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Endocrinology Vol. 142, No. 10 4414-4419
Copyright © 2001 by The Endocrine Society

ARTICLES

Hexosamines and Nutrient Excess Induce Leptin Production and Leptin Receptor Activation in Pancreatic Islets and Clonal ß-Cells

Valur Emilsson1, Jacqueline O’Dowd, Anna L. Nolan and Michael A. Cawthorne1

Clore Laboratory, University of Buckingham, Buckingham, United Kingdom MK18 1EG

Address all correspondence and requests for reprints to: Prof. M. A. Cawthorne, Clore Laboratory, University of Buckingham, Hunter Street, Buckingham, United Kingdom MK18 1EG. E-mail: mike.cawthorne{at}buckingham.ac.uk


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 References
 
Activation of the hexosamine biosynthesis pathway leads to insulin resistance in muscle and adipose tissue. In these tissues leptin gene expression is increased by glucosamine. In the present study we found that glucosamine rapidly activates the production of leptin and OB-Rb, which encodes the functional leptin receptor, in both primary pancreatic islets and clonal ß-cells. Secretion of leptin from clonal ß-cells into the medium was detected readily. In addition, the level of the transcripts encoding signal transducer and activator of transcription-3 and -5, both implicated in leptin signal transduction in islet ß-cells, was increased by glucosamine, although to a lesser degree than mRNA levels of leptin and OB-Rb. High glucose (16.7 mM) induced leptin biosynthesis in primary pancreatic islet cells, and the addition of 1 mM palmitate caused an additional incremental effect. The hexosamine-mediated induction of the leptin system in clonal ß-cells was associated with increased responsiveness to leptin, as demonstrated by a 2.6 ± 0.3-fold (P < 0.01) increase in tyrosine phosphorylation of signal transducer and activator of transcription-3. These findings are the first evidence of inducible leptin production in pancreatic islets and suggest that islet cells, like skeletal muscle, demonstrate a linkage between increased nutrient availability and both leptin expression and leptin responsiveness.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 References
 
GLUCOSE METABOLISM THROUGH the hexosamine biosynthesis pathway has been shown to mediate many of the adverse effects of hyperglycemia and has been claimed to act as a nutrient-sensing pathway. The hexosamine biosynthesis pathway uses 2–3% of the fructose-6-phosphate pool and yields uridine diphosphate-N-acetyl hexosamines (UDP-HexNac) as its major end product. Increased carbon flux through this pathway has been implicated in the desensitization of the glucose transport system in hyperglycemic conditions and in the development of insulin resistance (1, 2). Potentially, such effects are exaggerated further by increased concentrations of FFA (3) (Fig. 1Go), as FFA, by blocking glycolysis, increases the fructose-6-phosphate pool. Thus, both hyperglycemia and hyperlipidemia may act synergistically to expand the fructose-6-phosphate pools and thus increase carbon flux into the hexosamine biosynthesis pathway.



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  		Figure 1. Sites of entry of glucose, GlcN, and uridine into the hexosamine pathway and role of FFA. GFA is the rate-limiting enzyme for conversion of fructose-6-phosphate to GlcN-6-P. Inhibition of glycolysis increases the flux through the hexosamine pathway. O-Linked N-acetylglucosamine transferase (OGT) catalyzes the attachment of UDP-GlcNAc to various proteins and transcriptional activation.

 
The intracellular concentration of the first and rate-limiting enzyme for the hexosamine biosynthesis glutamine:fructose-6-phosphate amidotransferase (GFA) correlates with glucose disposal rates in humans (4). Furthermore, transgenic overexpression of GFA in muscle and adipose tissue results in decreased glucose uptake, which is mostly accounted for by disruption in the translocation of the glucose transporter GLUT4 (5, 6). The accumulated end product of the hexosamine pathway, UDP-HexNac, is used for O-linked glycosylation of transcription factors, RNA polymerase, and nuclear pore proteins altering their activity (7, 8, 9) (Fig. 1Go). This results in transcriptional activation of a number of growth factor and cytokine genes in muscle, including leptin (10), that are likely to cause some of the adverse metabolic effects of hyperglycemia and hyperlipidemia. Moreover, as most patients with noninsulin-dependent diabetes mellitus (NIDDM) present with both hyperglycemia and hyperlipidemia (11), and there is a strong inverse correlation between glucose oxidation and lipid oxidation in these patients (3, 12), it is likely that subsequent hyperstimulation of the hexosamine biosynthesis pathway can lead to a vicious cycle by worsening the insulin-resistant state in these patients.

