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Objective: DNAJC3, also known as P58IPK, is an Hsp40 family member that interacts with and inhibits PK R-like ER-localized eIF2α kinase (PERK). Dnajc3 deficiency in mice causes pancreatic β-cell loss and diabetes. Loss-of-function mutations in DNAJC3 cause early-onset diabetes and multisystemic neurodegeneration. The aim of our study was to investigate the genetic cause of early-onset syndromic diabe tes in two unrelated patients, and elucidate the mechanisms of β-cell failure in this syndrome. Methods: Whole exome sequencing was performed and identified variants we re confirmed by Sanger sequencing. DNAJC3 was silenced by RNAi in INS-1E cells, primary rat β-cells, human islets, and induced pluripotent stem cellderived β-cells. β-cell function and apoptosis were assessed, and potential media tors of apoptosis examined. Results: The two patients presented with juvenile-onset diabetes, short stature, hypothyroidism, neurodegeneration, facial dysmorphism, hypoacusis, microcephaly and skeletal bone deformities. They were heterozygous compound and homozygous for novel loss-of-function mutations in DNAJC3. DNAJC3 silencing did not impair insulin content or secretion. Instead, the knockdown induced rat and human β-cell apoptosis and further sensitized cells to endoplasmic reticulum stress, triggering mitochondrial apoptosis via the pro-apoptototic Bcl-2 proteins BIM and PUMA. Conclusions: This report confirms previously described features and expands the clinical spectrum of syndromic DNAJC3 diabetes, one of the five monogenic forms of diabetes per taining to the PERK pathway of the endoplasmic reticulum stress response. DNAJC3 deficiency may lead to β-cell loss through BIM- and PUMA-dependent activation of the mitochondrial pathway of apoptosis. [ABSTRACT FROM AUTHOR]

Type 1 diabetes (T1D) is an autoimmune disease leading to β-cell destruction. MicroRNAs (miRNAs) are small noncoding RNAs that control gene expression and organ formation. They participate in the pathogenesis of several autoimmune diseases, but the nature of miRNAs contributing to β-cell death in T1D and their target genes remain to be clarified. We performed an miRNA expression profile on human islet preparations exposed to the cytokines IL-1β plus IFN-γ. Confirmation of miRNA and target gene modification in human β-cells was performed by real-time quantitative PCR. Single-stranded miRNAs inhibitors were used to block selected endogenous miRNAs. Cell death was measured by Hoechst/propidium iodide staining and activation of caspase-3. Fifty-seven miRNAs were detected as modulated by cytokines. Three of them, namely miR-23a-3p, miR-23b-3p, and miR-149-5p, were downregulated by cytokines and selected for further studies. These miRNAs were found to regulate the expression of the proapoptotic Bcl-2 proteins DP5 and PUMA and consequent human β-cell apoptosis. These results identify a novel cross talk between a key family of miRNAs and proapoptotic Bcl-2 proteins in human pancreatic β-cells, broadening our understanding of cytokine-induced β-cell apoptosis in early T1D. [ABSTRACT FROM AUTHOR]

Objective: Friedreich ataxia (FRDA) is an autosomal recessive neurodegenerative disease caused in almost all cases by homozygosity for a GAA trinucleotide repeat expansion in the frataxin gene. Frataxin is a mitochondrial protein involved in iron homeostasis. FRDA patients have a high prevalence of diabetes, the pathogenesis of which is not known. We aimed to evaluate the relative contribution of insulin resistance and β-cell failure and the pathogenic mechanisms involved in FRDA diabetes. Methods: Forty-one FRDA patients, 26 heterozygous carriers of a GAA expansion, and 53 controls underwent oral and intravenous glucose tolerance tests. β-Cell proportion was quantified in postmortem pancreas sections from 9 unrelated FRDA patients. Using an in vitro disease model, we studied how frataxin deficiency affects β-cell function and survival. Results: FRDA patients had increased abdominal fat and were insulin resistant. This was not compensated for by increased insulin secretion, resulting in a markedly reduced disposition index, indicative of pancreatic β-cell failure. Loss of glucose tolerance was driven by β-cell dysfunction, which correlated with abdominal fatness. In postmortem pancreas sections, pancreatic islets of FRDA patients had a lower β-cell content. RNA interference-mediated frataxin knockdown impaired glucose-stimulated insulin secretion and induced apoptosis in rat β cells and human islets. Frataxin deficiency sensitized β cells to oleate-induced and endoplasmic reticulum stress-induced apoptosis, which could be prevented by the incretins glucagon-like peptide-1 and glucose-dependent insulinotropic polypeptide. Interpretation: Pancreatic β-cell dysfunction is central to diabetes development in FRDA as a result of mitochondrial dysfunction and higher sensitivity to metabolic and endoplasmic reticulum stress-induced β-cell death. ANN NEUROL 2012;72:971-982 [ABSTRACT FROM AUTHOR]

