Exercise and dexamethasone oppositely modulate beta-cell function and survival via independent pathways in 90% pancreatectomized rats

Long-term dexamethasone (DEX) treatment is well known for its ability to increase insulin resistance in liver and adipose tissues leading to hyperinsulinemia. On the other hand, exercise enhances peripheral insulin sensitivity. However, it is not clear whether DEX and/or exercise affect beta-cell mass and function in diabetic rats, and whether their effects can be associated with the modulation of the insulin/IGF-I signaling cascade in pancreatic beta-cells. After an 8-week study, whole body glucose disposal rates in 90% pancreatectomized (Px) and sham-operated male rats decreased with a high dose treatment of DEX (0.1mg DEX/kg body weight/day)(HDEX) treatment, while disposal rates increased with exercise. First-phase insulin secretion was decreased and delayed by DEX via the impairment of the glucose-sensing mechanism in beta-cells, while exercise reversed the impairment of first-phase insulin secretion caused by DEX, suggesting ameliorated beta-cell functions. However, exercise and DEX did not alter second-phase insulin secretion except for the fact that HDEX decreased insulin secretion at 120 min during hyperglycemic clamp in Px rats. Unlike beta-cell functions, DEX and exercise exhibited increased pancreatic beta-cell mass in two different pathways. Only exercise, through increased proliferation and decreased apoptosis, increased beta-cell mass via hyperplasia, which resulted from an enhanced insulin/IGF-I signaling cascade by insulin receptor substrate 2 induction. By contrast, DEX expanded beta-cell mass via hypertrophy and neogenesis from precursor cells, rather than increasing proliferation and decreasing apoptosis. In conclusion, the improvement of beta-cell function and survival via the activation of an insulin/IGF-I signaling cascade due to exercise has a crucial role in preventing the development and progression of type 2 diabetes.     

Determinants of pancreatic beta-cell regeneration

Recent studies have revealed a surprising plasticity of pancreatic beta-cell mass. beta-cell mass is now recognized to increase and decrease in response to physiological demand, for example during pregnancy and in insulin-resistant states. Moreover, we and others have shown that mice recover spontaneously from diabetes induced by killing of 70-80% of beta-cells, by beta-cell regeneration. The major cellular source for new beta-cells following specific ablation, as well as during normal homeostatic maintenance of adult beta-cells, is proliferation of differentiated beta-cells. More recently, it was shown that one form of severe pancreatic injury, ligation of the main pancreatic duct, activates a population of embryonic-type endocrine progenitor cells, which can differentiate into new beta-cells. The molecular triggers for enhanced beta-cell proliferation during recovery from diabetes and for activation of embryonic-type endocrine progenitors remain unknown and represent key challenges for future research. Taken together, recent data suggest that regenerative therapy for diabetes may be a realistic goal.     

Molecular control of cell cycle progression in the pancreatic beta-cell

Type 1 and type 2 diabetes both result from inadequate production of insulin by the beta-cells of the pancreatic islet. Accordingly, strategies that lead to increased pancreatic beta-cell mass, as well as retained or enhanced function of islets, would be desirable for the treatment of diabetes. Although pancreatic beta-cells have long been viewed as terminally differentiated and irreversibly arrested, evidence now indicates that beta-cells can and do replicate, that this replication can be enhanced by a variety of maneuvers, and that beta-cell replication plays a quantitatively significant role in maintaining pancreatic beta-cell mass and function. Because beta-cells have been viewed as being unable to proliferate, the science of beta-cell replication is undeveloped. In the past several years, however, this has begun to change at a rapid pace, and many laboratories are now focused on elucidating the molecular details of the control of cell cycle in the beta-cell. In this review, we review the molecular details of cell cycle control as they relate to the pancreatic beta-cell. Our hope is that this review can serve as a common basis and also a roadmap for those interested in developing novel strategies for enhancing beta-cell replication and improving insulin production in animal models as well as in human pancreatic beta-cells.     

GLP-1 receptor signaling: effects on pancreatic beta-cell proliferation and survival

Type 2 diabetes is a metabolic disorder characterized by insulin resistance as well as a progressive deterioration of pancreatic beta-cell mass and function. Glucagon-like peptide 1 (GLP-1), an incretin hormone secreted by intestinal L cells, is a promising therapeutic agent in the treatment of diabetes. GLP-1 analogs and enhancers constitute a novel class of anti-diabetes medications which address both the insulin secretion defect as well as the decline in beta-cell mass. GLP-1 improves glucose-stimulated insulin secretion, restores glucose competence in glucose-resistant beta-cells, and stimulates insulin gene expression and biosynthesis. Furthermore, GLP-1 acts as a growth factor by promoting beta-cell proliferation, survival and neogenesis. This review focuses on the molecular mechanisms by which GLP-1 signaling induces beta-cell mass expansion.     

