Prolonged elevation of insulin content in the unicellular tetrahymena after insulin treatment: induction of insulin production or storage?

In the unicellular organism, Tetrahymena, the first encounter with an exogeneously given hormone results in hormonal imprinting. This causes an increase of the binding capacity of receptors and the production of the appropriate hormone in the progeny generations of the treated cell. In the present experiments the quantity (using radioimmunoassay) and localization (using confocal laser scanning microscopy) of the immunologically insulin‐like material (hereafter insulin) were studied for 10 days after 4 h or 24 h 10⁻⁶ m insulin treatment (hormonal imprinting). Forty‐eight hours after both insulin treatments a high quantity of insulin was present in the cells. This value was also significantly increased after 96 h. After 8 days the difference to the control was significant only in the 24 h treated group. Confocal microscopy (using antibody to pig insulin) localized insulin in the cell body. The oral field contained extremely high quantities of the endogeneous hormone. Insulin treatment (after 48 and 96 h) caused an elevation of insulin content in general, and specific accumulation in the posterior sections of the cell, around the nucleus and in the periphery were observed. Ten days after both treatments only the peripheral region of the cell body and the ciliary row contained more insulin than the control. This means that after insulin treatment the quantity of insulin increases for a lengthy time period which is followed by the expression of insulin in the peripheral region. Insulin contained by Tetrahymena 48 h after imprinting stimulated glucose uptake of rat diaphragm. Copyright © 1999 John Wiley & Sons, Ltd.     

How α-crystallin prevents the aggregation of insulin

Using steady-state, polarized, and phase-modulation fluorometry, the dithiothreitol-induced denaturation of insulin and formation of its complex with alpha-crystallin in solution were studied. Prevention of the aggregation of insulin by alpha-crystallin is due to formation of chaperone complexes, i.e. interaction of chains of the denatured insulin with alpha-crystallin. The conformational changes in alpha-crystallin that occur during complex formation are rather small. It is unlikely that N-termini are directly involved in the complex formation. The 8-anilino-1-naphthalenesulfonate (ANS) is not sensitive to the complex formation. ANS emits mainly from alpha-crystallin monomers, dimers, and tetramers, but not from oligomers or aggregates. The possibility of highly sensitive detection of aggregates by light scattering using a spectrofluorometer with crossed monochromators is demonstrated.     

Enhanced modelling of the glucose–insulin system and its applications in insulin therapies

It is well known that Michaelis–Menten kinetics is suitable for the response function in chemical reaction, when the reaction rate does not increase indefinitely when an excess of resource is available. However, the existing models for insulin therapies assume that the response function of insulin clearance is proportional to the insulin concentration. In this paper, we propose a new model for insulin therapy for both type 1 and type 2 diabetes mellitus, in which the insulin degradation rate assumes Michaelis–Menten kinetics. Our analysis shows that it is possible to mimic pancreatic insulin secretion by exogenous insulin infusions, and our numerical simulations provide clinical strategies for insulin–administration practices.     

Maintenance of the postabsorptive plasma glucose concentration: insulin or insulin plus glucagon?

The prevalent view is that the postabsorptive plasma glucose concentration is maintained within the physiological range by the interplay of the glucose-lowering action of insulin and the glucose-raising action of glucagon. It is supported by a body of evidence derived from studies of suppression of glucagon (and insulin, among other effects) with somatostatin in animals and humans, immunoneutralization of glucagon, defective glucagon synthesis, diverse mutations, and absent or reduced glucagon receptors in animals and glucagon antagonists in cells, animals, and humans. Many of these studies are open to alternative interpretations, and some lead to seemingly contradictory conclusions. For example, immunoneutralization of glucagon lowered plasma glucose concentrations in rabbits, but administration of a glucagon antagonist did not lower plasma glucose concentrations in healthy humans. Evidence that the glycemic threshold for glucagon secretion, unlike that for insulin secretion, lies below the physiological range, and the finding that selective suppression of insulin secretion without stimulation of glucagon secretion raises fasting plasma glucose concentrations in humans underscore the primacy of insulin in the regulation of the postabsorptive plasma glucose concentration and challenge the prevalent view. The alternative view is that the postabsorptive plasma glucose concentration is maintained within the physiological range by insulin alone, specifically regulated increments and decrements in insulin, and the resulting decrements and increments in endogenous glucose production, respectively, and glucagon becomes relevant only when glucose levels drift below the physiological range. Although the balance of evidence suggests that glucagon is involved in the maintenance of euglycemia, more definitive evidence is needed, particularly in humans.     