Objective: Cytokines contribute to pancreatic beta-cell death in type 1 diabetes. This effect is mediated by complex gene networks that remain to be characterized. We presently utilized array analysis to define the global expression pattern of genes, including spliced variants, modified by the cytokines interleukin (IL)-1beta + interferon (IFN)-gamma and tumor necrosis factor (TNF)-alpha + IFN-gamma in primary rat beta-cells.Research Design and Methods: Fluorescence-activated cell sorter-purified rat beta-cells were exposed to IL-1beta + IFN-gamma or TNF-alpha + IFN-gamma for 6 or 24 h, and global gene expression was analyzed by microarray. Key results were confirmed by RT-PCR, and small-interfering RNAs were used to investigate the mechanistic role of novel and relevant transcription factors identified by pathway analysis. RESULTS Nearly 16,000 transcripts were detected as present in beta-cells, with temporal differences in the number of genes modulated by IL-1beta + IFNgamma or TNF-alpha + IFN-gamma. These cytokine combinations induced differential expression of inflammatory response genes, which is related to differential induction of IFN regulatory factor-7. Both treatments decreased the expression of genes involved in the maintenance of beta-cell phenotype and growth/regeneration. Cytokines induced hypoxia-inducible factor-alpha, which in this context has a proapoptotic role. Cytokines also modified the expression of >20 genes involved in RNA splicing, and exon array analysis showed cytokine-induced changes in alternative splicing of >50% of the cytokine-modified genes.Conclusions: The present study doubles the number of known genes expressed in primary beta-cells, modified or not by cytokines, and indicates the biological role for several novel cytokine-modified pathways in beta-cells. It also shows that cytokines modify alternative splicing in beta-cells, opening a new avenue of research for the field. [ABSTRACT FROM AUTHOR]

OBJECTIVE--Cytokines contribute to pancreatic β-cell death in type 1 diabetes. This effect is mediated by complex gene networks that remain to be characterized. We presently utilized array analysis to define the global expression pattern of genes, including spliced variants, modified by the cytokines interleukin (IL)-1β + interferon (IFN)-γ and tumor necrosis factor (TNF)-α + IFN-γ in primary rat β-cells. RESEARCH DESIGN AND METHODS--Fluorescence-activated cell sorter-purified rat β-cells were exposed to IL-1β + IFN-γ or TNF-α + IFN-γ for 6 or 24 h, and global gene expression was analyzed by microarray. Key results were confirmed by RT-PCR, and small-interfering RNAs were used to investigate the mechanistic role of novel and relevant transcription factors identified by pathway analysis. RESULTS--Nearly 16,000 transcripts were detected as present in β-cells, with temporal differences in the number of genes modulated by IL-1β + IFNγ or TNF-α + IFN-γ. These cytokine combinations induced differential expression of inflammatory response genes, which is related to differential induction of IFN regulatory factor-7. Both treatments decreased the expression of genes involved in the maintenance of β-cell phenotype and growth/regeneration. Cytokines induced hypoxia-inducible factor-α, which in this context has a proapoptotic role. Cytokines also modified the expression of >20 genes involved in RNA splicing, and exon array analysis showed cytokine-induced changes in alternative splicing of >50% of the cytokine-modified genes. CONCLUSIONS--The present study doubles the number of known genes expressed in primary β-cells, modified or not by cytokines, and indicates the biological role for several novel cytokine-modified pathways in β-cells. It also shows that cytokines modify alternative splicing in β-cells, opening a new avenue of research for the field. Diabetes 59:358-374, 2010 [ABSTRACT FROM AUTHOR]

