus on peptide growth factors that eventually may be deployed in the effort to augment islet mass and function in patients with diabetes.

Glucagon-like peptide 1 receptor signaling influences topography of islet cells in mice.

Glucagon-like peptide 1 (GLP-1) amplifies glucose-induced insulin release in vivo and in vitro. Activation of GLP-1 receptor (GLP-1R) signaling leads to differentiation of exocrine cells towards a beta-cell phenotype in vitro and stimulation of islet cell proliferation in vitro and in vivo, suggesting a potential role for GLP-1 in the modulation of islet growth and differentiation. To determine whether basal levels of GLP-1R signaling are essential for islet development, we examined islet cell composition and topography in GLP-1R-/- mice. Total beta-cell volume and number are not altered, but the topography of beta cells is markedly different in GLP-1R-/- mice compared with GLP-1R+/+ controls. The distribution of beta cells is shifted from large to small and medium-sized islets in the absence of GLP-1R signaling (large islets: 50 +/- 3% in GLP-1R+/+ vs 28 +/- 4% in GLP- 1R-/-, P < 0.01 and medium islets: 32 +/- 2% in GLP- 1R+/+ vs 48 +/- 3% in GLP-1R-/-, P < 0.001). Furthermore, GLP-1R-/- islets exhibit abnormalities in cell topography, with two to threefold more centrally located alpha cells detected in GLP-1R-/- islets. These alterations in alpha- and beta-cell topography indicate that basal levels of GLP-1 signaling in the normal rodent are involved in the normal cellular organization of the endocrine pancreas.

Elimination of glucagon-like peptide 1R signaling does not modify weight gain and islet adaptation in mice with combined disruption of leptin and GLP-1 action.

Leptin and glucagon-like peptide 1 (GLP-1) exhibit opposing actions in the endocrine pancreas. GLP-1 stimulates insulin biosynthesis, secretion, and islet growth, whereas leptin inhibits glucose-dependent insulin secretion and insulin gene transcription. In contrast, GLP-1 and leptin actions overlap in the central nervous system, where leptin has been shown to activate GLP-1 circuits that inhibit food intake. To determine the physiological importance of GLP-1 receptor (GLP-1R)-leptin interactions, we studied islet function and feeding behavior in ob/ob:GLP-1R(-/-) mice. Although GLP-1R actions are thought to be essential for glucose-dependent insulin secretion, the levels of fasting glucose, glycemic excursion after glucose loading, glucose-stimulated insulin, and pancreatic insulin RNA content were similar in ob/ob:GLP-1R(+/+) versus ob/ob:GLP-1R(-/-) mice. Despite evidence linking GLP-1R signaling to the regulation of islet neogenesis and proliferation, ob/ob:GLP-1R(-/-) mice exhibited significantly increased islet numbers and area and an increase in the number of large islets compared with GLP-1R(+/+) or (-/-) mice (P < -0.01 to 0.05). Similarly, growth rates and both shortand long-term control of food intake were comparable in ob/ob:GLP-1R(+/+) versus ob/ob:GLP-1R4(-/-) mice. Furthermore, leptin produced a similar inhibition of food intake in GLP-1R(-/-), ob/ob:GLP-1R(+/+), and ob/ob:GLP1R4(-/-) mice. These findings illustrate that although leptin and GLP-1 actions overlap in the brain and endocrine pancreas, disruption of GLP-1 signaling does not modify the response to leptin or the phenotype of leptin deficiency in the ob/ob mouse, as assessed by long-term control of body weight or the adaptive beta-cell response to insulin resistance in vivo.

Transcription factor expression during pancreatic islet regeneration

Recent studies by a number of laboratories have identified transcription factors that are involved in pancreatic development. Indeed, marked abnormalities in pancreatic development result from deficiencies in these molecules, which include, among others, PDX-1, islet-1 (Isl-1), and Pax-6. These studies have prompted us to evaluate the expression of Isl-1 and Pax-6 in the pancreas of the interferon-gamma (IFNγ) transgenic mouse, which exhibits new islet growth and expansion of ducts throughout the life of the animal. We have previously demonstrated that PDX-1 is strikingly expressed in the ducts of the IFNγ transgenic mouse. This latter observation compelled us to examine expression of hepatocyte nuclear factor-3beta (HNF3β), which mediates PDX-1 gene transcription, in the IFNγ transgenic pancreas as well. As a result of these studies, we now demonstrate marked expression of these transcription factors in the pancreatic ducts of IFNγ transgenic mice. These data suggest a role for these transcription factors during pancreatic regeneration in the IFNγ transgenic mouse.