Review: pancreatic beta-cell neogenesis revisited

Beta-cell neogenesis triggers the generation of new beta-cells from precursor cells. Neogenesis from duct epithelium is the most currently described and the best documented process of differentiation of precursor cells into beta-cells. It is contributes not only to beta-cell mass expansion during fetal and nonatal life but it is also involved in the maintenance of the beta-cell mass in adults. It is also required for the increase in beta-cell mass in situations of increase insulin demand (obesity, pregnancy). A large number of factors controlling the differentiation of beta-cells has been identified. They are classified into the following main categories: growth factors, cytokine and inflammatory factors, and hormones such as PTHrP and GLP-1. The fact that intestinal incretin hormone GLP-1 exerts a major trophic role on pancreatic beta-cells provides insights into the possibility to pharmacologically stimulate beta-cell neogenesis. This could have important implications for the of treatment of type 1 and type 2 diabetes. Transdifferentiation, that is, the differentiation of already differentiated cells into beta-cells, remains controversial. However, more and more studies support this concept. The cells, which can potentially "transdifferentiate" into beta-cells, can belong to the pancreas (acinar cells) and even islets, or originate from extra-pancreatic tissues such as the liver. Neogenesis from intra-islet precursors also have been proposed and subpopulations of cell precursors inside islets have been described by some authors. Nestin positive cells, which have been considered as the main candidates, appear rather as progenitors of endothelial cells rather than beta-cells and contribute to angiogenesis rather than neogenesis. To take advantage of the different differentiation processes may be a direction for future cellular therapies. Ultimately, a better understanding of the molecular mechanisms involved in beta-cell neogenesis will allow us to use any type of differentiated and/or undifferentiated cells as a source of potential cell precursors.     

Anti-diabetic actions of glucagon-like peptide-1 on pancreatic beta-cells

Glucagon-like peptide-1 (GLP-1), an incretin hormone, is released from intestinal L-cells in response to nutrients. GLP-1 lowers blood glucose levels by stimulating insulin secretion from pancreatic beta-cells in a glucose-dependent manner. In addition, GLP-1 slows gastric emptying, suppresses appetite, reduces plasma glucagon, and stimulates glucose disposal, which are beneficial for glucose homeostasis. Therefore, incretin-based therapies such as GLP-1 receptor agonists and inhibitors of dipeptidyl peptidase IV, an enzyme which inactivates GLP-1, have been developed for treatment of diabetes. This review outlines our knowledge of the actions of GLP-1 on insulin secretion and biosynthesis, beta-cell proliferation and regeneration, and protection against beta-cell damage, as well as the involvement of recently discovered signaling pathways of GLP-1 action, mainly focusing on pancreatic beta-cells.     

IGF-I and GH post-receptor signaling mechanisms for pancreatic beta-cell replication

Certain nutrients, pharmacological agents and growth factors can stimulate pancreatic beta-cell proliferation; however, mitogenic signal transduction pathways in beta-cells have not been particularly well characterized. As a model system we have focussed on characterizing the signal transduction pathways immediately downstream of the IGF-I and GH receptors in beta-cells. The original idea was to gain an idea of important elements in mitogenic signaling pathways which might then be exploited to generate a marked increase in beta-cell proliferation. Such an approach could eventually reveal a means to increase the number of human pancreatic endocrine cells in vitro, in order to obtain an abundant source of beta-cells for routine transplantation therapy of type-I diabetes. However, in the course of our studies, we have also unveiled an unexpected insight into the pathogenesis of obesity-linked type-II diabetes. It has been observed that free fatty acids inhibit glucose- and glucose-dependent IGF-I/GH-induced beta-cell proliferation. We hypothesize that a gradual accumulation of intracellular fat in beta-cells during obesity can eventually lead to an inhibition of beta-cell mass expansion and hence failure to compensate for peripheral insulin resistance, so that type-II diabetes ensues.     

Islet endothelial cells and pancreatic beta-cell proliferation: studies in vitro and during pregnancy in adult rats

The growth of both tumors and nonneoplastic tissues may be influenced by signals from the vascular endothelium. In the present investigation we show that purified proliferating endothelial cells from pancreatic islets can stimulate beta-cell proliferation through secretion of hepatocyte growth factor (HGF). This secretion could be induced by soluble signals from the islets, such as vascular endothelial growth factor-A (VEGF-A) and insulin. During pregnancy, the pancreatic beta-cells display a highly reproducible physiological proliferation. We show that islet endothelial cell proliferation precedes beta-cell proliferation in pregnant animals. Vascular growth was closely associated with endocrine cell proliferation, and prominent expression of HGF was observed in islet endothelium on d 15 of pregnancy, i.e. coinciding with the peak of beta-cell proliferation. In summary, our results suggest the existence of an endothelial-endocrine axis within adult pancreatic islets, which is of importance for adult beta-cell proliferation.     