Is the insulin resistance of patients with noninsulin-dependent diabetes mellitus secondary to insulin deficiency?

Defects in both insulin secretion and action have been documented in patients with noninsulin-dependent diabetes mellitus (NIDDM), leading to the suggestion that both fasting hyperglycemia and insulin resistance in NIDDM are secondary to insulin deficiency. In order to test this hypothesis, insulin secretion (plasma insulin response to oral glucose) and insulin action (insulin clamp) were determined in 25 patients with NIDDM. The results documented relationships between incremental plasma insulin response to glucose and degree of fasting hyperglycemia (r = -.045, P less than 0.05) and insulin-stimulated glucose utilization (r = 0.25, P = NS). These data indicate that differences in insulin secretory response accounted for only approximately 20% of the variance in fasting plasma glucose level and 6% of the variance in insulin resistance in NIDDM. Thus, differences in insulin-secretory response contribute modestly to magnitude of glycemia, and not at all to variations in insulin resistance in NIDDM, permitting rejection of the hypothesis that insulin resistance is secondary to insulin deficiency.     

Can insulin antibodies of diabetic patients distinguish human insulin from porcine insulin?

We examined antisera from patients treated with bovine-porcine mixture (hereafter referred to as bovine/porcine), porcine or human insulin, and compared their binding affinities to human insulin with those to porcine insulin. Patients treated with bovine/porcine insulin developed antisera with a higher affinity to porcine insulin compared with that to human insulin in five of nineteen cases. Furthermore, three of these five antisera had a comparable affinity to bovine and porcine insulin and appeared to recognize the amino acid residue at B-30. Treatment with porcine or human insulin, on the other hand, did not result in any significant difference in the affinity to porcine and human insulin in twenty-three patients. These results indicate the significant role of B-30 amino acid residue as an antigenic determinant, and suggest that the amino acid sequence of the A chain of bovine insulin may contribute to the development of antibody recognizing B-30 amino acid residue.     

Does the insulin-mimetic action of vanadate involve insulin receptor kinase?

Effects of vanadate administration on the insulin receptor status in liver were examined in streptozotocin-induced diabetic rats. Diabetic rats were characterized by hyperglycemia (4-fold increase), hypoinsulinemia (81% decrease) and a significant (P<0.01) increase in hepatic insulin receptor numbers. Autophosphorylation of the subunit of insulin receptor and its tyrosine kinase activity towards the synthetic peptide (poly glut₄tyr₁) decreased by approximately 60% as a result of diabetes. After chronic treatment of these rats with sodium orthovanadate, the plasma glucose levels were normalized to near control values with the hypoinsulinemia remaining unaltered. The insulin-stimulated phosphorylation of the subunit increased significantly (P<0.001) in diabetic rats after treatment with vanadate. However, the improvement in the tyrosine kinase activity was marginal.In vitro, vanadate prevented the dephosphorylation of the phosphorylated insulin receptor and increased its tyrosine kinase activity in the absence as well as presence of insulin. The findings of this study further support the view that insulin receptor is one of the sites involved in the insulin-mimetic actions of vanadate.     

A kinetic core model of the glucose-stimulated insulin secretion network of pancreatic β cells

The construction and characterization of a core kinetic model of the glucose-stimulated insulin secretion system (GSIS) in pancreatic β cells is described. The model consists of 44 enzymatic reactions, 59 metabolic state variables, and 272 parameters. It integrates five subsystems: glycolysis, the TCA cycle, the respiratory chain, NADH shuttles, and the pyruvate cycle. It also takes into account compartmentalization of the reactions in the cytoplasm and mitochondrial matrix. The model shows expected behavior in its outputs, including the response of ATP production to starting glucose concentration and the induction of oscillations of metabolite concentrations in the glycolytic pathway and in ATP and ADP concentrations. Identification of choke points and parameter sensitivity analysis indicate that the glycolytic pathway, and to a lesser extent the TCA cycle, are critical to the proper behavior of the system, while parameters in other components such as the respiratory chain are less critical. Notably, however, sensitivity analysis identifies the first reactions of nonglycolytic pathways as being important for the behavior of the system. The model is robust to deletion of malic enzyme activity, which is absent in mouse pancreatic β cells. The model represents a step toward the construction of a model with species-specific parameters that can be used to understand mouse models of diabetes and the relationship of these mouse models to the human disease state. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Dissociation of the insulin receptor from caveolae during TNFα-induced insulin resistance and its recovery by D-PDMP

