Regulation of RNA metabolism in relation to insulin production and oxidative metabolism in mouse pancreatic islets in vitro

This study was undertaken to investigate the long-term effects of different substrates, in particular glucose, on the regulation of islet RNA metabolism and the relationship of this regulation to the metabolism and insulin production of the islet B-cell. For this purpose collagenase-isolated mouse islets were used either in the fresh state or after culture for 2 or 5 days in RPMI 1640 plus 10% calf serum supplemented with various test compounds. Islets cultured with 16.7 mM glucose contained more RNA than those cultured with 3.3 mM glucose. Culture of islets in glucose at low concentrations inhibited glucose-stimulated RNA synthesis and this inhibitory effect was reversed by prolonged exposure to high glucose concentrations. Culture with 10 mM leucine and 3.3 mM glucose or with 10 mM 2-ketoisocaproate and 3.3 mM glucose increased the total RNA content of islets as compared to that of islets cultured with 3.3 mM glucose alone. Islets cultured with 5 mM theophylline maintained a high RNA content in the presence of 3.3 mM glucose. Theophylline also increased the islet RNA content when added together with 16.7 mM glucose, as compared to 16.7 mM glucose alone. Theophylline probably exerted this effect by decreasing the rate of RNA degradation. Changes in islet RNA metabolism showed a close correlation to changes in islet total protein biosynthesis, whereas islet (pro)insulin biosynthesis and insulin release exhibited different glucose-dependency patterns. The response of islet oxygen uptake to glucose was similar to that of islet RNA and protein biosynthesis. It is concluded that the RNA content of the pancreatic islets is controlled at the levels of both synthesis and degradation. Glucose stimulates the RNA synthesis and inhibits its degradation. Moreover, the results suggest that regulation of RNA synthesis may be mediated through islet metabolic fluxes and the cAMP system.     

Expression of glucagon-like peptide-1 (GLP-1) receptor and the effect of GLP-1-(7-36) amide on insulin release by pancreatic islets during rat ontogenic development.

The expression of glucagon-like peptide-1 (GLP-1) receptor and the effects of GLP-1-(7-36) amide (t-GLP-1) on glucose metabolism and insulin release by pancreatic islets during rat development were studied. GLP-1 receptor mRNA was found in significant amounts in pancreatic islets from all age groups studied, GLP-1 receptor expression being maximal when pancreatic islets were incubated at physiological glucose concentration (5.5 mM), but decreasing significantly when incubated with either 1.67 or 16.7 mM glucose. Glucose utilization and oxidation by pancreatic islets from fetal and adult rats rose as a function of glucose concentration, always being higher in fetal than in adult islets. The addition of t-GLP-1 to the incubation medium did not modify glucose metabolism but gastric inhibitory polypeptide and glucagon significantly increased glucose utilization by fetal and adult pancreatic islets at 16.7 mM glucose. At this concentration, glucose produced a significant increase in insulin release by the pancreatic islets from 10-day-old and 20-day-old suckling rats and adult rats, whereas those from fetuses showed only a significant increase when glucose was raised from 1.67 to 5.5 mM. t-GLP-1 elicited an increase in insulin release by pancreatic islets from all the experimental groups when the higher glucose concentrations were used. Our findings indicate that GLP-1 receptors and the effect of t-GLP-1 on insulin release are already present in the fetus, and they therefore exclude the possibility that alterations in the action of t-GLP-1 are responsible for the unresponsiveness of pancreatic beta cells to glucose in the fetus, but stimulation of t-GLP-1 release by food ingestion in newborns may partially confer glucose competence on beta cells.     

Cyproheptadine and beta cell function in the rat: insulin secretion from pancreas segments in vitro.

Pancreatic islet cell vacuolization, hyperglycemia, and glucose intolerance develop in rats after oral administration of cyproheptadine (CPH). In order to determine whether these effects were associated with abnormal insulin secretion, pancreas segments from CPH-treated and control rats were compared for their ability to secrete insulin in response to several stimuli. Oral administration of CPH (45 mg/kg/day) to rats for 1 or 8 days inhibited glucose-mediated insulin secretion from pancreas segments obtained 3 and 24 hr after the last dose of the drug. Insulin secretion had returned to normal by 48 hr after drug administration. Intraperitoneal administration of the drug was less effective than oral administration in inhibiting in vitro insulin secretion. Other stimuli for insulin secretion (tolbutamide, glucagon, L-leucine, and dibutyryl 3',5'cyclic AMP), like glucose, were incapable of releasing normal amounts of insulin from pancreas segments of CPH-treated rats. CPH and a metabolite, desmethyl-CPH, inhibited glucose-stimulated insulin secretion when added in vitro to pancreas segments from control rats. This suggests that the inhibition of insulin secretion in pancreas segments taken from animals treated with CPH could be due, at least in part, to the presence of drug and its metabolite in the tissue. A previously observed reduction in the pancreatic content of insulin in CPH-treated rats may also contribute to the abnormal insulin release in animals given the drug.     