Objective: Chronic exposure of pancreatic beta-cells to saturated free fatty acids (FFAs) causes endoplasmic reticulum (ER) stress and apoptosis and may contribute to beta-cell loss in type 2 diabetes. Here, we evaluated the molecular mechanisms involved in the protection of beta-cells from lipotoxic ER stress by glucagon-like peptide (GLP)-1 agonists utilized in the treatment of type 2 diabetes.Research Design and Methods: INS-1E or fluorescence-activated cell sorter-purified primary rat beta-cells were exposed to oleate or palmitate with or without the GLP-1 agonist exendin-4 or forskolin. Cyclopiazonic acid was used as a synthetic ER stressor, while the activating transcription factor 4-C/EBP homologous protein branch was selectively activated with salubrinal. The ER stress signaling pathways modulated by GLP-1 agonists were studied by real-time PCR and Western blot. Knockdown by RNA interference was used to identify mediators of the antiapoptotic GLP-1 effects in the ER stress response and downstream mitochondrial cell death mechanisms.Results: Exendin-4 and forskolin protected beta-cells against FFAs via the induction of the ER chaperone BiP and the antiapoptotic protein JunB that mediate beta-cell survival under lipotoxic conditions. On the other hand, exendin-4 and forskolin protected against synthetic ER stressors by inactivating caspase 12 and upregulating Bcl-2 and X-chromosome-linked inhibitor of apoptosis protein that inhibit mitochondrial apoptosis.Conclusions: These observations suggest that GLP-1 agonists increase in a context-dependent way the beta-cell defense mechanisms against different pathways involved in ER stress-induced apoptosis. The identification of the pathways modulated by GLP-1 agonists allows for targeted approaches to alleviate beta-cell ER stress in diabetes. [ABSTRACT FROM AUTHOR]

OBJECTIVE--Chronic exposure of pancreatic β-cells to saturated free fatty acids (FFAs) causes endoplasmic reticulum (ER) stress and apoptosis and may contribute to β-cell loss in type 2 diabetes. Here, we evaluated the molecular mechanisms involved in the protection of B-cells from lipotoxic ER stress by glucagon-like peptide (GLP)-1 agonists utilized in the treatment of type 2 diabetes. RESEARCH DESIGN AND METHODS--INS-1E or fluorescence-activated cell sorter-purified primary rat β-cells were exposed to oleate or palmitate with or without the GLP-1 agonist exendin-4 or forskolin. Cyclopiazonic acid was used as a synthetic ER stressor, while the activating transcription factor 4-C/EBP homologous protein branch was selectively activated with salubrinal. The ER stress signaling pathways modulated by GLP-1 agonists were studied by real-time PCR and Western blot. Knockdown by RNA interference was used to identify mediators of the antiapoptotic GLP-1 effects in the ER stress response and downstream mitochondrial cell death mechanisms. RESULTS--Exendin-4 and forskolin protected β-cells against FFAs via the induction of the ER chaperone BiP and the antiapoptotic protein JunB that mediate β-cell survival under lipotoxic conditions. On the other hand, exendin-4 and forskolin protected against synthetic ER stressors by inactivating caspase 12 and upregulating Bcl-2 and X-chromosome-linked inhibitor of apoptosis protein that inhibit mitochondrial apoptosis. CONCLUSIONS--These observations suggest that GLP-1 agonists increase in a context-dependent way the β-cell defense mechanisms against different pathways involved in ER stress-induced apoptosis. The identification of the pathways modulated by GLP-1 agonists allows for targeted approaches to alleviate β-cell ER stress in diabetes. Diabetes 58:2851-2862, 2009 [ABSTRACT FROM AUTHOR]

The UPR (unfolded protein response) or ER (endoplasmic reticulum) stress response was first described 20 years ago. The field of ER stress has expanded tremendously since, moving from basic biology in yeast to human neurodegenerative, inflammatory, cardiovascular and neoplastic diseases. The ER stress response has also been implicated in diabetes development, affecting both insulin production by pancreatic β-cells and insulin sensitivity in peripheral tissues. In the present mini-review, we focus on recent progress in the field of ER stress in pancreatic β-cells. Recent advances in the understanding of lipotoxic ER stress and β-cell recovery from ER stress are discussed. [ABSTRACT FROM AUTHOR]