The effects of hexosamine biosynthesis on pancreatic islet function are less well known, although the rate-limiting enzyme GFA is relatively abundant in islet cells (13). Overexpression of GFA specifically in ß-cells has recently been shown to result in hyperinsulinemia and peripheral insulin resistance in mice, but a reduction in insulin mRNA (14).

The obesity gene product leptin is produced predominantly by adipose tissue (15), although the mRNA transcript has also been identified in stomach (16) and placenta (17). Leptin and its functional leptin receptor isoform OB-Rb, a product of the diabetes gene (db) (18, 19), are closely related to class I cytokines and cytokine receptors, which signal through the Janus kinase and signal transducers and activators of transcription (STAT) (20). The leptin system is an effector in islet ß-cell function. Thus, leptin has been shown to directly inhibit insulin production and secretion in islet ß-cells (21, 22) and to stimulate lipid oxidation in islet ß-cells (23, 24). The latter effect suggests that leptin could antagonize hyperlipidemia-mediated lipotoxicity and thus promote islet ß-cell survival. Paradoxically, inappropriate inhibition of insulin secretion and insulin biosynthesis by leptin, when leptin levels increase with obesity (25), could result in the development of glucose intolerance or diabetes, as has been suggested previously (21). However, regardless of which are the dominating effects of leptin on islet ß-cell function, the accumulated evidence suggests that the leptin system is an important component in islet ß-cell physiology and pathophysiology.

In muscle and adipose tissue it has been argued that the hexosamine biosynthetic pathway is a cellular sensor of energy availability and mediates the effect of glucose on the expression of several genes, including leptin (10). In the present study we have examined whether a similar nutrient-sensing pathway might be present in pancreatic islets, leading to an increase in the local concentration of leptin and its activity in this tissue.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 References
 
Cell culture and isolation of pancreatic islets
RINm5F cells were a gift from Dr. S. Islam, (Rolf Luft Center, Karolinska Institute, Stockholm, Sweden). Cells were cultured routinely in 25-cm2 flasks in RPMI 1640 medium (Life Technologies, Inc., Paisley, UK) supplemented with 100 U/ml penicillin, 100 µg/ml streptomycin, 2 mM L-glutamine, 5.6 mM glucose, and 10% FCS at 5% CO2 in a humidified 37 C cabinet. Cells were treated with or without glucosamine (GlcN; 0.1–5 mM; Sigma, Poole, UK) in the fully supplemented medium for the indicated times. Flasks were washed in serum-free (0.1% BSA) RPMI 1640 before exposure to recombinant murine leptin (PeproTech, London, UK) in the same medium to measure phosphorylation of STAT-3. Islets were isolated from the pancreas of fed male Wistar rats (Harlan Olac, Bicester, UK) by collagenase digestion (collagenase type VI, Sigma) at 37 C using Gey and Gey buffer (26) supplemented with 1 mM CaCl2 and 4 mM glucose and equilibrated with 5% CO2-95% O2, pH 7.4. Islets were handpicked using a binocular microscope, and groups of 200 islets were incubated in Gey and Gey buffer supplemented with CaCl2 (1 mM) with or without GlcN (0.1–5 mM), 8.3 or 16.7 mM glucose, or 16.7 mM glucose plus palmitate (1 mM). In some studies the GFA inhibitor, 6 diazole-5-oxo-L-norleucine (20 µM) was also included.

All animals were maintained according to the United Kingdom Home Office Code of Practice for the housing and care of animals. Experimental procedures were approved by the local ethical committee and complied with all national regulations.