Islet growth and development in the adult.

Diabetes mellitus, whether type 1 (insulindependent diabetes mellitus; IDDM) or type 2 (noninsulin-dependent diabetes mellitus; NIDDM) results from an inadequate mass of functional pancreatic -cells. In the first case, the -cell mass is reduced by the autoimmune destruction of the -cells, while, in the latter, there is incomplete compensation to meet the demand often imposed by insulin resistance. This latter point is not always appreciated. However, only 20% of those people with severe insulin resistance due to obesity, Cushing’s disease, or acromegaly become diabetic, with the other 80% being able to compensate to

Transgenic mice overexpressing insulin-like growth factor-II in beta cells develop type 2 diabetes.

During embryonic development, insulin-like growth factor-II (IGF-II) participates in the regulation of islet growth and differentiation. We generated transgenic mice (C57BL6/SJL) expressing IGF-II in beta cells under control of the rat Insulin I promoter in order to study the role of islet hyperplasia and hyperinsulinemia in the development of type 2 diabetes. In contrast to islets from control mice, islets from transgenic mice displayed high levels of IGF-II mRNA and protein. Pancreases from transgenic mice showed an increase in beta-cell mass (about 3-fold) and in insulin mRNA levels. However, the organization of cells within transgenic islets was disrupted, with glucagon-producing cells randomly distributed throughout the core. We also observed enhanced glucose-stimulated insulin secretion and glucose utilization in islets from transgenic mice. These mice displayed hyperinsulinemia, mild hyperglycemia, and altered glucose and insulin tolerance tests, and about 30% of these animals developed overt diabetes when fed a high-fat diet. Furthermore, transgenic mice obtained from the N1 backcross to C57KsJ mice showed high islet hyperplasia and insulin resistance, but they also developed fatty liver and obesity. These results indicate that local overexpression of IGF-II in islets might lead to type 2 diabetes and that islet hyperplasia and hypersecretion of insulin might occur early in the pathogenesis of this disease.

Constitutive expression of placental lactogen in pancreatic beta cells: effects on cell morphology, growth, and gene expression.

To explore the roles of lactogens in islet function, we generated a stable line of rat insulinoma (INS-1) cells that express rat placental lactogen II (rPLII) constitutively in culture. We used this cell line (Ins-rPLII) to examine the effects of endogenous rPLII on beta-cell growth, islet formation, and the expression of glucose transporter 2 (glut-2) and insulin mRNA. Growth and maturation of Ins-rPLII cells were compared with that of cells transfected stably with an empty expression plasmid (control) and of INS-1 cells treated with exogenous prolactin. The Ins-rPLII cells proliferated more rapidly than control cells in serumfree medium and showed distinct morphologic characteristics in culture. Whereas the control cells flattened readily on plastic and formed a branching monolayer, the Ins-rPLII cells remained more rounded, sent out fewer projections, and formed more numerous (p<0.01) and larger (p<0.01) beta-cell clusters. Larger clusters assumed a spherical form with well-delineated smooth borders and detached more readily from the culture plates. Maturational progression of the Ins-rPLII cells was associated with a 40% increase in preproinsulin mRNA (p<0.05) and a 2-3-fold increase in glut-2 mRNA (p<0.01). Induction of glut-2 mRNA was accompanied by a 1.4-2.4-fold increase (p< 0.01) in the uptake of radiolabeled 2-deoxyglucose. Similar effects were observed in INS-1 cells exposed for 48 h to exogenous prolactin. These findings suggest novel roles for the lactogenic hormones in the maturation and growth of pancreatic islets. Lactogen induction of beta-cell aggregation coupled with localized beta-cell growth may contribute to the expansion of islet mass that occurs in pregnancy and during the perinatal period. The induction of insulin and glut-2 mRNA provides a mechanism by which the lactogens may increase fetal and maternal insulin production and enhance the sensitivity of the pancreas to glucose.