OBJECTIVE--Exposure of β-cells to inflammatory cytokines leads to apoptotic cell death through the activation of gene networks under the control of specific transcription factors, such as interferon-γ-induced signal transducer and activator of transcription (STAT)-1. We previously demonstrated that β-cells lacking STAT-1 are resistant to cytokine-induced cell death in vitro. The aim of this study was to investigate the effect of STAT-1 elimination on immune-mediated β-cell destruction in vivo. RESEARCH DESIGN AND METHODS--Multiple low-dose streptozotocin (STZ) was given to C57BL/6 mice after syngeneic STAT-1[sup -/-] or wild-type islet transplantation. STAT-1[sup -/-] and wild-type islets were also transplanted in alloxan-diabetic BALB/c and spontaneously diabetic nonobese diabetic (NOD) mice. Additionally, mice were treated with interleukin (IL)-1 blockade (IL-1 receptor antagonist [IL-1ra]) and low-dose T-cell suppression (cyclosporine A [CsA]). RESULTS--When exposed to multiple low-dose STZ in an immune-competent host, STAT-1[sup -/-] islets were more resistant to destruction than wild-type islets (28 vs. 100% diabetes incidence, P ≤ 0.05). STAT-1 deletion also protected allogeneic islet grafts against primary nonfunction in autoimmune NOD mice (0 vs. 17% using wild-type islets). However, no difference in survival time was observed. Additionally, treating recipients with IL-1ra and CsA prolonged graft survival in chemically diabetic BALB/c mice, whereas no difference was seen between STAT-1[sup -/-] and C57BL/6 grafts. CONCLUSIONS--These data indicate that STAT-1 is a key player in immune-mediated early [β-cell dysfunction and death. When considering the many effector mechanisms contributing to β-cell death following islet transplantation, multiple combined interventions will be needed for prolongation of β-cell survival in the autoimmune context of type 1 diabetes. Diabetes 56: 2169-2173, 2007 [ABSTRACT FROM AUTHOR]

beta-cells under immune attack are destroyed by the aberrant activation of key intracellular signaling cascades. The aim of the present study was to evaluate the contribution of the signal transducer and activator of transcription (STAT)-1 pathway for beta-cell apoptosis by studying the sensitivity of beta-cells from STAT-1 knockout (-/-) mice to immune-mediated cell death in vitro and in vivo. Whole islets from STAT-1-/- mice were completely resistant to interferon (IFN)-gamma (studied in combination with interleukin [IL]-1beta)-mediated cell death (92 +/- 4% viable cells in STAT-1-/- mice vs. 56 +/- 3% viable cells in wild-type controls, P < or = 0.001) and had preserved insulin release after exposure to IL-1beta and IFN-gamma. Moreover, analysis of cell death in cytokine-exposed purified beta-cells confirmed that protection was due to absence of STAT-1 in the beta-cells themselves. Deficiency of STAT-1 in islets completely prevented cytokine-induced upregulation of IL-15, interferon inducible protein 10, and inducible nitric oxide synthase transcription but did not interfere with monocyte chemoattractant protein 1 and macrophage inflammatory protein 3alpha expression. In vivo, STAT-1-/- mice were partially resistant to development of diabetes after multiple low-dose streptozotocin injections as reflected by mean blood glucose at 12 days after first injection (159 +/- 28 vs. 283 +/- 81 mg/dl in wild-type controls, P < or = 0.05) and diabetes incidence at the end of the follow-up period (39 vs. 73% in wild-type controls, P < or = 0.05). In conclusion, the present results indicate that STAT-1 is a crucial transcription factor in the process of IFN-gamma-mediated beta-cell death and the subsequent development of immune-mediated diabetes. [ABSTRACT FROM AUTHOR]