Previously, we demonstrated that an inhibitor of ganglioside biosynthesis, d-PDMP, could restore impaired insulin signaling in tumor necrosis factor α (TNFα)-treated adipocytes by blocking the increase of GM3 ganglioside. Here, we analyzed the interaction between insulin receptor (IR) and GM3 in the plasma membranes using immunoelectron microscopy. In normal adipocytes, most GM3 molecules localized at planar and non-caveolar regions. Approximately 19% of IR molecules were detected in caveolar regions. The relative ratio of IRs associated with caveolae in TNFα-treated adipocytes was decreased to one-fifth of that in normal adipocytes, but this decrease was restored by d-PDMP. Thus, we could obtain direct evidence that insulin resistance is a membrane microdomain disorder caused by aberrant expression of ganglioside.     

Insulin resistance in the liver: Deficiency or excess of insulin?

In insulin-resistant states (obesity, pre-diabetes, and type 2 diabetes), hepatic production of glucose and lipid synthesis are heightened in concert, implying that insulin deficiency and insulin excess coexists in this setting. The fact that insulin may be inadequate or excessive at any one point in differing organs and tissues has many biologic ramifications. In this context the concept of metabolic compartmentalization in the liver is offered herein as one perspective of this paradox. In particular, we focus on the hypothesis that insulin resistance accentuates differences in periportal and perivenous hepatocytes, namely periportal glucose production and perivenous lipid synthesis. Subsequently, excessive production of glucose and accumulation of lipids could be expected in the livers of patients with obesity and insulin resistance. Overall, in this review, we provide our integrative perspective regarding how excessive production of glucose in periportal hepatocytes and accumulation of lipids in perivenous hepatocytes interact in insulin resistant states.     

Review of the fauna of Rotatoria,Cladocera, and Copepoda of the basin of the Anadyr’ River

The zooplankton composition is studied in the thermokarst, glacial and meteorite lakes, channels, former riverbeds, and hollows in the basin of Anadyr’. We found 174 taxa: 78, Rotatoria, 55, Cladocera, and 41, Copepoda. The most diverse is the lake fauna: 51 taxa of Rotatoria, 48, Cladocera, and 37, Copepoda. The thermokarst Lake Maiorskoe hosts 68 taxa: 31, Rotatoria, 14, Cladocera, and 23, Copepoda, wheras the cold ultraoligotrophic Lake El’gygytgyn features only one species of Cyclop of the group scutifer Cyclops neymanae Strel., and Rotatoria and Cladocera are present as allochtonous forms. The Copepoda illustrate the relations of the Anadyr’ fauna with those of Europe, North America, and Japan.     

NAD kinase regulates the size of the NADPH pool and insulin secretion in pancreatic β-cells

NADPH is an important component of the antioxidant defense system and a proposed mediator in glucose-stimulated insulin secretion (GSIS) from pancreatic β-cells. An increase in the NADPH/NADP(+) ratio has been reported to occur within minutes following the rise in glucose concentration in β-cells. However, 30 min following the increase in glucose, the total NADPH pool also increases through a mechanism not yet characterized. NAD kinase (NADK) catalyzes the de novo formation of NADP(+) by phosphorylation of NAD(+). NAD kinases have been shown to be essential for redox regulation, oxidative stress defense, and survival in bacteria and yeast. However, studies on NADK in eukaryotic cells are scarce, and the function of this enzyme has not been described in β-cells. We employed INS-1 832/13 cells, an insulin-secreting rat β-cell line, and isolated rodent islets to investigate the role of NADK in β-cell metabolic pathways. Adenoviral-mediated overexpression of NADK resulted in a two- to threefold increase in the total NADPH pool and NADPH/NADP(+) ratio, suggesting that NADP(+) formed by the NADK-catalyzed reaction is rapidly reduced to NADPH via cytosolic reductases. This increase in the NADPH pool was accompanied by an increase in GSIS in NADK-overexpressing cells. Furthermore, NADK overexpression protected β-cells against oxidative damage by the redox cycling agent menadione and reversed menadione-mediated inhibition of GSIS. Knockdown of NADK via shRNA exerted the opposite effect on all these parameters. These data suggest that NADK kinase regulates intracellular redox and affects insulin secretion and oxidative defense in the β-cell.     