Combined effect of adenosine,alpha adrenergic and adenosine antagonists on serum insulin and insulin secretion from rat pancreatic islets

• 1.1. The effect of adenosine separately or in combination with alpha-1 adrenergic antagonist prazosin and alpha-2 adrenergic antagonist yohimbine as well as adenosine antagonists 8-phenyltheophylline and xanthine amine conjugate on glucose-induced insulin secretion from isolated rat pancreatic islets was studied.
• 2.2. Their in vivo effects on serum glucose and insulin levels were also investigated. Adenosine at 10 and 100 μM inhibited significantly, insulin secretion from the isolated islets whereas at 10 mM slightly increased the secretion of insulin.
• 3.3. Prazosin used at 100 μM inhibited insulin secretion. When it combined with adenosine (10 μM) it augmented the inhibitory effect of adenosine.
• 4.4. In vivo prazosin (21 mg/kg bodywt) caused a hyperglycaemia which was accompanied by hypoinsulinaemia.
• 5.5. Concurrent administration of this drug with adenosine neither affect the hyperglycaemic nor the hypoinsulinaemic effects of adenosine.
• 6.6. On the other hand, yohimbine (100 μM) has no effect neither separately nor in combination with adenosine (10 μM) in modulating the inhibitory effect of adenosine on insulin secretion.
• 7.7. When Yohimbine administered at 19.5 mg/kg body wt it did not alter serum glucose but it markedly increased the serum insulin level. Its combined administration with adenosine reduced the hyperglycaemic effect of adenosine with a remarkable increase in serum insulin.
• 8.8. Both adenosine-antagonists were ineffective in alteration of insulin secretion.
• 9.9. However, combination of 8-phenyltheophylline with adenosine (10 μM) totally blocked the inhibitory effect of adenosine on insulin secretion while xanthine amine conjugate failed to prevent this effect of adenosine.
• 10.10. These results indicate that the inhibitory effect of adenosine on insulin secretion is neither mediated via alpha-1 nor alpha-2 adrenoceptors. It might be via activation of specific adenosine receptors on rat islets which are sensitive to blockade by 8-phenyltheophylline.

     

Type 2 diabetes disrupts circadian orchestration of lipid metabolism and membrane fluidity in human pancreatic islets

Recent evidence suggests that circadian clocks ensure temporal orchestration of lipid homeostasis and play a role in pathophysiology of metabolic diseases in humans, including type 2 diabetes (T2D). Nevertheless, circadian regulation of lipid metabolism in human pancreatic islets has not been explored. Employing lipidomic analyses, we conducted temporal profiling in human pancreatic islets derived from 10 nondiabetic (ND) and 6 T2D donors. Among 329 detected lipid species across 8 major lipid classes, 5% exhibited circadian rhythmicity in ND human islets synchronized in vitro. Two-time point-based lipidomic analyses in T2D human islets revealed global and temporal alterations in phospho- and sphingolipids. Key enzymes regulating turnover of sphingolipids were rhythmically expressed in ND islets and exhibited altered levels in ND islets bearing disrupted clocks and in T2D islets. Strikingly, cellular membrane fluidity, measured by a Nile Red derivative NR12S, was reduced in plasma membrane of T2D diabetic human islets, in ND donors’ islets with disrupted circadian clockwork, or treated with sphingolipid pathway modulators. Moreover, inhibiting the glycosphingolipid biosynthesis led to strong reduction of insulin secretion triggered by glucose or KCl, whereas inhibiting earlier steps of de novo ceramide synthesis resulted in milder inhibitory effect on insulin secretion by ND islets. Our data suggest that circadian clocks operative in human pancreatic islets are required for temporal orchestration of lipid homeostasis, and that perturbation of temporal regulation of the islet lipid metabolism upon T2D leads to altered insulin secretion and membrane fluidity. These phenotypes were recapitulated in ND islets bearing disrupted clocks.