β-Cells under immune attack are destroyed by the aberrant activation of key intracellular signaling cascades. The aim of the present study was to evaluate the contribution of the signal transducer and activator of transcription (STAT)-1 pathway for β-cell apoptosis by studying the sensitivity of β-cells from STAT-1 knockout (-/-) mice to immune-mediated cell death in vitro and in vivo. Whole islets from STAT-1-/- mice were completely resistant to interferon (IFN)-γ (studied in combination with interleukin [IL]-1β)-mediated cell death (92 ± 4% viable cells in STAT-1-/- mice vs. 56 ± 3% viable cells in wild-type controls, P ≤ 0.001) and had preserved insulin release after exposure to IL-1β and IFN-γ. Moreover, analysis of cell death in cytokine-exposed purified β-cells confirmed that protection was due to absence of STAT-1 in the β-cells themselves. Deficiency of STAT-1 in islets completely prevented cytokine-induced upregulation of IL-15, interferon inducible protein 10, and inducible nitric oxide synthase transcription but did not interfere with monocyte chemoattractant protein 1 and macrophage inflammatory protein 3α expression. In vivo, STAT-1-/- mice were partially resistant to development of diabetes after multiple low-dose streptozotocin injections as reflected by mean blood glucose at 12 days after first injection (159 ± 28 vs. 283 ± 81 mg/dl in wild-type controls, P ≤ 0.05) and diabetes incidence at the end of the follow-up period (39 vs. 73% in wild-type controls, P ≤ 0.05). In conclusion, the present results indicate that STAT-1 is a crucial transcription factor in the process of IFN-γ-mediated β-cell death and the subsequent development of immune-mediated diabetes. Diabetes 54:2396-2403, 2005 [ABSTRACT FROM AUTHOR]

Under conditions of sustained hyperglycemia, glycogen accumulates in pancreatic islets, but not so in acinar pancreatic cells. We investigated whether advantage could be taken of such a situation in the perspective of the noninvasive imaging of the endocrine pancreas. Control rats or animals injected with streptozotocin (STZ) were infused with solutions of d-glucose mixed with a tracer amount of d-[U-C]glucose, and the radio-active glycogen content of both liver and pancreas was then measured. After 48 h of infusion, the radio-active glycogen content of the pancreas was 30 times lower in STZ rats than in control animals, coinciding with a 50 times lower insulin content. In the control rats, a sizable labeling of pancreatic glycogen was also recorded when d-[U-C]glucose was infused for only the last 4 h of unlabeled d-glucose infusion; such a labeling was not decreased when the animals were further infused for 1 h with only the unlabeled hexose. Moreover, a pronounced difference in the pancreatic gland and blood radioactive content of control rats was still observed when the hyperglycemic animals were killed only 40 min after the iv injection of d-[U-C]glucose. In STZ rats transplanted with islets and later infused with d-[U-C]glucose, the total radioactive content and radioactive glycogen content were both much higher in the transplanted islets than in the pancreatic gland. These results allow one to define the conditions under which the administration of either 2-deoxy-2-[F]fluoro- d-glucose or C-labeled d-glucose could conceivably be used to favor the selective labeling of the endocrine, as distinct from exocrine, pancreas. [ABSTRACT FROM AUTHOR]

The role currently ascribed to the accumulation of l-arginine in the pancreatic islet B-cell as a determinant of its insulinotropic action was reevaluated by comparing the uptake and the metabolic, ionic, electric, and secretory effects of the cationic amino acid with those of its more positively charged methyl ester in rat pancreatic islets. The response to l-arginine methyl ester differed from that evoked by the unesterified amino acid by a lower uptake and oxidation, lack of inhibitory action on d-glucose metabolism, more severe inhibition of the catabolism of endogenous l-glutamine, inhibition of Ca net uptake, decrease in both Rb outflow from prelabeled islets perifused at normal extracellular Ca concentration and Ca efflux from prelabeled islets perifused in the absence of extracellular Ca, and delayed and lesser insulinotropic action. These findings reinforce the view that the carrier-meadiated entry of l-arginine into the islet B-cells, with resulting depolarization of the plasma membrane, represents the essential mechanism for stimulation of insulin release by this cationic amino acid. [ABSTRACT FROM AUTHOR]

The fate of 6-deoxy-6-[I[iodo- d-glucose (6-DIG), injected intravenously, was compared in control rats and animals that had received streptozotocin and were then treated with insulin or not. In the control rats, the measurement of plasma radioactivity suggested that, after an initial and rapid (up to min 10) distribution phenomenon ( K value: 12.2 × 10 min), the clearance of the iodinated hexose occurred mainly by glomerular filtration ( K value: 0.2 × 10 min). Three minutes after the injection of 6-DIG, the radioactive content of muscle, liver, and pancreas, relative to the paired value in blood, was lower in untreated diabetic rats than in control animals. In the case of muscle and liver, such a difference was no longer observed when the treatment of the diabetic rats by insulin resulted in restoration of normoglycemia. In the pancreas, however, the radioactive content, whether expressed relative to the paired blood or liver value, remained significantly lower in the insulin-treated diabetic rats than in the control animals. No significant difference between control and diabetic rats, in terms of pancreatic radioactivity, was observed 10 min after the injection of 6-DIG. These findings indicate that advantage can be taken from the vastly different time course for 6-DIG uptake by pancreatic acinar and islet cells, as recently documented in vitro, to label preferentially the endocrine moiety of the pancreatic gland shortly after 6-DIG injection. [ABSTRACT FROM AUTHOR]