RNA analysis
Total RNA was isolated using Tri-Reagent (Amersham Pharmacia Biotech, Little Chalfont, UK) and isopropanol precipitation. RNA was treated with ribonuclease-free deoxyribonuclease I (Life Technologies, Inc.), and the integrity and loading of the RNA were studied by detection scanning of ribosomal RNA bands (28S and 18S) in agarose gels before and after treatment with deoxyribonuclease I. Single stranded cDNA synthesis was performed from approximately 2 µg total RNA using oligo(deoxythymidine)15–18 (Invitrogen, Groningen, Holland) in a first strand synthesis kit (Amersham Pharmacia Biotech), and negative controls were included in which the reverse transcriptase had been heat inactivated. mRNA expression was determined by quantitative PCR as described previously (27) using the following primer sequences: 5'-ACCCCATTCTGAGTTTGTCC-3' and 5'-GTCCAACTGTTGAAGAATGTCC-3' to amplify a 286-bp fragment of leptin (GenBank D49653), 5'-GCTGGATGAAAGGGGACTTG-3' and 5'-GTGACTTCCATACGCAAACC-3' to amplify a 348-bp fragment of OB-R (common to all isoforms) (GenBank U53144), 5'-AACTGATGAAGAGCAAGGGG-3' and 5'-GAGACAGTGAGCTGGGAATG-3' to amplify a 332-bp fragment of OB-Rb (GenBank D84551), 5'-CAGAAAGTGTCCTACAAGGGCG-3' and 5'-CGTTGTTAGACTCCTCCATGTTC-3' to amplify a 239-bp fragment of STAT-3 ({alpha}- and ß-isoforms) (GenBank U06922), 5'-CATCACGGACATCATCTCAGC-3' and 5'-GACATGTTTCTGAAGTGGGCG-3' to amplify a 302-bp fragment common to STAT-5a and STAT-5b (GenBank Z48538), 5'-CTACTGGGGTTCCATTACAG-3' and 5'-CCACGTAATGCTGCACAAG-3' to amplify a 246-bp fragment of cytokine inducible SH2-containing protein (cytokine inhibitory substance) (GenBank AF065161), 5'-CGATTCTACTGGAGTGCCGTA-3' and 5'-GTGCACCAACTTGAGTACACA-3' to amplify a 241-bp fragment of suppressor of cytokine signaling 3 (SOCS-3) (GenBank AJ249240.1), and finally 5'-CTCTTTAATGTCACGCACGAT-3' and 5'-AGTGCTGTGGGTGTAGGTACT-3' to amplify a 534-bp fragment of ß-actin (GenBank J00691). PCR products were then cloned directly into a pCR-TRAP cloning system (GeneHunter Corp., Nashville, TN), and the identity of PCR products was confirmed by sequencing using a ThermoSequenase terminator cycle sequencing kit (Amersham Pharmacia Biotech). PCR products were fractionated in either 5% (Bio-Rad Laboratories, Inc., Hercules, CA) PAGE system or 1.2% agarose, and bands were visualized by ethidium bromide staining. Bands were quantified using an AlphaImager 1200 system (Flowgen, Ashby de la Zouch, UK). Quantification of mRNA expression was performed by quantitative RT-PCR as described previously (27), and the levels of each mRNA species were normalized to ß-actin mRNA to provide information on the relative changes in tissue gene expression. Results are expressed as the mean ± SEM, and statistical significance between groups was determined by unpaired t test (n = 3–5).

Detection of leptin secretion
Clonal ß-cells (RINm5F) were cultivated in serum-free RPMI medium containing 0.1% BSA and 5.6 mM glucose for up to 6 h with or without 3 mM GlcN, and the medium was collected for leptin ELISA (Crystal Chem, Inc., Chicago, IL). The leptin ELISA kit detects leptin in a range of concentrations between 200 and 12,800 pg/ml. Thus, leptin standards were used in the range between 0 and 12,800 pg/ml to provide a leptin standard curve for determinations of the leptin concentration in the sample. Results are expressed as the mean ± SEM (n = 3).

Western blot analysis of leptin-mediated STAT-3 tyrosine phosphorylation
A RIPA buffer [PBS (pH 7.4), 1% IGEPAL, 0.5% SDS, 1 mM Na3VO4, 10 mM NaF, and 10 mM Na2MoO4] and an equal volume of 2x sample-loading buffer [100 mM Tris (pH 6.8), 10% SDS, 20% glycerol, 2% ß-mercaptoethanol, and 0.2% bromophenol blue] were mixed and added to aspirated culture flasks, and the RINm5F cells were lysed. The samples were sonicated with an ultrasonic cell disrupter and finally boiled for 3 min. Equivalent amounts of protein were resolved on a 7.5% SDS polyacrylamide gel (Bio-Rad Laboratories, Inc. Hemel Hempstead, UK) and then transferred to polyvinylidene difluoride membranes (Millipore Corp., Bedford, MA) by electrotransfer blotting. Immunoblotting was performed by blocking in 3% nonfat milk powder, 10 mM Tris (pH 7.5), 100 mM NaCl, 0.1% Tween 20, either N-terminal-directed STAT-3 or anti-phospho-STAT3 Y705 (diluted 1/1000) antibodies (Santa Cruz Biotechnology, Inc., Santa Cruz, CA), and horseradish peroxidase-conjugated antirabbit antibodies (Santa Cruz Biotechnology, Inc.). Enhanced chemiluminescence (Amersham Pharmacia Biotech) was used to develop immunoblots, which were exposed to Biomax ML film (Eastman Kodak Co., Rochester, NY).