A study of circadian regulation of lipid metabolism in human pancreatic islets reveals that Type 2 Diabetes leads to global and temporal alterations of phospholipid and sphingolipid metabolism in islets, resulting in decreased membrane fluidity and insulin secretion defects.     

Dehydroepiandrosterone increases beta-cell mass and improves the glucose-induced insulin secretion by pancreatic islets from aged rats

The effect of dehydroepiandrosterone (DHEA) on pancreatic islet function of aged rats, an animal model with impaired glucose-induced insulin secretion, was investigated. The following parameters were examined: morphological analysis of endocrine pancreata by immunohistochemistry; protein levels of insulin receptor, IRS-1, IRS-2, PI 3-kinase, Akt-1, and Akt-2; and static insulin secretion in isolated pancreatic islets. Pancreatic islets from DHEA-treated rats showed an increased beta-cell mass accompanied by increased Akt-1 protein level but reduced IR, IRS-1, and IRS-2 protein levels and enhanced glucose-stimulated insulin secretion. The present results suggest that DHEA may be a promising drug to prevent diabetes during aging.     

Hexose metabolism in pancreatic islets. Effect of (-)-hydroxycitrate upon fatty acid synthesis and insulin release in glucose-stimulated islets.

Anaplerotic reactions leading to the de novo synthesis of fatty acids, were recently proposed to participate in the coupling of metabolic to secretory events in the process of glucose-stimulated insulin release. In an attempt to validate such a proposal, the effect of (-)-hydroxycitrate upon fatty acid synthesis and insulin release was investigated in glucose-stimulated rat pancreatic islets. The inhibitor of ATP citrate-lyase, when tested in the 1.0-2.0 mM range, failed to affect glucose-stimulated insulin release, but also failed to inhibit the incorporation of 14C-labelled acetyl residues derived from L-[U-14C]leucine into islet lipids. A partial inhibition of fatty acid labelling by 3H2O was only observed in islets incubated for 120 min in the presence of 5.0 mM (-)-hydroxycitrate and absence of CaCl2. These findings suggest that (-)-hydroxycitrate is not, under the present experimental conditions, a useful tool to abolish fatty acid synthesis in intact pancreatic islets.     

Lipoxygenase-generated icosanoids inhibit glucose-induced insulin release from rat islets

Lipoxygenase-pathway metabolites of arachidonic acid are produced in pancreatic islets. They are are implicated in insulin release, since nonselective inhibitors of lipoxygenases inhibit glucose-induced insulin release. We studied the interplay in insulin release between glucose and selected icosanoids formed in 5-, 12- and 15-lipoxygenase pathways. Effects on immunoreactive insulin release of 10(7) to 10(6)-12-(R)-HETE, 12-(S)-HETE, hepoxilin A3, lipoxin B4, LTB4 or LTC4 were tested individually in 30-min incubations of freshly isolated young adult Wistar rat pancreatic islets, in the presence of 5.6 mM or 23 mM glucose. Basal insulin release (at 5.6 mM glucose) was stimulated by LTC4 and hepoxilin A3 (304% and 234% of controls at 5.6 mM glucose alone, respectively), inhibited by 12-(S)-HPETE (56%), and was not affected by 12-(R)-HETE, 12-(S)-HETE, lipoxin B4 or LTB4 (111%, 105%, 106% and 136%, respectively). Insulin release evoked by 23 mM glucose (190-320%) was inhibited (50-145%) by all icosanoids tested, except LTC4 (162%). We conclude that, among the lipoxygenase products tested, only leukotrienes and hepoxilin are candidates for a tonic-stimulatory influence on basal insulin release. Since glucose promotes icosanoid formation in islets, the observed inhibition of glucose-induced insulin release by lipoxygenase products suggests the existence of a negative-feedback system.     

Acute exposure of beta-cells to troglitazone decreases insulin hypersecretion via activating AMPK

Background

It has been recognized that insulin hypersecretion can lead to the development of insulin resistance and type 2 diabetes mellitus. There is substantial evidence demonstrating that thiazolidinediones are able to delay and prevent the progression of pancreatic β-cell dysfunction. However, the mechanism underlying the protective effect of thiazolidinediones on β-cell function remains elusive.