The apparent distribution space of 6-[I]iodi- d-glucose, recently proposed as a tracer of d-glucose transport, was measured in rat isolated islets, acinar tissue, and pieces of pancreas. While such a space reached a steady-state value corresponding to the HOH volume in pancreatic islets within 5 min, it slowly increased in pieces of pancreas and, even after 60-min incubation, remained lower than the HOH volume. Moreover, the net uptake of 6-deoxy-6-[I]iodo- d-glucose by pancreatic pieces was inhibited by unlabeled 6-deoxy-6-iodo- d-glucose, d-glucose, and cytochalasin B, while being less or not affected by these agents in isolated islets. A preferential labeling of the endocrine, relative to exocrine, moiety of the pancreas was documented both by comparing, after 2 min incubation, the uptake of 6-deoxy-6-[I]iodo- d-glucose by pieces of pancreas from normal vs streptozotocin-injected rats and by comparing the radioactive content of pancreatic islets and acinar tissue obtained from normal rats injected intravenously 3 min before sacrifice with 6-deoxy-6-[I]iodo- d-glucose. It is proposed, therefore, that advantage could conceivably be taken from the vastly different time course for the uptake of selected monosaccharides by pancreatic islets vs acinar cells in the perspective of imaging of the endocrine pancreas by a non invasive method. [ABSTRACT FROM AUTHOR]

Tritiated glibenclamide binds to specific receptors and is internalized in pancreatic insulin-producing B-cells. We investigated, therefore, whether tritiated glibenclamide could be used to preferentially label the endocrine, as distinct from exocrine, pancreas. In isolated rat pancreatic islets, the net uptake of H-glibenclamide reached within 30 min of incubation a near-equilibrium value, corresponding to an apparent distribution space close to three to four times the islet volume. In pieces of pancreas exposed up to 1 h to H-glibenclamide, however, its apparent distribution space progressively increased and, even at the min 60 of incubation, did not exceed a third of the wet weight of the pieces. Yet, no significant difference could be detected between the time course for H-glibenclamide uptake by pancreatic pieces from either control animals or rats injected with streptozotocin a few days before the experiments. Likewise, no significant difference in the paired ratio between the radioactive content of the pancreas and plasma could be found between the control and diabetic rats when examined 1, 5, or 24 h after the IV administration of H-glibenclamide. These findings indicate that the sulfonylurea does not represent a suitable tool for preferential labeling of the endocrine pancreas in the perspective of its imaging by a noninvasive procedure. [ABSTRACT FROM AUTHOR]

Tritiated glibenclamide binds to specific receptors and is internalized in pancreatic insulin-producing B-cells. We investigated, therefore, whether tritiated glibenclamide could be used to preferentially label the endocrine, as distinct from exocrine, pancreas. In isolated rat pancreatic islets, the net uptake of H-glibenclamide reached within 30 min of incubation a near-equilibrium value, corresponding to an apparent distribution space close to three to four times the islet volume. In pieces of pancreas exposed up to 1 h to H-glibenclamide, however, its apparent distribution space progressively increased and, even at the min 60 of incubation, did not exceed a third of the wet weight of the pieces. Yet, no significant difference could be detected between the time course for H-glibenclamide uptake by pancreatic pieces from either control animals or rats injected with streptozotocin a few days before the experiments. Likewise, no significant difference in the paired ratio between the radioactive content of the pancreas and plasma could be found between the control and diabetic rats when examined 1, 5, or 24 h after the IV administration of H-glibenclamide. These findings indicate that the sulfonylurea does not represent a suitable tool for preferential labeling of the endocrine pancreas in the perspective of its imaging by a noninvasive procedure. [ABSTRACT FROM AUTHOR]