Parathyroid hormone-related peptide stimulates DNA synthesis and insulin secretion in pancreatic islets.

Parathyroid hormone (PTH)-related protein (PTHrP) is present in the pancreatic islet. Recent data in transgenic mice suggest that PTHrP might modulate islet mass and insulin secretion. In the present study, we assessed the effect of the N-terminal PTH-like region of PTHrP on DNA synthesis in isolated rat islets. PTHrP (1-34), between 1 pM and 10 nM, for 48 h stimulated []thymidine incorporation into rat islets. This effect was maximally induced, about 2.5-fold over control, by 10 pM of this peptide, decreasing thereafter. In contrast, PTHrP (38-64) amide or PTHrP (107-139) were ineffective in increasing DNA synthesis in islets. Using reverse transcription followed by PCR, we confirmed that rat islets express PTHrP and the type I PTH/PTHrP receptor. Addition of a neutralizing anti-PTHrP antibody to the incubation medium of proliferating islets decreased islet DNA synthesis by 30%. The effect of a submaximal dose (30 pM) of PTHrP (1-34) on DNA synthesis in rat islets was abolished by 25 nM bisindolylmaleimide I, a protein kinase C (PKC) inhibitor, but not by 25 microM adenosine 3',5'-cyclic monophosphorothioate, Rp-isomer, a protein kinase A inhibitor. Moreover, 100 nM phorbol-12-myristate-13-acetate for 48 h also increased DNA synthesis 2-fold over controls in islets. PTHrP (1-34), at 100 nM, in contrast to 50 microM forskolin or 10 mM NaF, failed to affect adenylate cyclase activity in islet membranes. PTHrP, at 30 pM, was also found to increase 2-fold insulin released into the islet-conditioned medium within 24-48 h. Our results suggest that PTHrP is a modulator of pancreatic islet growth and/or function by a PKC-mediated mechanism.

Transgenic expression of epidermal growth factor and keratinocyte growth factor in beta-cells results in substantial morphological changes.

The upregulation of a limited number of growth factors in our interferon-gamma transgenic model for regeneration within the pancreas lead us to propose that these factors are important during pancreatic regeneration. In this study, we have assessed the influence of two growth factors within the pancreas, epidermal growth factor (EGF) and keratinocyte growth factor (KGF), by ectopically expressing these proteins under the control of the human insulin promoter in transgenic mice. This beta-cell-targeted expression of either EGF or KGF resulted in significant morphological changes, including cellular proliferation and disorganized islet growth. Intercrossing the individual Ins-EGF and Ins-KGF transgenic mice resulted in more profound changes in pancreatic morphology including proliferation of pancreatic cells and extensive intra-islet fibrosis. Insulin-producing beta-cells were found in some of the ducts of older Ins-EGF and Ins-EGFxKGF transgenic mice, and amylase-producing cells were observed within the islet structures of the double transgenic mice. These data suggest that both EGF and KGF are capable of affecting pancreatic differentiation and growth, and that co-expression of these molecules in islets has a more substantial impact on the pancreas than does expression of either growth factor alone.

Changing distribution of islets in the developing human pancreas: a computer-assisted three-dimensional reconstruction study.

Tissue specimens of nine normal human pancreata from fetuses, neonates, and adults were subjected to serial sectioning and computer-assisted three-dimensional (3-D) reconstruction in an effort to study the growth of Langerhans' islets based on their distribution in the lobule and their relationship with the ducts and arterioles. Also, islet volumes were obtained by 3-D morphometry. In 24-week-old fetuses, the islets were shown to be in direct contact with the ducts, whereas in the neonates, they gradually became separated from the ducts, finally dispersing throughout the lobule in the adults. This transition seemed to allow islet hormones to have physiological effects on the exocrine function of the acinar tissue, making the pancreas achieve the endocrine-exocrine correlation. However, half of the islets remained next to the ducts even in the adult pancreata. With regard to the relationship between the islets and arterioles, "arteriolar" islets having an afferent arteriole accounted for approximately 20% in number, but their volume cumulated to approximately 70% of the total, comprising a greater part of the pancreatic endocrine gland.

Initiation of increased pancreatic islet growth in young normoglycemic mice (Umeå +/?).