Methods

We synchronously detected the effects of troglitazone on insulin secretion and AMP-activated protein kinase (AMPK) activity under various conditions in isolated rat islets and MIN6 cells.

Results

Long-term exposure to high glucose stimulated insulin hypersecretion and inhibited AMPK activity in rat islets. Troglitazone-suppressed insulin hypersecretion was closely related to the activation of AMPK. This action was most prominent at the moderate concentration of glucose. Glucose-stimulated insulin secretion was decreased by long-term troglitazone treatment, but significantly increased after the drug withdrawal. Compound C, an AMPK inhibitor, reversed troglitazone-suppressed insulin secretion in MIN6 cells and rat islets. Knockdown of AMPKα2 showed a similar result. In MIN6 cells, troglitazone blocked high glucose-closed ATP-sensitive K⁺ (K_ATP) channel and decreased membrane potential, along with increased voltage-dependent potassium channel currents. Troglitazone suppressed intracellular Ca^2 + response to high glucose, which was abolished by treatment with compound C.

Conclusion

Our results suggest that troglitazone provides β-cell “a rest” through activating AMPK and inhibiting insulin hypersecretion, and thus restores its response to glucose.

General significance

These data support that AMPK activation may be an important mechanism for thiazolidinediones preserving β-cell function.     

The effect of sugars on (pro)insulin biosynthesis

Rates of incorporation of [4,5-(3)H]leucine into insulin plus proinsulin, designated ;(pro)insulin', and total protein in rat pancreatic islets were measured. Glucose stimulates rates of total protein and (pro)insulin biosynthesis, but (pro)insulin biosynthesis is stimulated preferentially. Mannose and N-acetylglucosamine also stimulate (pro)insulin and total protein biosynthesis; inosine and dihydroxyacetone stimulate (pro)insulin biosynthesis specifically. Fructose does not stimulate (pro)insulin biosynthesis when tested alone, but does so in the presence of low concentrations of glucose, mannose or N-acetylglucosamine. Many glucose analogues do not stimulate (pro)insulin biosynthesis. Mannoheptulose inhibits synthesis of (pro)insulin and total protein stimulated by glucose or mannose but not by dihydroxyacetone, inosine or N-acetylglucosamine; phloretin (9mum) inhibits N-acetylglucosamine-stimulated (pro)insulin biosynthesis preferentially. The data are in agreement with the view that the same glucose-sensor mechanism may control both insulin release and biosynthesis, and ;substrate-site' model is suggested. The threshold for stimulation of biosynthesis of (pro)insulin and total protein is lower than that found for glucose-stimulated insulin release; moreover the biosynthetic response to an elevation of glucose concentration is slower than that found for insulin release. The physiological implication of these findings is discussed. Caffeine and isobutylmethylxanthine, at concentrations known to increase islet 3':5'-cyclic AMP and potentiate glucose-induced insulin release, were without effect on rates of glucose-stimulated (pro)insulin biosynthesis.     

Cell swelling-induced signaling for insulin secretion bypasses steps involving G proteins and PLA2 and is N-ethylmaleimide insensitive.

BACKGROUND: This study was undertaken to examine putative mechanisms of calcium independent signal transduction pathway of cell swelling-induced insulin secretion. METHODS: The role of phospholipase A(2), G proteins, and soluble N-ethylmaleimide-sensitive-factor attachment protein receptor (SNARE) in insulin secretion induced by 30% hypotonic medium was studied using isolated rat pancreatic islets. RESULTS: In contrast to glucose stimulation, osmotically induced insulin secretion from pancreatic islets was not inhibited by 10 micromol/l bromoenol lactone, an iPLA(2) (Ca(2+) independent phospholipase) inhibitor. Similarly, preincubation of islets for 20 hours with 25 microg/ml mycophenolic acid to inhibit GTP synthesis fully abolished glucose-induced insulin secretion but was without effect on hypotonicity stimulated insulin release. Glucose-induced insulin secretion was prevented by preincubation with 20 nmol/l tetanus toxin (TeTx), a metalloprotease inactivating soluble SNARE. Cell swelling-induced insulin secretion was inhibited by TeTx in the presence of calcium ions but not in calcium depleted medium. The presence of N-ethylmaleimide (NEM, 5 mmol/l, another inhibitor of SNARE proteins) in the medium resulted in high basal insulin secretion and lacking response to glucose stimulation. In contrast, high basal insulin secretion from NEM treated islets further increased after hypotonic stimulation. CONCLUSION: G proteins and iPLA(2) - putative mediators of Ca(2+) independent signaling pathway participate in glucose but not in hypotonicity-induced insulin secretion. Hypotonicity-induced insulin secretion is sensitive to clostridial neurotoxin TeTx but is resistant to NEM.     