The calcium-sensing receptor gene was recently shown to be expressed in rat pancreatic islets and purified islet B-cells. In this study, we investigated the possible role of this receptor in the regulation of insulin release from isolated rat pancreatic islets. Poly- l-arginine (0.2-0.3 μ M) and poly- l-lysine (0.03-0.1 μM) increased insulin output evoked by d-glucose (8.3 m M). This positive effect faded out at higher concentrations of the basic peptides. Likewise, the release of insulin evoked by 8.3 m M d-glucose was significantly lower at high (1.0 m M) than low (0.05-0.1 m M) concentrations of neomycin. The insulinotropic action of Ba in Ca-deprived islets was potentiated in rats pretreated with pertussis toxin. However, Gd inhibited insulin release evoked by dd-glucose in islets prepared from normal rats or animals pretreated with pertussis toxin and incubated in the absence or presence of either theophylline or forskolin. Gd (0.3 m M) failed to affect effluent radioactivity from islets prelabeled with myo-[2-H]inositol and cyclic AMP net production in islets incubated in the absence or presence of forskolin. Gd decreased, however, Ca efflux from prelabeled islets perifused in the absence or presence of extracellular Ca. It is speculated that a negative insulinotropic action mediated by the calcium-sensing receptor, and possibly attributable to a fall in cytosolic Ca concentration, may prevent excessive insulin secretion in pathological situations of hypercalcemia. [ABSTRACT FROM AUTHOR]

Selected esters of D-glucose were recently proposed as tools to provide the sugar to cells, whilst bypassing the carrier system for hexose transport across the plasma membrane. In the present study, α-D-glucose pentaacetate, β-D-glucose pentaacetate, α-D-mannose pentaacetate and, to a lesser extent, 6-O-acetyl-D-glucose, all tested at a 1.7 mM concentration, were found to increase lactate production above basal value in rat erythrocytes. Over 90 min incubation, the increment in lactate production ranged from about 1.2 (α-D-glucose pentaacetate) to 0.6 (6-O-acetyl-D-glucose) μmol/μl of erythrocytes. Little or no change in lactate production was observed in cells exposed to β-L-glucose pentaacetate, α-D-glucose pentaethylsuccinate, α-D-galactose pentaacetate or β-D-galactose pentaacetate. The metabolic response to α-D-glucose pentaacetate was resistant to 3-O-methyl-D-glucose (10-80 mM) which suppressed, however, that evoked by D-glucose. D-mannoheptulose (10 mM) virtually failed to affect the response to D-glucose and its pentaacetate ester. On the contrary, 2-deoxy-D-glucose (10.6 mM) inhibited to the same relative extent (55% decrease) lactate production in erythrocytes exposed to either unesterified D-glucose or α-D-glucose pentaacetate. The tetraacetic ester of 2-deoxy-D-glucose was more efficient than unesterified 2-deoxy-D-glucose in inhibiting lactate production from α-D-glucose pentaacetate. It is proposed that selected esters of saccharides represent useful tools to bypass defects in hexose transport, and to increase their nutritional or therapeutic efficiency. [ABSTRACT FROM AUTHOR]

This study investigates the insulin secretory responsiveness of pancreatic islets to repaglinide in an experimental model of B-cell glucose incompetence. Rats were infused for 2 d with a 1.67 M solution of d-glucose administered at a rate close to 2.8 mL/h. This resulted in a modest rise in glycemia, a severe increase in plasma insulin concentration, an increased sensitivity of B-cells to adrenergic stress, an abnormally high insulin output from isolated islets perifused in the presence of 16.7 m M d-glucose, and a paradoxical transient increase in insulin release from the islets in response to a fall in hexose concentration. The early increment in insulin output evoked by repaglinide, in the presence of 16.7 m M d-glucose, was not lower in the islets from glucose-infused rats than in those from control rats. Moreover, when the meglitinide analog was administered concomitantly with the removal of d-glucose from the perifusion medium, the early response to repaglinide was further increased. Even after 24 min of glucose deprivation, the output of insulin by the islets from glucose-infused rats was higher in the presence of repaglinide than in its absence. These findings indicate that, in this model of B-cell dysfunction, the secretory responsiveness to repaglinide, as distinct from that to glucose, is fully preserved. Therefore, when taken into consideration together with prior observations, these findings argue in support of the use of this insulinotropic agent in the treatment of noninsulin-dependent diabetes. [ABSTRACT FROM AUTHOR]