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 References
 
The hexosamine biosynthesis pathway induces leptin biosynthesis and leptin secretion in clonal ß-cells
Leptin and the leptin receptor OB-Rb have been implicated in the normal physiology and, under certain circumstances, pathophysiology of the pancreatic ß-cell function (21, 22, 23). RINm5F cells cultured in a medium containing 10% FBS and 5.6 mM glucose were incubated with GlcN (5 mM), uridine (5 mM), or both for 24 h, and mRNA transcripts were quantified using RT-PCR as described previously (27). The leptin transcript was detected readily in GlcN- or uridine-treated cells, but was not detected in the cells cultured in the basal medium or in the presence of both metabolites (Fig. 2AGo). Culture of islets with glucosamine (5 mM) for 48 h led to an increase in the number of dead cells, possibly through ATP depletion. Using a shorter duration of GlcN (5 mM) treatment, we found that the leptin transcript was detected readily after 6 h (Fig. 2BGo). When both glucosamine and uridine were present in the medium, the leptin transcript was detected weakly after 2 h of treatment and was strongly detected after 4 and 6 h (Fig. 2BGo). Similar results were obtained in the ß-cell line INS-1 (data not shown). It is possible that by using a concentration of GlcN in the millimolar range, the cellular UTP concentration is rapidly limited by increased rates of nucleotide-hexosamine formation. By the addition of uridine, this limitation is overcome, resulting in more rapid accumulation of the end-product UDP-HexNac. To establish whether leptin was secreted from the clonal ß-cells after 6-h treatment with GlcN, we measured leptin levels by ELISA in cells cultured in serum-free medium with 5 mM GlcN for 6 h. Leptin was not detected in the medium between 0 and 4 h of GlcN treatment, but after 6 h of treatment leptin was detected readily at a concentration of 3 ng/ml (Fig. 3Go).



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  		Figure 2. Hexosamine-mediated induction of leptin transcription in clonal ß-cells RINm5F. A, Cells were cultured for 24 h with or without the addition of GlcN (5 mM), uridine (5 mM), or GlcN plus uridine, and leptin mRNA was detected using RT-PCR (37 cycles). B, Cells were cultured with GlcN (5 mM) or GlcN plus uridine (5 mM) for shorter durations of incubation, and leptin mRNA was detected with RT-PCR (37 cycles).

 


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  		Figure 3. Hexosamine-induced leptin secretion. Clonal RINm5F ß-cells were cultured in serum-free medium with GlcN (5 mM) for 6 h. Leptin was detected by a leptin ELISA kit (see Materials and Methods). Results are expressed as the mean ± SEM (n = 3).

 
Hexosamine stimulates levels of the transcripts encoding OB-R and STAT
We examined whether GlcN induced other molecules involved in the leptin action cascade or molecules that have been implicated in the physiology or pathophysiology of the islet ß-cell. When RINm5F cells were cultured with 3 mM GlcN for 48 h, the transcripts encoding the common leptin receptor OB-R increased by 4.8-fold (from 0.64 ± 0.07 to 3.10 ± 0.25; P < 0.001; n = 5), and the functional leptin receptor isoform OB-Rb increased by 4.6-fold (from 1.40 ± 0.12 to 6.40 ± 0.50; P < 0.002; n = 5; Fig. 4Go). The similar degree of stimulation of the common OB-R and the alternatively transcribed OB-Rb isoform implies regulation at the level of transcription initiation.



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  		Figure 4. GlcN-induced gene expression of leptin and the functional leptin receptor OB-Rb mRNA in rat isolated pancreatic islets. Groups of 200 islets were incubated with 5 mM GlcN for 48 h, and RNA was detected with RT-PCR.