Synergistic potent insulin release by combinations of weak secretagogues in pancreatic islets and INS-1 cells

Insulin secretion by the beta cell depends on anaplerosis in which insulin secretagogues are metabolized by mitochondria into molecules that are most likely exported to the extramitochondrial space where they have signaling roles. However, very little is known about the products of anaplerosis. We discovered an experimental paradigm that has begun to provide new information about these products. When various intracellular metabolites were applied in combination to overnight-cultured rat or human pancreatic islets or to INS-1 832/13 cells, they interacted synergistically to strongly stimulate insulin release. When these same metabolites were applied individually to these cells, insulin stimulation was poor. Discerning the contributions of the individual compounds to metabolism has begun to allow us to dissect some of the pathways involved in insulin secretion, which was not possible from studying individual secretagogues. Monomethyl succinate (MMS) combined with a barely stimulatory concentration of alpha-ketoisocaproate (KIC) (2 mm) stimulated insulin release in cultured rat islets 18-fold (versus 21-fold for 16.7 mm glucose). MMS plus low glucose (2 mm) or pyruvate (5 mm) gave 11- and 9-fold stimulations. These agents also potentiated MMS-induced insulin release in fresh islets, and KIC plus MMS gave synergistic insulin release in cultured human islets. In INS-1 cells, neither MMS nor KIC (10 mm) was an insulin secretagogue, but when added together KIC (2 mm) and MMS stimulated insulin release 7-fold (versus 12-fold for glucose). In islets and INS-1 cells, conditions that stimulated insulin release caused large relative increases in acetoacetate, which is a precursor of pathways to short chain acyl-CoAs. Liquid chromatography-tandem mass spectrometry measurements of acetyl-CoA, acetoacetyl-CoA, succinyl-CoA, hydroxymethylglutaryl-CoA, and malonyl-CoA confirmed that they were increased by insulin secretagogues. The results suggest a new mechanism of insulin secretion in which anaplerosis increases short chain acyl-CoAs that have roles in insulin exocytosis.     

Stimulation of proinsulin biosynthesis and insulin release by pyruvate and lactate.

Increasing concentrations of pyruvate failed to stimulate proinsulin biosynthesis and insulin release in freshly isolated islets. Glycolytic flux (3H2O from [5-3H]glucose) decreased by 80-85%, but decarboxylation of [1(-14)C]pyruvate was unaffected in islets tested immediately after alloxan exposure. This strongly suggested that in freshly isolated islets, beta-cells, in relation to other islet cells, hardly contribute to the decarboxylation of pyruvate. Non-alloxan-treated cultured islets decarboxylated 2-2.5 times as much pyruvate as did alloxan-treated islets cultured for 15-18h. Thus the contribution of beta-cells to the metabolism of pyruvate after culturing markedly increased. Concomitantly beta-cells became responsive to pyruvate. At 20mM-pyruvate, release of prelabelled proinsulin and insulin and incorporation of [3H]leucine into proinsulin reached values approximately half of those obtained with 20mM-glucose. Lactate was as effective as pyruvate in inducing responses in cultured islets. The experiments indicate that a critical degree of substrate utilization is necessary for the generation of signals for insulin release and proinsulin biosynthesis.     

Interleukin-1beta-induced nitric oxide production and inhibition of insulin secretion in rat islets of langerhans is dependent upon the nitric oxide synthase cofactor tetrahidrobiopterin