Xenin: the oldest after insulin?

Xenin is a regulatory peptide first isolated from the human gastric mucosa. Using an open-access protein database MEDLINE (33 million molecules; 11 billion amino acid residues) and our original computer program, we conducted a search for the xenin motifs in the primary structure of proteins across almost the entire taxonomic range of evolution. Motifs with 40% homology to human xenin are already present in prokaryotes. Homology reaches 84–96% in single-cell algae and plants, becoming complete since bony fishes. We suppose that this regulatory peptide is more ancient and significant than is usually thought.     

Increased Aβ production prompts the onset of glucose intolerance and insulin resistance

Type 2 diabetes (T2D) mellitus and Alzheimer's disease (AD) are two prevalent diseases with comparable pathophysiological features and genetic predisposition. Patients with AD are more susceptible to develop T2D. However, the molecular mechanism linking AD and T2D remains elusive. In this study, we have generated a new mouse model to test the hypothesis that AD would prompt the onset of T2D in mice. To test our hypothesis, we crossed Alzheimer APPswe/PS1dE9 (APP/PS1) transgenic mice with mice partially deficient in leptin signaling (db/+). Body weight, plasma glucose, and insulin levels were monitored. Phenotypic characterization of glucose metabolism was performed using glucose and insulin tolerance tests. β-Cell mass, islet volume, and islet number were analyzed by histomorphometry. APP/PS1 coexpression in mice with intact leptin receptor signaling did not show any metabolic perturbations in glucose metabolism or insulin sensitivity. In contrast, APP/PS1 coexpression in db/+ mice resulted in nonfasting hyperglycemia, hyperinsulinemia, and hypercholesterolemia without changes in body weight. Conversely, fasting blood glucose and cholesterol levels remained unchanged. Coinciding with altered glucose metabolism, APP/PS1 coexpression in db/+ mice resulted in glucose intolerance, insulin resistance, and impaired insulin signaling. In addition, histomorphometric analysis of pancreata revealed augmented β-cell mass. Taken together, these findings provide experimental evidence to support the notion that aberrant Aβ production might be a mechanistic link underlying the pathology of insulin resistance and T2D in AD.     

Stimulation of the insulin secretory mechanism following barium accumulation in pancreatic β-cells

Electrothermal atomic absorption spectroscopy was employed for measuring barium in β-cell-rich pancreatic islets microdissected from ob/ob-mice. Both the uptake and efflux of barium displayed two distinct phases. There was a 4-fold accumulation of barium into intracellular stores when its extracellular concentration was 0.26 mM. Unlike divalent cations with more extensive intracellular accumulation, the washout of Ba²⁺ was not inhibited by d-glucose. Ba²⁺ served as a substitute for Ca²⁺ both in maintaining the glucose metabolism after removal of extracellular Ca²⁺ and making it possible for glucose to stimulate insulin release. Furthermore, Ba²⁺ elicited insulin release in the absence of glucose and other secretagogues. The latter effect was reversible and was markedly potentiated under conditions known to increase the β-cell content of cyclic AMP. It is likely that the observed actions of Ba²⁺ are mediated by Ca²⁺, since Ca²⁺-dependent regulatory proteins, such as calmodulin, apparently cannot bind Ba²⁺ specifically.     

The effects of insulin,glucose and anti-insulin upon the activity of uridine diphosphate glucose glycogen α-4-glucosyl-transferase EC2.4.1.11 in the rat placenta

Summary The enzyme uridine diphosphate glucose glycogen -4-glyoosyl-transferase EC2.4.1.11 was found to be active in the rat placenta. The total activity of the enzyme, present from the earliest day investigated, day 12, increased significantly between days 14 and 16 and days 18 and 20. The enzyme was observed to be present in both the active and the inactive forms. In in vivo studies of the effects of insulin, glucose and anti-insulin serum it was observed that insulin and glucose produced an increase in the activity of the enzyme while anti-insulin serum inhibited its activity. Insulin was observed to exert its stimulatory effect also in vitro.The activity of the enzyme was observed to be localized strongly in the decidua basalis, the spongy zone, the labyrinth as well as in the yolk sac. There was a shift in the activity of the enzyme from the decidua basalis and the visceral layer of the yolk sac where it was strongest in the younger placentae (14 or 16 days) to the spongy layer where it was stronger towards the end of gestation. The activity of the enzyme was very weak, at 12 days, in all areas of the placenta.