 
Transcripts encoding STAT-3 and STAT-5, both of which have been implicated in mediating leptin signal transduction in islet ß-cells (28, 29), were increased moderately, yet significantly, 1.6-fold (from 3.90 ± 0.4 to 6.10 ± 0.58; P < 0.02; n = 5) and 1.5-fold (from 1.10 ± 0.11 to 1.68 ± 0.15; P < 0.02; n = 5), respectively, by treatment with GlcN (Table 1Go). The suppressors of cytokine signaling SOCS-3 and CIS have been implicated in terminating or dampening leptin-mediated signal transduction (30, 31). The transcripts encoding these factors were not affected by treatment with GlcN (Fig. 4Go). We also determined the effects of GlcN treatment on the levels of other molecules that have been implicated in islet ß-cell function, such as the transcripts encoding uncoupling protein-2 (32), caspase-3 (33), and the CCAAT/enhancer-binding protein ß (34). The levels of these transcripts were not altered by the 48-h GlcN (3 mM) treatment (data not shown).


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  		Table 1. The effect of glucosamine on mRNA species encoding leptin receptor, STAT, and SOCS in RINm5F cells

 
The hexosamine pathway induces leptin and OB-Rb expression in primary rat islet cells
The results from our studies of clonal ß-cells suggested that increased hexosamine metabolism might induce leptin and OB-Rb production in primary pancreatic islet cells. Groups of rat isolated islets were cultured in medium containing 10% FBS and 8.3 mM glucose with or without 5 mM GlcN for 48 h. The leptin mRNA was not detected in islets cultured in the basal medium, but in the GlcN-treated islets the leptin transcript was detected readily, and production of OB-Rb mRNA was stimulated by GlcN (Fig. 4Go). These results suggest that increased hexosamine metabolism has similar effects on the primary islets cell, as demonstrated for the clonal ß-cell RINm5F (and INS-1; data not shown). Isolated rat islets incubated with a high concentration of glucose [16.7 vs. 8.3 mM (control)] for 48 h contained readily detectable levels of leptin transcript (Fig. 5Go). The addition of 1 mM palmitate to the high glucose medium resulted in an additional increase (2.2-fold for palmitate vs. high glucose alone; P < 0.01; n = 3; relative to ß-actin) in leptin mRNA levels (Fig. 5). The inductive effect of 16.7 mM glucose on both leptin and OB-Rb transcripts was prevented by inclusion of the GFA inhibitor, 6-diazole-5-oxo-L-norleucine (20 µM), during the 48-h incubation. This GFA inhibitor did not affect the increase in leptin or OB-Rb transcripts caused by glucosamine (5 mM) in the presence of either 8.3 or 16.7 mM glucose. These results suggest that the hexosamine biosynthetic pathway is responsible for the increase in leptin and OB-Rb expression in rat islets and that chronic conditions of hyperglycemia and/or caloric excess can induce leptin production in rat pancreatic islets.



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  		Figure 5. High glucose (16.7 mM) and palmitate (1 mM) induced the expression of leptin mRNA in rat isolated pancreatic islets incubated for 48 h.

 
Leptin-mediated phosphorylation of STAT-3 is elevated in clonal ß-cells exposed to hexosamine
It has been shown that leptin activates the Janus kinase-2/STAT-3 signaling pathway in primary islet cells and clonal insulin-secreting cells (28). It was therefore reasoned that if increased hexosamine metabolism in ß-cells resulted in increased production of leptin and its receptor, then these same cells would be more sensitive to leptin by phosphorylation and activation of STAT-3. Clonal ß-cells were precultured in a medium containing 10% FBS and 5.6 mM glucose with or without 3 mM GlcN for 48 h. Cells were then washed in serum-free medium containing 0.1% BSA and cultured in the same medium containing 100 nM leptin for periods that were based on previously optimized time points for STAT-3 phosphorylation (28). Crude cell extracts were then fractionated in PAGE system and probed with anti-phospho-Y705 STAT-3 antibody. Leptin produced a time-dependent tyrosine phosphorylation of a single immunoreactive STAT-3 species at approximately 90 kDa, consistent with STAT-3{alpha} (Fig. 6AGo). When islet cells were preexposed to GlcN, they responded to leptin by increased tyrosine phosphorylation of STAT-3 at all time points except zero compared with cells that have not been treated previously with GlcN (Fig. 6BGo). The increase in STAT-3{alpha} tyrosine phosphorylation at 15 min relative to that at time zero normalized to the level of total STAT-3, detected with an N-terminal-directed STAT-3 antibody (data not shown) was 2.6 ± 0.3-fold (P < 0.01; n = 3) greater in cells previously exposed to glucosamine than in cells that had not been exposed to the metabolite.