Interleukin (IL)-1 beta-induced inhibition of glucose-stimulated insulin secretion in rat islets of Langerhans is mediated in part by nitric oxide (NO). The NO synthase cofactor 5,6,7,8-tetrahydrobiopterin (BH(4)) supports NO synthesis in many cell types and IL-1 beta-induced NO generation and inhibition of insulin secretion have been previously correlated with intracellular BH(4 )levels in rat insulinoma cells. Using rat islets and the beta cell line BRIN-BD11, we have investigated whether synthesis of BH(4) limits IL-1beta-induced NO generation and inhibition of glucose-induced insulin secretion. IL-1 beta-induced NO generation by BRIN cells and islets was reduced by 2,4-diamino-6-hydroxypyrimidine (DAHP), an inhibitor of de novo BH(4) synthesis. Sepiapterin, the substrate for salvage pathway BH(4) synthesis, reversed this inhibitory effect of DAHP in islets but not BRIN cells. DAHP reversed IL-1 beta-induced inhibition of islet insulin secretion, an effect prevented by sepiapterin. We conclude that BH(4) generation is necessary for IL-1 beta-induced NO generation in rat islets and BRIN cells. While a contribution of non-NO mediators cannot be excluded, our results support the proposal that IL-1 beta-induced, NO-mediated inhibition of insulin secretion in rat islets is dependent on the NOS cofactor BH(4).     

Modulation of the 6-position of benzopyran derivatives and inhibitory effects on the insulin releasing process

The synthesis of different series of 4- and 6-substituted R/S-3,4-dihydro-2,2-dimethyl-2H-1-benzopyrans is described. All of these new benzopyran derivatives were bearing, at the 4-position, a phenylthiourea moiety substituted on the phenyl ring by a meta or a para-electron-withdrawing group such as Cl or CN. The study aimed at exploring the influence of the nature of the substituent at the 6-position in order to develop new benzopyran-type K(ATP) channel activators exhibiting an improved selectivity towards the insulin secreting cells. The original compounds were examined in vitro on rat pancreatic islets (inhibition of insulin release) as well as on rat aorta rings (vasorelaxant effect) and their activity was compared to that of the reference K(ATP) channel activators (±)-cromakalim, (±)-pinacidil, diazoxide and to previously synthesized cromakalim analogues. Structure-activity relationships indicated that the inhibitory effect on the insulin secreting cells was related to the lipophilicity of the molecules and to the size of the substituent located at the 6-position. A marked inhibitory activity on the insulin secretory process was obtained with molecules bearing a bulky tert-butyloxycarbonylamino group at the 6-position (20-23). The latter compounds were found to have the same efficacy on the pancreatic endocrine tissue than some previously described molecules. Lastly, radioisotopic experiments further identified R/S-N-4-chlorophenyl-N'-(6-tert-butyloxycarbonylamino-3,4-dihydro-2,2-dimethyl-2H-1-benzopyran-4-yl)thiourea (23) as a K(ATP) channel opener.     

Changes in glucose-stimulated insulin secretion after long-term treatment of C57BL mice with glibenclamide

An insulin response to increased glucose concentrations could not be found in vivo and in vitro after long-term treatment of C57BL/KsJ and C57BL/6J mice with Glibenclamide. This missing stimulation of insulin secretion was not the result of an exhaustion of the islets or a disturbed (pro)insulin biosynthesis as demonstrated by measurements of insulin content of the islets and by in vitro (pro)insulin biosynthesis experiments. In the presence of glucose (15 mmol/l) theophylline increased the insulin secretion of isolated islets of Glibenclamide-treated mice to values similar to control islets. The insulin response to an i.p. glucose loading was found to be normal in comparison with control mice 1-2 weeks after the Glibenclamide treatment had been finished.     

Inhibition of insulin secretion from rat islets of Langerhans by interleukin-6. An effect distinct from that of interleukin-1.

Glucose-induced insulin secretion from islets cultured in the presence of interleukin-6 (IL-6) for 12-24 h was inhibited to a similar extent as when islets were treated with interleukin-1 beta (IL-1 beta). However, unlike IL-1 beta, IL-6 did not potentiate insulin secretion during an acute (30 min) exposure of islets to the cytokine, nor did it inhibit DNA synthesis during a 24 h culture period. A 12 h pretreatment of islets with tumour necrosis factor-alpha (TNF-alpha) combined with IL-1 beta potentiated the inhibitory effect of IL-1 beta on secretion, such that 20 mM-glucose-induced insulin secretion was abolished. No synergistic inhibition of secretion was observed with TNF-alpha and IL-6. However, IL-1 beta and IL-6 were found to inhibit insulin secretion in an additive manner. These results suggest that IL-6 inhibits insulin secretion in a manner distinct from that of IL-1 beta, and that IL-6 is unlikely to mediate the inhibitory effects of IL-1 beta or TNF-alpha on rat islets of Langerhans.