Pancreatic islets from obese hyperglycemic mice are large and contain a high proportion of normally functioning beta-cells. We have previously shown that young obese mice have an elevated beta-cell proliferation rate at 3 weeks of age. We now wanted to investigate possible factors involved in the initiation of islet growth, including blood glucose, C peptide, glucagon-like peptide-1, vasoactive intestinal polypeptide, and L-5-hydroxytryptophan. We found that the increased beta-cell proliferation on day 20 precedes the rise in blood glucose by 2 days. The islet cell proliferation, measured as the 5-bromo-2'-deoxyuridine labeling index, in 20-day-old lean mice, was enhanced in a dose-dependent manner when glucagon-like peptide-1 or C peptide was injected s.c. for 2 days. L-5-Hydroxytryptophan inhibited the proliferation. C Peptide also increased the islet cell labeling index during islet culture. We conclude that in addition to the effect of glucose, islet proliferation can be triggered by other factors involved in the physiological regulation of increased insulin release. Stimulation of islet proliferation may be related to the actual release of insulin, and C peptide may function as a mediator of such responses.

Initiation of Increased Pancreatic Islet Growth in Young Normoglycemic Mice (Umea +/?)

Initiation of Increased Pancreatic Islet Growth in Young Normoglycemic Mice (Umea +/?)

Expression of non-classical islet hormone-like peptides during the embryonic development of the pancreas.

Understanding of islet embryogenesis may prove to be key in the design of future therapies for diabetes directed at re-initiating islet growth, with the goal to replace and/or replenish the impaired beta-cell mass in the disease. In this context, studies of islet neurohormonal peptides, known to play a role in the local regulation of islet function, and their expression during islet embryogenesis are important. Here we review our studies on the embryonic islet expression of islet amyloid polypeptide (IAPP) and the PP-fold peptides pancreatic polypeptide (PP), peptide YY (PYY) and neuropeptide Y (NPY). IAPP, which is constitutively expressed in beta- and delta-cells in the adult rat, was found to occur in the assumed pluripotent islet progenitor cell, together with PYY, glucagon, and to a lesser extent with insulin. As development proceeds, the insulin/IAPP phenotype is segregated from that of PYY/glucagon; with the formation of islet-like structures, insulin/IAPP-expressing cells primarily occupy their central portions, while PYY/glucagon-expressing cells are found in their periphery. At the time of formation of islet-like structures, expression of NPY is induced in the insulin/IAPP-containing cells. Whereas NPY-expression ceases at birth, PYY is constitutively expressed in non-beta-cells in the mature rat. Expression of PP is induced just prior to birth in a separate population of islet cells, occasionally co-expressed with PYY. Although a clear role for these peptides during embryogenesis has not been identified, they conceivably could play a role in the control of insulin secretion, islet growth and islet blood flow.

Mapping of murine diabetogenic gene mody on chromosome 7 at D7Mit258 and its involvement in pancreatic islet and beta cell development during the perinatal period.

Mutation of the murine maturity-onset diabetes mellitus of the young (Mody) locus induces diabetes, but the effects of its homozygosity on the pancreas remain unknown. F2 mice were obtained by F1 (diabetic C57BL6 x normal Mus musculus castaneus) crosses. About 20% of the F2 progeny developed diabetes by 2 wk of age, 50% of the progeny were normal at 2 wk and developed diabetes between 5 and 8 wk of age, and the remaining 30% did not develop diabetes. Quantitative trait locus analysis using blood glucose levels of 118 F2 mice at 2 wk of age and 5-8 wk of age located Mody within 3 cM of D7Mit258. Histopathological investigation revealed hypoplastic islets (approximately 33% of that of wild-type mice) and a lower density of beta cells (approximately 20% of wild-type) with a reciprocal dominance of alpha cells (four times that of wild-type) in Mody homozygotes. Electron microscopic observations revealed a specific decrease in the number of insulin secretory granules and a lower density of beta cells. Ratios of insulin to glucagon contents confirmed specific decreases in insulin content: 0.01 for homozygotes, 0.54 for heterozygotes, and 1.11 for wild-type mice on day 14. These results suggest that Mody is involved in both islet growth and beta cell function.