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  		Figure 6. A, Leptin (100 nM)-induced time-dependent tyrosine phosphorylation of STAT-3{alpha} in cells that had been precultured in a medium containing GlcN. B, Representative blots showing comparison of leptin (100 nM)-mediated STAT-3 phosphorylation of cells precultured with or without GlcN.

 

   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
References
 
Hyperglycemia is the major cause of chronic complications of diabetes (35). Hyperglycemia has been strongly implicated in the mediation of toxic effects, termed glucose toxicity, on cellular function. However, some of the adverse effects of hyperglycemia might be mediated through routes of normal regulatory pathways (reviewed in Ref. 4). One potential pathway that has been suggested is the nutrient-sensing hexosamine biosynthesis pathway, which might mediate many of the adverse effects of hyperglycemia and caloric excess on insulin sensitivity during the development of NIDDM (4). Using immunohistochemical studies, GFA, the key enzyme in hexosamine biosynthesis, was detected readily in adipose and muscle tissues, but was also detected, albeit at somewhat lower levels, in endocrine tissues such as pancreatic islet cells (13). Thus, GFA is expressed in all the tissues that are involved in development of NIDDM. Moreover, the enzyme O-linked N-acetylglucosamine transferase, which catalyzes the attachment of the end-product UDP-HexNac to various proteins, thus altering their activity, is expressed abundantly in pancreatic islets (36). The accumulated data suggest that the hexosamine pathway might play an important role in the glucose-sensing mechanism of the pancreas.

Leptin has been shown to have peripheral metabolic and proliferative actions in addition to its interaction with satiety centers in the hypothalamus to inhibit food intake (37). In islet cells it has been shown to directly inhibit insulin biosynthesis and secretion (21, 22), to stimulate lipid oxidation in islet ß-cells (23, 24), and to stimulate the proliferation of fetal islets and clonal ß-cells (38, 39). Some of these effects are associated with the activation of STAT-3 and STAT-5 proteins in pancreatic islet ß-cells (28, 29). Recently, leptin expression has been shown to be induced in skeletal muscle and adipose tissues by hexosamine (10), thus linking the production of leptin to increased nutrient availability and possibly to the development of insulin resistance. In the present paper we demonstrate that hexosamine induces leptin and OB-Rb production in clonal ß-cells, which was associated with elevated response to leptin by increased phosphorylation of STAT-3{alpha}. The induced production of leptin and OB-Rb in clonal ß-cells by hexosamine was corroborated in native islet cells treated with high glucose concentrations and excess FFA, which had an incremental effect. Moreover, the effect of high glucose was blocked by the GFA inhibitor 6-diazole-5-oxo-L-norleucine, indicating a role for the hexosamine biosynthetic pathway. These findings present the first evidence of inducible leptin production in islet ß-cells and suggest that islet cells, like skeletal muscle and adipose tissues, demonstrate a linkage between increased nutrient availability and increased leptin responsiveness. Moreover, the induction of leptin and its reception in pancreatic islet cells by nutrient excess suggest the presence of a local feedback loop for the leptin system in this tissue. Given the known effects of leptin on the pancreatic islet, such a feedback loop might serve a role in the regulation of lipid metabolism, insulin production, and/or proliferation of the ß-cell during conditions of caloric excess. It is, however, as yet unclear whether such an effect might be a part of a ß-cell-destructive or ß-cell-protective mechanism during the pathological conditions of hyperglycemia and elevated fatty acid exposure.


   Acknowledgments
 
We are grateful for the assistance of David Hislop with the preparation of the figures, and to Julie Cakebread for help with the preparation of this manuscript.


   Footnotes
 
1 Present address, Decode Genetics, Inc., Lynghals 1, IS-110 Reykjavik, Iceland. Back

Abbreviations: GFA, Glutamine:fructose-6-phosphate amidotransferase; GlcN, glucosamine; NIDDM, noninsulin-dependent diabetes mellitus; OB-R, leptin receptor; STAT, signal transducer and activator of transcription; UDP-HexNac, uridine diphosphate-N-acetyl hexosamines.

Received January 23, 2001.

Accepted for publication June 11, 2001.


   References
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 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 References
 

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