Inhibition of cholesterol biosynthesis impairs insulin secretion and voltage-gated calcium channel function in pancreatic beta-cells

Insulin secretion from pancreatic beta-cells is mediated by the opening of voltage-gated Ca2+ channels (CaV) and exocytosis of insulin dense core vesicles facilitated by the secretory soluble N-ethylmaleimide-sensitive factor attachment protein receptor protein machinery. We previously observed that beta-cell exocytosis is sensitive to the acute removal of membrane cholesterol. However, less is known about the chronic changes in endogenous cholesterol and its biosynthesis in regulating beta-cell stimulus-secretion coupling. We examined the effects of inhibiting endogenous beta-cell cholesterol biosynthesis by using the squalene epoxidase inhibitor, NB598. The expression of squalene epoxidase in primary and clonal beta-cells was confirmed by RT-PCR. Cholesterol reduction of 36-52% was observed in MIN6 cells, mouse and human pancreatic islets after a 48-h incubation with 10 mum NB598. A similar reduction in cholesterol was observed in the subcellular compartments of MIN6 cells. We found NB598 significantly inhibited both basal and glucose-stimulated insulin secretion from mouse pancreatic islets. CaV channels were markedly inhibited by NB598. Rapid photolytic release of intracellular caged Ca2+ and simultaneous measurements of the changes in membrane capacitance revealed that NB598 also inhibited exocytosis independently from CaV channels. These effects were reversed by cholesterol repletion. Our results indicate that endogenous cholesterol in pancreatic beta-cells plays a critical role in regulating insulin secretion. Moreover, chronic inhibition of cholesterol biosynthesis regulates the functional activity of CaV channels and insulin secretory granule mobilization and membrane fusion. Dysregulation of cellular cholesterol may cause impairment of beta-cell function, a possible pathogenesis leading to the development of type 2 diabetes.     

Crucial role of insulin receptor substrate-2 in compensatory beta-cell hyperplasia in response to high fat diet-induced insulin resistance

In type 2 diabetes, there is a defect in the regulation of functional beta-cell mass to overcome high-fat (HF) diet-induced insulin resistance. Many signals and pathways have been implicated in beta-cell function, proliferation and apoptosis. The co-ordinated regulation of functional beta-cell mass by insulin signalling and glucose metabolism under HF diet-induced insulin-resistant conditions is discussed in this article. Insulin receptor substrate (IRS)-2 is one of the two major substrates for the insulin signalling. Interestingly, IRS-2 is involved in the regulation of beta-cell proliferation, as has been demonstrated using knockout mice models. On the other hand, in an animal model for human type 2 diabetes with impaired insulin secretion because of insufficiency of glucose metabolism, decreased beta-cell proliferation was observed in mice with beta-cell-specific glucokinase haploinsufficiency (Gck(+/) (-)) fed a HF diet without upregulation of IRS-2 in beta-cells, which was reversed by overexpression of IRS-2 in beta-cells. As to the mechanism underlying the upregulation of IRS-2 in beta-cells, glucose metabolism plays an important role independently of insulin, and phosphorylation of cAMP response element-binding protein triggered by calcium-dependent signalling is the critical pathway. Downstream from insulin signalling via IRS-2 in beta-cells, a reduction in FoxO1 nuclear exclusion contributes to the insufficient proliferative response of beta-cells to insulin resistance. These findings suggest that IRS-2 is critical for beta-cell hyperplasia in response to HF diet-induced insulin resistance.     

Long-term inhibition of protein tyrosine kinase impairs electrophysiologic activity and a rapid component of exocytosis in pancreatic beta-cells

Dysfunction of pancreatic beta-cells is a fundamental feature in the pathogenesis of type 2 diabetes. As insulin receptor signaling occurs via protein tyrosine kinase (PTK), we investigated the role of PTK activity in the etiology of beta-cell dysfunction by inhibiting PTK activity in primary cultured mouse pancreatic beta-cells and INS-1 cells with genistein treatment over 24 h. Electrophysiologic recordings showed genistein treatment significantly attenuated ATP-sensitive K(+) (K(ATP)) and voltage-dependent Ca(2+) currents, and depolarized the resting membrane potential in primary beta-cells. When stimulated by high glucose, genistein-treated beta-cells exhibited a time delay of both depolarization and Ca(2+) influx, and were unable to fire action potentials, as well as displaying a reduced level of Ca(2+) influx and a loss of Ca(2+) oscillations. Semiquantitative PCR analysis revealed decreased expression of K(ATP) and L-type Ca(2+) channel mRNA in genistein-treated islets. PTK inhibition also significantly reduced the rapid component of secretory vesicle exocytosis, as indicated by membrane capacitance measurements, and this is likely to be due to the reduced Ca(2+) current amplitude in these cells. These results illustrate that compromised PTK activity contributes to pancreatic beta-cell dysfunction and may be involved in the etiology of type 2 diabetes.     

Evidence of unrestrained beta-cell proliferation and neogenesis in a patient with hyperinsulinemic hypoglycemia after gastric bypass surgery

ABSTRACT

Hyperinsulinemic hypoglycemia syndrome (HIHG) is a rare complication of roux-en-Y gastric bypass surgery. The pathology is associated with an excessive function of pancreatic beta-cells, and requires pancreas resection in patients that are recalcitrant to nutritional and pharmacological interventions. The exact prevalence is not clearly understood and the underlying mechanisms not yet fully characterized. We herein sought to perform histological and molecular examination of pancreatic sections obtained from a patient who developed HIHG as a complication of gastric bypass compared to 3 weight-matched controls. We studied markers of cellular replication and beta-cell differentiation by immunohistochemistry and immunofluorescence. HIHG after gastric bypass was characterized by a profound increase in beta-cell mass. Cellular proliferation was increased in islets and ducts compared to controls, suggesting unrestrained proliferation in HIHG. We also detected beta-cell differentiation markers in duct cells and occasional duct cells displaying both insulin and glucagon immunoreactivity. These histological observations suggest that beta-cell differentiation from ductal progenitor cells could also underly beta-cell mass expansion in HIHG. Altogether, our results can be construed to demonstrate that HIHG after gastric bypass is characterized by abnormal beta-cell mass expansion, resulting from both unrestrained beta-cell replication and neogenesis.     

Acylated and unacylated ghrelin promote proliferation and inhibit apoptosis of pancreatic beta-cells and human islets: involvement of 3',5'-cyclic adenosine monophosphate/protein kinase A, extracellular signal-regulated kinase 1/2, and phosphatidyl inositol 3-Kinase/Akt signaling

Among its pleiotropic actions, ghrelin modulates insulin secretion and glucose metabolism. Herein we investigated the role of ghrelin in pancreatic beta-cell proliferation and apoptosis induced by serum starvation or interferon (IFN)-gamma/TNF-alpha, whose synergism is a major cause for beta-cell destruction in type I diabetes. HIT-T15 beta-cells expressed ghrelin but not ghrelin receptor (GRLN-R), which binds acylated ghrelin (AG) only. However, both unacylated ghrelin (UAG) and AG recognized common high-affinity binding sites on these cells. Either AG or UAG stimulated cell proliferation through Galpha(s) protein and prevented serum starvation- and IFN-gamma/TNF-alpha-induced apoptosis. Antighrelin antibody enhanced apoptosis in either the presence or absence of serum but not cytokines. AG and UAG even up-regulated intracellular cAMP. Blockade of adenylyl cyclase/cAMP/protein kinase A signaling prevented the ghrelin cytoprotective effect. AG and UAG also activated phosphatidyl inositol 3-kinase (PI3K)/Akt and ERK1/2, whereas PI3K and MAPK inhibitors counteracted the ghrelin antiapoptotic effect. Furthermore, AG and UAG stimulated insulin secretion from HIT-T15 cells. In INS-1E beta-cells, which express GRLN-R, AG and UAG caused proliferation and protection against apoptosis through identical signaling pathways. Noteworthy, both peptides inhibited cytokine-induced NO increase in either HIT-T15 or INS-1E cells. Finally, they induced cell survival and protection against apoptosis in human islets of Langerhans. These expressed GRLN-R but showed also UAG and AG binding sites. Our data demonstrate that AG and UAG promote survival of both beta-cells and human islets. These effects are independent of GRLN-R, are likely mediated by AG/UAG binding sites, and involve cAMP/PKA, ERK1/2, and PI3K/Akt.     

Cell therapy for type 2 diabetes: is it desirable and can we get it?

The functional mass of beta-cells is decreased in type 2 diabetes. Replacing missing beta-cells or triggering their regeneration may thus allow for improved treatment of type 2 diabetes, to the extent that this is combined with therapy for improved insulin sensitivity. Although progress has been made in deriving beta-cell-like cells from stem or precursor cells in vitro, these cannot yet be obtained in sufficient quantities or well enough differentiated to envisage their therapeutic use in beta-cell replacement therapy. Likewise, our very limited understanding of beta-cell regeneration in adult man does not yet allow for development of a valid strategy for kick-starting such a process in individuals with type 2 diabetes, whether by bona fide neogenesis or self-replication of existing beta-cells. Regardless of how beta-cell mass is restored in type 2 diabetes, it will be important to prevent any renewed decrease thereafter. Current understanding suggests that islet inflammation as well as signals from (insulin-resistant/inflamed) adipose tissue and skeletal muscle contribute towards decreased beta-cell mass in type 2 diabetes. It will likely be important to protect newly formed or implanted beta-cells from these negative influences to ensure their long-term survival.